AU5903100A - Vla-4 inhibitor compounds - Google Patents

Vla-4 inhibitor compounds Download PDF

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Publication number
AU5903100A
AU5903100A AU59031/00A AU5903100A AU5903100A AU 5903100 A AU5903100 A AU 5903100A AU 59031/00 A AU59031/00 A AU 59031/00A AU 5903100 A AU5903100 A AU 5903100A AU 5903100 A AU5903100 A AU 5903100A
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group
cooh
chosen
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pct
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AU781438B2 (en
Inventor
John J. Baldwin
Jun Chiba
Shin Iimura
Nobuo Machinaga
Edward Mcdonald
Kevin Joseph Moriarty
Atsushi Nakayama
Christopher Ronald Sarko
Yoshiyuki Yoneda
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Daiichi Pharmaceutical Co Ltd
Pharmacopeia Drug Discovery Inc
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Daiichi Pharmaceutical Co Ltd
Pharmacopeia Drug Discovery Inc
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Assigned to DAIICHI PHARMACEUTICAL CO., LTD., PHARMACOPEIA DRUG DISCOVERY, INC. reassignment DAIICHI PHARMACEUTICAL CO., LTD. Alteration of Name(s) of Applicant(s) under S113 Assignors: DAIICHI PHARMACEUTICAL CO., LTD., PHARMACOPEIA, INC.
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    • C07D207/04—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members
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    • C07D311/02—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings ortho- or peri-condensed with carbocyclic rings or ring systems
    • C07D311/04—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring
    • C07D311/06—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 2
    • C07D311/20—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 2 hydrogenated in the hetero ring
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Description

WO 01/00206 PCT/US00/18079 VLA-4 INHIBITOR COMPOUNDS FIELD OF THE INVENTION The present invention relates to compounds that selectively inhibit the binding of ligands to the adhesion receptor, ( 4 1 integrin, also known as VLA-4. Compounds of the present 5 invention are useful in the treatment and prevention of pathologies associated with VLA-4 mediated cell adhesion, such as inflammatory and autoimmune diseases, and tumor metastasis. BACKGROUND OF THE INVENTION A primary feature of such pathologies as inflammation and autoimmune diseases is the accumulation of activated leukocytes in affected tissues. The process by which leukocytes 10 transmigrate from the circulation at a site of inflammation involves a cascade of interactions that can be divided into four major steps: tethering and rolling, activation, firm adhesion, and transmigration (Springer, T., Ann. Rev. Physiol., 57:827 (1995)). Initially, leukocytes are lightly tethered to the endothelium and roll along its surface. This is followed by cell activation, mediated by soluble chemotactic stimuli, which initiates the development of a firmer bond between 15 individual leukocytes and endothelial cells. The firm bond then results in the successful adhesion and transmigration of the leukocytes through endothelial cell junctions. The steps occur in series and each is essential for transmigration to occur. This also means that transmigration can be modulated at each step, thus providing a number of potential targets for pharmacological inhibition. 20 The receptors involved in leukocyte migration have, to a large extent, been characterized as belonging to particular cell adhesion molecule families (Carlos and Harlan, Blood, 84:2068 (1994)). The initial attachment and rolling step is mediated by a family of adhesion receptors referred to as selectins. Firm adhesion is mediated by interaction of leukocyte surface integrins with molecules of the immunoglobulin superfamily expressed on the surface of the endothelium. 25 Both integrins and the immunoglobulin-type adhesion molecules are also primarily involved in leukocyte transmigration. After transmigration, the leukocytes rely on integrins to traverse through the extracellular matrix and remain at the site of inflammation. Integrins are a large family of heterodimeric glycoproteins composed of two noncovalently associated subunits, a and P (Hynes, R., Cell, 69:11 (1992)). There are at least 16 30 different a subunits (a 1 -a 9 , (XL, LM, OLD, X,, E o1b, ot) and at least 9 different P3 (31-[9) 1 WO 01/00206 PCT/US00/18079 subunits. Integrins are divided into sub-families, based upon the 13 subunit. Leukocytes express a number of different integrins, including a4I, s , a 6 1, a40 7 , aLP 2, aX2, and avCP 3 . 4 3 1 integrin, also known as very late antigen-4 (VLA-4) or CD49d/CD29, is expressed on monocytes, lymphocytes, eosinophils, and basophils, all of which are key effector cells in 5 various inflammatory disorders (Helmer, M., Ann. Rev. Inunmmunol., 8:365 (1990)). a43 1 integrin serves as a receptor for vascular cell adhesion molecule-1 (VCAM-1), as well as to the extracellular protein fibronectin (FN) (Elices et al., Cell, 60:577 (1990)). Anti-inflammatory effects and delayed disease progression have been demonstrated after in vivo monoclonal antibody blockade of the a 4 PI/VCAM-1 pathway (Lobb et al., J. Clin. Invest., 94:1722-28 (1994)). In a 10 guinea pig model of pulmonary inflammation, anti-a 4 inhibited both antigen-induced bronchial hyperreactivity and leukocyte recruitment in bronchoalveolar lavage fluid (Pretolani et al., J. Ex. Med., 180:795 (1994)). Antibodies to a 4 or VCAM-1, prevented antigen-induced eosinophil infiltration of the mouse trachea (Nakajima et al., J. Exp. Med., 179:1145 (1994)). a 4 or VCAM 1 monoclonal antibody treatment also delayed or prevented cutaneous delayed hypersensitivity 15 response in mice and monkeys (Chisholm et al., Eur. J. Immunol., 23:682 (1993); Silber et al., J. Clin. Invest., 93:1554 (1993); cardiac allograft rejection in mice, accompanied by specific immunosuppression (Isobe et al., J. Immunol., 153:5810 (1994); graft-versus-host disease in mice after bone marrow transfer (Yang et al., Proc. Natl. Acad. Sci. USA, 90:10494, (1993); and experimental autoimmune encephalomyelitis in rats and mice (Yednock et al., Nature, 356:63 20 (1992); Baron et al., J. Exp. Med., 177:57 (1993)). Rational drug design studies have produced soluble VCAM-Ig fusion protein containing the two N-terminal domains of human VCAM-1 fused to a human IgG1 constant region. In vivo administration of the fusion protein significantly delays the onset of adoptively transferred autoimmune diabetes in nonobese diabetic mice (Jakubowski et al., J. Immunol. 155:938 (1995)). 25 Another approach has used three-dimensional crystallographic structures of VCAM-1 fragments to synthesize cyclic peptide antagonists that closely mimicked the a4 integrin binding loop in domain 1 of VCAM-1. Synthetic VCAM-1 peptide CQIDSPC, was able to inhibit the adhesion of VLA-4 expressing cells to purified VCAM-1 (Wang et al., Proc. Natl. Acad. Sci. USA, 92:5714 (1995)). An additional strategy is to block the binding of a 4 1 to its other counter receptor, that is, 30 an alternatively spliced region of fibronectin containing the connecting segment-1 (CS-1) motif 2 WO 01/00206 PCT/US00/18079 (E.A. Wayner, J. Cell. Biol., 116:489 (1992)). A synthetic CS-1 tetrapeptide (phenylacetic acid Leu-Asp-Phe-d-Pro-amide) inhibited VLA-4-mediated lymphocyte adherence in vitro and reduced accelerated coronary arteriopathy in rabbit cardiac allografts (Molossi et al., J. Clin. Invest., 95:2601 (1995)). Each of these studies provide evidence that selective inhibition of G 4 1/VCAM-1 5 mediated adhesion is a proven strategy in the treatment of autoimmune and allergic inflammatory diseases. Moreover, while United States Patent 5,821,231 and PCT Applications WO 96/22966, WO 97/03094, WO 98/04247 and WO 98/04913 describe compounds exhibiting VLA-4 inhibitory activity in in vitro binding assays, none of the described compounds have exhibited efficacy in oral 10 administration. Accordingly, despite these advances, there remains a need for small, non-peptidic, specific inhibitors of VLA-4 dependent cell adhesion that are orally bioavailable and that are suitable for the long-term treatment of chronic inflammatory diseases and other pathologies associated with leukocyte migration and adhesion. 15 SUMMARY OF THE INVENTION The compounds of the present invention selectively inhibit the binding of ligands to 4 3 1 and therefore, are useful for inhibition, prevention and suppression of VLA-4-mediated cell adhesion and the pathologies associated with that adhesion, such as, for example, inflammation, asthma, arthritis, diabetes, autoimmune responses, multiple sclerosis, psoriasis, transplantation 20 rejection, and tumor metastasis. In one embodiment, the present invention provides a compound represented by Formula I, or a salt thereof, w W1 W N N R M H H wherein 25 W is chosen from aryl group, substituted aryl group, heteroaryl group and substituted heteroaryl group; W is chosen from arylene group, substituted arylene group, heteroarylene group and substituted heteroarylene group; A is chosen from =0O, =S and =NH; 3 WO 01/00206 PCT/US00/18079 R is chosen from a direct bond, alkyenylene group and -(CH 2 )n-, wherein n is chosen from 1 and 2; X is chosen from -C(O)-, -CH 2 - and S(O) 2 ; 5 M is chosen from Q R 1$31-AV 5 wherein Q N is a divalent 4-, 5-, 6- or 7-membered heterocyclic moiety, wherein the nitrogen atom is the point of attachment to X; 10 R1 ,
R
2 and R 3 are independently chosen from -H, -OH,-NH 2 , halogen atom, alkyl group, substituted alkyl group, aryl group, substituted aryl group, alkoxy group, substituted alkoxy group, monoalkylamino group, substituted monoalkylamino group, dialkylamino group, substituted dialkylamino group, cycloalkylamino group, substituted cycloalkylamino group, 15 alkylsulfonylamino group, substituted alkylsulfonylamino group, arylsulfonylamino group, substituted arylsulfonylamino group, aryloxy group, substituted aryloxy group, heteroaryloxy group, substituted heteroaryloxy group, benzyloxy group and substituted benzyloxy group, or 20 two of R 1 , R 2 and R 3 taken together may form a 3-, 4-, 5-, 6-, or 7-membered carbocyclic or heterocyclic residues optionally substituted with from 1 to 3 substituents chosen independently from -OH, halogen atom, -NH 2 , alkyl group, alkoxy group, aryl group, aryloxy group, alkylamino group, benzyloxy group and heteroaryl group; 25 R 4 is chosen from -H and lower alkyl group; Y is a direct bond or a divalent radical chosen from -C(O)-, -C(O)NH-, alkenylene group, alkynylene group and -(CH2)kY 2 , wherein k is chosen from 1, 2 and 3; and 30 Y2 is a direct bond or a divalent radical chosen from -0-, -S-, -S(O), -S(O) 2 - and -NY 3 -, 4 WO 01/00206 PCT/US00/18079 wherein Y 3 is chosen from -H and lower alkyl group; Z is chosen from arylene group, substituted arylene group, heterocyclylene group, substituted 5 heterocyclylene group, cycloalkylene group and substituted cycloalkylene group; Al is a direct bond or a divalent radical chosen from alkenylene group, alkynylene group, -(CH2) t - and -O(CH 2 ), wherein 10 t is chosen from 1, 2 and 3; and v is chosen from 0, 1, 2, and 3; and H H KN N
R
5 is chosen from -OH, lower alkoxy group, -N(H)OH, N-N" and H H KN N
N-N
15 wherein is a divalent 4-, 5-, 6- or 7--membered heterocyclic moiety, wherein the nitrogen atom is the point of attachment to X;
R
6 and R 7 are independently chosen from -H, -OH, halogen atom, alkyl group and alkoxy group; 20 Yl is a divalent radical chosen from -0-, -S-, -S(O)-, -S(O) 2 - and -NY 4 -, wherein
Y
4 is chosen from -H and lower alkyl group; Zi is a divalent radical chosen from arylene group, substituted arylene group, heterocyclylene group, substituted heterocyclylene group, 25 cycloalkylene group and substituted cycloalkylene group; A2 is a direct bond or a divalent radical chosen from alkenylene group, 5 WO 01/00206 PCT/US00/18079 alkynylene group and -(CH 2 )e wherein e is chosen from 1, 2 and 3; and H H 8 NKN N R is chosen from -OH, lower alkoxy group, -N(H)OH, N - and H H N N) 5 N-N 1 (CHR 9 )qRio wherein L is O wherein 10 is a divalent 4-, 5-, 6- or 7-membered heterocyclic moiety, optionally substituted with from 1 to 3 substitutents chosen independently from alkyl group, alkoxy group, hydroxyalkyl group, -OH, benzyloxy group, -NH 2 , halogen atom, aryl group and heteroaryl group, said moiety may be 15 fused to 1 or 2 additional carbocyclic or heterocyclic residues optionally substituted with from I to 3 substitutents chosen independently from alkyl group, aryloxy group, alkoxy group, hydroxyalkyl group, -OH, benzyloxy group, -NH 2 , , halogen atom, aryl group and heteroaryl group; 20 m and q are independently chosen from 0, 1, 2 and 3; Xi is chosen from -CH= and -N=;
R
9 is chosen from -H and lower alkyl group; H H
R
I0 is chosen from -COOH, lower alkoxycarbonyl group, N H H H N an N NN N and N4 ; and 6 WO 01/00206 PCT/US00/18079
Z
2 is chosen from -H, COOH and lower alkoxycarbonyl group; and -R 1 -Z3-Q 2
-L
1 wherein
R
11 is chosen from -0-, \ - - and -NR 2 5 wherein
R
12 is chosen from -H, alkyl group, substituted alkyl group, cycloalkyl group, substituted cycloalkyl group, aryl group, substituted aryl group, benzyl group, substituted benzyl group, lower alkenyl group, substituted lower alkenyl group and lower 10 alkynyl group the left hand bond is the point of attachment to -X- and the right hand bond is the point of attachment to -Z ;
Z
3 is chosen from a direct bond, a divalent aliphatic hydrocarbon moiety having 1 to 12 carbon atoms, 15 wherein one or more carbon atoms may be replaced with -0- or -NR 13 wherein
R
13 is chosen from -H and lower alkyl group, and 20 one or more hydrogen atoms attached to an aliphatic carbon atom may be replaced with lower alkyl group; and wherein x is chosen from 0 and 1; 25 y is chosen from 1, 2, and 3; and
R
14 is chosen from -H, -OH and halogen atom, 0, , and, when 7 WO 01/00206 PCT/US00/18079
R"
I is -NR12, I-ZI wherein 1 -z -I wherein 14a 5 Z 4 is chosen from wherein
R
14 a is chosen from -H, -OH, lower alkyl group and halogen atom; 10 and , wherein the left hand bond is the point of attachment to R" 1 and the right hand bond is the point of attachment to Q2 Q2 is a divalent radical chosen from arylene group, substituted arylene group, heterocyclylene group, substituted heterocyclylene group, 15 cycloalkylene group, substituted cycloalkylene group, 15 wherein R" and R 16 are independently chosen from -H, halogen atom and lower alkyl group; and
R
17
R
8 wherein R 1 7 and R 18 are independently chosen from -H, lower alkyl group, substituted lower alkyl 20 group and lower alkenyl group; and LI is chosen from -COOH and -COOR' 9 wherein
R
19 is a lower alkyl group. 8 WO 01/00206 PCT/US00/18079 R 2 R3 R In a preferred embodiment of Formula I, M is . Y-Z-A R In this embodiment, more preferred compounds are those wherein A is =0O, R is -(CH2) n - and X is -C(O)-. Y is preferably chosen from alkenylene group, alkynylene group, -(CH 2 )kY 2 , -CH 2 S(O)- and
-CH
2 0-, and more preferably, Y is -CH 2 0-. 5 Preferred compounds of this embodiment are those wherein W is unsubstituted phenyl group or phenyl group having one or two substituents chosen from lower alkyl group and halogen atom at the ortho positions thereof. W' is preferably unsubstituted phenylene group or phenylene group having a substituent chosen from methoxy group, lower alkyl group and halogen atom at the ortho position to -NH-. 10 In preferred compounds of this embodiment, A is preferably =0 and A' is a direct bond or
-(CH
2 )t-. More preferred compounds are those wherein Al is a direct bond and R 5 is -OH. R 4 R $ \LQ 4R Preferred compounds of Formula I, wherein M is N and A is =0O are represented in Table 1. With respect to the representation of -W 1 , the lower bond is the point of attachment to -NH- and the upper bond is the point of attachment to -R-. The entry entitled 15 -- R---R s depicts that portion of the particular compound represented by 3 - R 4 R - N Y-Z-A
R
5 9 WO 01/00206 PCT/US00/18079 TABLE: MassT spectrum : W- :W : ~ ~ ~ ~ ~ ~ ~ ~ ~ ~ ~ ~. .:":- ":I.;::: . .::' ..... ::-.: /.:::.. ... :-...-........ 5 502.6
H
3 C CO2H 516.7 507.0 o HC O CO 2 H
H
3 16 0 lo 486.6 7coH
H
3 C0 2 H 553.6 - HN-N lOl 508.7
H
3 C N 0C0 2 H 507.0 0-6 N 0 C0 2 H 10 501.6
H
3 C N N H 486.6 C0 2 H
H
3 553.6 0 HN-N
H
3 C N N 'N 10 WO 01/00206 PCT/US00/18079 TABLE 1 .(M .
..,...... •: Mass j_________. W- j 486.6 H OC2
H
3 IN 579.7
H
3 C- I %] H3H H
H
3
C
O O . N N 0 N- N 552.7 0 HN
H
3 I N N 496.6 H3C
CO
2 H 5 500.6 HC~ C0 2 H
H
3 C 0 512.6 o
H
3 C IN > 0. C0 2 H 527.7 H3HCO OCO2
H
3 C 11 I H 3 COX 4 C0 2
H
WO 01/00206 PCT/US00/18079 I. TABLE:1 Mass ectum " V -W -R ::::,L ~ ~ ~~~~ ~~~ .... .. .. .. ...... .. ... .. .... ... 484 N CO 2 H 486 l?-Y 0
CO
2 H 488 O O 504CO2H O 5 518 0 2 H 0 /00 50 0- N \ C2 500 CO 2 H Cl 498 /O \ CO 2 H 12 WO 01/00206 PCT/US00/18079 TABLE 1 Mass Spectrum W- :-W ( M + 1) . .... .. 512 N 526 O
CO
2 H 532 O CO
HO
2 C 514 O0
CO
2 H 5 500N
CO
2 H 516 < N 02C O 0 CO2H 13 WO 01/00206 PCT/US00/18079 TABLE 1: [Mass- .. . Spectrum .... ...- R 566 0O C02 KN 566 0K 0 0 CO 2 H 0 C0 2 H 0 -O-QCO 2 H 70 Co 2 H 548 V- _N\'- 2H 0 562 0 C0 2 H 552 C0 2 H A0 c 14 WO 01/00206 PCT/US00/18079 Mass TABLE 1 Spectrum : I .... . .. .. 586 A c 563 Yl 0 0 \ C0 2 H 0 2 N F0 /-- \ /CO 2 H
H
2 C 0 C0 2 H 5625 WO 01/00206 PCT/US00/18079 [.TABLE:L ... .. ........... .. ......... ~. ..... ... ..... ... ' ?. .. y. .: .. . .:. .: . ' . Mass: pectrum w -M a s: ::: .::.:"... .. .:... .. I : .... : :/::. . ::: '. .. . .:" :::: ' "".. .:: " 561 x- ~ 00 O . CO 2 H N 536 o F 552 \YN- OMe F O OCO 2 H 519CO2H O 5 654 OBn 0 0 0
CO
2 H 577 VN. N 577 CO 2 H
NO
2 564 PH /oV Q N O- CO 2 H 549 OH <N NH 2 M C0 2 H 16 WO 01/00206 PCT/US00/18079 . Mass: Sp5ectruk -R 566F 0 / - CO 2 H 575 ,ryNQ~ \ / CO 2 H 110 HN 0 0 \ C0 2 H N 523N /0 0 2 H F 1 5 524N 0 Nv_/ - O 517 /~ KKXOH F Me 521 17 WO 01/00206 PCT/US00/18079 M ss..I 580 Me 0 2 550 <NNQMe F
H
2 N Ol / NNHOH 582 NR _ Br / H 1~0 5 553COOH cI OMe 0 09
NH
2 674 CI OMe 0 ~ \/O-,COOH -0 598 BrOMe0 0
NH
2 ____ _ _ ___ ___ ___ ___ ___COOH 18 WO 01/00206 PCT/US00/18079 ..... . - :.TABLE 1 Mass Sp -ec- : , 626 Br OMe
NH
2 COOH 5 599 O O 5 4O O COOH Br OMe OO
NH
2 554 582 H /r \" O O 535eO COOH OMe O O_0 N ONH 2 C O O H 5376 cO Me 582 Br Me q 535 O COOH OH OMe0
NH
2 10 639 -- ~ 9-11 OMe ,COOH
H
2 646 -- ~ -~ N F 'l0 -- OOH 19 WO 01/00206 PCT/US00/18079 TABLE 1 "" . .........
: M ass pec . : . . .. ... ... .. .. ...... ' ' -' : : : r .. ','. ::, .:.: : : , . " " " ' '. ' '.. ' , , . '. ' -v R : . -- ] .. ,,.. .. . .. : .. .: : . .: . .: (M'+1) J W W- - -R 629 \-OMe NPBn / N OMe COOH 538 O COOH 583 BO o COOH 5 536 F 36I OMe N _ 5 _COOH 556 ,F 518 OMe - OOH 601 F / Br OMe N O -O -COOH 20 WO 01/00206 PCT/US00/18079 TABLE 1 Mass Spec1 M + - 1) . J -- '. ." 516 OO 54YN _aCOOH 542 F/\ \\ ' % OMe OOH 5 561 L Me 0 \-- OOH 556 OMe O O OOH 556 [F OMe OOH 5 561 M e NI 556 l ry -O-( -COOH 556 f2 Ci OMe 0 j f .CO 561 -N Me 21 WO 01/00206 PCT/US00/18079 TABLE 1 4MassSpec .JR. ' .:W-. -w .- . .. . R R .- :. .. .. 572 l CI Me \N
-
OOH 652 OMe F ON~ 5N O COOH 555 F cI OMe HOO \ OOH 672 O I F F OMe COOH 5 487 COOH 600 COOH 600 F Br Me NH 5No-- COOH 536 Me N OCOOH 554 - 9H Me N 022 /COOH 22 WO 01/00206 PCT/US00/18079 TABLE 10 :Mass Spec . (M,+ 1) : -- R , " : / : : 534 9H OM COOH 5 502 OMe \/COOH 574 OF / "F OMe OOH 580 F cI OMe \ Nio COOH 581 10 547 N/ \ H 548 OMMe o o'% ooH OMe pMe 0 OOH 548 -QMe OMe 0 OCH 552 - N oMe Me N3_ 23 WO 01/00206 PCT/US00/18079 TABLE 1 Mass Spec 1 -W -- R---R 539 O CI Me / OOH 584I Br Me / OOH 523 OMe 0N 568 N Me CI Me \ O OOH 5 613 NOMe r/Me \-y NQ O OOH 602 Br OMe N O\/COOH 592 CI OMe ,INO-QaCOOH 637 Br OM e x - 4- C O 24 WO 01/00206 PCT/US00/18079 TABLE. Mass Spec 1 .- . W - -w "- Q ... -- R---Rs 717 F Br C 0 651 / \K~ \.F Br MeF N vCOOH 651 F 611/ K OMe OF N COOH 716 F Br OMe F N "COOH 671 F Me F N COOH 5 524 C / K -<&N -OO 544 C OMe COOH 506 F N / COOH 25 WO 01/00206 PCT/US00/18079 TABLE I. Mass S pec. . 537 C OMe N O COOH 5 648I OMe 0 / COCH 581 \ CI OMe O N COOH 589 C S1COOH OMe XCOOH 537 557 0 H Me XN , H&COOH 10 616 0-0 OMe N .,COOH 63626 CI OMe N, 26 WO 01/00206 PCT/US00/18079 TABLE 1 Mass Spec -. . (M + 1) .
R. 681 Br OMe N O e-COOH 522 F OMe /COOH 590 OMe \/ N COOH 624 OMe O /COOH 5 5 3 4 C O 494 H COOH 5 50 NO)H Me COOH 570 QH Me N C S/ COOH 624 COOH -&COOH 27 WO 01/00206 PCT/US00/18079 TABLE 1 Mass Spec[ R ... . .. ..- - . 674 C / COCH Cl OMe -&/COOH 661 N OMe NCOOH N 0-01 654 COOH 66 / \K \ OMe 2COOH 670 OMe N COOH O 5 680 CI Me COOH N 636 / COOH 666 P/ K OMe O 0/COOH 28 WO 01/00206 PCT/US00/18079 TABLE 1 520 F OMe N _< /-C O 5 540 N O\/COOH 598 CI Me 0O\/ COOH 643 CI Me N \/COOH 585 NF Br -yN o 0O\: /COOH 601 N r~~ OMe 0& o 10 719 - ~e a -&C-COOH N OOH BrOMe \yN 0 OOH 29 WO 01/00206 PCT/US00/18079 .,~ . . . ., .. . .... TABL I... TABLE 1. - . .- 77 . . ., Mass Spec].. .+ _W -- R --- R + 1 686 OMe o OOH 731 573 N C/ OMeO H / H 617 \ Cl Br N O 5 492 \F OMe OO Br OMe F OOH 552 F O M e M e O 30 WO 01/00206 PCT/US00/18079 TAB LE I I MassS J J ~R -- R:. 743 -- N cI OMeH l 0~ OOH 548 NOMe OMe ryI0 0 568 -NOMe cI MeN0 613 . OMe rOMeN0 5 506 -~IF 780 r NOMe HN 0 \ OOH 831 N BrMe HN 11 0 \~OOH 31 WO 01/00206 PCT/US00/18079 TABLE: 1 Mas Spec. -W- : -R (M.R+ : 1).: 676 \ H HNiI Br OMe O: OOH 0N 64~4OOH 5 524 /F /H 644 - .OBn 690 OOH 689 --OBn CI OMe 0 O /
C
)H 0 OOH 681 -~N, 59IC F /N 10 726 N ,.. r Me 0 qo OOH 590 r- \ F cI OMe 32 WO 01/00206 PCT/US00/18079 'TABLE I.. W -. 'W - . -- Mas, Spec.. J-W ". J -- R -- R.. .(M + 1.. ) .-., w -. -. .. " . ." ... 594 OMe 0 - OOH 614 OMe O OOH 659 Br OMe O OOH 534 O OOH MeO OOH Br 5 554 OH IMe 0 OOH 599 Ni OH / K \ \ Br Me OOH 571 -F Br NJ0 648 -CN ci e0 OOH 33 WO 01/00206 PCT/US00/18079 I______TABLEA 1 Mass Jpe .. ... .... J -R - 643 Br OMe 0~ OOH 532 OMe N)~ It 552 -~OMe 597 -~OMe Nt 5 602 .- C OMe OMe -/ 562 - Me OMe N 550 .- IKF Me NJ0== / 532 K OMe 0 \OOH 552I Me 0 ~ OOH 34 WO 01/00206 PCT/US00/18079 TABLE 1 M assS c: " . .: ' J 528 O CI 0 OOH 5 573 OO o [OOH 578 0 OOH 516 OOH -Z 534 N F O O N-/O H 520 F 0 0 10 536 O 0 \ OOH 546 N F 0 o-(:-<H 540 F Cl OONj H 0 35 WO 01/00206 PCT/US00/18079 TABLE 1 M :ass SpIec-.-R -- R 560 F 605 -NF 562 N 0o 0-OOH 582 \o0 OOH 5 627 /r N o1 O OOH 508 -1 1 /0 OOH 558 o OOH 522 . CI0 OOH 522 I NF Br / ~-&-COOH 36 WO 01/00206 PCT/US00/18079 TABLE 1 Mass Spec _ _ _ --- -R. 526 CI \ F C OMeO 0 l--'
O
H 591 \ F OMe O0 600 513 F Me O Me O O H 0H 5 605 \ F CI OMe N<O H 0 650 \ F 617 O K 544 \F 37 WO 01/00206 PCT/US00/18079 TABLE.1 MassSec "1 -Wi- I S -- R -.. R: 589 F /' K Br O 5 526 F Cl OH 632 F cI OMe 0 OOH 616~ N \ YN _ 0 OOH 616 F Cl 0 OOH 574 F
CF
3 \ O 0 585 Br F 10 524 F F O H 0 541 F 0 H 38 WO 01/00206 PCT/US00/18079 l:.i :-.::. ": " E.::. , " : :..... .. ." " " , ..... .TABLE. I . :Mass Spec ". W -W -- R - R 542 F NJ -,//o N O O 0 0- F' O 542 H 607N 607 F F Br ,& ,NjII L F0 571 O H 5 6 1 9 N F Br VO 560 NIF Cl 0 -0" H 0CI N\H 39 WO 01/00206 PCT/US00/18079 R Z'-A2 R In another preferred embodiment of Formula I, M is In this embodiment, more preferred compounds are those wherein A is =0, R is -(CH2)
.
- and X is -C(0)-. Y is preferably chosen from -O-,-S-, -S(O)-, -S(0) 2 - and -NY 4 ,and more preferably, is -0-. Preferred compounds of this embodiment are those wherein W is unsubstituted phenyl 5 group or phenyl group having one or two substituents chosen from lower alkyl group and halogen atom at the ortho positions thereof W l is preferably unsubstituted phenylene group or phenylene group having a substituent chosen from methoxy group, lower alkyl group and halogen atom at the ortho position to -NH-. In this embodiment of Formula I, A is preferably =0 and A 2 is a direct bond or 10 -(CH 2 )e-. More preferred compounds are those wherein A 2 is a direct bond and R is -OH. R Q Z1-A2 R8 Preferred compounds of Formula I, wherein M is and A is =0, are represented in Table 2. With respect to the representation of -W 1 , the lower bond is the point of attachment to -NH- and the upper bond is the point of attachment to -R-. The entry entitled -R---R 8 depicts that portion of the particular compound represented by RR RZ-A2&R8 15 40 WO 01/00206 PCT/US00/18079 TABLE2 M i. s Sp .ec .. ... ..... MaWs-ec-Wi-R 534 OH CH3 OMe O COOH 0 5 504 P-C COOH H3 OMe 524 P COOH c, OMe 569 P COOH Br OMe ( (CHR 9
),R
10 In another preferred embodiment of Formula I, M is L N Z2 . In this embodiment, preferred compounds are those wherein A is =O, R 10 is -CO 2 H and q is 0 or 1. More 10 preferred are compounds wherein R 1 0 is -CO 2 H, q is 0 or 1, most preferably 0, and m is 2. When A is =0O, L is preferably chosen from H / N HO,,N 0 N % ^0 O - 0 ^AN O -0 HO OH H 2 N O O NN N N N 0, -e" 0 O O . 0 ,0 41 WO 01/00206 PCT/US00/18079 PhO Ph S 0 0 O0 0 0 p-MeOPh Oh BOno 0 N N N N -0 0 -0 0
H
2 N HN N N 0 O 0 and O HO, NNN I Y Y More preferably, L is chosen from O 1 O O ,p-CIPh HO OH 0 PhO N N N N 5 - 0 -0 0 1 0 P p-MeOPh Ph 0 BnO S 2 0 O 0 H 2 N N N N N - 0 W0 .W0 and- O 42 WO 01/00206 PCT/US00/18079 HO, N N N Most preferably, L is chosen from 1 0 1 0 0 ,p-CIPh HO OH H 2 N N N N N - 0 - 0 - 0 - 0 ,p-MeOPh PhO Ph O S L,,-oO -.- o oI o . K-" 0 0 0 0 -0 N N 0 and ~ O 5 Preferred compounds of Formula I, wherein R is -CH 2 -and X is =0, are those wherein W is unsubstituted phenyl group or phenyl group having one or two substituents chosen from lower alkyl group and halogen atom at the ortho positions thereof W' is preferably unsubstituted phenylene group or phenylene group having a substituent chosen from methoxy group, lower alkyl group and halogen atom at the ortho position to -NH-. (- 1,(CHR 9)qRio 10 Preferred compounds of Formula I, wherein M is "L* - 'Z2 , A is =0, R is -CH 2 -and X is =0O, are represented in Table 3. With respect to the representation of-W 1 , the lower bond is the point of attachment to -NH- and the upper bond is the point of attachment to -R-. The entry entitled -N---Z 2 depicts that portion of the particular compound represented by (M 1,(CHR 9 )qR O 0 43 WO 01/00206 PCT/US00/18079 TABLE 3 [Mass Spec W- -W- -.. L. .- N Z 5 554.7 HO,, CO2H
H
3 C / N
HH
3 CO N O N 493.6
CO
2 H
H
3 N I1 N 540.6 HO,, CO2H N. ~ N ()~~ y OC . N)
H
3 CO N O N --I 523.7
CO
2 H
H
3 N '
H
3 CO 555.7 CO 2 H
H
3 C N N 1 I 10 583.8 CO 2 H
H
3 N I -I N 44 WO 01/00206 PCT/US00/18079 TABLE 3> SM ass. ...... .... .. ..... 524.7 HO, CO 2 H H3C 3 N I 509.7 HO, CO 2 H
H
3 CO O I 538.7 HO, CO 2 H
H
3 '3
-~H
3 C 0N OON 554.7 HO, Co2H H 3 , N N )
HH
3 CO 0 N O I 5 569.7 CO 2 H
H
3 C ON ~I 666.9 H H
H
3 N I N-N 45 O N 45 WO 01/00206 PCT/US00/18079 Spe ... . 2W-. 567.8 C0 2 H [H3 N (.-CO 2 H
H
3 00 O 0 N 570.4 HCHO OH rC2 H3CC l N 552.7 H 2 N CO 2 H H31 1 1 N . H 3 CO 0 N 619.5 HO,, KC 2 H BrN Y N)
-~H
3 CO 0~. KN 5 575.1 HO,' KC0 2 H CI N ~- H 3 CO N X 0N 553.6 H30 C0 2 H
H
3 C 4 .. ?.
H
3 00 NN 664.1 ,p-CIPh CQ 2 H
H
3 0~
SH
3 004 N N 46 WO 01/00206 PCT/US00/18079 TABLE:3 [Mass W- -WI- -L 629.7 HCPhO CO 2 H H3CC j N 555.6 C0 2 H
H
3 C I -~ N ~Lo N 613.7 PhCO 2 H
H
3 I 3C N
H
3 00 0 555.7 S C0 2 H
H
3 C N 7-- H 3 CO4 0% N 5 585.7 C2
H
3 C I N
-~H
3 CO4~ 0N 591.7 CO 2 H
H
3 0 N 660.7 ,pMeOPh CO 2 H
H
3 C 0 .7 H 3 CO NN --1 N 47 WO 01/00206 PCT/US00/18079 TABLE::3. 3 [Mass. Spec. . -W I- .2. . ., _(_ 1) 555.6 HO,,, CO2H
H
2 N N H2H 3 CO N N O 556.6 HO, CO2H HO H3ON OH 3 CO N 570.7 CH30HO CO 2 H
CH
3 0 H3COK ON NI
H
3 CO 0L INI 570.7 H HO,, CO 2 H
H
3 CO N N 5 673.8 BnO C0 2 H
H
3
CO
2 H
H
3 CO N N I4I 48 WO 01/00206 PCT/US00/18079 TABLE 3 3 Mass: .W ~. L.....N- 552.7 -~- 'CO 2 H
H
3 C ~NN .' H 3 CO 0 N 668.8 Bnlt HN -N N'
H
3 CO N N 583.7 HO--HO OH C0 2 H
H
3 N
.H
3 CO4 '1. 704.9 K3CC02H ~- H 3 CO KN) N N -0 514.6 ~ -- ~C0 2 H
H
3 C N IN S js 0 7' N 49 WO 01/00206 PCT/US00/18079 I. TABLE 3.: Mass: Spec W- W -L- -N---Z 2 j 562.6 -. -CO 2 H
H
3 C N NS N o Yet another preferred embodiment of Formula I includes compounds wherein M is
-R
11
-Z--Q
2 -L1. Preferably, A is =0O, R is -CH 2 - and X is =0O. Preferably W is unsubstituted phenyl group or phenyl group having one or two substituents chosen from lower alkyl group and 5 halogen atom at the ortho positions thereof. W 1 is preferably unsubstituted phenylene group or phenylene group having a substituent chosen from methoxy group, lower alkyl group and halogen atom at the ortho position to -NH-. In compounds wherein Q 2 is R R and Z 3 is a divalent aliphatic hydrocarbon moiety, 121 preferred compounds are those wherein R" is or -NRI , more preferably NR2 , wherein 10 R2 is chosen from - H, lower alkyl group and substituted lower alkyl group, most preferably dihydroxy lower alkyl group. Preferred choices for Z 3 is a divalent aliphatic hydrocarbon moiety having 4, 5 or 6 carbon atoms. A preferred choice for W' is phenylene group having a substituent chosen from methoxy group, lower alkyl group and halogen atom at the ortho position to -NH-. In compounds wherein Q 2 is R1R' 8 and Z 3 is R 14 , R is preferably 15 -NR 2-. In these compounds, x and y are preferably 1. Preferred choices for R 14 include -H. -OH and -F. A preferred choice for W1 is phenylene group having a substituent chosen from methoxy group, lower alkyl group and halogen atom at the ortho position to -NH-. 50 WO 01/00206 PCT/US00/18079 In compounds wherein Q 2 is R 17 R 8 and Z 3 is o , R" 11 is preferably chosen from -0- and -NR 1 2 , preferably wherein R 12 is chosen from -H and lower alkyl group. Preferably, R 17 and R 8 are each - H. A preferred choice for W 1 is phenylene group having a substituent chosen from methoxy group, lower alkyl group and halogen atom at the ortho position 5 to -NH-. In compounds wherein Q 2 is R 17 R'18 and Z 3 is , R" is preferably
-NR
12
-
, wherein R 12 is preferably lower alkyl group. Preferred compounds of this embodiment also include those wherein at least one of R 17 and R 18 is lower alkyl group or substituted lower alkyl group. 10 In compounds wherein Q 2 is "R 18 and Z is , R is preferably -NH- and R 17 and R 18 are each preferably -H. A preferred choice for W1 is phenylene group having a substituent chosen from methoxy group, lower alkyl group and halogen atom at the ortho position to -NH-. 15 In compounds wherein Q 2 is chosen from aryl group, substituted aryl group and R , and more preferably from phenyl group and phenyl group substituted at the point of attachment to Z ,Z 3 is preferably a divalent aliphatic hydrocarbon moiety. Yet another embodiment of the invention is a compound represented by Formula II, O O N N^\ R Z3\ L 3 H H II 51 WO 01/00206 PCT/US00/18079 wherein the substituents W, W', R 1 " and Z 3 are defined as in Formula I, and L 3 is chosen from O :Q4-,COOR20, wherein R20 is preferably chosen from -H and lower alkyl, CH3, J6and & Still another embodiment of the invention is a compound represented by Formula EI, 0 0 W ) N W1 R1 d (CH 2 ) 5 H H m wherein the substituents W, W 1 , and R" 1 are defined as in Formula I, and d is chosen from 0 and 1, and f is chosen from 1 and 2. Preferred compounds of Formula I, wherein M is 11
ZL
2 - 1 , A is =0O, R is
-CH
2 -and X is =0, are represented in Table 4. With respect to the representation of -W 1 , the 10 lower bond is the point of attachment to -NH- and the upper bond is the point of attachment to -R-. The entry entitled --R 11 ---L' depicts that portion of the particular compound represented by 11 -Z3-Q 2
-L
1 52 WO 01/00206 PCT/US00/18079 TABLE 4 M ass s[ec W W -. R 466.22
H'
2 H
CH
3 O O 5 496.22 H0-.C 2 H CH3 OCH3
CH
3 OCH 3 445.19 H 2H
CH
3 475.20 N-HN '--CO 2 H I K
CH
3 OCH 3 497.22 \
CO
2 H
CH
3 OCH 3 0 510.42 \
CO
2 H
CH
3 OCH 3 0 10 517.62 N-)D
\CO
2 H
CH
3
OCH
3 473.56 CO 2 H 504.1 HN CO2H CH3
OCH
3 529.17 HC H CH3 OCH 3 N 'CO 2 H 53 WO 01/00206 PCT/US00/18079 TABLE.4 Mag§Spec:j~ -I 501.16 HO j
CH
3 OCH 3 ~ ~ C 2 427.20
CH
3 OCH 3 441.22 WK I NN C 2 H
CH
3 OCH 3 468.1 P NC0 2 H
CH
3 OCH 3 5 532.2 ' PvN C 0 2 H
CH
3 OCH 3 624.2 1 0
CH
3 OCH 3 N
CO
2 H 1 11 455.16 HC2
CH
3 OCH 3 483.70 P9 K \I '- Nl-
C
2 H
CH
3 OCH 3 45.5 C0 2 H
CH
3 54 WO 01/00206 PCT/US00/18079 W7,. ... 0w 1 489.56 H C02H
CH
3 OCH 3 487.59 C2
CH
3 469.24 N'"C2
CH
3 OCH 3 C 2 489.56 N.C0 2 H
CH
3 OCH 3 5 489.56 IN
CH
3
OCH
3 C2 459.54 N. C0 2 H
CH
3 483.26 P C"~ C~ \-q I
CH
3 OCH 3 441.23 9~jWHN- C02H
CH
3 OCH 3 503.59 'N O
CO
2 H
CH
3 OCH 3 10 47.5 C02H
CH
3 55 WO 01/00206 PCT/US00/18079 507.55 N C0 2 H
CH
3 OCH 3 521.58 C2
CH
3
OCH
3 F Hy--'-CO 2 H
CH
3 OCH 3 9K HN--0-0 2 H
CH
3 OCH 3 414.10 9 N0 2
CH
3 OCH 3 441.2 12 IH
CQ
2 H
CH
3 OCH 3 5 511.47 9 /'~~Y7~7.N~
CH
3 OCH 3 459.11 9~ ~HN 0
'-O-CO
2 H
CH
3 OCH 3 504.6 H I<N C0 2 H
CH
3
OCH
3 l 500.20 I /I~ ~N
CH
3
H
1 56 WO 01/00206 PCT/US00/18079 M assp Spec 1 1 1 M~ssS ec W- -R --- L 530.2 3 0 CH3 OCH3H CH3 5 4 4 . 2 O H CH3 OCH3 4980.2 N -) 506.2 O SAOH
CH
3
OCH
3 498.2 ,
CH
3 OCH 3 HOOH 475.3 57
CH
3 OH 498.29~>~ N 57 WO 01/00206 PCT/US00/18079 1 TABLE 4 Mass S j_______J-pe - " L 502.20
HO
CH
3 N < 506.3 Ile N OH
CH
3
OCH
3 t 518.1 0> P--"" I / HO
CH
3
OCH
3 IN K 530.1 K> A A HO
CF-
3
OCH
3 N 5 526 N'
CH
3
OCH
3 COOH 602 A N
CH
3 OCH 3 \ LN SCOOH
CH
3
OCH
3 -' N-COOH 622 IJX C 3OCH 3 ' 58 WO 01/00206 PCT/US00/18079 T-.ABLE:4 MAss $Oqs [ _____.J: ~ - J ~RI 646 0
CH
3 OCH 3 N 5 518 IN ci OCH 3 -CH 563 IN BrOCH 3 ,N COOH cI
OCH
3 NC0 638 BrOCH 3 M- COOH 639 BrOCH 3 A ,,.C00H 10 594 1\ I~~~> cl ~OCH 3 NCO clOCH 3 M'N -COOH 513 [N 'N
CH
3
OCH
3 CO 59 WO 01/00206 PCT/US00/18079 TABLJE:A 4 638<N BrOCH 3 1' COOH 593 c I OCH 3 \/N-N COOH 624 BrOCH 3 _COOH 515 IN CH3 OCH 3 CH 5 595 I N
CH-
3 ONH TCOOH 615 . I N ciOCH 3 O'N COOH 514 OH C H "< ICOOH
C
3 OCH 3 596 N
CH
3
OCH
3 0 -CO 616k I N cIOCH 3 N COOH 60 WO 01/00206 PCT/US00/18079 TABLE 4 Mass Sec .L 516 yCOOH
CH
3 OCH 3 486 COOH
CH
3 521.56 O
OCH
3 OH
CH
3
OCH
3
OCH
3 535.55 O 9K OH
CH
3
OCH
3 COOH 5 507.54 O OOH
CH
3
OCH
3 OCH 3 535.59 O CH3 OH
CH
3
OCH
3
OCH
3 522.51 0 H OH 539.55 0 rCH3 OH F
OCH
3 OCH 3 502.56 0 .- ~ OH
CH
3 OCH 3 10 521.56 0 N " ( OH
CH
3
OCH
3
CH
3
OCH
3 61 WO 01/00206 PCT/US00/18079 TABLE 4,:. Mass Spec . M+ 1) 547.6 CH 2 0OH VN C?-OH
CH
3 OCH 3 OCH 3 620.69 O
CH
3
OCH
3 O OH
OCH
3 633.73
CH
3 590.67 I .. k 5 0HA O ~OH
CH
3 OCH 3 OH 5 495.5 0
OH
3 OH F OCH3 3 OCH 3 529.94 0
CH
3 OH
SOCH
3 Cl 491.54 0
CH
3 48.' OH
CH
3 OCH 3 525.98 0 CH3 Y OH
CH
3
OCH
3 Cl 489.56
OH
3 0 \'N OH
CH
3
OCH
3 62 WO 01/00206 PCT/US00/18079 TABLE .4: Mass Spec ... +-+ 1). - - R : --- ' I .v -. ,ia . v.:. .... : Tv.--.::.":/ : ..... '':. .... . :" . ..... ... 461.51 O
H
3 OH
CH
3 495.95 0 I)~ jCH 3 -'-OH
CH
3 <~. 4 9 1 .5 4 ^ 0 H _ I' -OH
CH
3 OCH 3 CH 3 461.51 O 0 H OH
CH
3 OH 3 5 495.95 0 525.98 N - O-.H O OH
CH
3
OCH
3
H
3 CI 534.6 H 538.57 0 525.98 k0 H -OH \ oN CH3 OCH 3
H
3
CH
3 534.6 0 CH3504.58 OH
CH
3
N
0 N C3 OCH3 H3C.-I NCH 3 538.57 \ , 0 CH3 " "O H OCH3
H
3 CI 'CH 3 63504.58
CH
3 H3 c -
-CH
3 63 WO 01/00206 PCT/US00/18079 TABLE4 ass S c . . .. . . .. . . .: , . , . .. . . . .: . ., ... .. .. ... . , . , - ... ... 552.59 0
CH
3 OH F OCH 3
H
3 C NH
CH
3 523.55 O CH3H 3 C CH 3 OH F
OCH
3 529.94 O H I OH FN F OCH 3 CH 3 CI 4 9 0 .5 5 YH N O H CH O
CH
3
OCH
3 H 5 543.97 O H OH F
OCH
3 CH 3 CI 540.01 O H O OH
CH
3 OCH 3 6H 3 Cl 5 4 8 .6 3 C3 I - , O 3H 3 " " O H
CH
3 OCH 3 H 3 C, NH
CH
3 504.58 3 O0 CH3 O H
H
3 OCH 3 CH 3 620.07 O O IIOH F
OCH
3 CH 3 CI 64 WO 01/00206 PCT/US00/18079 TABLE 4 Mass S c .W.
562.66 O 0 CHH OH
C
3 OCH 3 H 3 CyN
'
CH
3
CH
3 578.63 0 F
CH
3 OH
OCH
3 N 506.55 0
CH
3 O OH
CH
3 OCH 3 NH 2 574.67 0
CH
3 j:AOH
CH
3
OCH
3 O 5 510.51 0
CH
3 -- "OH F OCH 3 NH 2 506.55 0O 3CH 3 OH 3 OC H3
CH
3
NH
2 505.56 0 HOH
CH
3 OCH 3 H 3 C CH 3 556.41 0O H YI'OH Br
OCH
3 CH 3 596.67
H
2 N OH
CH
3 OCH 3 (H 3 65 WO 01/00206 PCT/US00/18079 TABLE 4 *MassSpec : .. . + ... .... . .: : .. ::. W . - . -. . .i. : ,, : . I::::. . .- :::: ::. ." " ... . . . . -.. !.....,..... . ..-..... : . ... ai.......:. 532.63 H3O K~i ICH 3 OH
H
3 OCH 3
CH
3 5 691.53
O
2 N I-; o -'OH Br \ 0_()
OCH
3 CH 3 54 1.0 O H -- 'OH CI OCH 3
H
3 C CH 3
NH
2 607 0cooH Me O
ONH
2 497 COOH H OMe H 525 - \.--COOH CI OMe IN 10 527 COOH Cl Me
NH
2 542 O H 9 / -~ r ,, COOH 498 H C O OH OMe 543 H COOH 66 WO 01/00206 PCT/US00/18079 ed .. ..
AB E. M a.. Spe .. .__. .... . .. R .... 611 / I N-V COOH Cl OMe Me H 512 -/ KV -ICOO cl OMe 518 C O O H M Me VN 5 569 / ., COOH OMe0 569 N M e 592 N0 / K COCH OMe 591 569 N I I CO OH CI "[Me \N ~ ' 10 538 Ci / K ~e N ja COOH 583-N Q/ K OO Br M 67 WO 01/00206 PCT/US00/18079 M sspec ...... 561 - c - COGH -j cI OMe 0 NH 2 549 -N / Me , 604 -N r- / N. ~N/Nr-,COQH 9-11 OMe 589 N /N N 2 COOH OMe N : 5 506 COOH M e 607 9/ COO OMe 575\~ COOH Me ~ / 0 COOH OMe ~ 10 6 1 \ N -1 COCH OMe / 603 NCN- CO 9/ OMe 68 WO 01/00206 PCT/US00/18079 ...... ... -.. ~~ ~... "" ".... .... " "... " - •. . .. • - . . " • - . ". : . . -- " :. . .-. ,.- - " 5 .: .:. 617 M e IN- COOH 5 625O / ~OMe N~ 0 c 611 OMe /- COOH 522 H COOH 95/ oMe\ N N O Mee 565 MeO 9 /O COOH OMeVN -a 593 F 93)/ COOH Me 10 518 y yCOOH OMe 538 C ICOOH CI Me 627. 627 ./ N KICOOH CIl MeO" 585 \ MeO 69 COOH Me _of 69 WO 01/00206 PCT/US00/18079 ..... TABLE.4 Mas .
1 .... ... L. 624
-
-\N 609 I CN-- COOH 639 OMe CO 623 I N- COH c I OMe ~ N--NQI< 5 637 ~ Q/ I LD CO c Me C OO 0 JII 59 ~COOH MI Oe N oa. c I yCOOH OMe
N-N
/ ~'~COOH OlMe L cMe Me 70 WO 01/00206 PCT/US00/18079 TAIBLE14. MassSpe J - j -I 611 O OF /F ~ -COOH 5 631 F O C O /IF COOH OMe v'O 488 O OO 504 , 5 'O. COOH 522..O.COOH 548-N COOH-O OMe H 640 ....... . .. ...
oo. ... .... . -, j" -O . . ... . .. . . . 548 / \/ ....-... COO OMe 1 OOH / I L"NI < COOH Br Me 10 6361 F F666 ' COOH 7Me1 488 -vCOOH 50 0/ MeCO 522 9-/ ~OMe N OaCO Br Me - 0 548 COOH Br OMe H 71 WO 01/00206 PCT/US00/18079 MassSpecJ WI-Ru L' 614 . COOH 626 \ H Br Me NCOOH 630 \ MeO Br .. Me ....... o . 676 -/ \ Q r OMe O CO 5 506 'O COOH 526... 'O COOH 502 .C O 514 COO Br- MeCOOH 630 MMe 484 'COOH 10 579 ' - .COOH /72 r~ e 72 WO 01/00206 PCT/US00/18079 ......... TA BL 4.... .. .. .. .. ... S ......... ~ 585I rJ~?XCOOH Br M 512 (I~CO 482 . K~QfCOOH 532 -I OMe CO 5 560 9-11 ~Me N< CO 625 583I Yy COOH Br Me 568 COOI OMeN 10 574 / ~fOOH 536 C / Me 73 WO 01/00206 PCT/US00/18079 IX 1...... MN AssSO .... . -. ... 581 o COO Br Me 522 COOH 520 COOH C-1 VNL
O<X
CO O H 520 Cl \, O The principles of the present invention also encompass prodrugs in the scope of Formula 5 I, and compounds representative thereof include those wherein R 5 or R 8 is a lower alkoxy group, and those wherein Rio or R 19 is a lower alkoxycarbonyl group. The principles of the present invention also provide a method for inhibiting cell adhesion, and in particular, VLA-4 mediated cell adhesion at a4p1 receptor sites in a mammal, wherein the method comprises administering an effective amount of a compound represented by Formula I. As 10 used herein, inhibiting cell adhesion is intended to include inhibiting, suppressing and preventing VLA-4 mediated cell adhesion-associated conditions, including but not limited to, inflammation and cell adhesion-associated immune or autoimmune responses. The principles of the present invention therefore also provide a method of treating a condition associated with VLA-4 mediated cell adhesion, wherein the method comprises 15 administering to a mammal in need of such treatment, an effective amount of a compound represented by Formula I. Such conditions include for example, but are not limited to, inflammatory and autoimmune responses, diabetes, asthma, arthritis, psoriasis, multiple sclerosis, inflammatory bowel disease, transplantation rejection, and tumor metastasis. As used herein, "treatment" of a mammal is intended to include prophylaxis as well. 74 WO 01/00206 PCT/US00/18079 The compounds of the present invention may be administered as a monotherapy, or in combination with antiinflammatory or immunosuppressive agents. Such combination therapies can involve the administration of the various pharmaceuticals as a single dosage form or as multiple dosage forms administered at the same time or at different times. 5 Any suitable route of administration may be employed for providing a patient with an effective amount of a compound of the present invention. Suitable routes of administration may include, for example, oral, rectal, nasal, buccal, parenteral (such as, intravenous, intrathecal, subcutaneous, intramuscular, intrasternal, intrahepatic, intralesional, intracranial, intra-articular, and intra-synovial), transdermal (such as, for example, patches), and the like. Due to their ease of 10 administration, oral dosage forms, such as, for example, tablets, troches, dispersions, suspensions, solutions, capsules, soft gelatin capsules, and the like, may be preferred. Administration may also be by controlled or sustained release means and delivery devices. Methods for the preparation of such dosage forms are well known in the art. Pharmaceutical compositions incorporating compounds of the present invention may 15 include excipients, a pharmaceutically acceptable carrier, in addition to other therapeutic ingredients. Excipients such as starches, sugars, microcrystalline cellulose, diluents, lubricants, binders, coloring agents, flavoring agents, granulating agents, disintegrating agents, and the like may be appropriate depending upon the route of administration. Because of their ease of administration, tablets and capsules represent the most advantageous oral dosage unit forms. If 20 desired, tablets may be coated by standard aqueous or nonaqueous techniques. The compounds of the present invention may be used in the form of pharmaceutically acceptable salts derived from inorganic or organic bases. Suitable pharmaceutically acceptable base addition salts include, but are not limited to, ammonium salts, alkali metal salts, metallic salts made from aluminum, calcium, lithium, magnesium, potassium, sodium and zinc, organic salts 25 made from chloroprocaine, choline, N,N'-dibenzylethylenediamine, dicyclohexylamine, diethanolamine, ethylenediamine, lysine, meglumine (N-methylglucamine) and procaine, as well as salts with amino acids, such as arginine, lysine, and so forth. Where the compounds of the invention have a basic moiety, such as an amino group, the compounds may be used in the form of pharmaceutically acceptable non-toxic organic or inorganic 30 acids. Such acids include acetic, benzenesulfonic, benzoic, camphorsulfonic, citric, 75 WO 01/00206 PCT/US00/18079 ethanesulfonic, methanesulfonic, fumaric, gluconic, glutamic, hydrobromic, hydrochloric, lactic, maleic, malic, mandelic, nitric, pamoic, pantothenic, phosphoric, succinic, sulfuric, tartaric, p toluenesulfonic acids, and the like. Particularly preferred are citric, hydrochloric, maleic, fumaric, phosphoric, sulfuric, tartaric and p-toluenesulfonic acids. Compounds of the invention may also 5 be in the form of hydrates. DETAILED DESCRIPTION OF THE INVENTION Abbreviations & Definitions The following terms and abbreviations have the indicated meaning throughout this disclosure. 10 293E HEK cells = 293E human embryonic kidney cells Ac = acetyl anti-(4-PE conjugated = monoclonal antibody against integrin 24 subunit, phycoerythrin conjugated anti-P 1 -FITC conjugated = monoclonal antibody against integrin P3 1 subunit, fluorescein 15 conjugated 5pl3 = integrin 5 031, fibronectin receptor, VLA-5 xvP33 = integrin vi33, vitronectin receptor a4P37 = integrin a437 Bn = benzyl 20 Boc = t-butoxycarbonyl BSA = bovine serum albumin c- = cyclo cDNA = complementary DNA CHO cells = Chinese Hamster Ovary cells 25 p-CIPh = para-chlorophenyl CMV promoter = cytomegalovirus promoter m-CPBA = 3-chloroperoxybenzoic acid DAST = diethylaminosulfur trifluoride DCM = dichloromethane = methylene chloride = CH 2 C1 2 30 DELFIA = dissociation enhanced lanthanide fluor-immune assay DIAD = diisopropyl azodicarboxylate DIC = diisopropylcarbodiimide DIEA = N,N-diisopropylethylamine 76 WO 01/00206 PCT/US00/18079 DMAP = 4-N,N-dimethylaminopyridine DMEM = Dulbecco's Modified Eagle's Medium DMF = N,N-dimethylformamide DTPA = Diethylenetriaminepentaacetic acid 5 EDC = 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide Et 2 O = ethyl ether FACS = fluorescence cell sorting Fmoc = 9-fluorenylmethoxycarbonyl GPIIb/IIIa = integrin cdlIb33, fibrinogen receptor 10 HEPES = N-(2-hydroxyethyl)piperazine-N'-(2-ethanesulfonic acid) HMDS = 1,,1,3,3,3-hexamethyldisilzane HOAc = acetic acid HOBt = 1-hydroxybenzotriazole human IgG1 = human immunoglobulin GI 15 ICAM = intracellular adhesion molecule LDV = Leu-Asp-Val LFA-1 and Mac-1 = Lymphocyte function-related antigen LiHMDS = lithium 1,1,1,3,3,3-hexamethyldisilazane Me = methyl 20 p-MeOPh = para-methoxyphenyl nM = nanomolar PBS = phosphate buffered saline PEG = polyethylene glycol Ph = phenyl 25 PhOH = phenol PyBroP = bromo-tris-pyrrolidinphosphonium hexafluorophosphate RPMI medium = Russell Park Memorial Institute medium TFA = trifluoroacetic acid TFAA = trifluoroacetic acid anhydride 30 THF = tetrahydrofuran TLC = thin-layer chromatography TMS = trimethylsilyl Ts = toluenesulfonyl VCAM-1 (D1D7) = vascular cell adhesion molecule (containing one to seven 77 WO 01/00206 PCT/US00/18079 immuloglobulin domains) VCAM-IgG fusion protein = a VCAM IgG fusion protein containing the one to seven immunoglobulin domains of human VCAM-1 (D1D7) attached above the hinge region of an IgG1 molecule 5 "Alkyl group" is intended to include linear or branched hydrocarbon radicals and combinations thereof of 1 to 20 carbons. "Lower alkyl group" means alkyl groups of from 1 to about 10, preferably from 1 to about 8, and more preferably, from 1 to about 6 carbon atoms. Examples of such radicals include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, iso-amyl, hexyl, octyl groups and the like. 10 "Alkylene group" means a divalent radical formed by removing a hydrogen atom from an "alkyl group." "Aryl group" means a radical formed from an aromatic hydrocarbon ring of 4 to about 16 carbon atoms, preferably of 6 to about 12 carbon atoms, and more preferably of 6 to about 10 carbon atoms. The rings may optionally be substituted with 1-3 substituents selected from alkyl, 15 halogen, hydroxy, alkoxy, aryloxy, haloalkyl, phenyl and heteroaryl. Examples of aryl groups are phenyl, biphenyl, 3,4-dichlorophenyl and naphthyl. "Arylene group" means a divalent radical formed by removing a hydrogen atom from an "aryl group." "Arylalkyl group" denotes a structure comprising an alkyl attached to an aryl ring. 20 Examples include benzyl, phenethyl, 4-chlorobenzyl, and the like. "Cycloalkyl group" refers to a saturated hydrocarbon ring radical of from 3 to 12 carbon atoms, and preferably from 3 to 8 carbon atoms. Examples include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, norbornyl, adamantyl, myrtanyl groups and the like. "Lower cycloalkyl group" refers to cycloalkyl of 3 to 6 carbons. 25 "Cycloalkylene group" means a divalent radical formed by removing a hydrogen atom from a "cycloalkyl group." 78 WO 01/00206 PCT/US00/18079 "Divalent C 1 to C 20 aliphatic hydrocarbon moiety" includes alkylene, cycloalkylene, alkenylene, alkynylene groups and combinations thereof. Examples include ethylene, propylene, propynylene, 2,4-heptadienylene groups and the like. "Heterocyclyl group" refers to a cyclic radical having from 1 to 6 carbon atoms, preferably 5 3 to 6 carbon atoms, and from 1 to 4 heteroatoms chosen from O, N and S. Examples include: pyrrolyl, pyridinyl, pyrazolyl, triazolyl, pyrimidinyl, pyridazinyl, oxazolyl, thiazolyl, imidazolyl, indolyl, thienyl, furyl, azetidiyl, tetrazolyl, 2-pyrrolinyl, 3-pyrrolinyl, pyrrolindinyl, 1,3-dioxolanyl, imidazolinyl, imidazolidinyl, pyrazolinyl, pyrazolidinyl, isoxazolyl, isothiazolyl, 1,2,3-oxadiazolyl, 1,2,3-triazolyl, 1,3,4-thiadiazolyl, 2H-pyranyl, 4H-pyranyl, piperidinyl, 10 1,4-dithianyl, morpholinyl, thiomorpholinyl, pyrazinyl, piperazinyl, 1,3,5-triazinyl, 1,2,5-trithianyl, benzo(b)thiophenyl, benzimidazolyl, quinolinyl groups and the like. "Heterocyclylene group" means a radical formed by removing a hydrogen atom from a "heterocyclyl group." "Heteroaryl group" refers to an aromatic cyclic radical having from I to 12 carbon atoms, 15 preferably 1 to 6 carbon atoms, and from I to 4 heteroatoms chosen from O, N and S; or a bicyclic 9- or 10-membered heteroaromatic ring system containing 1-4 heteroatoms selected from O, N and S. The methine H atoms of a heterocyclyl or heteroaryl structure may be optionally substituted with alkyl, alkoxy or halogen. Examples include: imidazolyl, pyridyl, indolyl, thienyl, benzopyranyl, thiazolyl, furyl, benzimidazolyl, quinolinyl, isoquinolinyl, quinoxalinyl, 20 pyrimidinyl, pyrazinyl, tetrazolyl, pyrazolyl groups and the like. "Heteroarylene group" means a divalent radical formed by removing a hydrogen atom from a "heteroaryl group." "Alkoxy group" means a straight, branched or cyclic hydrocarbon configuration and combinations thereof, including from 1 to 20 carbon atoms, preferably from 1 to 8 carbon atoms, 25 more preferably from 1 to 4 carbon atoms, and an oxygen atom at the point of attachment. Suitable alkoxy groups include methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, iso-butoxy, sec butoxy, tert-butoxy, cyclopropoxy, cyclohexyloxy groups and the like. "Lower alkoxy group" refers to alkoxy groups having from 1 to 4 carbon atoms. 79 WO 01/00206 PCT/US00/18079 "Alkenyl group" refers to an unsaturated acyclic hydrocarbon radical in so much as it contains at least one double bond. "Lower alkenyl group" refers to such radicals containing from 2 to 10 carbon atoms, preferably from 2 to 8 carbon atoms and more preferably from 2 to 6 carbon atoms. Examples of suitable alkenyl radicals include propenyl, buten-1-yl, isobutenyl, penten-1-yl, 5 2-methylbuten-1-yl, 3-methylbuten-l-yl, hexen-l-yl, hepten-l-yl, and octen-1-yl groups and the like. "Alkenylene group" means a divalent radical formed by removing a hydrogen atom from an "alkenyl group." "Alkynyl group" refers to an unsaturated acyclic hydrocarbon radical containing at least 10 one triple bond. Examples include ethynyl, propynyl groups, and the like. "Alkynylene group" means a divalent radical formed by removing a hydrogen atom from an alkynyl group." "Substituted alkyl group" means a linear or branched alkyl group wherein at least one hydrogen atom attached to an aliphatic carbon is replaced with a substituent such as alkyl, amino, 15 alkoxy, hydroxy, aryl, cyano, carboxy, alkoxycarbonyl, monoalkylamino, alkyloxy, cyanoalkyl, cycloalkyl, alkylthio, alkylsulfinyl, alkylsulfonyl, arylthio, carboxyalkyl, alkoxycarbonylalkyl, haloalkyl, acylamino, dialkylamino, cyclicamino groups, halogen atom and nitro. Examples of such substituent groups include methyl, isopropyl, methoxy, ethoxy, propoxy, amino, methylamino, phenyl, naphthyl groups, chlorine, fluorine and the like. 20 "Substituted alkylene group" means a linear or branched alkylene group wherein at least one hydrogen atom attached to an aliphatic carbon is replaced with a substituent such as alkyl, amino, alkoxy, hydroxy, aryl, cyano, carboxy, alkoxycarbonyl, monoalkylamino, alkyloxy, cyanoalkyl, cycloalkyl, alkylthio, alkylsulfinyl, alkylsulfonyl, arylthio, carboxyalkyl, alkoxycarbonylalkyl, haloalkyl, acylamino, dialkylamino, cyclicamino groups, halogen atom and 25 nitro. "Substituted cycloalkyl group" means a cycloalkyl group wherein at least one hydrogen atom attached to a ring carbon atom is replaced with a substituent such as alkyl, amino, alkoxy, hydroxy, aryl, cyano, carboxy, alkoxycarbonyl, monoalkylamino, alkyloxy, cyanoalkyl, cycloalkyl, 80 WO 01/00206 PCT/US00/18079 alkylthio, alkylsulfinyl, alkylsulfonyl, arylthio, carboxyalkyl, alkoxycarbonylalkyl, haloalkyl, acylamino, dialkylamino, cyclicamino groups, halogen atom and nitro. "Substituted cycloalkyene group" means a cycloalkylene group wherein at least one hydrogen atom attached to a ring carbon is replaced with a substituent such as alkyl, amino, 5 alkoxy, hydroxy, aryl, cyano, carboxy, alkoxycarbonyl, monoalkylamino, alkyloxy, cyanoalkyl, cycloalkyl, alkylthio, alkylsulfinyl, alkylsulfonyl, arylthio, carboxyalkyl, alkoxycarbonylalkyl, haloalkyl, acylamino, dialkylamino, cyclicamino groups, halogen atom and nitro. "Substituted aryl group" means an aryl group wherein at least one methine hydrogen atom attached to an aromatic carbon is replaced with a substituent such as alkyl, amino, alkoxy, 10 hydroxy, aryl, cyano, carboxy, alkoxycarbonyl, monoalkylamino, alkyloxy, cyanoalkyl, cycloalkyl, alkylthio, alkylsulfinyl, alkylsulfonyl, arylthio, carboxyalkyl, alkoxycarbonylalkyl, haloalkyl, acylamino, dialkylamino, cyclicamino groups, halogen atom and nitro. Substituted arylene group" means an arylene group wherein at least one hydrogen atom attached to an aromatic carbon is replaced with a substituent such as alkyl, amino, alkoxy, 15 hydroxy, aryl, cyano, carboxy, alkoxycarbonyl, monoalkylamino, alkyloxy, cyanoalkyl, cycloalkyl, alkylthio, alkylsulfinyl, alkylsulfonyl, arylthio, carboxyalkyl, alkoxycarbonylalkyl, haloalkyl, acylamino, dialkylamino, cyclicamino groups, halogen atom and nitro. "Substituted heteroaryl group" or "substituted heterocyclyl group" means a heteroaryl or heterocyclyl group wherein at least one hydrogen atom attached to a ring thereof is replaced with a 20 substituent such as alkyl, amino, alkoxy, hydroxy, aryl, cyano, carboxy, alkoxycarbonyl, monoalkylamino, alkyloxy, cyanoalkyl, cycloalkyl, alkylthio, alkylsulfinyl, alkylsulfonyl, arylthio, carboxyalkyl, alkoxycarbonylalkyl, haloalkyl, acylamino, dialkylamino, cyclicamino groups, halogen atom and nitro. "Substituted heteroarylene group" or "substituted heterocyclylene group" means a 25 heteroarylene or heterocyclylene group wherein at least one hydrogen atom attached to a ring thereof is replaced with a substituent such as alkyl, amino, alkoxy, hydroxy, aryl, cyano, carboxy, alkoxycarbonyl, monoalkylamino, alkyloxy, cyanoalkyl, cycloalkyl, alkylthio, alkylsulfinyl, alkylsulfonyl, arylthio, carboxyalkyl, alkoxycarbonylalkyl, haloalkyl, acylamino, dialkylamino, cyclicamino groups, halogen atom and nitro. 81 WO 01/00206 PCT/US00/18079 "Substituted arylalkyl group" means an arylalkyl having one or more substituents such as alkyl, amino, alkoxy, hydroxy, aryl, cyano, carboxy, alkoxycarbonyl, monoalkylamino, alkyloxy, cyanoalkyl, cycloalkyl, alkylthio, alkylsulfinyl, alkylsulfonyl, arylthio, carboxyalkyl, haloalkyl, alkoxycarbonylalkyl, acylamino, dialkylamino, cyclicamino groups, halogen atom and nitro. 5 "Halogen" is intended to include for example, F, Cl, Br and I. The term "prodrug" refers to a chemical compound that is converted to an active agent by metabolic processes in vivo. [See, e.g., N. Boder and J.J. Kaminski, Ann. Rep. Med. Chem. 22:303 (1987) and H. Bundgarrd, Adv. Drug Delivery Rev., 3:39 (1989)]. The use of prodrug precursors of compounds of the present invention in any of the methods described herein is contemplated and 10 is intended to be within the scope of the invention. Terminology related to "protected," "protecting" and/or "deprotecting" functionalities is used throughout this application. Such terminology is well understood by persons of skill in the art and is used in the context of processes which involve sequential treatment with a series of reagents. In this context, a protecting group refers to a group which is used to mask a functionality during a 15 process step in which it would otherwise react, but in which reaction is undesirable. The protecting group prevents reaction at that step, but may be subsequently removed to expose the original functionality. The removal or "deprotection" occurs after the completion of the reaction or reactions in which the functionality would interfere. Thus, when a sequence of reagents is specified, as it is in the processes of the invention, the person of ordinary skill can readily envision 20 those groups that would be suitable as "protecting groups" for the functionalities involved. In the case of the present invention, the functionalities that must be protected are amines. Suitable groups for that purpose are discussed in standard textbooks in the field of chemistry, such as Protective Groups in Organic Synthesis by T.W.Greene [John Wiley & Sons, New York, 1991], which is incorporated herein by reference. Particular attention is drawn to the chapter entitled 25 "Protection for the Amino Group" (pages 309-405). Preferred protecting groups include BOC and Fmoc. Exemplary methods for protecting and deprotecting with these groups are found in Greene and Wuts on pages 318 and 327. The materials upon which the syntheses described herein are performed are referred to as solid supports, beads, and resins. These terms are intended to include: (a) beads, pellets, disks, 30 fibers, gels, or particles such as cellulose beads, pore-glass beads, silica gels, polystyrene beads optionally cross-linked with divinylbenzene and optionally grafted with polyethylene glycol, poly 82 WO 01/00206 PCT/US00/18079 acrylamide beads, latex beads, dimethylacrylamide beads optionally cross-linked with N,N'-bis acryloyl ethylene diamine, glass particles coated with hydrophobic polymer, etc., i.e., material having a rigid or semi-rigid surface; and (b) soluble supports such as polyethylene glycol or low molecular weight, non-cross-linked polystyrene. The solid supports may, and usually do, have 5 functional groups such as amino, hydroxy, carboxy, or halo groups; where amino groups are the most common. TentagelTM NH 2 (Rapp Polymere, Tubingen, Germany) is a preferred amine functionalized polyethylene glycol- grafted polystyrene resin. Tentagel"-S-PHB resin has a para hydroxy benzyl linker which can be cleaved by the use of 90% trifluoroacetic acid in 10 dichloromethane. Techniques for functionalizing the surface of solid phases are well known in the art. Attachment of lysine to the amino groups on a bead (to increase the number of available sites) and subsequent attachment of linkers as well as further steps in a typical combinatorial synthesis are described, for example, in PCT application WO95/30642, the disclosure of which is incorporated herein by reference. In the synthesis described in WO95/30642, the linker is a 15 photolytically cleavable linker, but the general principles of the use of a linker are well illustrated. Optical Isomers - Diastereomers - Geometric Isomers Some of the compounds described herein contain one or more asymmetric centers and may thus give rise to enantiomers, diastereomers, and other stereoisometric forms which may be defined in terms of absolute stereochemistry as (R)- or (S)-, or as (D)- or (L)- for amino acids. 20 The present invention is meant to include all such possible diastereomers as well as their racemic and optically pure forms. Optically active (R)- and (S)-, or (D)- and (L)- isomers may be prepared using chiral synthons or chiral reagents, or resolved using conventional techniques. When the compounds described herein contain olefinic double bonds or other centers of geometric asymmetry, and unless specified otherwise, it is intended to include both (E)- and (Z)- geometric 25 isomers. Likewise, all tautomeric forms are intended to be included. The configuration of any carbon-carbon double bond appearing herein is selected for convenience only and is not intended to designate a particular configuration; thus a carbon-carbon double bond depicted arbitrarily herein as trans may be cis, trans, or a mixture of the two in any proportion. In view of the above definitions, other chemical terms used throughout this application 30 can be easily understood by those of skill in the art. Terms may be used alone or in any combination thereof. The preferred and more preferred chain lengths of the radicals apply to all such combinations. 83 WO 01/00206 PCT/US00/18079 Utility The compounds of the present invention have demonstrated utility as selective inhibitors at VLA-4 receptors. The inhibitory concentration (IC 50 so) and the VLA-4 selectivity of test compounds for an 4f41 receptor using in vitro assays are determined in direct binding assays and 5 competitive assays with other integrin receptors such as 032 (LFA-1 and Mac-1), P3 (GPIIb/IIIa and avcp3) and P1 (a407). Compounds of the present invention have I, values < 1 ptM. Preferred compounds of the invention are those having K, values < 300 nM, more preferably < 100nM, even more preferably < 50 nM, and most preferably, < 12nM. Examples of preferred compounds having a KI, value < 50 nM are shown below. These 10 examples are provided by way of illustration only, and are not intended to limit the invention thereto. y\/ H0 / H 0 0 / 0' H / H N H HO NH O-OH N NN OH \ O O H / N' /H N H MeO H HN-NI N H ON H ON H H/ CO N N0O COOH N N o cozN? 0 ~ ~ N'H N N)'JQO 0 C02H ~ 15 H -H0 2 H 84 WO 01/00206 PCT/US00/18079 N 'k N- ' lH H 911Nr H H 0 2 H 0 2 H 0 0 N k H H H H C0 2 H 0 2 H '~ NN ~ <'OC 0 2 H N N N~ 0 N l?0 Hj ~ / 0 2 H j) C2 H ~ H H 0, {& QC 2 0 N ~NqN 0 o 0 2
H
1 5? N C H H qN Nj 0\ CO 2 H 5c0 H H 0 C02 AN'[;) o -P 02 0:/ C2 H H C 2 N0 NYN N N"'N 0-0 , -H H ii N'N~~~ C H H H0 2 C 02 00 2 85 WO 01/00206 PCT/US00/18079 N ONC C -YNy'C2 Me H H r H H , 0 CO 2 H
H
N N NCO2 CO2H O2 H o F0 N , - "'oN Me02 H H CN' N' OBn I0 NJ N N N H H H /Oy " /'O
HH
0 H,0
CO
2 H
CO
2 H 02 .OH OH NO% 0 C0 2 H N NH 2 H H 0 5 M H H CO2H
SN
Q
O C2H O C N O CO 2 H CO2 H HNHl0 N CNN2 N? N N CO 2 H H N I8 6 H 0 MeH H 86 WO 01/00206 PCT/US00/18079 NNaCO2H NNCO2H NN NON O2H H H N I H H / I H H /I 02H OC2HQf 5J OH N N N OO0H 0 / - ,M e NN . N N O H NOOH OF H H NO 2 r H H
*
r - - 02H I Q 1 y A O 2 H ' H Me NH2 M N'O1{NI r Me H NH2 HH Me r N'N NN iH2 Me QH H H Me SN O H N COOH 87r HH Me COOH N'N 0 ' ,IH 2 0 -' H H - 2 H H -l e M NMe c o H . N N NO C-HN'L N 0 N-( OOH I H H Me e H H OMe , O eN e (OCOOH Me H H H e H7 WO 01/00206 PCT/US00/18079 \ OH OH HH OMe F ~HH Me ~ C OH 9J:XN)9 y OH OH H HOOH N OOH 0COOH 6e H H Me KCOOH N <NCOOH ~[ N\J~OOH HOOH NIN-Q q-CO ( NQCO MeMO HJ-COOH H Me 9INY. NQ-OGHCO? ~HH Q ,COOH Me Me I H H 6Me ON2QCOOH e H HHKH -NXN)-OCOOH N'N 'QyC COOH 88 6M WO 01/00206 PCT/US00/18079
H
2 NIN-Q OOH 9: NXNIQ"YO-O-COOH H H 6Me Me H H M K~XN 0r OOH Z 0 4-iNOOH Me N Y~ H MeN F K~HH e 0OOH c:NYN ICOOH MeeH H Me NmN N CO Z ~Me C0H ~ N Me LC OH Y ~ H Me _,rNON r _, 0H HQC0 Hr H H 6Me H H MeMe~ HMe H H OMe
N
OOH YCOOH ,? HH OMe CIH H Me KHMe 'rN Me H H M 89 WO 01/00206 PCT/US00/18079 o-0 N N ~OOH H N OOH HH Me OMe OOH NQ OH N OOH NI Ne H O N° NMe H)M Me Me COOH COOH °t 5 C NOOH N N C OOH OOH CH HOOOOH -- cooH o--cOH M -e NO O NJ.-%) I /jr COOH N 3 e /P COOH o COOH o-OCOOH NN K(NN I /OH ' COOH I?..-H H cooH MeH H Me OOH NNN COOH -H H Me Me HMe H Me OOH 1COOH MeH H Me I Y'HH Me N& 90 WO 01/00206 PCT/US00/18079 OOH YN COOH Br H H Me r HH Me O-Q- COOH Y ,NMe Me N'N / y COOH N Nq/ OOH COOH OOH O OMe NCOOH N NOOH SOOH N N COOH Br H H OMe r H H Me Hr eOO N 9AQCOOH ONO N COOH me COH Me H HMe Me OMe N NO aCOOH ?NXN KO OOH N9 H 00 NH Me N -aOH K 0 &N -COOH q -eKO-QcooH l Me O-COOH 91 WO 01/00206 PCT/US00/18079 5K O-Q-COOH aCH ee 6HH Me N? ' 0-O-{j NIN COOH Me MeH Me oQc H Me H Me .405, KO e 0 -COOH <lI J:aCO H Me rJH H M MeM M MeO-COO Me FO ,,mq -a OOH I q oa OOH 5 ZHH MZeHH e / M N Me -Q/COOH I 0 OOH MeH H Me M I I OH I I--- Q- 92 WO 01/00206 PCT/US00/18079 FF ? I O-a(~COOH (?NLI COOH ri NloaCOOHOOH M M~H H Me HO H MeOH me Me'N' Q 6H HH Me N Me '? MeCOOH N Me H N & C H(I - 'Nr NJ CarCOOH H HiH Hr F F NN N INOa OH NINN O-CJYCOOH H H r rH H Me F F3 WO 01/00206 PCT/US00/18079 F NN I -- COOH N rCOOH o -COO Me H H F O COOH N O-~~COOH F F SN NJT O NlCOOH |l" N N N' aCOH M e H H rMe H H F F N NeOOH N N HCOOH e J -- COOH 'a-COOH O5H H H rO OH T COOM HN HN H NH NHHH Me 4P-Q-COOH HO 5-OH NOH O 0 0 HN HN 0~ NNH 9NHN\\/-NH 94 WO 01/00206 PCT/US00/18079 HN-I N NAN OH H N OH o OHH N NN 0 Nl 0 0 SeOH OMeO H HN N HN N HN N OO .. y. _ y o HQ OH OH H2 O H HQ OH NJ N MeO MeO MeO Br HN N 6 H N Hy N_6 0 00 Ph OH 0 OH Ph NO OH N r,5 ClQ N NJ N N 0 0 Me MeO MeO HN N NHH-N 95 0 0 Ph N O P OH N NJ N 0y OH NN , 0 Meo H0 Meo HN HNN' -NHH ~ -NH HyN 95 WO 01/00206 PCT/US00/18079 r~ry OH N ,_N , 000 O No_ OH O 0 0 lp p z M H H HN Hy /NHO NH p-MeOPh HI.HHQ"N, O QOH N, r N N NO 0 O MeO He H 2 Meo H OH H ,6 yN- HN yN _ HN y H Bn-O 4
.
N 0 HN MeO HMeO0 H N H H N H~< ( ~HN N1 HY J NO K- O SN Njl' N1 0~Y COOH
H
3 H H 6CH 3
CH
3 H H OCH 3
CH
3 0 I J QkCOO KCOOH iHPHH CH 3
CH
3 H H OCH 3
CH
3
CH
3 H 3 1 CH
CH
3 KNNNCOGH 0COOH 2H H 6CH 3
CH
3 H H 96 WO 01/00206 PCT/US00/18079 HO HO H 0 ~ .- CO Y~~ HO 0 , N-COOH 'H H 6CH, CH 3 H H OCH 3 H Ho CH N O .NCOOH O N)'N N N
H
3 H H CH3 H3H H OCH3 NOH COOH NNCOO H3H 3 H H OCH 3
H
3 C3 O N . OCOOH SOOHOH
CH
3 H H OCH 3 3H H
OCH
3 H C- 3
NH
3 N Ol" O H - OOH I' N N COOH ' -N -Q 0,, O N"'Nj
CH
3 H H OCH 3
CH
3 H H OCH 3
CH
3 , COOH y 3 S N N 3 N 0 COOH PT.N N- ' N N r 5 H H OCH 3
CH
3 H H H H COOH N NNH3 CH3 2HH CH 3
CH
3 H H OCH 3
H
3 CF F-,,¢ .9H 3 - 1 CH 3 Q ~ N.N OOHLJ. - I ''OOH H CH 3 OH OCH 3
OH
3 H H OCH 3 C \ N N OO 300-cOH H 3H H OCH 3 CH3 H H OCH 3 N N - COOH 0 NIcOOH 2H 3 H H CH 3
H
3 H H OCH 3
H
3
CH
3 NXN "O v COOH 10 CeH H COMeOH 97 WO 01/00206 PCT/US00/18079 F Me N N N N'kI 1M 'N'O * 1 N NT IIN-"-0 fie H H 6Me CeH HH Me KN ,COOH (C N N 0N Y(-NYN 0l Cl H H OMe 'N.,.NCOOH Br H H OMe TNCO I xN -N N (CiNQ Y H H e NN:C00 0 ~ C0 cl H H OMe TNCOOH 0 H Me 1 N,-COOH NN I(J-eHH Me1 N HY N O~ NCO 5 l H Bre H H Me C0 ~Ye N N Nj 0 0 I~N MeHH Me~-C00H -QeH O0 Q-)COOH Me H-- Hr M eH H Me yMe NN q~ Me C00COH r~ Me T ~COOH 10 Me HH Me MeCH H H 6Me COOH e C98 WO 01/00206 PCT/US00/18079 Hj jCOOH IH2 COCH O~ IIN EMe OMe MMeHM
YH
3 N N COOH COOH N- 6CH 3 Y'-NZNY'
OCH
3 H1H OCH 3
CH
3 H H OCH 3 HNA COH H 3 $COOH Y~~'NY OCH3 0j~~
CH
3 H H OCH 3 H H OCH 3
YH
3 O-COOH
H
3 -~COOH 9-NY-Nr'M c
CH
3 H H OCH 3
CH
3 H H OCH 3 9~N ~ N Y"HJ3 9H 3 c~COOH P-YH'(ry 'OH 3 3 5 CH 3 H H OCH 3 CHH H COOH COOH N ~ 6 H 3 I H 3 ORH
CH
3 H H OCH 3
CH
3 H H OH ?H COOH 9H 3 COQH N3 N HH 6CH 3
CH
3 H H0H NX(H 3 CkH YH3 OH YH C99 WO 01/00206 PCT/US00/18079
COH
3
CH
3 H N
H
3 NH NZ H H H 6CH 3
C
6
H
3
CH
3 H H OCH 3
OH
3 'COOH
CH
3 ~COOH N ( N NH 6i$'.~ OH 3 C HCH HCHHH 3 H
OH
3 ' 1vCO HCH 3 <COO HH H 6CH 3
CH
3 H H CH 3 C3OOH
CH
3 ~-COOH II I N ) N H3OCN H -- N_ 0 C
OH
3 HH OH 3
H
3 CH OCH 3 jH 3 OOH
CH
3 Oq OOH PN'N H 7NX N
H
3 H H 3 H 3CH9 3 C3 H H OCH 3 (iH 3 >OOOH I'NNH 3XCOOH 5 3H H OH C H 3 H OCH 3 T H3 OOH H COOH ~IXN ~ 0 0~ CH H 6CH H 3 H H CH 3 H2NOQ CH ICOOH 9 N'N I0 H 3 \Hk r'-Nzj C3
CH
3 H H OOH H3 HH OCH 3
CH
3 100 WO 01/00206 PCT/US00/18079 02N-, COOH H COOH .0 coo. 'N r oo. OMe QN Nr e~A~ N N OiHH Me e NH 2 jO COOH MecCOOH NN N O NH 2 - ~ MH H Me H H M P COOH COOH N NH 2 O0 C H H OMe HH Me
S
COOH H COOH NeO N H2 NN N H ClH OMe MeHH Me e - COOH H COOH N NN CI H HOMe MeH H OMe H COOH H2 COOH NY N N Na SCl H H OMe MeH H OMe NM~CO h COO .COOHCOH o~re Me H H OMe MeHH OMe Mee N COOH MN COOH 101 MeHH OMe MeHH Me Oe O e-101 Q-NYN- oY NN10! WO 01/00206 PCT/US00/18079 y~e OOH SJJ <-COOH -ja N ,-K Ke 0N'' Me H H 6Me Me H H OMe KZN-D Oj:CCOOH )N Z1r7JCfCOOH Me HH 6Me Me HH OMe 0jCOOH - ~ cI~>Y COOH MH H 6Me MeH H OMe met N - jCOOH MefT(jCOOH Me H H 6Me me H H 6Me M e-O HO 01 0 C00H Me c NYN 0' NYN-P 5 e Me 0Me H H OMe -COOH 7COOH Me H H 6&e MeH H 7 - f"COOH NO, CO 8r H H 6MeH H Me ( COOH N~>~ COOH N-Y N( 0 5 'N~ CH H OMe Me H H OMe Y Jy COOH
-
7-,0COOH Me H H 6Me- I H H 6Me 102 WO 01/00206 PCT/US00/18079 COOH Me-)N , COOH 1 1 H Me HH6M MeNJP-\ COOH KZV COOH I HH 6Me MH H 6Me MeH_,HMMe -HH -\ F __OHN- C0QH NN - H__j~CO Me 0 5 rHMe Me H H Me FN)COGH MFj3N--) COOH MeH He~ HVK 6M Me 9 -COOH yeqCOH Me rH H 6eMe H H 6Me BrHH MeMeH H COOH MeNO Nay N " a re H e MerH M COOH I? 103 WO 01/00206 PCT/US00/18079 In vitro Assays A direct binding assay was used to quantify the inhibitory activity of the compounds. In this assay, VLA-4-expressing cells were seeded in a 96-well microtiter plate. The cells were allowed to grow for 2 days until confluent. Various concentrations of the test compound were 5 added together with 2 nM of the europium-labeled, VCAM-IgG fusion protein. The cells were allowed to incubate at room temperature in the microwells for at least 30 minutes. Following incubation, the microwells were emptied and washed. The amount of europium-labeled VCAM IgG fusion protein bound was determined by time-resolved fluorescence measurement. Inhibition of binding was determined by quantifying the fluorescence bound to the plate for each of the 10 various concentrations of test compound, as well as for controls containing no test compound. The VLA-4-expressing cells used in this assay was a CHO cell line stably transfected with the cDNA of the human cx4 and P 1 subunits. Construction and maintenance of the cell line are described in the assay procedures. A VCAM IgG fusion protein containing the one to seven immunoglobulin domains of human VCAM-1 (D1D7) attached above the hinge region of an IgG1 15 molecule was labeled with europium chelates. The preparation and labeling of the fusion protein are described in the assay procedures. The cell adhesion inhibitory activity of the test compound was determined by blocking the Jurkat cell attachment to the D 1D7-VCAM IgG fusion protein. Jurkat cell is a human lymphocytic cell line expressing VLA-4 on cell surface. In this assay, each of the 96-well microtiter wells was 20 coated with 75 ng of the VCAM IgG fusion protein. The wells were then blocked by the addition of 1% bovine serum albumin to remove nonspecific adhesive sites. Varying concentrations of the test compound were added together with the calcein-labeled Jurkat cells. The cells were allowed to adhere to the VCAM coated wells at room temperature for 1 hour in the dark. Following incubation, the plate was washed by immersing face down into a container filled with phosphate 25 buffered saline. The wells were blotted dry on paper towel. Quantitation of the adhered cell was determined by fluorescence measurement. Decreased fluorescence indicated inhibition of cell adhesion by the test compound. Specificity for a43 1P of each test compound among other integrin receptors, namely, 02 (LFA-1 and Mac-1), 033 (GPIIb/IIIa and cavP3), 31 (c5p31) and 37 (Ca437) was examined. LFA-1 30 binds to ICAM-1 and mediates the emigration of leukocytes into inflammatory sites. Mac-1 binds to a number of ligands, including ICAM-1 and fibrinogen, and plays an important role in 104 WO 01/00206 PCT/US00/18079 neutrophil phagocytosis and oxygen free radical generation. GPIIb/IIIa on platelet surface binds to fibrinogen in plasma and induces platelet aggregation. avP3 binds to a number of extracellular matrix proteins, including vitronectin and mediates cell migration and prevents cell apoptosis. a4P7 shares the same ligands as VLA-4 (VCAM-1, MAdCAM-1,and fibronectin), but with 5 different preference. This receptor is expressed on lymphoid cells and is involved in lymphocyte migration to mucosal tissues. Assays of LFA-1, Mac-1, GPIIb/IIIa and avP3 involved coating the purified receptor on a 96-well microtiter plate. The specific ligands for these receptors were labeled with europium chelates. In the assays of LFA-1 and Mac-1, an ICAM-1 IgG fusion protein containing the one to 10 five immunoglobulin domains of human ICAM-1 (D1D5) attached above the hinge region of an IgG1 molecule, was used. In the assays of GPIIb/IIIa and avP3, europium-labeled fibrinogen and vitronectin, respectively, was used. The purified receptors were allowed to incubate in the wells with various concentrations of test compound, in the presence of europium-labeled ligands. Following incubation, the wells are emptied and washed. The amount of europium-labeled ligand 15 bound was determined by time-resolved fluorescence measurement. Assay of a4P7 is similar to the adhesion inhibition assay of VLA-4 described above, and uses the a4p7-expressing cell, RPMI-8886. A MAdCAM-1 IgG fusion protein containing the one and two immunoglobulin domains of human MAdCAM- 1 and mucin-like repeat domain, is used as the corresponding ligand for a4P7. 20 Eu 3 +-VCAM-1 IgG binding to CHO/VLA-4 cells may be determined as follows. 4B4 cells (CHO/VLA-4 cells) are distributed into each well of a 96-well microtiter plate at 3 x 10 4 /well. The plate is incubated at 37 0 C, 5% CO 2 for 48 hours and then washed twice with washing buffer, then blot dried. 50 Ml of the inhibitor solution diluted with assay buffer (2% DMSO final) is added to each well, followed by 50 p,1 of Eu 3 '-VCAM-1 IgG diluted with assay buffer at 2 nM. The plate is 25 incubated at room temperature for at least 30 min. Each well is then washed four times with washing buffer and blot dried. 100 p,1 of DELFIA Enhancement solution is added to each well, followed by agitation of the plate at room temperature for 5 min. Fluorescence of each sample is then measured (e.g., DELFIA Fluorometer 1234, Wallace, Inc., USA). In this assay, the washing buffer comprises 25 mM HEPES (pH 7.5), 150 mM NaCl, 1 mM CaCl 2 , 1 mM MgCl 2 ,and 4 mM 30 MnC1 2 ; the assay buffer comprises 25 mM HEPES (pH 7.5), 150 mM NaC1, 1 mM CaCl 2 , 1 mM MgCl 2 , 4 mM MnCl 2 , 1% BSA, and 20 pM DTPA. In Vivo Assays 105 WO 01/00206 PCT/US00/18079 The VLA-4 inhibitors may be further characterized in in vivo assays. One such assay examines the inhibition of eosinophil infiltration into the bronchoalveolar lavage fluid in the mouse (murine) model. In this assay, the animals are treated with cyclophosphamide on day 0. On days 2 and 14, the animals are immunized intraperitoneally with Ascaris suum extract. Seven 5 days later, the animals are treated with various doses of the VLA-4 inhibitor. Shortly after drug administration, the animals are challenged with Ascaris suum extract by instillation into the trachea. Bronchoalveolar lavage of the animal is performed by instilling saline into the lung, 48 hours later. Total cell and eosinophil counts in the lavage are determined. In the murine model ofAscaris-induced bronchial inflammation, one of the representative 10 compounds (example number 32) inhibited eosinophil infiltration by 49% at an oral dosage of 30 mg/kg. By contrast, a representative prior art compound, 4-(N'-2-methylphenylurea)phenylacetyl LDVP-OH, described in WO 97/03094, did not inhibit eosinophil infiltration (% inhibition = -2%) at an oral dosage of 50 mg/kg. Other representative compounds were also tested in mice. The dosage, route of 15 administration and inhibitory effect of representative compounds (hereinafter, all tested compounds are referenced by compound number provided in the Synthetic Examples) are shown in Table 5. TABLE 5 Dosage/Regimen Route % Inhibition of Eosinophil Infiltration Compound 79 30 mg/kg b.i.d i x 2days i.v. 35.9 20 Compound 90 10 mg/kg t.i.d 2 x 2 days p.o. 18.1 Compound 90 30 mg/kg t.i.d. x 2 days p.o. 39.1 Compound 90 50 mg/kg t.i.d. x 2 days p.o. 45.9 Compound 90 30 mg/kg b.i.d. x 2 days i.v. 47.3 Compound 314 50 mg/kg t.i.d x 2days p.o. 18.9 25 Compound 311 50 mg/kg t.i.d x 2 days s.c. 80.2 Compound 311 50 mg/kg t.i.d x 2 days p.o. 42.5 Compound 311 50 mg/kg t.i.d x 2 days s.c. 49.3 two times a day (0 and 8 hrs) 2 three times a day (0, 8 and 16 hrs) 106 WO 01/00206 PCT/US00/18079 The compounds of the present invention may also be further characterized in other in vivo assays, such as the eosioophil accumulation model tested in the rat. Fifty pg of Compound 48/80 was injected into the pleural cavities of male Sprague Dawley rats. After 24 hrs, each cavity was washed twice with Hank's Balanced Salt Solution containing 0.2% EDTA. Total cell and 5 eosinophil counts were determined. Test compounds were given intraveneously, orally or subcutaneously, b.i.d. at 0 and 8 hours. The dosage, route of administration and inhibitory effect for the test compounds are shown in Table 6. TABLE 6 Dosage/Regimen Route % Inhibition of Eosinophil Infiltration Compound 90 3 mg/kg 2days i.v. 25.4 10 mg/kg 2 days i.v. 46.7 30 mg/kg 2days i.v. 83.7 10 Compound 90 50 mg/kg 2days p.o. 50.5 Compound 80 50 mg/kg 2days s.c. 65.3 Compound 92 50 mg/kg 2days s.c. 43.1 Compound 95 50 mg/kg 2days s.c. 40.9 Mouse Bio-Assay Method 15 A compound was dissolved or suspended with an appropriate solvent at 1 mg/mL. Female Balb/c mice (7-9 weeks old) were given the compound orally. Blood samples were collected from the postcaval vein of the anesthetized mice after fifteen minutes. Serum was prepared and stored at -20 oC. Serum concentration of the compound was determined from inhibitory activities of the diluted serum by a direct binding assay using VLA-4-expressing cells and VCAM-IgG fusion 20 protein. Serum concentration determined by this method correlated well with the concentration determined by LC/MS/MS methodologies. The dosage, route of administration and resulting inhibitory effect for the test compounds are shown in Table 7. TABLE 7 Dosage Minutes Post- Serum Concentration Administration (ng/ml) Compound 58 50 mg/kg 30 3614 25 Compound 68 10 mg/kg 15 261 Compound 78 10 mg/kg 15 368 Compound 79 10 mg/kg 15 618 107 WO 01/00206 PCT/US00/18079 Compound 80 10 mg/kg 15 693 Compound 90 10 mg/kg 15 3659 Compound 91 10 mg/kg 15 2523 Compound 92 10 mg/kg 15 2162 5 Compound 96 10 mg/kg 15 3514 Compound 97 10 mg/kg 15 1733 Compound 98 10 mg/kg 15 2796 Compound 102 10 mg/kg 15 503 Compound 124 10 mg/kg 15 841 10 Compound 134 10 mg/kg 15 224 Compound 146 10 mg/kg 15 527 Compound 156 10 mg/kg 15 285 Compound 158 10 mg/kg 15 301 Compound 166 10 mg/kg 15 360 15 Compound 179 10 mg/kg 15 428 Compound 309 10 mg/kg 15 669 Compound 311 10 mg/kg 15 467 Compound 314 50 mg/kg 30 2309 Compound 318 50 mg/kg 30 2105 20 Compound 319 50 mg/kg 15 603 Compound 323 50 mg/kg 15 1423 Pharmakokinetic Evaluations Pharmacokinetic parameters of exemplary compounds, in mouse, rat and monkey models, are shown in Tables 8, 9 and 10. 25 TABLE 8 MOUSE Dosage AUC 1
MRT
2
CL
3 (ng-h/mL) (hr) (mL/min/kg) Compound 68 10 mg/kg (i.v.) 1595 (0-6 hr) 0.2 104.5 1595 (0-o) 0.2 104.5 20 mg/kg (p.o.) 1307 (0-6 hr) 1.6 637.6 108 WO 01/00206 PCT/US00/18079 1751 (0-o) 4.1 476.0 Compound 90 10 mg/kg (i.v.) 8995 (0-6 hr) 0.5 18.53 9219 (0-) 0.7 18.08 10 mg/kg (p.o.) 2540 (0-6 hr) 1.2 65.62 2950 (0-) 2.5 56.50 Compound 80 10 mg/kg (i.v.) 5259 (0-6 hr) 0.4 31.69 5389 (0-m) 0.6 30.93 20 mg/kg (p.o.) 2190 (0-6 hr) 2.3 152.24 3615 (0-o) 6.5 92.21 total area under the plasma concentration (measured by LC/MS/MS method) versus time curve 2 mean residence time 5 apparent plasma clearance TABLE 9 RAT Dosage AUC' MRT 2
CL
3 (ng.h/mL) (hr) (mL/min/kg) Compound 90 10 mg/kg (i.v.) 31915 (0-6 hr) 0.8 5.23 33488 (0-o) 1.1 4.98 10 mg/kg (p.o.) 11454 (0-6 hr) 1.7 17.85 19968 (0-o) 8.8 9.99 Compound 79 10 mg/kg (i.v.) 5259 (0-6 hr) 0.4 31.69 5389 (0-c) 0.6 30.93 10 mg/kg (p.o.) 22119 (0-6 hr) 0.5 8.5 24867 (0-c) 0.7 6.7 10 Compound 68 10 mg/kg (i.v.) 6345 (0-6 hr) 0.63 26.45 6405 (0-o) 0.67 26.20 20 mg/kg (p.o.) 1867 (0-6 hr) 1.1 181.1 2086 (0-o) 2.0 162.7 total area under the plasma concentration (measured by LC/MS/MS method) versus time curve 2 mean residence time 3 apparent plasma clearance Pharmnnacokinetic parameters and the time course of the serum concentration of a single 109 WO 01/00206 PCT/US00/18079 intravenous dosage (2mg/kg) of a representative compound and ATENOLOL {4-[2'-hydroxy-3' isopropylamino) proppyxy]phenylacetamide; Sigma Chemical Co., code no. A-7655}are summarized in Tables 10 and 11 for the monkey model. TABLE 10 5 MONKEY AUCI (ng-h/mL) Cltot 2 (mL/min/kg) Compound 195 8405 4.0 ATENOLOL 5021 6.7 1 total area under the plasma concentration (measured by the LC/MS/MS method) versus time curve 10 2 apparent plasma clearance TABLE 11 TIME 5 min 30 min I hr 2 hr 4 hr 8 hr Serum conc I (ng/ml) Compound 195 65447 3900 2225 772 194 28 15 Serum conc 1 (ng/ml) ATENOLOL 4057 1189 771 479 348 137 measured by the LC/MS/MS method Binding assay of VCAM-1 to VLA-4 expressing cells Preparation of VCAM IgG fusion protein 20 A VCAM IgG fusion protein containing the one to seven immunoglobulin domains of VCAM-1 (D1D7) ligated to the hinge (H), CH2 and CH3 regions of human IgG1 was used in the binding assay. Construction of a stable cell line expressing D 1D7-VCAM IgG fusion protein An Epstein-Barr virus based, episomal plasmid containing a DID7-VCAM IgG fusion 25 gene under transcriptional control of the CMV promoter, was transfected into 293E human embryonic kidney cells. Stably transfected cells were selected using 250 pg/mL hygromycin in DMEM with 10% fetal calf serum. The cells secreted D 1D7 VCAM IgG fusion protein into the medium cumulatively for up to 9 days. Purification of D 1D7 VCAM IgG fusion protein 30 The cells were cultured in DMEM with 10% fetal calf serum for 2 days, then changed to CCM5 medium and cultured for a further 10 days. The medium was centrifuged, filtered and then 110 WO 01/00206 PCT/US00/18079 incubated overnight with Protein A Sepharose 4. The Protein A Sepharose was washed extensively and the D 1D7 VCAM IgG fusion protein bound was eluted using 100 mM citric acid, pH 3. Preparation of europium labeled-DID7 VCAM IgG fusion protein The D1D7-VCAM IgG fusion protein, at 1 mg/mL, was dialyzed against 50 mM 5 NaHCO 3 , 0.9% NaC1, pH 8.5. The fusion protein was added to one vial of europium-labeling reagent (DELFIA labeling kit from Wallac, Gaithersberg, MD; catalog no. 1244-302) and incubated at room temperature in the dark overnight. The labeled protein was purified using a Sepharose G10 column and assayed for the europium content and protein concentration. The protein was stored at minus 80 0 C until used. 10 Construction of cell line expressing VLA-4 (CHO/VLA-4) A CHO cell line stably transfected with the cDNA of a4 and P 1 was used in the binding assay. The gene for human (x4 was obtained from the American Type Culture Collection and recloned between the Xhol and Xba sites of the mammalian expression vector pCI-neo (Promega, Madison, WI). The P 1 gene was amplified by PCR from human peripheral leukocyte cDNA and 15 engineered such that the start codon was placed in the context of a consensus Kozak sequence. The gene was recloned into pCI-neo downstream of the CMV promoter and chimeric intron. CHO-K1 cells were stably co-transfected with plasmids encoding the aX4 and 131P genes, and single cells expressing high levels of VLA-4 were selected by fluorescence cell sorting (FACS). The antibodies used in FACS analysis were: anti-a4-PE conjugated (PharMingen, San 20 Diego, CA) and anti-P 1-FITC conjugated (Biosource, Camarillo, CA). A cell line 4B4, which expresses 400,000 and 300,000 sites/cell of the a4 and 31 subunit, respectively, was used in the binding assay. The subunit numbers were determined by FACS analysis, using Quantum Simply Cellular microbeads (Flow Cytometry Standards Corporation, Puerto Rico) as standards. The cells were maintained in F12 medium, containing 10% fetal bovine serum, 10 mM HEPES, pH 7.5, 0.5 25 mg/mL G418, using a 1:48 passage/week. Binding Assay The CHO/VLA-4 cells were seeded in a 96-well microtiter plate at 30,000 cells/well and incubated at 37oC, 5% CO 2 for 48 hours until confluent. On the day of assay, the wells were emptied and washed twice with 350 Al of a washing buffer containing 25 mM HEPES, pH 7.5, 150 30 mM NaC1, 1 mM MgC 2 , 111 WO 01/00206 PCT/US00/18079 1 mM CaCl2, 2 mM MnCl 2 . The plate was then drained and blotted dry on paper towels to remove buffer. The test compound was serially diluted in assay buffer (washing buffer together with 0.1% bovine serum albumin, 20 jM DTPA and 1% dimethysulfoxide), in the presence of 2 nM of 5 europium-labeled D1D7-VCAM IgG fusion protein. Final concentrations used ranged from 0.1 nM-10 gM. 50 p aliquot of the test compound mixture was added to duplicate wells in the plate. Control wells for total binding received no test compound. Non-specific binding wells contained an anti-x4 monoclonal antibody (L25.3, Becton Dickinson, Bedford, MA). The cells were allowed to incubate with the test compound mixture, in the presence of 10 europium-labeled D1D7-VCAM IgG fusion protein at room temperature for at least 30 minutes. The cells were then washed three times with 350 Ml of washing buffer, using a Skatron plate washer and blot dry. An 100 /l aliquot of DELFIA Enhancement solution was added to each well, followed by gentle agitation at room temperature for 10 minutes. The amount of europium-labeled VCAM-IgG fusion protein bound was determined by time-resolved fluorescence measurement 15 (Model: Victor
T
, Wallac Inc., Gaithersberg, MD). Percent binding was calculated as: [(FT-FNs) - (FI-FNs)] / (FT-FNs) x 100 wherein FT and FNs is the fluorescence signal of the europium labeled D 1D7-VCAM IgG fusion protein bound to cells, in the absence of test compound and containing an anti-a4 monoclonal antibody, respectively. F, is the fluorescence in wells containing a test compound. The IC 50 (concentration 20 of the inhibitor to inhibit 50% binding of VACM to CHO/VLA-4 cell) was determined by a curve fitting routine, PRIZM (GraphPad Software, Inc., San Diego, CA). Adhesion of VLA-4 expressing cell to VCAM-1 This secondary functional assay was used to determine the potency of a test compound in inhibiting VLA-4 mediated cell adhesion. 25 Preparation of VCAM coated plate A 50 Ml aliquot of the D 1D7-VCAM IgG fusion protein (1.5 gg/mL in phosphate buffered saline, PBS) was added to each well of a 96-well Costar flat bottom plate (Costar, Franklin Lakes, NJ, catalog no. 2580). The plate was then incubated overnight at 4oC. On the day of assay, the wells were emptied and washed twice with 350 /l of PBS. The plate was then blocked with 100 /l 112 WO 01/00206 PCT/US00/18079 of 1% bovine serum albumin (BSA, Sigma, cat# A9418) in PBS at room temperature for at least a hour. Cell preparation Jurkat cell (clone E6-1) was obtained from American Type Cultured Collection and was 5 maintained in RPMI medium, 10 mM HEPES, pH 7.5, 1 mM sodium pyruvate, 10% FCS, using a 1:64 passage/week. Just prior to running the assay, Jurkat cells were labeled with 5 4M of calcein AM (Molecular Probe, Eugene, OR, catalog no. C1430) in RPMI medium, at room temperature for 30 min in the dark. Following labeling, cells were washed twice with RPMI medium and resupended at 1 x 106 cells/mL. 10 Cell adhesion assay Immediately before the assay, the BSA solution was emptied from the VCAM-coated plate. The plate was then washed twice with RPMI medium. A 100 p1 aliquot of the labeled Jurkat cells was added to each well, followed by the addition of 50 pl of the inhibitor solutions. Final inhibitor concentrations range from 1 nM to 10 pM and each concentration was tested in 15 triplicates. The inhibitor and cells were allowed to incubate at room temp for 1 hr in the dark. Following the incubation, the plate was immersed gently into a container filled with PBS, then inverted face down under PBS. The wells were drained and blotted dry on a layer of paper towel. A 50 pl aliquot of 0.1% Triton X-100 was added to each well. The plate was incubated in the dark for 10 min. Adhesion of Jurkat cell was quantitated in a Millipore Cytofluor 2300 System plate 20 reader set at 485 nM excitation and 530 nM emission. The IC 5 o (concentration of the inhibitor to inhibit 50% Jurkat cell adhesion) was determined by a curve fitting routine, PRIZM (GraphPad Software, Inc., San Diego, CA). Methods of Synthesis Compounds of the present invention may be prepared by standard chemical synthesis 25 methods, as well as by methods of combinatorial chemistry, such as that described in Published PCT application, WO 95/30642. Synthetic Examples General methods of synthesis are illustrated by the following examples. The specific embodiments are presented by way of illustration only, and are not intended to limit the invention. 30 Modifications and variations in any given material or process step will be readily apparent to one 113 WO 01/00206 PCT/US00/18079 of skill in the art. Unless otherwise indicated, the solid-phase support used in certain examples is Tentagel'-S-PHB resin. This resin has a para-hydroxy benzyl linker which can be cleaved by the use of 90% trifluoroacetic acid in dichloromethane. The loading for this resin varies between 0.27 and 0.30 mmol/g and is not double loaded. 5 Example 1 I N Boc{ A A three-necked 500 mL round-bottomed flask was charged with 200 mL of THF and NaH (1.5 g, 62.9 mmol). A solution of 1-vinyl-2-pyrrolidinone (6.9 g, 62.9 mmol) and methyl 3 iodobenzoate (15.0 g, 57.3 mmol) in THF (100 mL) was added dropwise to the flask over 15 min. 10 After the addition was complete the reaction mixture was heated to reflux for 1 hr. The reaction vessel was allowed to cool to room temp and then 6 N HCI (100 mL) was carefully added. The reaction was concentrated in vacuo to remove the THF and then an additional aliquot of 6 N HCI (100 mL) was added and the reaction was refluxed for 14 hr. The reaction was quenched by the addition of NaHCO 3 until pH 9 and then the mixture was extracted 3x with EtOAc. The combined 15 organics were dried over MgSO 4 and concentrated in vacuo to afford a yellow oil. This oil was then placed in MeOH (100 mL) and cooled to minus 78 oC. NaBH 4 (3.5 g, 96.5) was then added portionwise and the reaction was allowed to warm to room temp over 2 hr. The reaction was quenched by the addition of 6 N HCI until acidic and then made basic by the addition of 40% aqueous NaOH. The solution was extracted 3x with CH 2 C12, the combined 20 organics were dried over MgSO 4 , and then concentrated in vacuo to afford 11.4 g as a yellow oil. The above amine was then Boc-protected by placing the amine in 50% dioxane:H 2 0 (100 mL) and adding K 2
CO
3 until basic. To this solution was added Boc-anhydride (9.1 g, 41 mmol) and then allowed to stir for 14 hr at room temp. The reaction was quenched by the addition of 1 N HCI until acidic. The solution was extracted 3x with EtOAc, dried over MgSO 4 and concentrated 25 in vacuo to afford a yellow viscous oil. The oil was chromatographed (25% EtOAc:hexanes) to afford 7.0 g A. Br \/Boc A A' Hydrochloric acid (gas) was bubbled through 15.8 g (73.5 mmol) 2-bromophenylacetic 114 WO 01/00206 PCT/US00/18079 acid in 100 mL of methanol for 10 min. The resulting solution was partitioned between 100mL water and 100 mL CH 2 C1. The organic layer was dried over MgSo 4 and the solvent was removed under reduced pressure to give 16.8 g (73.5 mmol) of methyl-2-brompophenylacetate which was combined with 9.0 g (80.8 mmol) of 1-vinyl-2-pyrrolidinone, and 100 mL of dry THF under argon 5 in a 250 mL round-bottomed flask. To this flask was added 3.5 g (147 (mmol) sodium hydride (95%) and the solution was stirred for 10 min at room temp. A reflux condenser was added and the mixture was heated to reflux for 1 hr. The solution was cooled to room temp and the solvent was removed under reduced pressure. A solution of 30 mL aqueous hydrochloric acid and 50 mL water was added to the 10 resultant mixture and was heated to reflux with no condenser until the solution temperature reached 96 0 C at which time a condenser was added and the solution was allowed to reflux for 16 hr. The solution was cooled to room temp, made basic with 150 mL of an aqueous solution of 40% sodium hydroxide, extracted with 3 x 125 mL CH 2 C1 2 , and the combined organic layers were washed with a saturated aqueous solution of sodium chloride, dried over magnesium sulfate, and 15 the solvent was removed under educed pressure to give 15.0 g (63.0 mmol, 86%) of 2-(2 bromobenzyl)-1-pyrolline. To a solution of 15 g (63 mmol) of 2-(2-bromobenzyl)-l-pyrolline in a solution of 80:20 methanol:aqueous acetic acid cooled to minus 78 0 C was added, in portions over a 15 min period, 5.3 g (140.0 mmol) sodium borohydride. The mixture was allowed to stir for 1 hr warming to 20 room temp at which time the solvent was removed under reduced pressure, 150 mL of water was added and the solution was made basic with an aqueous solution of sodium hydroxide which was extracted 10 x 100 mL CH 2 C1 2 which resulted in emulsions. The combined organic layers were washed with a saturated aqueous solution of sodium bicarbonate, dried over magnesium sulfate and the solvent was removed in vacuo to give 14.6 g (60.8 mmol, 97%) of the benzyl proline. 25 To a solution of 14.6 g( 60.8 mmol) of the benzyl proline in a solution of 70 mL of saturated aqueous sodium bicarbonate and 70 mL dioxane was added 15.1 g (67.0 mmol) of di-t butyl-dicarbonate and the mixture was stirred for 16 hr at room temp. The solution was then partitioned between a 200 mL aqueous solution of hydrochloric acid and 200 mL ethyl acetate. The ethyl acetate layer was washed with 200 mL of a saturated aqueous solution of sodium 30 chloride, dried over MgSO 4 and the solvent was removed under reduced pressure to give a residue which was purified by flash column chromatography (20%-100% ethyl acetate/hexane) to give 11.5 g (33.8 mol, 56%) pure A'. 115 WO 01/00206 PCT/US00/18079 OH Fmoc 1 A (1.88 g, 5.0 mmol) was placed in a 100 mL round-bottomed flask and dissolved in DMF (50 mL). To this solution was added Pd(OAc) 2 (23 mg, 0.3 mmol), P(o-Tol) , (12 mg, 0.3 mmol), methyl acrylate (0.47 g, 5.5 mnmol), and NaOAc (0.5 g, 5.5 mmol). This mixture was then 5 heated to 80 oC for 14 hr. The reaction mixture was then cooled to room temp and 1 N HCI (100 mL) was added. The solution was then extracted 3x with EtOAc, dried over MgSO 4 , and then concentrated in vacuo to afford a brown oil. This oil was chromatographed with 25% EtOAc:hexanes to afford 1.32 g of the alkene ester as a colorless viscous oil. The alkene ester (1.32 g, 4.1 mmol) was then subjected to hydrogenation. The alkene was 10 placed in a Parr hydrogenation bottle, EtOAc (10 mL) and 10% Pd/C (100 mg) was added under inert atmosphere. The bottle was then pressurized with hydrogen at 45 psi and shaken for 4 hr at room temp. The solution was then filtered through celite and concentrated in vacuo to afford 1.29 g of the alkane ester. The alkane ester (1.29 g, 4.0 mmol) was dissolved in THF (30 mL), MeOH (20mL), and 15 water (10 mL) and saponified with LiOH (200 mg, 8.0 mmol). The reaction was stirred at room temp for 3 hr and then poured into 1 N HCI (50 mL). This solution was then extracted 3x with EtOAc, dried over MgSO 4 , and then concentrated in vacuo to afford 1.02 g of the alkane acid as a yellow solid. The alkane acid (1.02 g, 3.3 mmol) was then deprotected by the addition of a 25% 20 TFA/CH 2 C1 2 solution and stirred for 2 hr at room temp. The resulting mixture was then concentrated in vacuo and immediately protected by dissolving the deprotected acid in 50% dioxane/water, adding K 2
CO
3 (1.2 g), and Fmoc-Cl (1.08 g, 4.0 nmmol). This mixture was stirred at room temp for 14 hr and then poured in 1 N HCI (100 mL). The solution was then extracted 3x with EtOAc, dried over MgSO 4 , and then concentrated in vacuo to afford 495 mng 1 as a white 25 crystalline solid. 116 WO 01/00206 PCT/US00/18079 HON o HN-/ 0 1' The dried resin (500 mg, 0.14 mmol) was placed into a small shaker vessel. The vessel was then charged with 9 mL of DMF (2 mg, 0.42 mmol), DIC (102 mg, 0.84 mmol), and DMAP (17 mg, 0.14 mmol). The vessel was subsequently shaken for 16 hr at room temp. The contents 5 were drained and the resin was washed 3x with DMF, MeOH, and CHC1 2 . The Fmoc group was then removed by the addition of 10 mL of 50% piperidine/DMF to the shaker vessel and shaking for 2 hr at room temp. The resulting amine resin was washed 3x with DMF, MeOH, and CH 2
CI
2 . To the above resin was added 9 mL of DMF, 4-[N'-(o-Tolylurea)]-phenylacetic acid (132 mg, 0.42 mmol), PyBroP (196 mg, 0.42mmol), and DIEA (107 mg, 0.84 mmol). The contents 10 were shaken for 14 hr at room temp and then drained and washed 3x with DMF, MeOH, and
CH
2 C1 2 . The compound was then cleaved from the resin and the filtrate was collected and then concentrated in vacuo. The resulting oil was triturated by taking up the oil in MeOH and slowly adding in Et 2 0O until a precipitate formed. This precipitate was collected and dried in vacuo to afford 67 mg 1' as a white crystalline solid. 15 Example 2 TMS N Boc 2 A (4.2 g, 11.3 mmol) was placed in a 100 mL round-bottomed flask and dissolved in NEt 3 (50 mL). To this solution was added Pd(PPh 3
)
2 C1 2 (0.16 g, 0.23 mmol), CuI (21 mg, 0.12 mmol), and trimethylsilylacetylene (1.38 g, 13.5 mmol). This mixture was stirred at room temp for 14 hr. 20 The reaction mixture was quenched by the addition of 1 N HCI (100 mL). The solution was then extracted 3x with EtOAc, dried over MgSO 4 , and then concentrated in vacuo to afford a yellow oil. This oil was chromatographed with 15% EtOAc:hexanes to afford 3.8 g 2 as a colorless viscous oil. HO 0 N Fmoc 3 117 WO 01/00206 PCT/US00/18079 A solution of dicyclohexylborane was generated by the addition of borane-THF (12.0 mL, 12 mmol), at 0oC to a solution of cyclohexene (2.3 mL) in 6 mL of anhydrous THF. This solution was stirred for an additional 1 hr at 0oC. The acetylene (2) (2.0 g, 5.84 mmol) was then added dropwise over 15 min at o0 C and then allowed to warm to room temp over 1 hr. The reaction 5 mixture was then diluted with MeOH (20 mL) and then recooled to 0oC. A solution of 2 N NaOH (6 mL) and 30% H 2 0 2 (3.5 mL) was then added dropwise. The reaction mixture was then stirred at 0 0 C for 1 hr and then warmed to 40 0 C for 2.5 hr. The mixture was then cooled to room temp and an additional 6 mL of 2 N NaOH was added. The organics were removed in vacuo and the remaining aqueous solution was extracted 3x Et 2 O and the organics were discarded. The aqueous 10 extracts were then acidified with 1 N HCI and extracted with EtOAc dried over MgSO 4 , and then concentrated in vacuo to afford 1.7 g of the phenylacetic acid as a tan crystalline solid. The acid (1.7 g, 5.6 mmol) was then deprotected by the addition of a 25% TFA/CH 2 C1 2 solution and stirred for 2 hr at room temp. The resulting mixture was then concentrated in vacuo and immediately protected by dissolving the deprotected acid in 50% dioxane/water, adding K 2 CO3 15 (15g), and Fmoc-Cl (1.4 g, 5.5 mmol). This mixture was stirred at room temp for 14 hr and then poured in 1 N HCI (100 mL). The solution was then extracted 3x with EtOAc, dried over MgSO 4 , and then concentrated in vacuo to afford 1.7 g 3 as a white crystalline solid. N HO 3' The dried resin (500 mg, 0.14 mmol) was placed into a small shaker vessel. The vessel 20 was then charged with 9 mL of DMF, 3 (180 mg, 0.42 mmol), DIC (102 mg, 0.84 mmol), and DMAP (17 mg, 0.14 mmol). The vessel was subsequently shaken for 16 hr at room temp. The contents were drained and the resin was washed 3x with DMF, MeOH, and CH 2 C1 2 . The Fmoc group was then removed by the addition of 10 mL of 50% piperidine/DMF to the shaker vessel and shaking for 2 hr at room temp. The resulting amine resin was washed 3x with DMF, MeOH, and 25 CH 2 C1 2 . To the above resin was added 9 mL of DMF, 4-[N'-(o-Tolylurea)]-phenylacetic acid (132 mg, 0.42 mmol), PyBroP (196 mg, 0.42 mmol), and DIEA (107 mg, 0.84mmol). The contents were shaken for 14 hr at room temp and then drained and washed 3x with DMF, MeOH, and
CH
2 C1 2 . The compound was then cleaved from the resin and the filtrate was collected and then 118 WO 01/00206 PCT/US00/18079 concentrated in vacuo. The resulting oil was triturated by taking up the oil in MeOH and slowly adding in Et 2 0 until a precipitate formed. This precipitate was collected and dried in vacuo to afford 52 mg 3' as a white crystalline material. Example 3 0 HO N 5 Fmoc 4 The acetylene (2) was deprotected by placing 2 (0.75 g, 2.1 mmol) in MeOH (25 mL) and adding to this solution KOH (1.4 g). The resulting solution was stirred at room temp for 1 hr. The reaction mixture was then concentrated in vacuo and acidified with 1 N HC1. The resulting aqueous solution was extracted 3x EtOAc, the combined organics were dried over 10 MgSO 4 , and then concentrated in vacuo to afford 0.56 g of the deprotected acetylene as a brown oil. The deprotected acetylene (0.56 g, 2.0 mmol) was then placed into THF (50 mL) and cooled to -78 0 C. LiHMDS (1 M soln, 4.7 mL) was then added dropwise and the reaction was stirred for 30 min. CO 2 gas was then bubbled through the reaction mixture for 15 min and the 15 reaction was then poured onto CO 2 solid. The reaction was quenched by the addition of 1 N HCI (100 mL) and the aqueous solution was extracted 3x EtOAc, the combined organics were dried over MgSO 4 , and then concentrated in vacuo to afford the propionic acid (0.71 g) as a white solid. The alkane acid (0.71 g ) was then deprotected by the addition of a 25% TFA/CH 2
C
2 solution and stirred for 2 hr at room temp. The resulting mixture was then concentrated in vacuo 20 and immediately protected by dissolving the deprotected acid in 50% dioxane/water, adding K 2
CO
3 (15 g), and Fmoc-Cl (1.29 g, 4.9 mmol). This mixture was stirred at room temp for 14 hr and then poured in 1 N HC1 (100 mIL). The solution was then extracted 3x with EtOAc, dried over MgSO 4 , and then concentrated in vacuo to afford 4 as a brown oil. The oil was then chromatographed with 5% MeOH/ dichloromethane to afford 110 mg of the desired compound. HO N -~0 25 0 4' 119 WO 01/00206 PCT/US00/18079 The dried resin (500 mg, 0.14 mmol) was placed into a small shaker vessel. The vessel was then charged with 9 mL of DMF, 4 (184 mg, 0.42 mmol), DIC (102 mg, 0.84 mmol), and DMAP (17 mg, 0.14 mmol). The vessel was subsequently shaken for 16 hr at room temp. The contents were drained and the resin was washed 3x with DMF, MeOH, and CH 2
CI
2 . The Fmoc 5 group was then removed by the addition of 10 mL of 50% piperidine/DMF to the shaker vessel and shaking for 2 hr at room temp. The resulting amine resin was washed 3x with DMF, MeOH, and
CH
2 C1 2 . To the above resin was added 9 mL of DMF, 4-[N'-(o-Tolylurea)]-phenylacetic acid (132 mg, 0.42 mmol), PyBroP (196 mg, 0.42 mmol), and DIEA (107mg, 0.82 mmol). The contents 10 were shaken for 14 hr at room temp and then drained and washed 3x with DMF, MeOH, and
CH
2
C
2 . The compound was then cleaved from the resin and the filtrate was collected and then concentrated in vacuo. The resulting oil was triturated by taking up the oil in MeOH and slowly adding in Et 2 0 until a precipitate formed. This precipitate was collected and dried in vacuo to afford 27 mg 4' as a white crystalline material. 15 Example 4 HO N Fmoc 5 The iodide (A) (0.5 g, 1.3 mmol) was placed into THF (20 mL) and cooled to minus 78 0 C. Butyllithium (2.21 mL, 1.6M soln) was added dropwise and then the cooling bath was removed and gaseous CO 2 was bubbled through for 10 min. The reaction mixture was poured onto 20 dry ice and then 1 M HCI (100 mL) was added. The mixture was extracted 3x EtOAc, the combined organics were dried over MgSO 4 , and then concentrated in vacuo to afford 0.32 g of the benzoic acid as a white crystalline solid. The benzoic acid (0.32 g, 1.68 mmol) was then deprotected by the addition of a 25%
TFA/CH
2 Cl 2 solution and stirred for 2 hr at room temp. The resulting mixture was then 25 concentrated in vacuo and immediately protected by dissolving the deprotected acid in 50% dioxane/water, adding K 2
CO
3 (15 g), and Fmoc-Cl (0.44 g, 1.67 mmol). This mixture was stirred at room temp for 14 hr and then poured in 1 N HC1 (100 mL). The solution was then extracted 3x with EtOAc, dried over MgSO 4 , and then concentrated in vacuo to afford 0.38 g 5 as a white crystalline solid. 120 WO 01/00206 PCT/US00/18079 HO N 0 5' The dried resin (500 mg, 0.14 mmol) was placed into a small shaker vessel. The vessel was then charged with 9 mL of DMF, 5 (173 mg, 0.42 mmol), DIC (102 mg, 0.84 mmol), and DMAP (17 mg, 0.14 mmol). The vessel was subsequently shaken for 16 hr at room temp. The 5 contents were drained and the resin was washed 3x with DMF, MeOH, and CH 2 C1 2 . The Fmoc group was then removed by the addition of 10 mL of 50% piperidine/DMF to the shaker vessel and shaking for 2 hr at room temp. The resulting amine resin was washed 3x with DMF, MeOH, and
CH
2 C12. To the above resin was added 9 mL of DMF, 4-[N'-(o-Tolylurea)]-phenylacetic acid (132 10 mg, 0.42 mmol), PyBroP (196 mg, 0.42 mmol), and DIEA (107 mg, 0.84 mmol). The contents were shaken for 14 hr at room temp and then drained and washed 3x with DMF, MeOH, and
CH
2 Cl 2 The compound was then cleaved from the resin and the filtrate was collected and then concentrated in vacuo. The resulting oil was triturated by taking up the oil in MeOH and slowly adding in Et 2 0 until a precipitate formed. This precipitate was collected and dried in vacuo to 15 afford 51 mg 5' as a white crystalline material. Example 5 (E)-4-[2-[1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl]ethenyl] benzoic acid 0
CO
2 H 6 To a cold (minus 78 oC), stirred solution of triethyl 4-phosphonomethylbenzoate (904 mg, 20 3.01 mmol) in THF (20 mL) was added LiHMDS (1.0 M in THF, 3 mL, 3.00 mmol) and the stirring was continued for 1 hr at the same temp. N-Boc prolinal (500 mg, 2.51 mmol) in THF (10 mL) was added to this mixture and the mixture was allowed to warm to room temp for over 1 hr. After being stirred for 2 hr, the mixture was quenched by water and extracted with EtOAc. The extract was washed with brine (200 mL), dried over MgSO 4 , and evaporated. The residue was 25 chromatographed on silica-gel with n-hexane-EtOAc (8:1, v/v) as eluent to give 713 mg (82%) ethyl (E)-4-[2-[1-(tert-butoxycarbonyl)-2-pyrrolidinyl]ethenyl]benzoate as a colorless crystalline solid. mp 68-70 'C; IR (KBr) 1710, 1697, 1681 cm'; 1 H-NMR (CDCl 3 ) 81.39 (12 H, series of min), 1.77-1.93 (3 H, min), 2.11 (1 H, min), 3.47 (2 H, min), 4.34-4.54 (total 3 H, min), 6.22 (1 H, min), 6.43 (1 121 WO 01/00206 PCT/US00/18079 H, d, J = 14.2 Hz), 7.39 (2 H, J = 8.3 Hz), 7.97 (2 H, d, J = 8.3 Hz); MS (FAB) m/z 346 (M'+1); Anal. Calcd for C 2
H
27
NO
4 : C, 69.54; H, 7.88; N, 4.05. Found: C, 69.52; H, 8.08; N, 4.07. To a stirred solution of ethyl (E)-4-[2-[1-(tert-butoxycarbonyl)-2-pyrrolidinyl]ethenyl] benzoate (700 mg, 2.03 mmol) in CH 2 Cl 2 (3 mL) was added TFA (3 mL) and the resulting mixture 5 was stirred for 3 hr. The mixture was concentrated and the residue was made basic by the addition of sat. NaHCO 3 . The mixture was extracted with CHC1 3 (2 x 100 mL). The combined extracts were dried over Na 2
CO
3 and concentrated in vacuo to give 434 mg (87%) ethyl (E)-4-[2-(2 pyrrolidinyl) ethenyl]benzoate as a brown oil. 'H-NMR (CDC13) 8 1.39 (3 H, t, J= 7.3 Hz), 1.52 2.06 (4 H, series of m), 2.93-2.99 (1 H, m), 3.07-3.13 (1 H, m), 3.74 (1 H, q, J= 7.3 Hz), 4.37 (2 10 H, q, J= 7.3 Hz), 6.34 (1 H, dd, J= 15.6, 7.3 Hz), 6.54 (1 H, d, J= 15.6 Hz), 7.41 (2 H, d, J= 8.3 Hz), 7.97 (2 H, d, J= 8.3 Hz). A mixture of ethyl (E)-4-[2-(2-pyrrolidinyl)ethenyl]benzoate (434 mg, 1.77 mmol), pentafluorophenyl 4-[N'-(2-methylphenyl)ureido]phenylacetate (797 mg, 1.77 mmol), Et 3 N (0.37 mL, 2.66 mmol) in DMF (15 mL) was stirred for 15 hr. The mixture was diluted with EtOAc (300 15 mL). The solution was washed with brine (2 x 200 mL), dried over MgSO 4 , and evaporated off in vacuo. The residue was chromatographed on silica-gel with CHCl 3 -EtOAc (4:1) as eluent to give 906 mg (q.y.) ethyl (E)-4-[2-[1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl] ethenyl]benzoate as a brown oil. 'H-NMR (CDCl 3 ) 8 1.39 (3 H, t, J 7.3 Hz), 1.83-2.20 (4 H, series of m), 2.24 (3 H, d, J= 4.9 Hz), 3.63 (4 H, m), 4.36 (2 H, q, J= 7.3 Hz), 4.62 and 4.84 (total 20 1 H, m), 6.18-6.47 (2 H, m), 7.03-8.02 (14 H, series of m). A stirred mixture of ethyl (E)-4-[2-[1-[4-[N'-(2-methylphenyl)ureido] phenylacetyl]-2 pyrrolidinyl]ethenyl]benzoate (906 mg, 1.77 mmol) in 0.25 N NaOH (14 mL) and THF (14 mL) was heated under reflux for 3 days. The mixture was poured into ice-IN HCI (200 mL) and the precipitate was collected with suction. The solid was recrystallized from CHCl 3 -MeOH-n-hexane to 25 give 453 mg (53%) 6 as a light yellow crystalline powder. mp 165-168 oC; IR (KBr) 3282, 2974, 2663, 2537, 1700, 1685 cm-; 'H-NMR (DMSO-d 6 ) 8 1.74-2.12 (4 H, m), 2.24 (3 H, d, J= 4.9 Hz), 3.35-3.66 (4 H, m), 4.67-4.74 (1 H, m), 6.25-6.41 (1 H, m), 6.53 (1 H, s), 6.93 (1 H, t,J= 7.3 Hz), 7.08-7.92 (12 H, series of m), 9.00 (1 H, m), 12.87 (1 H, br s); MS (FAB) m/z 484 (M +1); Anal. Calcd for C 29
H
29
N
3 0 4 "0.5H 2 0: C, 70.71; H, 6.14; N, 8.39. Found: C, 70.46; H, 6.07; N, 8.39. 30 Example 6 122 WO 01/00206 PCT/US00/18079 4-[2-[1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl]ethyl]benzoic acid N N2H H H 7 A mixture of (E)-4-[2-[1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl] ethenyl]benzoate (200 mg, 0.414 mmol) and 5% Pd/C (200 mg) in MeOH (20 mL) was 5 hydrogenated at 1 atm for 1 hr with vigorously stirring. The mixture was filtered and the filtrate was concentrated. The residue was chromatographed on silica-gel with CHCl 3 -MeOH (4:1) as eluent to give 201 mg (q.y.) 7 as a colorless crystalline powder. mp 180-190 oC; IR (KBr) 3345, 3124, 3060, 3027, 2960, 2927, 2875, 1706, 1672 cm'; 'H-NMR (DMSO-d 6 ) 8 1.04-3.96 (total 16 H, series of min), 6.91-7.41 (9 H, min), 7.79-7.90 (3 H, min), 8.23 (1 H, br s), 9.31 (1 H, br s); MS 10 (FAB) m/z 486 (M*+1); Anal. Calcd for C 29
H
31
N
3 0 4 2.25H 2 0: C, 66.21; H, 6.80; N, 7.99. Found: C, 65.97; H, 6.20; N, 7.72. Example 7 (S)-4-[2-[ 1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]pyrrolidinyl]methoxy]benzoic acid N N OCCO2H H H 8 15 (S)-4-[2-[ 1-[3-methoxy-4-(N'-phenylureido)phenylacetyl]pyrrolidinyl]methoxy]benzoic acid N- O
--
CO 2 H H H 9 (S)-4-[2-[1-[4-[N-(2-chlorophenyl)ureidolphenylacetyl]pyrrolidinyl]methoxy]benzoic acid N NO -/ -\ -CO 2H IH H 10 8,9 and 10 To a stirred mixture of Boc-prolinol (3.00 g, 14.9 mmol), ethyl p 20 hydroxybenzoate (2.40 g, 14.5 mmol), and triphenylphosphine (3.91 g, 14.9 mol) in THF (80 mL) was added dropwise diethyl azodicarboxylate (2.86 g, 16.4 mmol) at room temp. After the addition was completed, the resulting mixture was heated under reflux for 2 hr. After cooling, the mixture was concentrated in vacuo. The residue was dissolved in EtOAc and washed successively with 1 N NaOH, water, brine. The EtOAc layer was dried over MgSO 4 and evaporated in vacuo. 25 The residue was purified by column chromatography on silica gel with EtOAc-n- hexane (1:4, v/v) as eluent to give 4.88 g (93 %) ethyl (S)-4-(1-tert-butoxycarbonyl-2-pyrrolidinyl)methoxybenzoate 123 WO 01/00206 PCT/US00/18079 as an oil. To the above ethyl (S)-4-(1-tert-butoxycarbonyl-2-pyrrolidinyl)methoxybenzoate was added MeOH (100 mL) and 1 N NaOH (50 mL). The mixture was stirred for 15 hr at room temp. After removal of MeOH under a reduced pressure, water (50 mL) was added to the residual 5 solution. The aqueous solution was washed with Et 2 O (x2 ) and then acidified by the addition of 1 N HCL. The mixture was extracted with EtOAc., washed with water, brine, dried over MgSO 4 and evaporated in vacuo to afford 4.26 g (95 %) (S)-4-(1-tert-butoxycarbonyl-2-pyrrolidinyl) methoxybenzoic acid as a crystalline solid. To the above (S)-4-(1-tert-butoxycarbonyl-2-pyrrolidinyl)methoxybenzoic acid was added 10 CH 2
C
2 (10 mL) and TFA (10 mL). The mixture was stirred at room temp for 1 hr Et 2 O was added to the mixture and resulting solid was collected. The solid was dissolved in water (100 mL), dioxane (50 mL) and NaHCO 3 (4.4 g). Fmoc-Cl (3.34 g, 12.9 nmmnol) was added to the solution, and the resulting mixture was stirred for 20 hr at room temp. The mixture was washed with Et20 (x2) and aqueous layer was separated. The layer was acidified by the addition of 1 N HC1. The 15 mixture was extracted with EtOAc. The extract was washed with water, brine, dried over MgSO 4 and evaporated in vacuo to afford 5.36 g (91%) (S)-4-(1-Fmoc-2-pyrrolidinyl)methoxybenzoic acid as a viscous oil, which was crystallized on standing. Wang resin (0.71 mmol/g, 400 mg) was suspended in a solution of (S)-4-(1-Fmoc-2 pyrrolidinyl) methoxybenzoic acid (500 mg, 1.13 mol), DMAP (35 mg, 0.29 mmol), HOBt (40 mg, 20 0.30 mmol) and DIC (0.45 mL, 2.9 mmol) in a mixture of DMF (3 mL) and CH 2 C1 2 (7 mL). The mixture was shaken for 20 hr and drained. The resin was washed with DMF (x3), MeOH (x3),
CH
2 C1 2 (x3) and dried under a reduced pressure to give 522 mg of resin, which was used to prepare 8, 9 and 10. 8 To the above resin (115 mg) was added a solution of piperidine-DMF 25 (50 % v/v, 4 mL) and the mixture was shaken for 1 hr. The resin was washed with DMF (x3), MeOH (x3), CH 2 C1 2 (x3). To the resin was added DMF (4 mL), CH 2 C1 2 (2 mL), 4-[N'-(2 methylphenyl)ureido] phenylacetic acid (70 mg, 0.25 mmol), PyBrop (115 mg, 0.25 mmol) and DIEA (0.13 mL, 0.75 mmol). The mixture was shaken for 21 hr and drained. The resin was washed with DMF (x3), MeOH (x3), CH 2 C1 2 (x3). To the resin was added a solution of TFA in 30 CH 2 C1 2 (50 % v/v, 4 mL) and the mixture was shaken for 2 hr. The mixture was filtered and the 124 WO 01/00206 PCT/US00/18079 filtrate was concentrated in vacuo. The residue was purified by Sep-Pak column. After removal of the solvent, Et 2 0 was added to the residue and resulting solid was collected to afford 25 mg 8 as a pale yellow crystalline material. MS (FAB) m/z 488 (M +1) 9 To the above resin (60 mg) was added a solution of piperidine in DMF (50 % 5 v/v, 3 mL) and the mixture was shaken for 2 hr. The resin was washed with DMF (x3), MeOH (x3), CH 2 C1 2 (x3). To the resin was added DMF (2 mL), CH 2 C1 2 (1 mL) 3-methoxy-4-(N' phenylureido) phenylacetic acid (40 mg, 0.13 mmol), PyBrop (60 mg, 0.13 mmol) and DIEA (0.060 mL, 0.34 mmol). The mixture was shaken for 40 hr and drained. The resin was washed with DMF (x3), MeOH (x3), CH 2
C
2 (x3). To the resin was added a solution of TFA in CH 2 C1 2 (30 10 % v/v, 3 mL) and the mixture was shaken for 5 hr. The mixture was filtered and the filtrate was concentrated in vacuo. The residue was purified by Sep-Pak column. After removal of the solvent, Et 2 O was added to the residue and the solid was collected to afford 8 mg 9 as a crystalline solid. MS (FAB) m/z 504 (M'+1) 10 To the above resin (637 mg) was added a solution of piperidine in DMF (50 % 15 v/v, 20 mL) and the mixture was shaken for 4 hr. The resin was washed with DMF (x3), MeOH (x3), CH 2 C1 2 (x3). To the resin was added DMF (12 mL), CH 2 C1 2 (8 mL), 4-(Fmoc amino)phenylacetic acid (530 mg, 1.42 mmol), PyBrop (660 mg, 1.43 mmol) and DIEA (0.62 mL, 3.56 mmol). The mixture was shaken for 60 hr and drained. The resin was washed with DMF (x3), MeOH (x3), CH 2 C1 2 (x3) and dried under a reduced pressure to afford 617 mg of the resin. 57 mg 20 of this resin was added Piperidine in DMF (40 % v/v, 2 mL). The mixture was shaken for 1 hr. The resin was washed with DMF (x3), MeOH (x3), CH 2 Cl 2 (x3). 2-chlorophenyl isocyanate (0.050 mL, 0.41 mmol) was added to a suspension of resin in THF (1 mL) and CH 2 Cl 2 (1 mL). The mixture was shaken for 20 hr and drained. The resin was washed with DMF (x3), MeOH (x3),
CH
2 Cl 2 (x3). To the resin was added a solution of TFA in CH 2 C1 2 (25 % v/v, 2 mL) and the 25 mixture was shaken for 1.5 hr. The mixture was filtered and the filtrate was concentrated in vacuo. The residue was purified by Sep-Pak column. After removal of the solvent, Et 2 O was added to the residue and the solid was collected to afford 2 mg 10 as a crystalline solid. MS (FAB) m/z 508 (M++I) Example 8 30 (S)-3-[2-[1-[3-methoxy-4-(N'-phenylureido)phenylacetyl]pyrrolidinyl]methoxy] phenylacetic acid 125 WO 01/00206 PCT/US00/18079 N O _
CO
2 H N 'K N H H 11 (S)-3-[2-[1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]pyrrolidinyl]methoxy] phenylacetic acid N N O CO2 H H H 12 11 and 12 To a stirred mixture of Boc-prolinol (3.51 g, 17.5 mmol), methyl 5 m-hydroxyphenylacetate (2.90 g, 17.5 mmol), triphenylphosphine (4.60 g, 17.6 mol) in THF (50 mL) was added dropwise diethyl azodicarboxylate (3.05 g, 17.5 mmol) at room temp. After the addition was completed, the mixture was heated under reflux for 3 hr. After cooling, the mixture was concentrated in vacuo. The residue was dissolved in EtOAc, washed successively with 1 N NaOH, water, brine and dried over MgSO 4 . After removal of the solvent, the residue was purified 10 by column chromatography on silica-gel with EtOAc- hexane (1:4, v/v) as eluent to give 5.49 g (90 %) methyl (S)-3-(1-tert-butoxycarbonyl-2-pyrrolidinyl) methoxyphenylacetate as an oil. A mixture of the above methyl (S)-3-(1-tert-butoxycarbonyl-2-pyrrolidinyl) methoxy phenylacetate in MeOH (60 mL) and 1 N NaOH (20 mL) was stirred for 8 hr at room temp. After removal of the solvent under a reduced pressure, water (50 mL) was added to the residue. The 15 mixture was extracted with Et 2 O (x2), and the aqueous layer was acidified by the addition of 1 N HCL. The mixture was extracted with EtOAc. The extract was washed with water, brine, dried over MgSO 4 and then concentrated in vacuo to afford 4.43 g (88 %) (S)-3-(1-tert-butoxycarbonyl -2 pyrrolidinyl)methoxy phenylacetic acid as a viscous oil. A mixture of the above (S)-3-(1-tert-butoxycarbonyl-2-pyrrolidinyl) methoxyphenyl acetic 20 acid in CH 2 C1 2 (10 mL) and TFA (10 mL) was stirred for 1 hr at room temp. Et 2 O was added to the mixture and allowed to stand. Upper layer was removed by decantation to give an oil. A mixture of this oil in water (100 mL), dioxane (30 mL) and NaHCO 3 (6.0 g) was added Fmoc-Cl (2.86 g, 11.1 mmol) and the mixture was stirred for 20 hr at room temp. The mixture was extracted with Et 2 0 (x2), and the aqueous layer was acidified by the addition of 1 N HCL. The mixture was extracted 25 with EtOAc. The extract was washed with water, brine, dried over MgSO 4 and concentrated in vacuo to afford 5.08 g (81 %) (S)-3-(1-Fmoc-2-pyrrolidinyl)methoxyphenylacetic acid as a viscous oil. 126 WO 01/00206 PCT/US00/18079 Wang resin (0.71 mmol/g, 400 mg) was suspended in a solution of (S)-3-(1-Fmoc-2 pyrrolidinyl) methoxyphenylacetic acid (520 mg, 1.14 mol), DMAP (35 mg, 0.29 mmol), HOBt (40 mg, 0.30 mmol) and DIC (0.45 mL, 2.9 mmol) in a mixture of DMF (3 mL) and CH 2
C
2 (7 mL). The mixture was shaken for 20 hr and drained. The resin was washed with DMF (x3), MeOH 5 (x3), CH 2 C1 2 (x3) and dried under a reduced pressure to give 593 mg of resin, which was used for the preparation of 11 and 12. 11 A mixture of the above resin (70 mg) in piperidine-DMF (40 % v/v, 3 mL) was shaken for 1 hr. The resin was washed with DMF (x3), MeOH (x3), CH 2 Cl 2 (x3). To the resin was added DMF (1.5 mL), CH 2 C1 2 (1.5 mL) 3-methoxy-4-(N'-phenylureido) phenylacetic acid (42 mg, 10 0.14 mmol), PyBrop (70 mg, 0.15 mmol) and DIEA (0.065 mL, 0.37 mmol). The mixture was shaken for 15 hr and drained. The resin was washed with DMF (x3), MeOH (x3), CH 2
CI
2 (x3). To the resin was added a solution of TFA in CH 2 C1 2 (25 % v/v, 2 mL) and the mixture was shaken for 3 hr. The mixture was filtered and the filtrate was concentrated in vacuo. The residue was purified by Sep-Pak column. After removal of the solvent, Et 2 0O was added to the residue and the solid was 15 collected to afford 8 mg 11 as a crystalline solid. MS (FAB) m/z 518 (M +1) 12 A mixture of the above resin (70 mg) in piperidine - DMF (40 % v/v, 3 mL) was shaken for 1 hr. The resin was washed with DMF (x3), MeOH (x3), CH 2
CI
2 (x3). To the resin was added DMF (1.5 mL), CH 2 C1 2 (1.5 mL), 4-[N'-(2-methylphenyl)ureido] phenylacetic acid (40 mg, 0.14 mmol), PyBrop (70 mg, 0.15 mmol) and DIEA (0.065 mL, 0.37 mmol). The mixture was 20 shaken for 15 hr and drained. The resin was washed with DMF (x3), MeOH (x3), CH 2 Cl 2 (x3). A mixture of the resin in TFA-CH 2 Cl 2 (25 % v/v, 2 mL) was shaken for 3 hr. The mixture was filtered and the filtrate was concentrated in vacuo. The residue was purified by Sep-Pak column. After removal of the solvent, Et 2 0 was added to the residue and the solid was collected to afford 11 mg 12 as a crystalline solid. MS (FAB) m/z 502 (M+1) 25 Example 9 4-[2-[1-[3-methoxy-4-(N'-phenylureido)phenylacetyl]-2-pyrrolidinyl]ethynyl] benzoic acid NN H Hj ! "N COOH 13 127 WO 01/00206 PCT/US00/18079 To a stirred cold (minus 50 oC) solution of N-boc-prolinal (5.98 g, 30 mmol) and PPh 3 (62.95 g, 240 mmol) in CH 2 C12 (200 mL) was slowly added a solution of CBr 4 (39.80 g, 120 mmol) in CH 2 C1 2 (50 mL), and the stirring was continued for 1 hr at 0 'C. To this mixture was added sat. NaHCO 3 and the mixture was extracted with CHCl 3 . The extract was washed with H 2 0, dried over 5 MgSO 4 , and evaporated. The residue was chromatographed on silica-gel with CHC1 3 and n hexane-AcOEt (4:1, v/v) as eluent to give 7.84 g (74%) 1-(tert-butoxycarbonyl)-2-(2,2 dibromoethenyl) pyrrolidine as colorless plates. mp 61-63; IR (KBr) 1693 cm-'; 'H-NMR (CDCl 3 ) 8 1.46 (9 H, s), 1.72-2.19 (4H, m), 3.35-3.45 (2H, m),4.35 (1H, br s), 6.36 (1H, br s); MS (FAB) m/z 352, 354, 356, 358;Anal. Calcd for CH 1 7
NO
2 Br 2 : C, 37.21; H, 4.83; N, 3.94. Found: C, 10 37.14; H, 4.83; N, 4.00. To a stirred cold (minus 78 oC) solution of 1-(tert-butoxycarbonyl)-2-(2,2 dibromoethenyl) pyrrolidine (7.81 g, 22 mmol) in THF (200 mL) was added n-BuLi (1.59 M in hexane, 28 mL, 44 mmol) over 10 min, and the stirring was continued for 2 hr at the same temp. The reaction was quenched by the addition of sat. NH 4 Cl and extracted with EtOAc. The extract 15 was washed with brine, dried over MgSO 4 , and evaporated. The residue was chromatographed on silica-gel with n-hexane-AcOEt (10:1, v/v) as eluent to give 4.15 g (97%) 1-(tert-butoxycarbonyl) 2-ethynyl pyrrolidine as a light yellow oil. 'H-NMR (CDCl 3 ) 8 1.48 (9 H, s), 1.82-2.21 (4 H, m), 3.30-3.45 (2 H, m), 4.41-4.52 (1 H, m). A suspension of ethyl 4-iodobenzoate (1.7 mL, 10 mmol), Pd(PPh 3
)
4 (578 mg, 0.5 mmol), 20 and CuI (190 mnmol, 1 mmol) in i-Pr 2 NH (20 mL) was stirred for 0.5 hr under N 2 . To this mixture was added a solution of 1-(tert-butoxycarbonyl)-2-ethynylpyrrolidine (1.95 g, 10 mmol) in i-Pr 2 NH (20 mL) for over 10 min. After stirring for 3 hr at room temp, the mixture was poured into 1H120 and extracted with EtOAc. The extract was washed with brine, dried over MgSO 4 , and evaporated. The residue was chromatographed on silica-gel with n-hexane-AcOEt (10:1, v/v) as eluent to give 25 2.77 g (81%) 1-(tert-butyloxycarbonyl)-2-(2-(4-ethoxycarbonylphenyl)ethynyl) pyrrolidine as a colorless oil. 1 H-NMR (CDC13) 8 1.37 (3 H, t, J=6.8 Hz), 1.49 (9 H, s), 1.85-2.12 (4 H, m), 3.37 3.51 (2 H, m), 4.37 (2 H, q, J=6.8 Hz), 4.54-4.77 (1 H, m), 7.44 (2 H, d, J=7.8 Hz), 7.96 (2 H, d, J=7.8 Hz) To a stirred solution of 1-(tert-butoxycarbonyl)-2-[2-(4-ethoxycarbonylphenyl) ethynyl] 30 pyrrolidine (2.75 g, 8 mmol) in CH 2 C12 (5 mL) was added TFA (5 mL), and the resulting mixture was stirred overnight. The mixture was concentrated in vacuo and made basic with sat. NaHCO 3 128 WO 01/00206 PCT/US00/18079 and extracted with CHC1 3 . The extract was washed with brine, dried over MgSO 4 , evaporated to give 1.95 g (q.y.) 2-[2-(4-ethoxycarbonylphenyl) ethynyl]pyrrolidine as a light yellow oil.' H-NMR (CDCl 3 ) 8 1.38 (3 H, t, J=6.8 Hz), 1.82-2.16 (4 H, m), 3.01-3.48 (2 H, m), 4.00-4.11 (1 H, m), 4.37 (2H, q, J=6.8 Hz), 4.54-4.77 (1 H, m), 7.44-7.46 (2 H, m), 7.95-7.97 (2 H, m). 5 A mixture of 3-methoxy-4-(N'-phenylureido)phenylacetic acid (180 mg, 0.6 mmol), 2-(2 (4-ethoxycarbonylphenyl)ethynyl)pyrrolidine (146 mg, 0.6 mmol), EDC (173 mg, 0.9 mmol), DMAP (73 mg, 0.6 mmol), and cat. HOBt in DMF (10 mL) was stirred overnight. The mixture was poured into 1 N HCI and the solid was collected with suction. The residue was dissolved in
CHCI
3 and dried over MgSO 4 . After removal of the solvent, the residue was chromatographed on 10 silica-gel with CHCl 3 -MeOH (10:1, v/v) as eluent to give 192 mg (61%) ethyl 4-[2-[1-[3-methoxy 4-(N'-phenylureido)phenylacetyl]-2-pyrrolidinyl] ethynyl]benzoate as a light yellow amorphous solid. 'H-NMR (CDCl 3 ) 8 1.38 (3 H, t, J=6.8 Hz), 1.98-2.24 (4 H, m), 3.48-3.89 (2 H, m), 3.53 (2 H, s), 3.62 (3 H, s), 4.33-4.40 (2 H, m), 4.78-5.04 (1 H, m), 6.77-8.00 (14 H, m). To a stirred solution of ethyl 4-[2-[1-[3-methoxy-4-(N'-phenylureido) phenylacetyl]-2 15 pyrrolidinyl] ethynyl]benzoate (184 mg, 0.35 mmol) in THF (5 mL) was added 0.25 N NaOH (4 mL). The resulting mixture was stirred overnight. The mixture was poured into H 2 0 and made acidic by the addition of 1 N HCI (1 mL). The solid was collected with suction and dissolved in CHC1 3 . The solution was dried over MgSO 4 and evaporated. The residue was recrystallized from CHCl 3 -n-hexane to give 65 mg (37%) 13 as a white crystalline powder. mp 154-157; IR (KBr) 20 3346, 2952, 2615, 1712, 1693cm'; 'H-NMR (CDCl 3 ) 8 1.92-2.29 (4 H, m), 3.32-3.82 (2 H, m), 3.78 (2 H, s), 3.80 (3 H, s), 4.87-5.11 (1 H, m), 6.77-9.26 (14 H, m), 13.10 (1 H, br s); MS (FAB) m/z 498 (M+1). Example 10 4-[2-[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl]lethynyl] benzoic 25 acid N N N COOH 14 A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (1.45 g, 4.6 mmol), 2-(2-(4-ethoxycarbonylphenyl)ethynyl)pyrrolidine (1.12 g, 4.6 mmol), EDC 1.32 g (6.9 129 WO 01/00206 PCT/US00/18079 mmol), DMAP (562 mg, 4.6 mmol) in DMF (20 mL) was stirred overnight. The mixture was poured into 1 N HCI and the solid was collected with suction. The solid was dissolved in CHCl 3 and dried over MgSO 4 . After removal of the solvent, the residue was chromatographed on silica gel with CHCl 3 -MeOH (100:1, v/v) as eluent to give 2.20 g (89%) ethyl 4-[2-[1-[3-methoxy-4-(N' 5 (2-methylphenyl)ureido] phenylacetyl]-2-pyrrolidinyl]ethynyl]benzoate as a white amorphous solid. 'H-NMR (CDCI 3 ) 8 1.37-1.41 (3 H, m), 1.94-2.22 (4 H, m), 2.29 (3 H, s), 3.41-3.89 (2 H, m), 3.62 (3 H, s), 3.69 (2 H, s), 4.34-4.40 (2 H, m), 4.72-5.01 (1 H, m), 6.33 (1 H, br s), 6.80-8.06 (12 H, m). 10 To a stirred solution of ethyl 4-[2-[1-[3-methoxy-4-(N'-(2-methylphenyl)ureido]phenyl acetyl]-2-pyrrolidinyl]ethynyl]benzoate (2.16 g, 4 mmol) in THF (30 mL) was added 0.25 N NaOH (32 mL) and the stirring was continued overnight. The mixture was poured into H 2 0 and acidified by the addition of 1 N HCI (8 mL). The resulting precipitate was collected with suction and dissolved in CHCI 3 . The solution was dried over MgSO 4 and evaporated. The residue was 15 recrystallized from CHCl 3 -n-hexane to give 555 mg (27%) 14 as a white crystalline powder. mp 161-164; IR (KBr) 3338, 2954, 2875, 1707, 1691 cm-'; 'H-NMR (CDCl 3 ) 8 1.96-2.10 (4 H, m), 2.24 (3 H, s), 3.32-3.81 (2 H, m), 3.62 (2 H, s), 3.81 (3 H, s), 4.87-5.10 (1 H, m), 6.76-8.58 (13 H, m); MS (FAB) m/z 512 (M'+1) Example 11 20 4-[2-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinylethynyl] phenylacetic acid '?-N'N H H COOH 15 A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (141 mg, 0.45 mmol), 2-[2-(3-ethoxycarbonylmethylphenyl)ethynyl]pyrrolidine (116 mg, 0.45 mmol), EDC (130 25 mg, 0.68 mmol), DMAP (55 mg, 0.45 mmol), cat. HOBt in DMF (10 mL) was stirred overnight. The mixture was poured into 1 N HCI and the solid was collected by filtration. The solid was dissolved in CHCI 3 , dried over MgSO 4 , and evaporated. The residue was chromatographed on silica -gel with CHCl 3 -MeOH (100:1 v/v) as eluent to give an oil, which was dissolved in THF (5 mL). 0.25 N NaOH (4 mL) was added to this solution with stirring. After stirring overnight, the 30 mixture was poured into IN HC1 (20mL). The resulting precipitate was collected with suction and dissolved in CHCI The solution was dried over MgSO 4 and evaporated. The residue was 130 WO 01/00206 PCT/US00/18079 chromatographed on silica-gel with CHCl 3 -MeOH (5:1, v/v) as eluent to give 92 mg (39%) 15 as a white amorphous solid. 'H-NMR (CDCI) 6 1.96-2.18 (7 H, min), 3.50-3.88 (9 H, min), 4.78-4.98 (2 H, min), 6.72-7.99 (14 H, min); MS (FAB) m/z 526 (MW+1). Example 12 5 4-[ 1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinylmethoxy]isophthalic acid 0 N NOOH H H COOH 16 To a stirred solution of pentafluorophenyl ester of 4-[N'-(2-methylphenyl)ureido] phenylacetic acid (2.32 g, 5.15 mmol), dimethyl (S)-4-(2-pyrrolidinylmethoxy)isophthalate (1.51 g, 5.15 mmol) in DMF (20 mL) was added Et 3 N (1.0 mL, 6.65 mmol), and the mixture was stirred 10 overnight. The resulting mixture was diluted with EtOAc. The solution was washed with brine, dried over MgSO 4 , and evaporated off in vacuo. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (10:1, v/v) as eluent to give 1.58 g (55%) dimethyl 4-[ 1-[4-[N'-(2-methylphenyl)ureido] phenylacetyl]-2-pyrrolidinylmethoxy]isophthalate as a yellow crystalline solid. 'H-NMR (CDCl 3 ) 8 1.87-2.25 (min, total 7 H), 3.50-3.65 (min, 4 H), 3.85 15 (s, 3 H), 3.89 (s, 3 H), 4.18-4.31 (min, 2 H), 4.44 (min, 1 H), 6.95-8.45 (min, total 13 H). To a stirred solution of dimethyl 4-[1-[4-[N'-(2-methylphenyl)ureido] phenylacetyl]-2 pyrrolidinylmethoxy]isophthalate (1.56 g, 2.79 mmol) in THF (30 mnL) was added 0.25 N NaOH (20 mL), and the reaction mixture was heated under reflux overnight. The resulting mixture was poured into 1 N HCI, and solid was collected. The crude solid was washed with Et 2 0 to give 574 20 mg (39%) 16 as a yellow amorphous solid. IR (KBr) 1710 cm-'; 'H-NMR (DMSO-d 6 ) 8 1.83-2.18 (min, 4 H), 2.24 (s, 3 H), 3.36-4.28 (min, 8 H) 6.91-9.02 (series of m, total 13 H), 12.89 (br s, 1 H); MS (FAB) m/z 532 (M+1). Example 13 4-[2-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl] ethenyl] benzoic 25 acid NNN COOH 17 131 WO 01/00206 PCT/US00/18079 To a stirred solution of the 4-[2-[2-(N-tert-butoxycarbonyl)pyrrolydinylethenyl] benzonitrile (2.26g, 7.57mmol) in CH 2
CI
2 (23 mL) was added dropwise a 1.5M solution of diisopropylaluminum hydride (toluene solution) (6.06mL, 9.09mmol) at 0oC for over 15 min. The resulting solution was stirred for3 hr at 0OC. The solution was quenched by the addition of 5 sat.NH 4 C1. The resulting mixture was filtered through Celite, and the filtrate was extracted with EtOAc. The filtrate was washed with brine, dried over Na 2
SO
4 and evaporated in vacuo to afford 1.89 g (83%) 4-[2-[2-(N-tert-butoxy carbonyl)pyrrolidinyl]ethenyl] benzaldehyde as a yellow syrup. To a stirred solution of NaOH (1.00 g, 25.1 mmol) in water(10 mL) was added a solution 10 of AgNO 3 (2.13g, 12.5mmol) in CH 3 CN (10 mL) at 0oC for over 0.5 hr. To the stirred above mixture was added dropwise a solution of 4-[2-[2-(N-tert- butoxycarbonyl) pyrrolydinyl]ethenyl] benzaldehyde (1.89g, 6.27mmol) in CH 3 CN (O10mL) at 0 0 C for over 20 min. After the resulting mixture was stirred for a further 3 hr at room temp. The mixture was filtered with suction, and then washed with hot water. After the filtrate was washed with EtOAc, the aqueous layer was 15 acidified by carefully adding 1 N HCI, and then extracted with CHC 3 . The extract was dried over Na 2
SO
4 and evaporated in vacuo to afford 0.700g (35% for 2 steps) 4-[2-[2-(N-tert-butoxycarbonyl) pyrrolidinyl]ethenyl] benzoic acid as a pale yellow crystalline material. To a stirred solution of 4-[2-(2-(N-tert-butoxycarbonyl)pyrrolidinyl) ethenyl]benzoic acid (0.700g, 2.2 1mmol) in MeOH-benzene(1:4, v/v, 30mL) was added dropwise a 2 M-n-hexane 20 solution of TMSCHN 2 (1.32mL, 2.65mmol) at room temp. After the solution was stirred for 0.5 hr at room temp, the solution was evaporated in vacuo. The resulting oily residue was chromatographed on silica-gel with EtOAc-n-hexane(1:6, v/v) as eluent to afford 0.64g (88%) methyl 4-[2-[2-(N-tert-butoxycarbonyl) pyrrolidinyl] ethenyl]benzoate as a pale yellow crystalline material. 25 To a stirred solution of methyl 4-[2-[2-(N-tert-butoxycarbonyl)pyrrolidinyl] ethenyl] benzoate (0.64g, 1.93mmol) in CH 2 C1 2 (5mL) was added TFA (5mL) at room temp. After the mixture was stirred for 1 hr at room temp, the mixture was evaporated in vacuo, The residue was treated with sat. NaHCO 3 and extracted with CHC1 3 . The extract was dried over Na 2
SO
4 and evaporated in vacuo to afford 0.45g (100%) methyl 4-[2-(2-pyrrolidinyl)ethenyl]benzoate as a 30 yellow crystalline material. 132 WO 01/00206 PCT/US00/18079 To a stirred mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (285mg, 0.906mmol), methyl 4-[2-(2-pyrrolydinyl)ethenyl]benzoate (210mg, 0.906mmol) in DMF (4mL) was added 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) (209mg, 1.09mmol), 1 hydroxybenzotriazole (HOBt) (147mg, 1.09mmol) and 4-dimethylaminopyridine (DMAP) (11 mg, 5 0.0906mmol) at room temp. After the resulting mixture was stirred for 48 hr at room temp, the mixture was poured into ice-1 N HCI and extracted with EtOAc. The extract was dried over Na 2
SO
4 and evaporated in vacuo. The residue was chromatographed on silica-gel with acetone toluene (1:4 to 1:1, v/v) as eluent to afford 0.47g (98%) methyl 4-[2-[1-[3-methoxy-4-[N'-(2 methylphenyl)ureido] phenylacetyl]-2-pyrrolidinyl]ethenyl]benzoate as a white crystalline material. 10 To a stirred solution of methyl 4-[2-[1-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenyl acetyl]-2-pyrrolidinyl]ethenyl]benzoate (0.47g, 0.891mmol) in THF (5mL) was added 0.25 N NaOH (5.36mL). The resulting mixture was stirred at room temp for 20 hr. The mixture was acidified with 1N HCL. The mixture was extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 and evaporated in vacuo to afford 430mg (94%) 17 as a white crystalline 15 material. 'H-NMR (400MHz, DMSO-d 6 ) 8 1.78-2.13 (4H1, m), 2.50 (3H, s), 3..44 -3.68 (4H, m), 3.75 and 3.82 (3H, s), 4.71 (1H11, m), 6.26-8.59 (15H, m). Example 14 4-[2-[1-[3-methoxy-4-(N'-phenylureido)phenylacetyl]-2-pyrrolidinyl]ethenyl] benzoic acid N N N H H 0 COOH 18 20 To a stirred solution of 3-methoxy-4-(N'-phenylureido)phenylacetic acid (305mg, 1.0 lmmol), methyl 4-[2-(2-pyrolydinyl)ethenylJbenzoate (235mg, 1.0 lmmol) in DMF (4mL) was added 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (232mg, 1.21mmol), 1 hydroxybenzotriazole (HOBt) (164mg, 1.2 1mmol), and 4-dimethylaminopyridine (DMAP) (12mg, 0.101ommol) at room temp. After the mixture was stirred for 48 hr, the mixture was acidified by the 25 addition of 1 N HC1. The mixture was extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 and evaporated in vacuo to give an oily residue. The residue was chromatographed on silica-gel with acetone:toluene (1:4 to 1:1, v/v) as eluent to afford 0.43g (83%) methyl 4-[2-[1-[3-methoxy-4-(N'-phenylureido) phenylacetyl]-2-pyrrolidinyl]ethenyl] benzoate acid as a white crystalline material. 133 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[2-[1-[3-methoxy-4-(N'-phenylureido) phenylacetyl]-2 pyrrolidinyl]ethenyl]benzoate (0.43g, 0.837mmol) in THF (5mL) was added a solution of 0.25 N NaOH (5.04mL) at room temp. After the resulting mixture was stirred for 20 hr, the mixture was acidified by the carefully addition of 1 N HC1. The mixture was extracted with EtOAc. The extract 5 was washed with brine, dried over Na 2
SO
4 and evaporated in vacuo to afford 397mg (95%) 18 as a white crystalline material. 'H-NMR (400MHz, DMSO-d 6 ) 8 1.78-2.13 (4H, min), 3.17-3.68 (4H, min), 3.74, 3.82 (3H11, s), 4.71 (1H, min), 6.27-9.28 (16H, min). Example 15 4-[2-[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl]ethyl]benzoic acid N N H H 10 OOH 19 A mixture of 4-[2-[1-[3-methoxy-4-[N'(2-methylphenyl)ureido]phenylacetyl]-2 pyrrolidinyl] ethenyl]benzoic acid (184mg, 0.358mmol) and 5% Pd-C(368mg) in MeOH was hydrogenated in an atmospheric pressure at room temp. After the mixture was stirred for 21 hr at room temp, insoluble catalyst was filtered off and the filtrate was evaporated in vacuo. The residue 15 was chromatographed on silica-gel with MeOH-CHCl 3 (1:4 to 1:3, v/v) as eluent to afford 123mg (66%) 19 as a white crystalline material. 'H-NMR (DMSO-d 6 ) 5 1.55-2.03 (mn, 6H), 2.24 (s, 3H), 2.60 (min, 2H), 3.17-3.59 (min, 4H), 3.83 (s, 3H11), 3.97 (min, 1H), 6.61-8.57 ( min, 13H); MS (FAB) m/z 516 (M +1). Example 16 20 3-[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl] methylthiobenzoic acid N N xq S OOH 2 H "j 20 To a solution of m-iodophenol (20.0 g, 90.9 mmol) in DMF (200mL) was added 1,4 diazabicyclo [2,2,2]octane (20.4g, 181.8 mmol) and dimethylthiocarbamoyl chloride (16.9 g, 136.4 25 mmol). The resulting cloudy solution was stirred for 0.5 hr at 35 0 C and then heated at 75 0 C for 0.5 hr. After cooling, 300 mL of water was added to the mixture. The solid was collected, washed with water and dried under a reduced pressure to give 27.63 g (99%) O-m-iodophenyl dimethylthio carbamate as a pale yellow crystalline powder. IR (KBr) 1540, 1463, 1278, 1193, 1166, 1124cm"'; 'H-NMR (400 MHz, CDCl 3 ) 8 3.33 (s, 3H), 3.45 (s, 3H), 7.05 - 7.14 (mn, 2H), 7.43 (d, J= 1.9 Hz, 134 WO 01/00206 PCT/US00/18079 1H), 7.58 (dd, J = 1.0, 7.8 Hz, 1H); MS (FAB) m/z 307 (M+1); Anal. Called for CH 1 IoINOS: C, 35.19; H, 3.28; N, 4.56. Found: C, 35.23; H,3.40; N, 4.41. A solution of O-m-iodophenyl dimethylthiocarbamate (10.0 g, 32.6 mmol) in Ph20 (25mL) was heated at 230 0 C for 10 hr. After cooling, the reaction mixture was chromatographed 5 on silica-gel with n-hexane-EtOAc (5:1, v/v)as eluent to give 9.31 g (93%) S-m-iodophenyl dimethylthio carbamate as a pale yellow oil. 'H-NMR (400 MHz, CDCl 3 ) 8 3.08 (br s, 6H), 7.11 (t, J= 7.8 Hz, 1H), 7.46 (d, J= 7.3 Hz, 1H), 7.71 (d, J= 7.3 Hz, 1H), 7.85 (s, 1H); MS (FAB) m/z 307 (M'+1). To a solution of S-m-iodophenyl dimethylthiocarbamnate (5.01 g, 16.31 mmol) in MeOH 10 (20 mL) was added 28%-MeONa in MeOH (3.46 mL, 17.94mmol). The resulting mixture was stirred at room temp for 3.5 hr and then heated at 70 0 C overnight. After cooling, IN HCI was added. The solvent was removed under a reduced pressure and the residue was diluted with EtOAc. The solution was washed with H 2 0, brine, and dried over Na 2 SO4. The organic layer was concentrated under a reduced pressure. The residue was chromatographed on silica-gel with n 15 hexane-AcOEt (10:1, v/v) as eluent to afford 3.42 g (89%) m-iodothiophenol as an oil. 'H-NMR (400 MHz, CDCl 3 ) 8 3.45 (s, 1H), 6.95 (t, J= 7.8 Hz,H),7.23 (d,J= 7.8 Hz 1H), 7.48 (d, J= 7.3 Hz,1 IH), 7.64 (t, J= 1.5 Hz,1 IH); MS(EI) m/z 236(M). To a stirred solution of N-(tert-butoxycarbonyl)-2-pyrrolidinylmethanol (4.30 g, 20.0 mmol) in pyridine (40 mL) was added p-TsCl (5.72 g, 30.0 mmol). The resulting mixture was 20 stirred at room temp for 3 hr. The reaction mixture was quenched with H 2 0, and evaporated off. The residue was diluted with EtOAc and washed with IN HCI, brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the residue was chromatographed on silica-gel with n-hexane-EtOAc (2:1, v/v) as eluent to afford 5.76 g (81%) N-(tert-butoxycarbonyl)-2 pyrrolidinylmethyl p-toluenesulfonate as a colorless oil. 'H-NMR (400 MHz, CDCl 3 ) 8 1.36 and 25 1.41 (s each, total 9H), 1.82 (br m, 2H), 1.96 (br m, 2H), 2.44 (s, 3H), 3.30 (br m, 2H), 3.89 (br s, 1H), 3.96 (br s, 1H), 4.09 (br m, 1H), 7.34 (br s, 2H), 7.77 (d, J= 8.3 Hz, 2H); MS (FAB) m/z 356 (M++I). To a stirred mixture of m-iodothiophenol (2.67 g, 11.31 mmol) and N-(tert-butoxy carbonyl)-2- pyrrolidinylmethyl p-toluenesulfonate (3.34 g, 9.43 mmol) in pyridine (9.4 mL) was 30 added 8N KOH (1.77 mL). The resulting mixture was stirred at room temp overnight. The reaction 135 WO 01/00206 PCT/US00/18079 mixture was diluted with EtOAc. The solution was washed with H 2 0, sat. NH 4 Cl solution, brine, and dried over Na 2
SO
4 . The organic layer was concentrated under a reduced pressure and the residue was chromatographed on silica-gel with n-hexane-EtOAc (5:1, v/v) as eluent to afford 1.79 g (45%) [N-(tert-butoxycarbonyl)-2-pyrrolidinyl]methyl 3-iodophenyl sulfide as an oil. 'H-NMR 5 (400 MHz, CDC13) 8 1.45 (s, 9H), 1.78 - 2.01 (br m, 4H), 2.71 (dt, 1H), 3.32 - 3.49 (br m, 3H), 3.90 - 4.02 (br m, 1H), 7.12 (d, J= 7.8 Hz, 1H), 7.18 (d, J= 7.8 Hz, 1H), 7.57 (dd, J= 2.0, 8.3 Hz, 2H); MS (FAB) m/z 420 (M'+1). To a stirred solution of [1-(tert-butoxycarbonyl)-2-pyrrolidinyl]methyl 3-iodophenyl sulfide (1.76 g, 4.20 mmnol) in DMSO (20 mL) and MeOH (16 mL) was added Et 3 N (1.28 mL, 9.24 10 mmol), Pd(OAc) 2 (47.1 mg, 0.21 mmol), and 1,3-bis(diphenylphosphino)propane (86.6 mg, 0.21mmol), then CO gas was bubbled for 5 min. The resulting mixture was stirred at 700 C overnight. After cooling, the mixture was concentrated. The residue was diluted with EtOAc and washed with brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the residue was chromatographed on silica-gel with n-hexane-EtOAc (5:1, v/v) as eluent to afford 15 1.28 g (87 %) methyl 3-[1-(tert-butoxycarbonyl)-2-pyrrolidinyl]methylthiobenzoate as an oil. 'H NMvR (400 MHz, CDCl 3 ) 8 1.42 and 1.45 (each s, 9H), 1.79 - 2.05 (br m, 4H) 2.83 (dt, J= 10.8, 30.3 Hz, 1H11), 3.34 - 3.54 (br m, 3H1), 3.92 (s, 3H), 3.92 and 4.05 (d, J = 7.8 Hz, 1H), 7.36 (t, J= 7.8 Hz, 1H11), 7.63 (br d, J= 14.7 Hz, 1H), 7.83 (br d, J= 12.7 Hz, 1H11), 8.04 (s, 1H1); MS (FAB) m/z 352 (M+1). 20 To a stirred solution of methyl 3-[1-(tert-butoxycarbonyl)-2-pyrrolidinyl] methylthio benzoate (1.46 g, 4.16 mmol) in CH 2 Cl 2 (30 mL) was added TFA (15 mL) at 0oC. The resulting mixture was stirred at room temp for 1 hr. The solvent was removed under a reduced pressure and the residue was treated with 1N NaOH and extracted with CHCI 3 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated under a reduced pressure to afford 947 mg (91%) 25 methyl 3-(2-pyrrolidinyl) methylthio benzoate as a brown oil. 'H-NMR (400 MHz, CDCl 3 ) 8 1.45 1.54 (m, 1H1), 1.72 - 2.00 (m, 4H) 2.88 - 3.10 (min, 4H), 3.30 (min, 1H), 3.92 (s, 3H), 7.34 (t, J= 7.8 Hz, 1H), 7.52 (min, 1H), 7.84 (min, 1H), 8.01 (t, J= 2.0 Hz, 1H); MS (FAB) m/z 252 (M'+1). The mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (1.18 g, 3.77 mmol), EDC (1.08 g, 5.65 mmol), DMAP (23 mg, 0.19 mmol) and HOBt (25 mg, 0.19 mmol) in 30 DMF (5 mL) was stirred at room temp for lhr methyl 3-(2-pyrrolidinyl)methylthio benzoate (947 ing, 3.77 mmol) was added to the mixture and the resulting mixture was stirred overnight. After 136 WO 01/00206 PCT/US00/18079 DMAP (460 mg, 3.77 mmol) and HOBt (835 mg, 6.18 mmol) was added and stirred for a further 5 hr. The reaction mixture was diluted with EtOAc. The solution was washed with brine and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure. The residue was chromatographed on silica-gel with n-hexane-EtOAc (2:3, v/v) a eluent to afford 294.3 mg (14%) 5 methyl 3-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido ]phenylacetyl]-2-pyrrolidinyl]methylthio benzoate as a pale yellowish amorphous. IR (KBr) 2875, 1724, 1620, 1284, 1182cm'; 'H-NMR (400 MHz, CDCl 3 ) 8 1.86 - 2.05 (m, 4H), 2.31(s, 3H), 2.84 (dd, J= 9.3, 13.7 Hz, 1H), 3.43 - 3.59 (m,5H), 3.73 (s, 3H), 3.92 (s, 3H), 4.33 (m, 1H), 6.16 (s, 1H), 6.77 -6.80 (m, 2H), 7.04 (s, 1H), 7.16 (t, J= 8.3 Hz, 1H11), 7.38 (t, J= 7.8 Hz, 1H), 7.49 (t, J= 7.8 Hz, 1H), 7.73 (dt, J= 1.0, 7.8 Hz, 10 1H), 7.79 (dd, J = 2.0, 6.8 Hz, 1H), 8.01 (d, J = 2.0 Hz, 1H), 8.05 (dd, J = 2.4, 7.8 Hz, 1H); MS (FAB) m/z 548 (M+I). To a stirred solution of methyl 3-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-2-pyrrolidinyl]lmethylthiobenzoate (148.2 mg, 0.271 mmol) in THF (7.4 mL) and
H
2 0 (1.8 mL) was added LiOH (19.4 mg, 0.812 mmol), and the resulting mixture was stirred for 9 15 hr at room temp. The mixture was treated with 1N HCI and extracted with CHCl 3 . The extract was washed with brine and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the residue was purified by preparative TLC eluting with CHC1 3 -MeOH (10:1, v/v), and crystallized from n-hexane-EtOAc to afford 89.7mg (62%) 20 as a white powder. IR (KBr) 2960, 1708cm'; 'H-NMR (400 MHz, DMSO-d6) 8 1.82 - 2.01 (m, 4H), 2.24 (s, 3H), 2.93 (dd, J= 9.3, 20 12.7 Hz, 1H), 3.40 - 3.54 (m, 5H), 3.86 (s, 3H), 4.13 (br m, 1H), 6.74 (d, J= 8.3 Hz, 1H), 6.87 (d, J= 1.5 Hz, 1H), 6.94 (t, J= 7.8 Hz, 1H), 7.10 -7.17 (m, 2H), 7.42 (t, J= 7.8 Hz, 1H), 7.70 (d, J 7.8 Hz, 1H), 7.74 (d, J= 8.3 Hz, 1H), 7.79 (d, J= 7.8 Hz, 1H), 7.84 (s, 1H), 8.00 (d, J= 8.3 Hz, 1H), 8.48 (s, 1H), 8.57 (s, 1H); MS (FAB) m/z 534 (M t +1). Example 17 25 3-[[ 1-[3 -methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl]methylsulfonyl] benzoic acid HC~ NT NCOOH C O 21 To a stirred solution of methyl 3-[[1-[3-methoxy-4-[N'-(2-methyl phenyl)ureido]phenyl acetyl]-2-pyrrolidinyl]methylthio]benzoate (131.8 mg, 0.241 mmol) in CH 2 C1 2 (3 mL) was added 30 m-CPBA (130.5 mg, 0.529 mmol) at 0oC. The reaction mixture was stirred at room temp for 0.5 hr. The mixture was diluted with CHCl, and quenched with sat. Na 2
S
2 03 solution The organic 137 WO 01/00206 PCT/US00/18079 layer was separated, washed with sat. NaHCO 3 solution, brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure to afford methyl 3-[[1-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenyl acetyl]-2-pyrrolidinyl]-methylsulfonyl]benzoate as an amorphous solid. To a stirred solution of this crude compound in THF (7.4 mL) and H 2 0 (1.8 mL) was added LiOH (17.3 mg, 5 0.723 mmol), and the stirring was continued overnight at room temp. The reaction mixture was diluted with CHCl 3 and washed with IN HCI, then brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the residue was purified by preparative TLC with CHC1 3 MeOH (10:1, v/v) as eluent, and the crude solid was recrystallized from n-hexane-EtOAc to afford 69.9 mg (51%/) 21 as a white crystalline powder. mp 243 - 245; IR (KBr) 3354, 2974, 1533cm-'; 10 'H-NMR (400 MHz, DMSO-d 6 ) 8 1.80 - 2.00 (min, 4H), 2.24 (s, 3H), 3.19 - 3.62 (min, 6H), 3.82 (s, 3H), 4.18 (min, 1H), 6.67 (d, J= 8.8 Hz, 1H), 6.80 (d, J= 1.0 Hz, 1H), 6.93 (t, J= 7.3 Hz, 1H), 7.10 - 7.17 (min, 2H), 7.66 (t, J= 7.8 Hz, 1H), 7.78 (d, J= 8.3 Hz, 1H), 7.91 - 7.99 (min, 2H), 8.20 (d, J= 7.3 Hz, 1H), 8.34 (s, 1H), 8.48 (s, 1H), 8.56 (s, 1H); MS (FAB) m/z 566 (M+1); Anal. Calcd for
C
2 9HI 31
N
3 0S'HCl'lH 2 0: C, 56.17; H, 5.53; N, 6.78. Found: C, 55.92; H,5.58; N, 6.71. 15 Example 18 4-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl] methylsulfonyl] benzoic acid 0 N N H H OOH 22 To a stirred solution of methyl 4-[[1-[3-methoxy-4-[N'-(2-methyl phenyl)ureido] 20 phenylacetyl]-2-pyrrolidinyl]methylthio]benzoate (300 mg, 0.548 mmol) in CH 2 C1 2 (6 mL) was added m-CPBA (297 mg, 1.206 mmol) at 0oC, and the reaction mixture was stirred at room temp for 1 hr. The mixture was diluted with CHCl3, and quenched with sat. Na 2
S
2 0 3 solution. The separated organic layer was washed with sat. NaHCO 3 solution, brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure to afford methyl 4-[[1-[3-methoxy-4-[N'-(2 25 methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl]-methylsulfonyl]benzoate as a crude yellow oil. To a stirred solution of this crude compound in THF (4.4 mL) and H 2 0 (1. ImL) was added LiOH (39.4 mg, 1.643 mmol), and the stirring was continued overnight at room temp. The reaction mixture was diluted with CHC1 3 and washed with IN HCI, brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the residue was purified by preparative TLC 30 eluting with CHCl 3 -MeOH (10:1, v/v), and crystallized from n-hexane-EtOAc to afford 128.0 mg (41%) 22 as a white powder. IR (KBr) 3388, 2974, 1537, 1298, 1155cm-'; 'H-NMR (400 MHz, DMSO-d 6 ) 8 1.80 - 1.98 (min, 4H), 2.24 (s, 3H11), 2.54 (s, 1H), 3.20 - 3.70 (mn, 5H), 3.82 (s, 3H), 4.16 138 WO 01/00206 PCT/US00/18079 (br m, H), 6.67 (dd, J= 1.5, 8.3 Hz, 1H), 6.80 (d, J= 1.5 Hz, 1H), 6.93 (d, J 7.3 Hz, 1H), 7.10 7.16 (m, 2H), 7.78 (d, J= 7.3 Hz, 1H), 7.95 (d, J= 8.3 Hz, 2H), 7.98 (d, J= 8.3 Hz, 1H), 8.14 (d, J= 8.3 Hz, 2H), 8.49 (s, 1H), 8.57 (s, 1H); MS (FAB) m/z 566 (M'+1);Anal. Calcd for
C
29
H,
31
N
3 0 7 S-3H 2 0: C,56.21; H, 6.02; N, 6.78. Found: C, 56.76; H,5.37; N, 6.70. 5 Example 19 4-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl] methylthio] benzoic acid 0 S H H NCOOH23 23 To a stirred solution of p-iodophenol (20.0 g, 90.9 mmol) in DMF (200 mL) was added 10 1,4-diazabicyclo [2,2,2]octane (20.4g, 181.8 mmol) and dimethylthiocarbamoyl chloride (16.9 g, 136.4 mmol). The resulting solution was stirred for 3.5 hr at 75 0 C. After cooling, 300mL of water was added. The solid was collected with suction and was dissolved in EtOAc. The EtOAc layer was washed with water, dried over Na 2
SO
4 , and evaporated under a reduced pressure. The crude solid was recrystallized from H 2 0 to give the 26.75 g (96%) O-p-iodophenyl 15 dimethylthiocarbamate as a pale a yellow crystalline powder. IR (KBr) 1479, 1207, 827cm-'; 'H NMR (400 MHz, CDCI 3 ) 8 3.37 (s, 3H), 3.45 (s, 3H), 6.83 (d, J= 8.8 Hz, 2H), 7.69 (d, J= 8.3 Hz, 2H); MS (FAB) m/z 307 (M'+1); Anal. Calcd for CgHio 0 INOS: C, 35.19; H, 3.28; N, 4.56. Found: C, 35.17; H,3.35; N, 4.44. A stirred solution of O-p-iodophenyl dimethylthiocarbamate (10.0 g, 32.6 mmol) in Ph 2 0 20 (25mL) was heated at 230 0 C for 5.5 hr. After cooling, the reaction mixture was chromatographed on silica-gel with n-hexane-EtOAc (3:1, v/v) as eluent to give 2.55 g (26%) S-p-iodophenyl dimethylthio carbamate as a white crystalline powder. IR (KBr) 3299, 1651, 1469, 1371cm-'; 'H NMR (400 MHz, CDC13) 8 3.03 (br s, 3H), 3.08 (br s, 3H), 7.21 (d, J= 8.3 Hz, 2H), 7.70 (d, J= 8.3 Hz, 2H); MS (FAB) m/z 308 (MW+1); Anal. Calcd for CH,o 0 INOS: C,35.19; H,3.28; N,4.56. 25 Found: C,35.49; H,3.28; N, 4.43. To a solution of S-p-iodophenyl dimethylthiocarbamate (2.55 g, 8.31 mmol) in MeOH (10 mL) was added MeONa (495 mg, 9.14mmol), and the resulting mixture was stirred at 70'C overnight. After cooling, IN HCI was added and the mixture was concentrated under a reduced pressure. The residue was diluted with EtOAc and washed with H 2 0, brine, and dried over 30 Na 2 SO4. The organic layer was concentrated under a reduced pressure and the residue was 139 WO 01/00206 PCT/US00/18079 chromatographed on silica-gel with n-hexane EtOAc (5:1, v/v) as eluent to afford 1.75 g (89%) p iodothiophenol as a pale yellow crystalline solid. IR (KBr) 2559, 1097, 1002, 806cm-'; 'H-NMR (400 MHz, CDC13) 8 3.43 (s, 1H), 7.10 (d, J= 8.3 Hz, 2H), 7.53 (d, J= 8.3 Hz, 2H); MS (FAB) m/z 236 (M +1); Anal Calcd for C 6 Hs 5 IS: C, 30.53; H, 2.13. Found: C, 30.57; H, 2.15. 5 To a stirred mixture ofp-iodothiophenol (1.75 g, 7.43 mmol) and N-(tert butoxycarbonyl)-2- pyrrolidinylmethyl p-toluenesulfonate (2.39 g, 6.75 mmol) in pyridine (12.7 mL) was added 8N KOH (1.27 mL) at room temp, and the resulting mixture was stirred for 4 hr at the same temp. The reaction mixture was diluted with EtOAc. The solution was washed with H 2 0, sat. NIH 4 C1, brine, and dried over Na 2
SO
4 . The organic layer was concentrated under a reduced 10 pressure. The residue was chromatographed on silica-gel with n-hexane-EtOAc (5:1, v/v) as eluent to afford 1.49 g (53%) [N-(tert-butoxycarbonyl)-2-pyrrolidinyl]methyl 4-iodophenyl sulfide as a pale yellowish oil. 'H-NMR (400 MIHz, CDC13) 5 (s, 9H), 1.78 - 2.01(br m, 4H), 2.71 (dt, 1H), 3.32 - 3.49 (br m, 3H), 3.90 - 4.02 (br m, 1H), 7.12 (d, J= 7.8 Hz,1H), 7.18 (d,J= 7.8Hz, 1),7.57(dd,J= 2.0,8.3Hz,2H);MS(FAB)m/z 420(M'+1). 15 To a stirred solution of [1-(tert-butoxycarbonyl)-2-pyrrolidinyl]methyl 4-iodophenyl sulfide (1.49 g, 3.56 mmol) in DMSO (16 mL) and MeOH (13mL) was added Et 3 N (1.09 mL, 7.84 mmol), Pd(OAc) 2 (40 mg, 0.178 mmol), and 1,3-bis(diphenylphosphino)propane (73.4 mg, 0.178 mmol). To the stirred resulting mixture was induced CO gas for 5 min, and the mixture was stirred at 70 0 C overnight. After cooling, the mixture was concentrated to a small volume. The residue was 20 diluted with EtOAc, washed with brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the residue was chromatographed on silica-gel with n-hexane-EtOAc (5:1, v/v) as eluent to afford 1.16 g (93%) methyl 4-[1-(tert-butoxycarbonyl)-2 pyrrolidinyl]methylthiobenzoate as an oil. 'H-NMR (400 MHz, CDCl 3 ) 8 1.51 and 1.47 (each s, 9H), 1.78 - 2.05 (br m, 4H) 2.77 (dt, J= 10.8, 37.1 Hz, 1H), 3.34 - 3.58 (m, 3H), 3.89 (s, 3H11), 4.03 25 (br d, J= 27.3 Hz, 1H), 7.38 (d, J= 7.3 Hz, 1H), 7.47 (d, J = 7.3 Hz, 1H), 7.92 (br s, 2H); MS (FAB) m/z 352 (M'+1). To a stirred solution of methyl 4-[ 1-(tert-butoxycarbonyl)-2-pyrrolidinyl]methylthio benzoate (1.16 g, 3.32 mmol) in CH 2 C1 2 (20 mL) was added TFA (4 mL), and the mixture was stirred at room temp for 1.5 hr. The solvent was removed under a reduced pressure and the residue 30 was treated with IN NaOH. The mixture was extracted with CHC1 3 . The extract was washed with brine, dried over KOH, and concentrated under a reduced pressure to afford 767 mg (92%) methyl 140 WO 01/00206 PCT/US00/18079 4-(2-pyrrolidinyl) methylthio benzoate as a yellow oil. 'H-NMR (400 MHz, CDCl 3 ) 8 (dt, J = 3.9, 12.7 Hz, 1H), 1.85 - 2.09 (m, 2H) 2.13 (m, 1H), 3.11 - 3.27 (m, 3H), 3.40 (dd, J= 6.8, 13.2 Hz, 1H), 3.54 (dd, J= 7.3, 15.1 Hz, 1H), 3.89 (s, 3H), 5.07 (br, 1H), 7.38 (d,J= 8.3 Hz, 2H), 7.91 (d, J= 8.3 Hz, 2H); MS (FAB) m/z 252 (M t +1). 5 To a stirred mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (1.30 g, 4.136 mmol), Et 3 N (0.63 mL, 4.549 mmol) in DMF (20 mL) was added pentafluorophenyl trifluoroacetate at 0 0 C. The resulting mixture was stirred at room temp for 1 hr. The mixture was poured into water (60 mL) and precipitate was collected with suction. The crude solid was washed with 0.1 N HCI, H 2 0, n-hexane, and dried at 40 0 C to afford 1.91 g (96%) pentafluorophenyl 3 10 methoxy-4-[N'-(2-methylphenyl) ureido]phenylacetate as a pale brownish crystalline powder. IR (KBr) 1785,1224, 1216cm'; 'H-NMR (400 MHz, CDCl 3 ) 8 2.29 (s, 3H), 3.76 (s,3H), 3.90 (s, 211), 6.49 (s, 11), 6.81 (d,J= 1.5 Hz, 1H11), 6.91 (dd, J= 1.5, 8.3 Hz, 1H), 7.15 (t, J= 7.3 Hz, 3H), 7.24 (m, 1H), 7.50 (d, J= 7.8 Hz, 1H), 8.17 (d, J= 7.8 Hz, 1H1); MS (FAB) m/z 481 (M+l); Anal. Calcd for C 30
H
33
N
3 0sS-1/4H 2 0: C, 57.51; H, 3.57; N, 5.83. Found: C, 57.40; H,3.75; N, 5.68. 15 A mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetate (1.47 g, 3.05 mmol), methyl 4-(2-pyrrolidinyl)methylthiobenzoate (767 mg, 3.05 mmol), Et 3 N (0.51 mL, 3.66 mmol) in DMF (15 mL) was stirred overnight at room temp. The reaction mixture was diluted with EtOAc, washed with brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the residue was chromatographed on silica-gel with n-hexane-EtOAc 20 (1:2, v/v) as eluent to afford 1.366 g (82%) methyl 4-[[1-[3-methoxy-4-[N'-(2 methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl] methylthio]benzoate as a white crystalline powder. IR (KBr) 1785,1224, 1216cm-'; 'H-NMR (400 MHz, CDC13) 8 1.88 - 1.99 (m, 4H), 2.30 (s, 3H), 2.75 (dd, J= 9.8, 13.2 Hz, 1H), 3.43 - 3.55 (m,3H), 3.56 (s, 2H11), 3.64 (dd, J= 1.1, 14.2 Hz, 1H11), 3.73 (s, 3H11), 3.88 (s, 3H1), 4.33 (m, 1H), 6.29 (s, 1H), 6.78 -6.81 (m, 2H), 7.11 -7.26 (m, 25 5H), 7.50 (d, J= 8.3 Hz, 3H), 7.93 (d, J= 8.8 Hz, 2H), 8.07 (d, J= 7.8 Hz, 1H); MS (FAB) m/z 548 (M'+I); Anal. Calcd for C 30
H
33
N
3 0sS-1/4H 2 0: C, 65.26; H, 6.12; N, 7.61. Found: C, 65.48; H,6.20; N, 7.47. To a stirred solution of methyl 4-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-2-pyrrolidinyl]methylthio]benzoate (300 mg, 0.548 mmol) in THF (5.5 mL) and 1H120 30 (1.1 mL) was added LiOH (39.4mg, 1.643 mmol), and the reaction mixture was stirred at room temp overnight and at 50 0 C for 9 hr. The mixture was diluted with CHC 3 . The solution was 141 WO 01/00206 PCT/US00/18079 washed with IN HC1, brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure, and the obtained crude solid was recrystallized from n-hexane-EtOAc-MeOH to afford 218.6 mg (75%) 23 as a white crystalline powder. IR (KBr) 3318,2952, 1596, 1536, 1299, 1155cm'; 'H-NMR (400 MHz, DMSO-d) 8 1.82 - 2.05 (m, 4H), 2.25 (s, 3H), 2.91 (dd, J= 9.8, 5 13.2 Hz, 1H), 3.47 - 3.52 (m, 3H), 3.57 (s, 2H), 3.87 (s, 3H), 4.14 (br m, 1H), 6.76 (d, J= 1.5, 8.3 Hzd, 1H), 6.89 (d, J= 1.5 Hz, 1H), 6.94 (t, J= 7.3 Hz, 1H), 7.11 - 7.19 (m, 2H), 7.57 (d, J= 8.3 Hz, 2H), 7.80 (d, J= 8.3 Hz, 1H), 7.83 (d, J = 8.3 Hz, 2H), 8.02 (d, J= 8.3 Hz, 1H), 8.49 (s, 1H), 8.58 (s, 1H); MS (FAB) m/z 534 (M'+1); Anal. Calcd for C 29
H
3 1
N
3 0sS-5/4H 2 0: C, 62.63; H, 6.07; N, 7.36; S, 5.77. Found: C, 62.62; H,5.74; N, 7.36; S, 5.67. 10 Example 20 4-[[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl] methylsulfinyl] benzoic acid kCOOH 24 To a stirred solution of methyl 4-[[1-[3-methoxy-4-[N'-(2-methyl phenyl)ureido] 15 phenylacetyl]-2-pyrrolidinyl]methylthio]benzoate (264 mg, 0.482 mmol) in CH 2 C1 2 (5.2 mL) was added m-CPBA (118.8 mg, 0.482 mmol) at 0 0 C, and the mixture was stirred at room temp for 1 hr. The mixture was diluted with CHCl 3 , and quenched with sat. Na 2
S
2
O
3 . The separated organic layer was washed with sat. NaHCO 3 , brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure to afford methyl 4-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] 20 phenylacetyl]-2-pyrrolidinyl]methylsulfinyl] benzoate as a crude amorphous solid. To a stirred solution of this crude compound in THF (4 mL) and H 2 0 (ImL) was added LiOH (34.6 mg, 1.45 mmol), and the stirring was continued overnight at room temp. The mixture was diluted with
CHCI
3 , washed with IN HC1, brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure, and the obtained crude solid was recrystallized from n-hexane-CHCl 3 -MeOH to 25 afford 193.2 mg (73%) 24 as a white crystalline powder. IR (KBr) 3338, 2956, 1708, 1529, 1299, 1207, 1155cm-'; 'H-NMR (400 MHz, DMSO-d) 8 1.70 - 2.06 (m, 4H), 2.24 (s, 3H), 2.90 (dd, J= 8.3, 13.2 Hz, 1H), 3.02 - 3.08 (m, 1H), 3.16 - 3.25 (m, 1H), 3.41 - 3.60 (m, 3H), 3.84 (s, 3H), 4.40 (br s, 1H), 6.74 (d, J= 7.8 Hz, lIH), 6.87 (s, 1H), 6.94 (d, J = 7.3 Hz, 1H), 7.11 - 7.17 (m, 2H), 7.75 - 7.81 (m, 3H), 7.98 - 8.05 (m, 1H), 8.10 (d, J= 8.3 Hz, 2H), 8.46 (s, 1H), 8.56 (s, IH); MS 30 (FAB) m/z 550 (M++1), 572 (M +Na); Anal. Calcd for C, 29
H
3 1
N
3 0S-3/2H 2 0: C, 60.40; H, 5.94; N, 7.29. Found: C, 60.15; H,5.82; N, 6.90. 142 WO 01/00206 PCT/US00/18079 Example 21 (S)- 4-[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]lphenylacethyl]-2-pyrrolidinylmethoxy] benzoic acid. NCOOH 25 5 To a stirred solution of methyl 4-hydroxybenzoate (1.96 g, 12.88 mmol), N-Boc-prolinol (2.59 g, 12.87 mmol) and PPh 3 (4.06 g, 15.48 mmol) in THF (40 mL) was added DIAD (3.10 mL, 15.74 mmol). The resulting mixture was heated under reflux for 14 hr. The mixture was evaporated off in vacuo and the residue was purified by column chromatography on silica-gel with n-hexane-EtOAc (6:1, v/v) as eluent to give 3.34 g (77%) methyl (S)-4-[1-(tert-butoxycarbonyl)-2 10 pyrrolidinylmethoxyl benzoate as an oil. 1 H-NMR (CDCl 3 ) 8 1.48 (s, 9 H), 1.67 (d, J=9.3 Hz, 1 H), 1.87-2.03 (m, 3 H), 3.36-3.43 (m,2H), 3.87-4.09 (m,1H), 4.13-4.20 (m,2 H, 6.94 (d, J=8.3 Hz, 2H), 7.98 (d, J=8.3 Hz, 2H). A mixture of methyl (S)-4-[ 1-(tert-butoxycarbonyl)-2-pyrrolidinylmethoxy] benzoate (3.34 g, 9.96 mmnol) in TFA (20 mL) and CH 2 C1 2 (35 mniL) was stirred at room temp for 15 hr. The 15 mixture was concentrated in vacuo and made basic with sat. NaHCO 3 . The mixture was extracted with CHC1 3 , washed with brine, and dried over Na 2
CO
3 . The organic layer was evaporated to give 1.70 g (73%) methyl (S)-4-(2-pyrrolidinylmethoxy)benzoate as a yellow oil. 'H-NMR (CDC13) 5 1.54-1.61 (m, 1 H), 1.77-1.86 (m, 2 H), 1.87-1.97 (m, 1 H), 2.00 (bs, 1 H), 2.93-3.06 (m, 2 H), 3.52-3.57 (m, 1 H), 3.88 (s, 3 H), 3.90-3.99 (m, 2 H), 6.92 (d, J=9.0 Hz, 2 H), 7.98 (d, J=9.0 Hz, 2 20 H) A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (428 mg, 1.36 mmol), methyl (S)-4-(2-pyrrolidinylmethoxy)benzoate (330 mg, 1.40 mmol), EDC (312 mg, 1.63 mg), HOBt (220 mg, 1.63 mmol), and a catalytic amount of DMAP in DMF (15 mL) was stirred for 6 hr. The resulting mixture was diluted with EtOAc, washed with 0.5 N HC1, sat. NaHCO 3 , 25 brine, and dried over MgSO 4 . The solvent was evaporated off in vacuo to give an oily residue, which was purified by column chromatography on silica-gel with CHCl 3 -MeOH (50:1, v/v) as eluent to give 540 mg (75%) methyl (S)-4-[1-[3-methoxy-4-[N'-(2 methylphenyl)ureido]phenylacetyl]-2-pyrrolidinylmethoxy] benzoate as an oil. 'H-NMR (CDC13) 6 1.81-2.12 (m, 4 H) 2.88 (bs, 3 H), 3.48-3.61 (m, total 7 H), 3.88 (s, 3 H), 4.10-4.21 (m, 2 H), 4.42 30 4.46 (m, 1 H), 6.75-8.08 (series of m, total 13 H). 143 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl (S)-4-[1-[3-methoxy-4-[N'-(2-methylphenyl) ureido] phenylacetyl]-2-pyrrolidinylmethoxy]benzoate (540 mg, 1.02 mmol) in THF (10 mL) was added 0.25 N NaOH (10 mL). The resulting mixture was heated under reflux for 16 hr. The mixture was poured into 1 N HCI and the solid was collected. The crude solid was washed with Et 2 O to give 5 278 mg (53%) 25 as a white amorphous solid. IR (KBr) 1708 cm-'; 'H-NMR (DMSO-d 6 ) 5 1.83 2.14 (m, 4 H), 2.21 (s, 3 H), 2.46 (s, 2 H), 3.78 (s, 3 H), 3.95-4.02 (m, 1 H), 4.13-4.16 (m, 1 H), 4.24 (bs, 1 H), 6.51-7.98 (series of m, 12 H), 8.43 (s, 1 H), 8.53 (s, 1 H), 12.57 (bs, 1 H); MS (FAB) m/z 517 (M). Example 22 10 1-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]benzoyl]-L-prolyl]-4-piperidinylacetic acid O N N' COOH 26 A mixture of 3-methoxy-4-nitrobenzoic acid (229mg, 1.16mmol), tert-butyl 4-(1 prolylpiperidinyl)acetate (344mg, 1.16mmol), HOBt (188mg, 1.39mmol), DMAP (14.2mg, 0.11 6mmol), and EDC (267mg, 1.39mmol) in DMF (7mL) was stirred for 22 hr at room temp. 15 The mixture was diluted with EtOAc (50mL) and washed successively with 1 N HC1, sat. NaHCO 3 , and H 2 0. The organic layer was dried over Na 2
SO
4 and evaporated in vacuo. The residue was chromatographed on silica-gel with MeOH:CHCl 3 (1:30, v/v) as eluent to afford 520mg (94%) tert buty 1-(3-methoxy-4-nitrobenzoyl)prolyl-4-(1-piperidinyl)acetate as a white crystalline material. 'H-NMR (CDCl 3 ) 8 1.12-1.33 and 1.62-2.23 (each m,9H), 1.44 (s,9H), 2.65,3.13,3.47, 3.67, 4.44, 20 and 4.61 (each m, 8H), 3.99 (s, 3H), 5.05 (m, 1H), 7.21 (d, J=8.3Hz, 1H), 7.31 (s, 1H), 7.86 (d, J=8.3Hz, 1H). A stirred mixture of tert-buty 1-(3-methoxy-4-nitrobenzoyl)prolyl-4-(1-piperidinyl) acetate (0.52g, 1.09mmol) and 5% Pd-C (2.08g) in MeOH (10mL) was hydrogenated under an atmospheric pressure for 94 hr at room temp. Insoluble catalyst was removed, and the filtrate was 25 evaporated in vacuo. The residue was chromatographed on silica-gel with MeOH:CHCl 3 (1:40 to 1:6 , v/v) as eluent to afford 279mg (57%) tert-buty 1-(4-amino-3-methoxybenzoyl)prolyl-4-(1 piperidinyl)acetate as a white crystalline material. 'H-NMR (CDCl 3 ) 8 1.16-2.17, 2.69, 3.06, 3.67, 4.12, and 4.59 (each m, 17H), 3.86 (s, 3H), 5.10 (m, 1H), 6.64 (m, 1H), 7.12 (each m, 2H). 144 WO 01/00206 PCT/US00/18079 To a stirred solution of tert-buty 1-(4-amino-3-methoxybenzoyl)-L-prolyl-4-(1-piperidinyl) acetate (279mg, 0.627mmol) and Et 3 N (0.0876mL, 0.627mmol) in THF (4mL) was added dropwise o-tolyl isocyanate (0.0777mL, 0.627mmol) at room temp, and the resulting mixture was stirred for a further 21 hr at room temp. Ice water was added to the mixture and the precipitate 5 was collected with suction. The crude solid was purified by silica-gel column chromatography with MeOH:CHCI 3 (1:40, v/v) as eluent to afford 254mg (70%) tert-butyl 1-[1-[3-methoxy-4-[N'-(2 methylphenyl)ureido] benzoyl]-L-prolyl]-4-(1-piperidinyl)acetate as a crystalline solid. 1 H-NMR (CDC13) 5 1.43 (s, 9H), 1.13-1.25 and 1.76-2.14 (each m, 9H), 2.60, 3.18, 3.71, 4.06, and 4.57 (each m, 8H), 3.67 (s, 3H), 5.06 (min, 1H), 6.63, and 6.90 (s, 2H), 6.98-7.23, and 7.64 (each m, 5H), 10 7.56 (d, J=7.8Hz, 1H), 8.21 (d, J=8.8Hz, 1H). A solution of tert-butyl 1-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] benzoyl]prolyl] 4-(1-piperidinyl)acetate (254mg, 0.440mmol) in CH2C1 2 (6mL) and TFA (6mL) was stirred for 5 hr at room temp. The mixture was poured into ice water. The solid was collected with suction, washed with water and air-dried to afford 179mg (78%) 26 as a white crystalline solid. 'H-NMR ( 15 DMSO-d 6 ) 5 0.47, 1.05, 1.44, and 1.62-1.99 (each m, 9H), 2.49 (s, 3H), 2.15-2.30, 2.35, 2.56, 2.78, 3.09, 3.04-3.80, 4.05, 4.15, and 4.32 (each m, 8H), 3.92 (s, 3H), 4.92(m, 1H), 6.82, 6.95, 7.11, 7.77, 8.20, 8.57, and 8.75 (min, 9H); MS (FAB) m/z 523 (M' +1); Anal. Calcd for C 28
H
3 4
N
4 0 6 : C, 64.35; H, 6.56; N, 10.72. Found: C, 55.58; H, 5.89; N, 8.75. Example 23 20 (S)-3-methoxy-4-[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl methoxy]benzoic acid S0 Nj 1?H H COOH 27 To a stirred solution of ethyl 4-hydroxy-3-methoxybenzoate (3.00 g, 15.29 mmol), (S)-N Boc-prolinol (3.08 g, 15.30 mmol), Ph 3 P (4.81 g, 18.34 mmol) in THF (50 mL) was added DIAD 25 (3.61 mL, 18.33 mmol) at 0 0 C. The resulting mixture was heated under reflux 6.5 hr. After cooling to room temp, the mixture was evaporated and purified by column chromatography on silica-gel with CHCl 3 -MeOH (50:1, v/v) as eluent to give ethyl (S)-3-methoxy-4-[1-(tert butoxycarbonyl)-2-pyrrolidinyl methoxy] benzoate as a gum. The above ethyl (S)-3-methoxy-4-[1 (tert-butoxycarbonyl)-2-pyrrolidinylmethoxy] benzoate was dissolved in CH 2 C1 2 (50 mL) and TFA 30 (45 mL). The mixture was stirred for 2 days at room temp. The resulting mixture was 145 WO 01/00206 PCT/US00/18079 concentrated in vacuo and made basic with sat. NaHCO 3 . The mixture was extracted with CH 2 C1 2 , washed with brine, and dried over MgSO 4 . The solvent was evaporated and the residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (20:1, v/v) as eluent to give 3.27 g (77% for 2 steps) ethyl (S)-3-methoxy-4-(2-pyrrolidinylmethoxy) benzoate as a yellow oil. 5 1 H-NMR (CDC1 3 ) 8 1.39 (t, 3 H, J=7.1 Hz), 1.52-1.59 (m, 1 H), 1.76-1.88 (m, 2 H), 1.92-2.01 (m, 1H), 2.92-3.06 (m, 2 H), 3.56-3.63 (m, 1 H), 3.90 (s, 3 H), 3.91-4.02 (m, 2 H), 4.35 (q, 2 H, J=7.1 Hz), 6.89 (d, 1 H, J=8.3 Hz), 7.54 (d, 1 H, J=2.0 Hz), 7.65 (dd, 1 H, J=2.0, 8.3 Hz). To a stirred solution of ethyl (S)-3-methoxy-4-(2-pyrrolidinylmethoxy)benzoate (424 mg, 1.52 mmol) in DMF (8 mL) was added pentafluorphenyl ester of 3-methoxy-4-[N'-(2 10 methylphenyl) ureido]phenylacetic acid (728 mg, 1.52 mmol) and Et 3 N (0.26 mL, 1.87 mmol). And the resulting mixture was stirred at room temp overnight. The mixture was diluted with EtOAc, washed with 1 N HC1, sat. NaHCO 3 , brine, and dried over MgSO 4 . The solvent was evaporated off in vacuo and the residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (50:1, v/v) as eluent to give 830 mg (95%) ethyl (S)-3-methoxy-4-[1-[3-methoxy-4 15 [N'-(2-methylphenyl)ureido]phenyl acetyl]-2-pyrrolidinyl methoxy]benzoate as an amorphous solid. 'H-NMR (CDCI 3 ) 5 1.38 (t, 3 H, J=7.3 Hz), 1.88-2.20 (m, 4 H, m), 2.24 (m, 3 H), 3.44-3.50 (m, 1 H), 3.53-3.58 (m, 7 H), 3.82 (s, 3 H), 4.09-4.17 (m, 1 H), 4.22-4.25 (m, 1 H), 4.35 (q, 2 H, J=7.3 Hz), 4.38-4.49 (m, 1 H), 6.71-6.78 (mi, 1 H), 6.99 (d, 1 H, J=8.3 Hz), 7.04-7.07 (m, 1H), 7.16-7.19 (m, 2 H), 7.49-7.66 (m, 3 H), 8.06 (d, 1 H, J=8.3 Hz). 20 To a stirred solution of ethyl (S)-3-methoxy-4-[[ 1-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenylacetyl]-2-pyrrolidinyl]methoxy]benzoate (760 mg, 1.32 mmol) in THF (10 mL) was added 0.25 N NaOH (10 mL), and the resulting mixture was heated under reflux overnight. After cooling to room temp, the mixture was poured into 1 N HCI (100 mL) and the solid was collected. The crude solid was washed with Et20 to give 429 mg (59%) 27 as a yellow amorphous solid. mp 25 132-135; IR (KBr) 1707 cm-'; 'H-NMR (DMSO-d 6 ) 8 1.84-2.18 (m, 4 H), 2.25 (s, 3 H), 2.49-2.51 (m, 2 H), 3.29-3.59 (m, 4 H), 3.80 (s, 3 H), 3.82 (s, 3 H), 4.00-4.05 (m, 1 H), 6.53-8.01 (m, 10 H), 8.45 (s, 1 H), 8.54 (s, 1 H), 12.63 (bs, 1 H); MS (FAB) m/z 548 (M'+I). Example 24 (S)-4-[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinylmethoxy] phthalic 30 acid 146 WO 01/00206 PCT/US00/18079 N N 0 H H (.).,. HCOOH COOH 28 To a stirred solution of dimethyl 4-hydroxyphthalate (3.00 g, 14.27 mmol), N-Boc prolinol (2.87 g, 14.26 mmol), Ph 3 P (4.49 g, 17.12 mmol) in THF (50 mL) was added DIAD (3.40 mL, 17.27 mmol) at 0 0 C. Then the resulting mixture was heated under reflux overnight. The 5 resulting mixture was evaporated and the residue was purified by column chromatography on silica-gel with n-hexane-EtOAc (3:1, v/v) as eluent to give 5.75 g (q.y.) dimethyl (S)-4-[1-(tert butoxycarbonyl)-2-pyrrolidinyl methoxy] phthalate as an oil. 'H-NMR (CDCl 3 ) 5 1.47 (s, 9 H), 1.86-2.05 (min, 4 H), 3.36-3.40 (min, 2 H), 3.87 (min, 3 H), 3.91 (s, 3 H), 3.96-4.19 (min, 3 H), 7.03-7.24 (mn, 2 H), 7.80 (min, 1H). 10 To a solution of dimethyl (S)-4-[1-(tert-butoxycarbonyl)-2-pyrrolidinylmethoxy] phthalate (5.75 g, 14.62 mmol) in CH 2 Cl 2 (25 mL) was added TFA (20 mL), and the resulting mixture was stirred for 50 min at room temp. The resulting mixture was concentrated in vacuo and made basic with sat. NaHCO 3 . The mixture was extracted with CH 2 Cl 2 , washed with brine, dried over MgSO 4 , and evaporated off in vacuo. The residue was purified by column chromatography on silica-gel 15 with CHC13-MeOH (50:1, v/v) as eluent to give 790 mg (18%) dimethyl (S)-4-(2 pyrrolidinylmethoxy) phthalate as a brown oil. 'H-NMR (CDC13) 8 1.48-1.57 (min, 1 H), 1.72-1.84 (min, 2 H), 1.89-1.98 (min, 2 H), 2.91-3.03 (min, 2 H), 3.48-3.54 (min, 1 H), 3.82-3.97 (mn, total 8 H), 6.98 (dd, 1 H, J=2.4, 8.8 Hz), 7.06 (d, 1 H, J=2.4 Hz), 7.78 (d, 1 H, J=8.8 Hz). To a stirred solution of dimethyl (S)-4-(2-pyrrolidinylmethoxy)phthalate (212 mg, 0.72 20 mmol) in DMF (8 mL) was added pentafluorophenyl ester of the 3-methoxy-4-[N'-(2 methylphenyl)ureido] phenylacetic acid (346 mg, 0.72 mmol) and Et 3 N (120 ml, 0.86 mmol), and the mixture was stirred overnight. The resulting mixture was diluted with EtOAc, washed with 1 N HC1, sat. NaHCO 3 , brine, and dried over MgSO 4 . The solvent was evaporated off in vacuo to give 413 ming (97%) dimethyl (S)-4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2 25 pyrrolidinylmethoxy]phthalate as an oil. 'H-NMR (CDCl 3 ) 8 1.92-2.12 (mn, 4 H), 2.29 (br s, 3 H), 3.51-3.64 (min, 7 H), 3.87 (s, 3 H), 3.89 (s, 3 H), 4.10-4.19 (min, 2 H), 4.44 (mn, 1 H), 6.73-8.02 (series of m, total 12 H). To a stirred solution of dimethyl (S)-4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] 147 WO 01/00206 PCT/US00/18079 phenylacetyl]-2-pyrrolidinylmethoxy]phthalate (413 mg, 0.70 mmol) in THF (10 mL) was added 0.25 N NaOH (10 mL) at room temp, and then the resulting mixture was heated under reflux overnight. After cooling to room temp, the reaction mixture was poured into 1 N HCI (100 mL). The solid was collected, washed with water and air-dried. The crude solid was washed with Et20 to 5 give 310 mg (79%) 28 as a yellow amorphous solid. IR (KBr) 1701 cm-'; 'H-NMR (DMSO-d 6 ) 8 1.87-2.18 (m, 4 H), 2.25 (s, 3 H), 2.50 (s, 2 H), 3.38-3.60 (m, 4 H), 3.83 (s, 3 H), 4.00-4.14 (m, 1 H), 6.74-8.02 (series of m, 10 H), 8.46 (s, 1 H), 8.54 (s, 1 H); MS (FAB) m/z 562 (M+1). Example 25 3-chloro-4-[[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl]methoxy] 10 benzoic acid 0 NNj HC COOH 29 To a stirred solution of methyl 3-chloro-4-hydroxybenzoate (600 mg, 3,215mmol), N-tert butoxycalbonylprolinol (647,1 mg, 3,215 mmol), and Ph 3 P (1.01 g, 3.858 mmol) in THF (10 mL) was added dropwise diisopropyl azodicarboxylate (DIAD) (0.8 mL, 3.890 mmol) at room temp and 15 the mixture was stirred for 3days at room temp, and for 18 hr at 70 0 C. The reaction mixture was evaporated off in vacuo, and the residue was chromatographed on silica-gel with n-hexane:EtOAc (5:1, v/v) as eluent to give 1.147g (97%) methyl 3-chloro-[1-(tert-butoxycarbonyl)-2 pyrrolidinyl]methoxy benzoate as an oil. 'H-NMR (400 MHz, CDCl 3 ) 5 1.46, 1.48 (s each, 9H), 1.59 - 1.63 (br, 1H), 1.88 (br s, 1H), 2.05 (s, 1H), 2.05 - 2.21 (m, 2H), 3.34 - 3.45 (br m, 1.5H), 20 3.89 (s, 3H), 3.97(br m, 0.5H), 4.21 (br s, 2H), 7.05 (d, J= 8.8 Hz, 1H), 7.90 (dd, J= 2.0, 8.8 Hz, 1H), 8.04 (d, J = 2.0 Hz, 1H); MS (FAB) m/z 370 (M+1). To a stirred solution of methyl 3-chloro-[1-(tert-butoxycarbonyl)-2-pyrrolidinyl]methoxy benzoate (1.14 g, 3.10 mmol) in CH 2
CI
2 (20 mL) was added TFA (5 mL) at 00C, and the reaction mixture was-stirred at room temp for 2 hr. The solvent was removed under a reduced pressure and 25 the residue was treated with IN NaOH. The mixture was extracted with CHC1 3 . The extract was washed with brine, dried over KOH, and concentrated under a reduced pressure to afford 741 mg (89%) methyl 3-chloro-4-(2-pyrrolidinyl)methoxybenzoate as a yellow oil. 'H-NMR (400 MHz, CDCl 3 ) 8 1.60 - 1.67 (m, 1H), 1.78 - 2.02 (m, 3H), 2.93 - 2.98 (m, 1H), 3.03 - 3.09 (m, 1H), 3.59 (dt, J= 2.0, 9.3 Hz, 1H), 3.89(s, 3H), 3.98(dd, J= 6.3, 8.8 Hz, 1H), 4.05 (dd, J= 4.9, 9.3 Hz, 1H), 30 6.93 (d, J= 8.8 Hz, 1H), 7.90 (dd, J = 2.0, 8.8 Hz, 1H), 8.04 (d, J = 2.0 Hz, 1H); MS (FAB) m/z 270 (M+1). 148 WO 01/00206 PCT/US00/18079 The mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetate (500 mg, 1.04mmol), methyl 3-chloro-4-(2-pyrrolidinyl)methoxybenzoate (281 mg, 1.04 mmol), Et 3 N (0.17 mL, 1.25 mmol) in DMF (5 mL) was stirred for 1 hr at room temp. The mixture was diluted with EtOAc, washed with brine, and dried over Na 2
SO
4 . The solvent was 5 removed under a reduced pressure, and the residue was chromatographed on silica-gel with n hexane - EtOAc (1:3, v/v) as eluent to afford methyl 3-chloro-4-[[1-[3-methoxy-4-[N'-(2 methylphenyl)ureidolphenylacetyl]-2-pyrrolidinyl] methoxy]benzoate (620 mg, 1.04 mmol) as a white crystalline solid. To a stirred solution of this compound in THF (8 mL) and H 2 0 (2 mL) was added LiOH (74.9mg, 3.126 mmol), and the mixture was stirred at room temp for 2 days. The 10 mixture was diluted with CHCI 3 , and treated with IN HC1. The solution was washed with brine, dried over Na 2
SO
4 , and evaporated in vacuo. The crude solid was recrystallized from n-hexane EtOAc-CHC13 to afford 561.2 mg (98%) 29 as a white crystalline material. IR (KBr) 1676, 1599, 1487, 1267, 758, 754cm'; 'H-NMR (400 MHz, DMSO-d 6 ) 8 1.82 - 2.24 (m, 4H), 2.25 (s, 3H), 3.48 -3.60 (m, 4H), 3.78 (s, 3H), 4.18 (m, 2H), 4.31 (m, 1H), 6.74 (dd, J= 1.5, 8.3 Hz, 1H), 6.84 15 (d, J= 2.0 Hz, 1H), 6.91 - 6.95 (m, 1H), 7.11 - 7.17 (m, 3H), 7.79 (dd, J= 2.0, 8.3 Hz, 2H), 7.85 (d, J = 2.0 Hz, 1H), 7.98 (d, J = 8.3 Hz, 1H), 8.53(s, 1H), 8.58 (s, 1H); MS (FAB) m/z 552 (M+1). Example 26 3,5-dichloro-4-[[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl] methoxy] benzoic acid NNN 20 C COOH 30 To a stirred solution of methyl 3,5-dichloro-4-hydroxybenzoate (600mg, 2.714 mmol), N tert-butoxycarbonylprolinol (546mg, 2.714mmol), and Ph 3 P (854 mg, 3.257 mmol) in THF (10mL) was added dropwise DIAD (0.68mL, 3.283 mmol) at room temp and the mixture was stirred for 3days at room temp, and for 18 hr at 70 0 C. The reaction mixture was concentrated and the residue 25 was chromatographed on silica-gel with n-hexane - EtOAc (6:1, v/v) as eluent to give 988.8 mg (90%) methyl 4-[1-(tert-butoxycarbonyl)-2-pyrrolidinyl]methoxy-3,5-dichlorobenzoate a pale yellowish oil. 'H-NMR (400 MHz, CDCl 3 ) 8 1.44 (s, 9H), 1.88 - 2.15 (br m, 3H), 2.34 (br s, 1H), 3.40 - 3.44 (m, 2H), 3.92 (s, 3H), 3.92, 4.14 (m, 1H), 4.18 (br s, 2H), 7.98 (s, 2H); MS (FAB) m/z 404 (M+1). 30 To a stirred solution of methyl 4-[1-(tert-butoxycarbonyl)-2-pyrrolidinyl]methoxy-3,5 dichlorobenzoate (988 mg, 3.248 mmol) in CH 2 Cl 2 (20 mL) was added TFA (5 mL) at 00C, and 149 WO 01/00206 PCT/US00/18079 the reaction mixture was stirred at room temp for 2 hr. The solvent was removed under a reduced pressure and the residue was treated with IN NaOH. The solution was extracted with CHCI 3 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated under a reduced pressure to afford 672 mg (68%) methyl 3,5-dichloro-4-(2-pyrrolidinyl)methoxybenzoate as a pale yellowish 5 oil. 'H-NMR (400 MHz, CDC13) 8 1.62 - 1.69 (m, 1H11), 1.78 - 1.86 (m, 2H), 1.89 - 1.99 (m, 1H), 2.92 -2.98 (m, 1H), 3.04 -3.09 (m, 1H), 3.55 -3.60 (m, 1H), 3.91 (s, 3H), 4.01 (dd, J= 6.8, 8.8 Hz, 1H), 4.08 (dd, J = 4.9, 8.8 Hz, 1H), 7.97 (s, 2H); MS (FAB) m/z 304 (M+1). A mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetate (385.8 mg, 0.803 mmol), methyl 3,5-dichloro-4-(2-pyrrolidinyl)methoxybenzoate (244.3 mg, 0.803 10 mmol), Et 3 N (0.13 mL, 0.964 mmol) in DMF (4 mL) was stirred for 1 hr at room temp. The mixture was diluted with EtOAc, washed with brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure, and the residue was chromatographed on silica-gel with n hexane:EtOAc (1:2, v/v) as eluent to afford methyl 3,5-dichloro-4-[[1-[3-methoxy-4-[N'-(2 methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl] methoxy]benzoate as an oil. To a stirred 15 solution of this compound in THF (8 mL) and H20 (2 mL) was added LiOH (57.7 mg, 2.409 mmol), and the mixture was stirred at room temp overnight. The mixture was concentrated in vacuo, and the residue was diluted with CHC 3 . The solution was washed with IN HCI, brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure, and the obtained crude solid was recrystallized from n-hexane-MeOH-CHCl 3 to afford 428.2 mg (91%) 30 as a white 20 crystalline powder. IR (KBr) 1618, 1535, 1454, 1257, 754cm-'; 'H-NMR (400 MHz, DMSO-d) 8 1.83 - 2.24 (m, 4H), 2.24 (s, 3H), 3.50 - 3.58 (m, 4H), 3.84 (s, 3H), 3.98 - 4.05 (m, 1H), 4.15 (dd, J = 2.9, 8.7 Hz, 1H), 4.29 (br m, 1H), 6.74 (d, J= 8.3 Hz, 1H), 6.87 (s, 1H), 6.93 (t, J= 7.3 Hz, 1H), 7.11 (d, J = 7.8 Hz, 11), 7.16 (d, J= 8.3 Hz, 1H), 7.79 (d, J= 8.3 Hz,1H), 7.86 (s, 1H), 7.87 (d, J = 9.8 Hz, IH), 7.99 (d, J= 8.3 Hz, 1H), 8.49 (s, 1H), 8.58 (s, 1H); MS (FAB) m/z 586 (M*+1) 25 Example 27 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrffolidinylmethoxy]-3 nitrobenzoic acid N NqT H H 00 2 N COOH 31 To a stirred solution of 4-hydroxy-3-nitrobenzoic acid (3.00g, 0.0164mol) in MeOH 30 benzene (1:4, v/v) was added dropwise 2.0 M-n-hexane solution of TMSCHN 2 (8.2mL, 0.0164mol) at room temp. After the resulting solution was stirred for 4 hr at room temp, the mixture was 150 WO 01/00206 PCT/US00/18079 evaporated off in vacuo. The oily residue was chromatographed on silica-gel with CHC1 3 as eluent to afford 4.23g (79%) methyl 4-hydoroxy-3-nitrobenzoate as a pale yellow crystalline material. To a stirred mixture of N-tert-butoxycarbonylprolinol (1.02g, 5.07mmol), methyl 4 hydoroxy-3-nitrobenzoate (1.00g, 5.07mmol), and Ph 3 P (1.46g, 5.58mmol) in THF (O10mL) was 5 added dropwise diisopropyl azodicarboxylate (DIAD) (95%) (1.16mL, 5.58mmol) at 0oC. The resulting mixture was heated under reflux for 46 hr. After cooling, the mixture was evaporated off in vacuo. The residue was dissolved in CH 2 C1 2 (O10mL) and added TFA (O10mL). After the solution was stirred for 0.5 hr at room temp, the solution was evaporated in vacuo. Water was added to the residue and washed with EtOAc. The aqueous layer was neutralized by the addition of sat. 10 NaHCO 3 and extracted with EtOAc. The extract was dried over Na 2
SO
4 and evaporated in vacuo to afford 0.698g (49%) methyl 3-nitro-4-(2-pyrrolidinylmethoxy) benzoate as a gum. A mixture of methyl 3-nitro-4-(2-pyrrolidinylmethoxy)benzoate (0.668g, 2.38mmol), 3 methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (1.12g, 3.57mmol), 1-hydroxybenzo triazole (HOBt) (0.482g, 3.57mmol), 4-dimethylaminopyridine(DMAP) (43.6mg, 0.357mmol), 15 and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) (0.684g, 3.57mmol) in DMF (10mL) was stirred for 15 hr at room temp. EtOAc was added to the mixture and washed successively with 1 N HC1, sat. NaHCO 3 , and brine. The organic layer was dried over Na 2
SO
4 and evaporated in vacuo. The residue was chromatographed on silica-gel with EtOH-CHC13 (1:20, v/v) as eluent to afford 0.927g (68%) methyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacethyl]-2 20 pyrrolidinylmethoxy]-3-nitrobenzoate as a yellow crystalline material. A mixture of methyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacethyl]-2 pyrrolidinylmethoxy]-3-nitrobenzoate (0.917g, 1.59mmol) in THF (O10mL) and 1 N NaOH (2.38mL, 2.38mmol) was heated under reflux for 2 hr. After cooling, the mixture was poured into ice water and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 and 25 evaporated in vacuo to afford 0.826g (92%) 31 as a yellow crystalline solid. 'H-NMR (400MHz, CDC1 3 ) 8 1.91, 2.09 (1H, 3H, each m), 2.28 (3H, s), 3.54-3.62 (4H, m), 3.64 (3H, s), 4.15, 4.59 (each 1H, each d, J=7.8Hz), 4.46 (1H, m), 6.66, 7.22 (each 1H, each s), 6.72 (lH, d, J=8.3Hz), 7.11-7.28 (4H, m), 7.46 (1H, d, J=7.8Hz), 7.74 (1H, d, J=7.8Hz), 7.85 (1H, s), 8.17 (1H, dd, J=2.0, 8.8Hz), 8.48 (1H, d, J=2.4Hz); MS (FAB) nm/z 563 (M++1). 30 Example 28 3-amino-4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinylmethoxy] 151 WO 01/00206 PCT/US00/18079 benzoic acid N ',N H H
H
2 N COOH 32 A stirred mixture of 4-[ 1 -[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacethyl]-2 pyrrolidinylmethoxy]-3-nitrobenzoic acid (101mg, 0.190mmol) and 5% Pd-C (0.247g) in 5 methanol was hydrogenated at 1 atm for 48 hr. Insoluble catalyst was removed, and the filtrate was evaporated in vacuo. The residue was chromatographed on silica-gel with EtOH-CHC 3 (1:1, v/v) as eluent to afford 61.0mg (60%) 32 as a crystalline material. 'H-NMR (400MHz, DMSO-d 6 ) 8 1.95 (4H, min), 2.23 (3H, s), 3.60, 3.91, 4.10, 4.34 (5H, each min), 3.81 (3H, s), 4.88 (2H1, min), 6.74 (1H, d, J=8.3Hz), 6.86-7.28 (5H, min), 7.78 (lH, d, J=7.8Hz), 7.99 (1H, d, J=8.3Hz), 8.30 (1H, s), 10 8.45, 8.55 (each 1H, each s); MS (FAB) m/z 533 (M++1). Example 29 4-[2-[1-[4-[N'-(2-fluorophenyl)ureido)-3-methoxyphenylacetyl]-2-pyrrolidinyl]ethynyl] benzoic acid 0 N N ) N H H COOH 33 15 To a stirred solution of benzyl 4-amino-3-methoxyphenylacetate (1.36 g, 5 mmol) in THF (20 mL) was added 2-fluorophenyl isocyanate (561 ul, 5 mimol) and a catalytic amount of Et 3 N. The resulting mixture was stirred for 3 hr. The mixture was quenched by the addition of H20 (10 mL) and extracted with EtOAc. The extract was washed with brine, dried over MgSO4, and evaporated. The residue was chromatographed on silica-gel with CHC 3 as eluent to give 2.06 g 20 (q.y.) benzyl 4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenylacetate as a green oil. 'H-NMR (CDC1) 3 8 3.63 (2H1, s), 3.82 (3H, s), 5.14 (2H, s), 6.79-7.37 (12H, min), 8.01 (1H, d, J=7.8 Hz), 8.09-8.14 (1H, min). To a stirred solution of benzyl 4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenylacetate (2.04 g, 5 mmol) in THF (40 mL) was added 0.25 N NaOH (40 mL). The resulting mixture was 25 stirred overnight. The mixture was poured into 1 N HCI (10 mL), and the resulting precipitate was collected with suction. The residue was recrystallized from CHCl 3 -EtOH to give 1.04 g (66%) 4 152 WO 01/00206 PCT/US00/18079 [N'-(2-fluorophenyl)ureido]-3-methoxyphenylacetic acid as a white crystalline powder. mp 185 188 (d); 'H-NMR (DMSO-d 6 ) 8 3.50 (2H, s), 3.82 (3H, s), 6.78 (1H, dd, J=1.4 and 8.3 Hz), 6.92 (1H, d, J=1.4 Hz), 6.95-7.01 (1H, m), 7.10-7.14 (1H, m), 7.19-7.24 (1H, m), 8.01 (1H, d, J=8.3 Hz), 8.14-8.18 (lH, m), 8.72 (1H, s), 9.17 (1H, s); MS (FAB) nm/z 319 (M'+1); Anal. Calcd for 5 C, 6
H,
5
N
2 0 4 F: C, 60.37; H, 4.75; N, 8.80. Found: C, 60.20; H, 4.82; N, 8.67. A mixture of 4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenylacetic acid (255 mg, 0.8 mmol), 2-[2-(4-ethoxycarbonylphenyl)ethynyl]pyrrolidine (195 mg, 0.8 mmol), EDC (230 mg, 1.2 mmol), DMAP (98 mg, 0.8 mmol) in DMF (20 mL) was stirred overnight. The reaction mixture 10 was poured into 1 N HCI and the resulting precipitate was collected with suction and dissolve in
CHCI
3 . The solution was dried over MgSO 4 and evaporated. The residue was chromatographed on silica gel with CHCl 3 -MeOH (100:1, v/v) as eluent to give the desired compound, which was dissolved in THF (8 mL). 0.25 N NaOH (8 mL) was added to this solution and the resulting mixture was stirred overnight. The mixture was poured into 1 N HCI and extracted with CHCI 3 . 15 The extract was washed with brine, dried over MgSO 4 , and evaporated. The residue was recrystallized from CHCl 3 -n-hexane to givel44 mg (37%) 33 as a pale yellow crystalline powder. mp 152-155 (d); 'H-NMR (DMSO-d 6 ) 8 1.92-2.27 (4 H, m), 2.50 (2 H, s), 3.33-3.78 (2 H, m), 3.80 and 3.82 (total 3 H, s, each), 4.88-5.12 (1 H, m), 6.77-7.24 and 7.99-8.20 (total 7 H, m), 7.48 and 7.52 (2 H, d, J=8.3 Hz, each), 7.91 (2H, d, J=8.3 Hz), 8.72 (1H, s), 9.18 (1H, s), 13.11 (1H, br-s); 20 MS (FAB) m/z 516 (M+1); Anal. Calcd for CH 26
N
3 0sF-2H 2 0: C, 63.15; H, 5.48; N, 7.62. Found: C, 63.58; H, 5.15; N, 7.22. Example 30 4-[[ -[3-methoxy-4-[N'-(2-methylphenyl)ureidojphenylacetyl]-2-pyrrffolidinyl]methoxy]-3 methylbenzoic acid q-N N N O 25 H H OOH 34 To a stirred solution of 4-iodo-2-methylphenol (465 mg, 1.987mmol), N-tert butoxycarbonylprolinol (400 mg, 1.987 mmol), and Ph 3 P (625 mg, 2.384 mmol) in THF (7 mL) was added dropwise DIAD (0.5 mL, 2.404 mmol) at room temp, and the mixture was stirred for 13 hr at 70oC. The reaction mixture was concentrated in vacuo and the residue was chromatographed 30 on silica-gel with n-hexane - EtOAc (9:1, v/v) as eluent to give 645.3 mg (78%) 4-[1-(tert butoxycarbonyl)-2-pyrrolidinyl]methoxy-1l-iodo-3-methylbenzene as a pale yellow oil. 'H-NMR 153 WO 01/00206 PCT/US00/18079 (400 MHz, CDCl 3 ) 8 1.47 (s, 9H), 1.83 - 1.89 (m, 1H), 1.96- 2.04 (m, 3H), 2.16 (s, 3H), 3.37 3.43 (br m, 2H), 3.81, 3.94 (br m each, 1H), 4.08 -4.18 (m, 2H), 6.62 (br s, 1H), 7.42 (s, 2H); MS (FAB) m/z 418 (M+1). To a stirred solution of 4-[ 1-(tert-butoxycarbonyl)-2-pyrrolidinyl]methoxy-1-iodo-3 5 methylbenzene (645.3 mg, 1.546 mmol) in DMSO (7 mL) and MeOH (6 mL) was added Et 3 N (0.47 mL, 3.401 mmol), Pd(OAc) 2 (17.4 mg, 0.077 mmol) and 1,3-bis(diphenylphosphino) propane (31.46 mg, 0.077mmol). To the stirrede mixture was induced CO gas for 10 min. The mixture was stirred at 70 0 C for 2 days and concentrated. The residue was diluted with EtOAc, washed with brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure, and 10 the residue was chromatographed on silica-gel with n-hexane - EtOAc (5:1, v/v) as eluent to afford 301.6 mg (56 %) methyl4-[ 1-(tert-butoxycarbonyl)-2-pyrrolidinyl] methoxy-3-methylbenzoate as an oil. 'H-NMR (400 MHz, CDCl 3 ) 8 1.47 (s, 9H1), 1.86 - 2.10 (br m, 4H), 2.33 (s, 3H11), 3.32 - 3.50 (br m, 2H), 3.88 (s, 3H), 3.88, 4.04 (br m each, 1H), 4.13-4.20 (m, 2H), 6.89 (br m, 1H), 7.82 (s, 1H), 7.85 (dd, J= 2.0, 8.8 Hz, 1H1); MS (FAB) m/z 350 (M'+l). 15 To a stirred solution of methyl 4-[1-(tert-butoxycarbonyl)-2-pyrrolidinyl]methyloxy-3 methylbenzoate (301.6 mg, 0.863 mmol) in CH 2 C1 2 (6 mL) was added TFA (1.2 mL) at 0oC, and the mixture was stirred at room temp for 1 hr. The solvent was removed under a reduced pressure, and the residue was made basic by the addition of IN NaOH. The mixture was extracted with CHCl 3 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated under a reduced 20 pressure to afford 192.5 mg (90%) methyl 3-methyl-4-(2-pyrrolidinyl)methoxybenzoate as an oil. 1 H-NMR (400 MHz, CDC13) 6 1.58 - 1.65 (m, 1H), 1.78- 2.00 (m, 3H11), 2.24 (s, 3H), 2.97 (dt, J 6.8, 10.2 Hz, 1H), 3.05 (dt, J= 5.9, 6.8 Hz, 1H), 3.54 - 3.58 (m, 1H), 3.87 (s, 3H), 3.92 (dd, J= 6.3, 9.3 Hz, 1H), 3.99 (dd, J= 4.9, 9.3 Hz, 1H), 6.81(d, J= 8.3 Hz, 1H), 7.83 (s, 1H), 7.85 (dd, J 2.0, 8.3 Hz, 1H); MS (FAB) m/z 250 (M+1). 25 A mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetate (211.3 mg, 0.44 mmol), methyl 3-methyl-4-(2-pyrrolidinyl)methoxybenzoate (109.7 mg, 0.44 nmmnol), Et 3 N (73.6 ul, 0.528 mmol) in DMF (2 mL) was stirred for 1.5 hr at room temp. The reaction mixture was diluted with EtOAc. The solution was washed with brine and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure, and the residue was chromatographed 30 on silica-gel with n-hexane - EtOAc (1:3, v/v) as eluent to afford methyl 4-[[1-[3-methoxy-4-[N' (2-methylphenyl)ureido] phenylacetyl]-2-pyrrolidinyl] methoxy]-3-methyl benzoate (241.6 mg, 154 WO 01/00206 PCT/US00/18079 q.y.) as an oil. To a stirred solution of this compound in THF (4.4 mL) and HO 2 0 (1.1 mL) was added LiOH (32mg, 1.32 mmol), and the reaction mixture was stirred at room temp overnight. The mixture was diluted with CHCl 3 , and acidified by the addition of IN HCl. The solution was washed with brine and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure, and the 5 obtained crude solid was recrystallized from n-hexane-EtOAc-CHCl 3 -MeOH to afford 126.3 mg (54%) 34 as a white crystalline powder. IR (KBr) 1685, 1606, 1454, 1257, 752cm-'; 'H-NMR (400 MHz, DMSO-d6) 8 1.87 - 2.10 (min, 4H), 2.12 (s, 3H), 2.25 (s, 3H), 3.51 - 3.71 (min, 4H), 3.76 (s, 3H), 4.08 - 4.18 (min, 2H), 4.34 (min, 1H), 6.74 (dd, J= 1.5, 9.8 Hz, 1H), 6.84 (d, J= 1.5 Hz, 1H), 6.94 (t, J= 6.8 Hz, 1H), 7.06 (d, J= 8.8 Hz, 1H), 7.12 (d, J= 7.8 Hz, 1H), 7.16 (d, J= 7.8 Hz, 10 1H11), 7.72 (s,1H), 7.76 (dd, J= 2.0, 8.3 Hz, 1H), 7.79 (d, J= 7.8 Hz, 1H), 7.99 (d, J= 8.3 Hz, 1H), 8.46 (s, 1H), 8.54 (s, 1H); MS (FAB) nm/z 532 (M+ 1); Anal. Calcd for C 3 oH 3 3
N
3 0 6 -1/2H 2 0: C, 66.65; H, 6.34; N, 7.77. Found: C, 66.16; H, 6.37; N, 7.50. Example 31 (S)-4-[ 1 -[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]J-2-pyrrolidinylmethoxy] 15 isophthalic acid ,0 COOH N J N H H - OOH 35 To a solution of dimethyl 4-acetoxyisophthalate (1.52 g, 6.03 mmol) in MeOH (70 mL) was added sat. NaHCO 3 , and the resulting mixture was stirred for 3 hr at room temp. The resulting mixture was poured into 1 N HCI and extracted with EtOAc. The extract was washed 20 with sat. NaHCO 3 , brine, and dried over MgSO 4 . The solvent was evaporated to give 1.27 g (q.y.) dimethyl 4-hydroxy isophthalate as a white crystalline powder. 'H-NMR (CDCl 3 ) 8 3.91 (s, 3 H), 3.99 (s, 3 H), 7.01 (d, 1 H, J=8.8 Hz), 8.11 (dd, 1 H, J=2.4, 8.8 Hz), 8.55 (d, 1 H, J=2.4 Hz) To a stirred solution of dimethyl 4-hydroxyisophthalate (1.27 g, 6.04 mmol), (S)-N-Boc Prolinol (1.22 g, 6.06 mmol), PPh 3 (1.90 g, 7.24 mmol) in THF (30 mL) was added DIAD (1.43 25 mL, 7.26 mmol)at room temp. The resulting stirred mixture was then heated under reflux for 15 hr. After cooling to room temp, the resulting mixture was evaporated and the residue was purified by column chromatography on silica-gel with n-hexane-EtOAc (3:1, v/v) as eluent to give 2.10 g (88%) dimethyl (S)-4-[1-(tert-butoxycarbonyl)-2-pyrrolidinylmethoxy]isophthalate as a yellow oil. 'H-NMR (CDC13) 8 1.26 (s, 9 H), 1.85-2.16 (mn, 3 H), 3.36-3.46 (mn, 2 H), 3.90 (s, 6 H), 4.11-4.31 30 (min, 2 H), 4.95-5.02 (min, 2 H), 7.09 (dd, 1 H, J=9.3, 24.9 Hz), 8.11-8.14 (min, 1 H), 8.46 (d, 1 H, 155 WO 01/00206 PCT/US00/18079 J=9.3 Hz) A mixture of dimethyl (S)-4-[1-(tert-butoxycarbonyl)-2-pyrrolidinylmethoxy] isophthalate (2.01 g, 5.11 mmol), TFA (20 mL), and CH 2 Cl 2 (25 mL) was stirred for 1.5 hr at room temp. The resulting mixture was concentrated in vacuo and made basic with sat. NaHCO 3 . The mixture was 5 extracted with CH 2 C1 2 , washed with brine, dried over Na 2
CO
3 , and evaporated. The residue was purified by column chromatography on silica-gel with CHC13-MeOH (9:1, v/v) as eluent to give 0.80 g (53%) dimethyl (S)-4-(2-pyrrolidinylmethoxy)isophthalate as a yellow oil. 'H-NMR (CDC13) 8 1.71 (m, 1 H), 1.89 (m, 2 H), 2.00 (m, 1 H), 3.05-3.13 (m, 2 H), 3.67 (m, 1 H), 3.90 (s, 3 H), 3.91 (s, 3 H), 4.05-4.18 (m, 2 H), 7.00 (d, 1 H, J=8.8 Hz), 8.14 (dd, 1 H, J=2.4, 8.8 Hz), 8.50 10 (d, 1 H, J=2.4 Hz). To a stirred solution of dimethyl (S)-4-(2-pyrrolidinylmethoxy)isophthalate (616 mg, 2.10 mmol) in DMF (13 mL) was added pentafluoro ester of 3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetic acid (1.00 g, 2.08 mmol) and Et 3 N (425 pl, 3.12 mmol), and the resulting mixture was stirred for 3.5 hr at room temp. The resulting mixture was diluted with EtOAc, washed with 1 15 N HCI, sat. NaHCO 3 , brine, and dried over Na 2
SO
4 . After removal of the solvent, the residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (50:1, v/v) as eluent to give 1.41 g(q.y.) dimethyl (S)-4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2 pyrrolidinylmethoxy] isophthalate as a yellow oil. 'H-NMR (CDC1 3 ) 8 1.86-2.29 (m, 4 H), 2.30 (s, 3 H), 3.47-3.57 (m, 2 H), 3.58 (s, 3 H), 3.59 (s, 2 H), 3.83 (s, 3 H), 3.91 (s, 3 H), 4.22-4.37 (m, 2 20 H), 4.42-4.47 (m, 1 H), 6.44-8.46 (series of m, 12 H). To a stirred solution of dimethyl (S)-4-[1-[3-methoxy-4-[N'-(2 methylphenyl)ureido]phenyl acetyl] -2-pyrrolidinylmethoxy]isophthalate (1.41 g, 2.39 mmol) in THF (20 mL) was added 0.25 N NaOH (20 mL), and the resulting mixture was then heated under reflux overnight. After cooling to room temp, the mixture was poured into 1 N HCI (150 mL) and 25 the solid was collected. The crude solid was recrystallized from CHCl 3 -MeOH to give 140 mg (10%) 35 as a white crystalline powder. 'H-NMR (CDCl 3 ) 8 1.83-2.18 (m, 4 H), 2.24 (s, 3H), 3.44-3.55 (m, 4H), 3.59 (s, 2 H), 3.80 (s, 3 H), 4.05-4.24 (m, 2 H), 4.28-4.32 (m, 1 H), 6.73-8.55 (series of m, total 12 H); MS (FAB) m/z 562 (MW+1); Anal Calcd. for C 3 0
H
31
N
3 0s-4H 2 0: C, 56.87; H, 6.20; N, 6.63. Found: C, 56.73; H, 5.56: N, 6.52. 30 Example 32 3-methoxy-4-[2-[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2- pyrrolidinyl] 156 WO 01/00206 PCT/US00/18079 ethynyl] benzoic acid N NN HH COOH 36 A stirred mixture of methyl 3-methoxy-4-nitrobenzoate (1.20 g, 5.7 mmol) and 5% Pd-C (1.0g) in EtOH (30 mL) and THF (20 mL) was hydrogenated overnight at 1 atm. The mixture was 5 filtered and the filtrate was evaporated. The residue was chromatographed on silica-gel with CHCl 3 as eluent, and the solid obtained was further purified by recrystallization from CHCl 3 -n-hexane to give 805 mg (78%) methyl 4-amino-3-methoxybenzoate as white plates. mp 126-128; IR (KBr) 3475, 1700 cm'; 1H-NMR (CDCl 3 ) 8 3.86 (3H, s), 3.89 (3H, s), 4.21 (2H, br s), 6.66 (1H, d, J=8.3 Hz), 7.45 (1H, d, J=1.9 Hz), 7.54 (1H, dd, J=1.9 and 8.3 Hz); MS (FAB) nm/z 182 (M+1);Anal. 10 Calcd for CIHNO 3 : C, 59.66; H, 6.12; N, 7.73. Found: C, 59.65; H, 6.15; N, 7.65. A stirred solution of methyl 4-amino-3-methoxybenzoate (725 mg, 4 mmol) in EtOH (10 mL) was added to dil.H 2
SO
4 (prepared from H 2
SO
4 0.5 mL and H 2 0 10 mL) at 0OC. A solution of NaNO 2 (331 mg, 4.8 mmol) in H 2 0 (10 mL) was added to the mixture. After stirring for 0.5 hr at the same temp, the mixture was poured into a cooled (0OC), stirred suspended solution of KI (1.83 15 g, 11 mmol) and cat. Cu in H 2 0 (100 mL). The mixture was vigorously stirred for 1 hr at room temp and extracted with CHC 3 . The extract was washed with brine, dried over MgSO 4 , and evaporated. The residue was chromatographed on silica-gel with n-hexane-EtOAc (10:1, v/v) as eluent to give a mixture of methyl 4-iodo-3-methoxybenzoate and methyl 3-methoxybenzoate (748 mg) as a colorless oil. 20 To this oil was added Pd(PPh 3
)
4 (150 mg, 0.13 mmol), Cul (57 mg, 0.3 mmol) and i Pr 2 NH (10 mL). The mixture was stirred for 1 hr under N 2 and a solution of 1-(tert butoxycarbonyl)-2-ethynylpyrrolidine (488 mg, 2.5 mmol) in i-Pr 2 NH (10 mL) was added to the mixture. After stirring for 2 hr, the mixture was poured into H 2 0 and extracted with EtOAc. The extract was washed with brine, dried over MgSO 4 , and evaporated. The residue was 25 chromatographed on silica-gel with n-hexane-EtOAc (5:1, v/v) as eluent to give 431 mng (48%) 1 (tert-butoxycarbonyl)-2-[2-(2-methoxy-4-methoxycarbonylphenyl) ethynyl]pyrrolidine as a yellow oil. 'H-NMR (CDCl 3 ) 8 1.49 (9 H, s), 1.77-2.14 (4 H, min), 3.36-3.51 (2 H, min), 3.90 (3 H, s), 3.91 (3 H, s), 4.60-4.81 (1 H, min), 7.36-7.39 (1 H, min), 7.51 ( 1 H, s), 7.55-7.57 (1 H, min). 157 WO 01/00206 PCT/US00/18079 To a stirred solution of 1-(tert-butoxycarbonyl)-2-[2-(2-methoxy-4-methoxycarbonyl phenyl)ethynyl]pyrrolidine (395 mg, 1.1 mmol) in CH 2 C0 2 (3 mL) was added TFA (3 mL). The resulting mixture was stirred for 1 hr. The mixture was concentrated in vacuo and made basic by the addition of sat. NaHCO 3 , and extracted with CHCl 3 . The extract was washed with H 2 0, dried 5 over MgSO 4 , and evaporated to give 238 mg (84%) 2-[2-(2-methoxy-4-methoxycarbonylphenyl) ethynyl]pyrrolidine as a yellow oil. 'H-NMR (CDCl 3 ) 8 1.81-2.16 (4 H, min), 2.97-3.17 (2 H, min), 3.91 (6 H, s), 4.13-4.15 (1 H, min), 7.41 (1 H, d, J=8.3 Hz), 7.51 (1 H, s), 7.56 (1 H, d, J=8.3 Hz). A mixture of 2-[2-(2-methoxy-4-methoxycarbonylphenyl)ethynyl]pyrrolidine (233 mg, 0.9 mmol), 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (314 mg, 1 mmol), EDC (268 10 mg, 1.4 mmol), DMAP (110 mg, 0.9 mmol) in DMF (10 mL) was stirred overnight. The mixture was poured into 1 N HCI and the resulting solid was collected with suction. The solid obtained was dissolved in CHCI 3 , and the solution was dried over MgSO 4 and evaporated. The residue was subjected to short column chromatography on silica-gel with EtOAc as eluent to give an oil. The oil was dissolved in THF (5 mL) and 0.25 N NaOH was added to this solution with stirring. The 15 solution was poured into ice-1 N HCI to give a solid. The solid was collected, washed with water, and air-dried. The crude solid was recrystallized from CHCl 3 -n-hexane to give 215 mg (44%) 36 as a white crystalline powder. mp 141-145; IR (KBr) 3338, 2956, 2935, 2875, 2593, 1711 cm-'; 'H NMR (DMSO-de) 8 1.91-2.14 (4 H, min), 2.24 (3 H, s), 3.38-3.68 (4 H, m), 4.88-5.08 (1 H, min), 6.76-8.56 (12 H, min); MS (FAB) m/z 542 (M+1). 20 Example 33 3-N,N-dimethylamino-4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-2 pyrrolidinylmethoxy]benzoic acid 0 N ) N N H H ~COOH 37 A stirred mixture of methyl 4-hydroxy-3-nitrobenzoate (3.22g, 0.0163mol) and 5% Pd-C 25 (12.9g) in MeOH (30mL) was hydrogenated under an atmospheric pressure for 70 hr at room temp. Insoluble catalyst was removed, and the filtrate was evaporated in vacuo. The residue was chromatographed on silica-gel with EtOH-CHCl 3 (1:20, v/v) as eluent to afford 1.89g (69%) methyl 3-amino-4-hydroxybenzoate as a pale brown syrup. A stirred mixture of methyl 3-amino-4-hydroxybenzoate (1.07g, 6.40mmol) and 5% Pd-C 158 WO 01/00206 PCT/US00/18079 (2.14g) in MeOH (20mL) and 37% aq. formaldehyde (1.08mL, 0.0122mol) and 1 N HCI (6. 1mL) was hydrogenated under an atmospheric pressure for 26 hr at room temp. Insoluble catalyst was removed, and the filtrate was evaporated in vacuo. The residue was chromatographed on silica-gel with EtOAc-n-hexane (1:10, v/v) as eluent to afford 0.817g (70%) methyl 3-(N, N 5 dimethylamino)-4-hydroxybenzoate as a syrup. To a stirred mixture of methyl 3-(N, N-dimethylamino)-4-hydroxybenzoate (0.817g, 4.18mmol), N-tert-butoxycarbonylprolinol (0.926g, 4.60mmol), Ph 3 P (1.21g, 4.60mmol) in THF (20mL) was added dropwise DIAD (95%) (0.953mL, 4.60mmol) at 0 0 C. The resulting mixture was heated under reflux for 41 hr. After cooling, the mixture was evaporated off in vacuo. The 10 residue was chromatographed on silica-gel with EtOAc -n-hexane (1:10 to 1:6, v/v) as eluent to give a syrup which was used for the subsequent reaction without further purification. This syrup was dissolved in CH2C1 2 (10mL) and added TFA (10mL). After the solution was stirred for 5 hr at room temp, the solution was evaporated in vacuo. Water was to the residue and washed with CHCl 3 . The aqueous layer was neutralized by the addition of sat. NaHCO 3 and extracted with 15 CHCl 3 . The extract was dried over Na 2
SO
4 and evaporated in vacuo to afford 1.03g (89%) methyl 3-(N, N-dimethylamino)-4-(2-pyrrolidinylmethoxy)benzoate as a gum. A mixture of methyl 3-(N, N-dimethylamino)-4-(2-pyrrolidinylmethoxy)benzoate (0.529g, 1.90mmol), 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (0.597g, 1.90mmol), HOBt (0.308g, 2.28mmol), 4-dimethylaminopyridine(DMAP) (23.2mg, 0.190mmol), and 1-ethyl 20 3-(3-dimethylaminopropyl)carbodiimide (EDC) (0.437g, 2.28mmol) in DMF (10mL) was stirred for 15 hr at room temp. The mixture was neutralized by the carefully addition of 1 N HCI and extracted with EtOAc. The extract was dried over Na 2
SO
4 and evaporated in vacuo to afford 0.607g (56%) methyl 3-N,N-dimethylamino-4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-2-pyrrolidinyl methoxy]benzoate as a white crystalline material. 25 A mixture of methyl 3-N,N-dimethylamino-4-[1-[3-methoxy-4-[N'-(2-methylphenyl) ureido]-phenylacetyl]-2-pyrrolidinylmethoxy]benzoate (0.600g, 1.04mmol) in THF (O10mL) and 0.25 N NaOH (5mL, 1.25mmol) was stirred for 21 hr at room temp. CHC1 3 was added to the mixture and extracted with a mixture of water(100mL)-1N NaOH (4mL). The extract was neutralized with sat. NH 4 Cl and extracted withCHC13. The extract was dried over Na 2
SO
4 and 30 evaporated in vacuo to afford 428mg (70%) 37 as a white crystalline solid. 'H-NMR (400MIHz, DMSO-d 6 ) 8 1.88, 1.99 and 2.11 (4H, each min), 2.24 (3H, s), 2.67 (6H, s), 3.33 (2H, min), 3.58 (2H, 159 WO 01/00206 PCT/US00/18079 m), 4.05-4.32 (3H, m), 6.75 (1H, d, J=7.3Hz), 6.92-6.95 (1H, m), 7.05 (1H, d, J=8.3Hz), 7.11-7.17 (2H, m), 7.42 (lH, s), 7.52 (1H, d, J=7.8Hz), 7.79 (1H, d, J=7.8Hz), 8.00 (1H, d, J=7.8Hz), 8.31 (1H, s), 8.46, 8.55 (each 1H, each s); MS (FAB) m/z 533 (M'+1). Example 34 5 3-fluoro-4-[[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl]methoxy] benzoic acid N N H H COOH 38 To a stirred solution of 4-bromo-2-fluorophenol (217 ul, 2.002 mmol), N-tert butoxycarbonyl prolinol (403 mg, 2.002 mmol), and Ph 3 P (630 mg, 2.403 mmol) in THF (7 mL) 10 was added DIAD (477 ul, 2.423 mmol) at room temp. The resulting mixture was stirred for 6 hr at room temp and then overnight at 70oC. The mixture was concentrated in vacuo and the residue was chromatographed on silica-gel with n-hexane - EtOAc (5:1, v/v) as eluent to give 549.4 mg (73%) 1-bromo-4-[1-(tert-butoxycarbonyl)-2-pyrrolidinyl]methoxy-3-fluorobenzene as an oil. 'H NMR (400 MHz, CDCl 3 ) 8 1.46 (s, 9H11), 1.85 (br m, 1H), 1.90 - 2.10 (br s, 3H), 3.30 - 3.47 (m, 15 2H), 3.85, 4.04 (br s each, 1H), 4.11 - 4.20 (m, 2H), 6.82 - 6.98 (m, 1H), 7.13 - 7.26 (m, 2H); MS (FAB) m/z 374 (M++1). To a stirred solution of 1-bromo-4-[1-(tert-butoxycarbonyl)-2-pyrrolidinyl] methoxy-3 fluorobenzene (549.4 mg, 1.468 mmol) in DMSO (6 mL) and MeOH (5 mL) was added Et 3 N (448 ul, 3.229 mmol), Pd(OAc) 2 (36.2 mg, 0.161 mmol), and 1,3-bis(diphenylphosphino)propane (66.4 20 mg, 0.161mmnunol). To the mixture was induced CO gas for 10 min. The resulting mixture was stirred at 70 0 C for 2 days under a current of CO. After the mixture was concentrated, the residue was diluted with EtOAc. The solution was washed with brine and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the residue was chromatographed on silica-gel eluting with n-hexane:EtOAc (5:1, v/v) as eluent to afford 323.0 mg (62 %) methyl 4-[1-(tert 25 butoxycarbonyl)-2-pyrrolidinyl] methoxy-3-fluorobenzoate as a pale yellow oil. 'H-NMR (400 MHz, CDC1 3 ) 5 1.47 (s, 9H), 1.87(br s, 1H), 1.95 - 2.10 (m, 3H), 3.34 - 3.44 (br m, 2H), 3.89 (s, 3H), 3.94 and 4.11 - 4.26 (br m each, 3H), 7.03 - 7.11 (m, 1H), 7.75 - 7.80 (m, 2H); MS (FAB) m/z 354 (M++1). To a stirred solution of methyl 4-[1-(tert-butoxycarbonyl)-2-pyrrolidinyl Imethoxy-3 30 fluorobenzoate (323.0 mg, 0.914 mmol) in CH 2 C1 2 (6.5 mL) was added TFA (1.3 mL) at 0OC, and 160 WO 01/00206 PCT/US00/18079 the mixture was stirred 1.5 hr at room temp. The solvent was removed under a reduced pressure and the residue was made basic by the addition of IN NaOH. The mixture was extracted with CHC1 3 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated under a reduced pressure to afford 174.8 mg (76%) methyl 3-fluoro-4-(2-pyrrolidinyl)methoxybenzoate as a 5 brownish oil. 'H-NMR (400 MHz, CDCl 3 ) 8 1.54 - 1.63 (m, 1H), 1.76 - 2.02 (m, 3H11), 2.93 - 3.07 (m, 2H), 3.57 (ddd, J = 4.9, 6.9, 14.3 Hz, 1H), 3.89 (s, 3H), 3.97 (dd, J= 6.8, 9.3 Hz, 1H), 4.04 (dd, J = 5.0, 8.8 Hz, 1H11), 6.98 (t, J = 17.6 Hz, 111), 7.73 (dd, J= 2.0, 11.7 Hz, 1H), 7.78 (dt, J = 2.0, 8.8 Hz,1H); MS (FAB) m/z 253 (M++1). A mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetate 10 (324.5 mg, 0.676 mmol), methyl 3-fluoro-4-(2-pyrrolidinyl)methoxybenzoate (171.1 mg, 0.676 mmol), Et 3 N (113 ul, 0.811 mmol) in DMF (5 mL) was stirred for 2 hr at room temp. The mixture was diluted with EtOAc, washed with brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the residue was chromatographed on silica-gel with n-hexane:EtOAc (1:2, v/v) as eluent to afford methyl 3-fluoro-4-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] 15 phenylacetyl]-2-pyrrolidinyl]methoxy] benzoate (365.1 mg, 0.664 mmol) as an oil. To a stirred solution of this compound in THF (4.4 mL) and H20 (1.1 mL) was added LiOH (46.3 mg, 1.932 mmol), and the reaction mixture was stirred at room temp overnight. The mixture was diluted with CHCl 3 and acidified by the addition of IN HCL. The separated organic layer was washed with brine and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure, and the obtained 20 crude solid was recrystallized from n-hexane-EtOAc-CHC1 3 to afford 102 mg (30%) 38 as a white crystalline powder. mp 123 - 126; IR (KBr) 1616, 1537, 1282, 756cm'; 1 H-NMR (400 MHz, DMSO-d 6 ) 8 1.87 - 2.09 (m, 4H11), 2.25 (s, 3H), 3.48 - 3.57 (m, 2H), 3.60 (s, 2H), 3.83 (s, 3H), 4.11 - 4.16 (m, 1H), 4.24 (dd, J= 2.9, 9.8 Hz, 1H), 4.28 - 4.34 (br s, 1H11), 6.74 (dd, J= 1.5, 8.3 Hz, 1H), 6.87 (s, 1H), 6.94 (t, J= 7.3 Hz, 1H), 7.12 (d, J= 7.8 Hz, 111), 7.15 (t, J= 8.3 Hz, 1H), 7.34 25 (t, J= 8.8 Hz, 1H11), 7.66 (dd, J= 2.0, 12.2 Hz, 1H), 7.73 (d, J= 9.3 Hz, 1H), 7.79 (d, J= 8.3 Hz, 1H), 7.99 (d, J= 7.8 Hz, 1H), 8.46 (s, 1H), 8.55 (s, 1H); MS (FAB) m/z 536 (M +1); Anal. Calcd for C 29
H
30
FN
3 06-0.5H 2 0: C, 63.96; H, 5.74; N, 7.72; F; 3.49. Found: C, 64.11; H, 5.80; N, 7.39; F, 3.54. Example 35 30 4-[ 1-[4-[N'-(2-fiuorophenyl)ureido]-3-methoxyphenylacetyl]-2-pyrrffolidinyllmethoxy-3-methoxy benzoic acid 161 WO 01/00206 PCT/US00/18079 N N COOH N ) N 39 A mixture of 4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenylacetic acid (318 mg, 1 mmol), 2-(2-methoxy-4-ethoxycarbonyl)phenoxymethylpyrrolidine (279 mg, 1 mmol), EDC (288 mg, 1.5 mmol), and DMAP (122 mg, 1 mmol) in DMF (20 mL) was stirred overnight. The 5 mixture was poured into 1 N HCI and the resulting solid was collected with suction. The solid was dissolved in CHCl 3 and dried over MgSO 4 . After removal of the solvent, the residue was chromatographed on silica-gel with CHCl 3 :MeOH (100:1, v/v) as eluent to give an oil, which was dissolved in THF:MeOH (4:1, v/v, 10 mL). 0.25 N NaOH (8 mL) was added to the solution and the resulting stirred mixture was heated under reflux for 3 hr. The mixture was poured into 1 N HC1. 10 The resulting solid was collected with suction, dissolved in CHClz, dried over MgSO 4 , and evaporated. The residue was recrystallized from CHCl 3 -n-hexane-ether to give 329 mg (60%) 39 as a white crystalline powder. mp 140-144; IR (KBr) 3338, 2956, 2875, 2607, 1709 cm-'; 'H-NMR (CDCl 3 ) 8 1.95-2.25 (4 H, m), 3.45-4.50 (12 H, m), 6.66-8.15 (12 H, m); MS (FAB) m/z 552 (M+1). 15 Example 36 2-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl]methoxy] pyridine-5 carboxylic acid 0~ 0 N 1 N" I H H COOH 40 To a stirred solution of 6-hydroxynicotinic acid (500 mg, 3.594 mmol) in benzene (8 mL) 20 and MeOH (2 mL) was added dropwise TMSCHN 2 (1.97 mL, 3.953 rmmol) at 0 0 C, and the mixture was stirred overnight at room temp. The reaction mixture was quenched by the addition of AcOH, and the resulting mixture was concentrated in vacuo. The residue was chromatographed on silica-gel with n-hexane-EtOAc (1:3, v/v) as eluent to give 269.8 mg (49%) methyl 2 hydroxypyridine-5-carboxylate as a white crystalline powder. IR (KBr) 3062, 1657, 1654, 1612, 25 1435, 1300, 1113,775, 642cm- 1 ; 1H-NMR (400 MHz, CDC1 3 ) 8 3.87 (s, 3H), 6.58 (d, J= 9.8 Hz, 1H), 7.99 (dd, J = 2.4, 9.8 Hz, 111), 8.19 (d, J = 2.4 Hz, 1I); MS (FAB) m/z 154 (M+1); Anal Called for C 1 HN03-1/4H 2 0: C, 53.33; H, 4.80; N, 8.89. Found: C, 53.58; H, 4.65; N, 8.87. To a stirred solution of methyl 2-hydroxypyridine-5-carboxylate (269.8 mg, 1.762mmol), 162 WO 01/00206 PCT/US00/18079 N-tert-butoxycarbonylprolinol (354.6 mg, 1.762 mmol), and Ph 3 P (554.6 mg, 2.114 mmol) in THF (10 mL) was slowly added DIAD (0.42 mL, 2.114 mmol) at room temp, and the resulting mixture was stirred for 6 hr at 700 C. The reaction mixture was concentrated and the residue was chromatographed on.silica-gel with n-hexane:EtOAc (5:1, v/v) as eluent to give 262.5 mg (44%) 5 methyl 2-[[1-(tert-butoxycarbonyl)-2-pyrrolidinyl]methoxy]pyridine-5-carboxylate as an oil. 'H NMR (400 MHz, CDCl 3 ) 8 1.47 (s, 9H1), 1.85 - 1.98 (m, 4H), 3.37 (br s, 2H), 3.92 (s, 3H), 4.12 4.33 (br m, 2H),4.4 (brs, 1H),6.75(m, 1H),8.15 (m, 1H), 8.79 (m, 1H); MS (FAB) m/z 337 (M'+1). To a stirred solution of methyl 2-[[1-(tert-butoxycarbonyl)-2-pyrrolidinyl]methoxy] pyridine-5-carboxylate (262.5mg, 0.870 mmol) in CH 2 C1 2 (5.3 mL) was added TFA (1.1 mL) at 10 0 0 C, and the resulting mixture was stirred at room temp for 1 hr. The solvent was removed under a reduced pressure and the residue was made basic by the addition of the IN NaOH, and extracted with CHCl 3 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated under a reduced pressure to afford 173.1 mg (94%) methyl 2-[(2-pyrrolidinyl)methoxy]pyridine-5 carboxylate as a pale yellowish oil. 'H-NMR (400 MHz, CDCl 3 ) 5 1.49 - 1.58 (ddt, J = 6.8, 8.8 Hz, 15 1H), 1.72 - 1.87 (m, 2H), 1.90 - 1.99 (m, 1H), 2.92 - 3.05 (m, 2H), 3.50 -3.57 (ddd,J=4.4,7.3, 15.1 Hz, 1H11), 3.91 (s, 3H11), 4.23 (dd, J= 7.8, 10.7 Hz, 1H1), 4.38 (dd, J= 4.4, 10.7 Hz, 1H), 6.78 (d, J 8.8 Hz, 1H11), 8.15 (dd, J= 2.4, 8.8 Hz, 111), 8.80 (d, J= 2.4 Hz, 1H); MS (FAB) m/z 237 (M t +1). A mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetate (351.7 mg, 0.732 mmol), methyl 2-[(2-pyrrolidinyl)methoxy]pyridine-5-carboxylate (173.0 mg, 20 0.732 mmol), Et 3 N (122.4 gl, 0.878 mmol) in DMF (5.2 mL) was stirred for 1 hr at room temp. The mixture was diluted with EtOAc, washed with brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the residue was chromatographed on silica-gel with n hexane:EtOAc (1:5, v/v) as eluent to afford methyl 2-[[1-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenylacetyl]-2-pyrrolidinyl]methoxy] pyridine-5-carboxylate (338.4 mg, 87%) as an oil. 25 To a stirred solution of this compound in THF (5.6 mL) and H20 (1.4 mL) was added LiOH (45.7 mg, 1.91 mmol), and the reaction mixture was stirred at room temp overnight. The mixture was diluted with CHC13, and treated with sat. NH 4 CI, washed with brine, dried over Na 2
SO
4 . The solvent was removed under a reduced pressure, and the obtained crude solid was recrystallized from n-hexane-Et 2 0-CHCl 3 -MeOH to afford 193.8 mg (59%) 40 as a white crystalline powder. mp 30 125 -128; IR (KBr) 1716, 1600, 1533, 1255cm'; 'H-NMR (400 MHz, DMSO-d 6 ) 8 1.67 - 2.03 (m, 4H11), 2.50 (s, 3H), 3.33 - 3.42 (m, 1H), 3.52 (m, 2H), 3.58 (d, J= 4.4 Hz, 1H), 3.83 (s, 3H), 4.27 4.31 (m, 2H1), 4.42 - 4.47 (m, 1H), 6.73 (d, J= 7.8 Hz, 1H), 6.87 - 6.95 (m, 3H), 7.11 - 7.17 (m, 163 WO 01/00206 PCT/US00/18079 2H), 7.79 (d, J= 8.3 Hz, 1H), 7.99 (d, J= 8.3 Hz, 1f), 8.14 (dd, J= 2.0, 8.8 Hz, 1H), 8.46 (s, 1H), 8.56 (s, 1H), 8.69 (d, J = 2.0 Hz, 1H1), 13.06 (br s, 1H); MS (FAB) m/z 519 (M'+l);Anal. Calcd for C 28
H
30
N
4 0 6 -1/2H 2 0: C, 63.75; H, 5.92; N, 10.62. Found: C, 63.61; H, 5.94; N, 10.27. Example 37 5 3-methoxy-4-[2-[4-[N-(2-methylphenyl)ureido]benzyl]-4-thiazolyl]methoxybenzoicacid COOH 41 To a stirred solution of phosphorous pentasulfide (27.4 g, 123.34 mmol) and freshly prepared anhydrous Na 2 S (4.8 g, 61.67 mmol) in THF (200 mL) was added p-nitrobenzyl cyanide (2.0 g, 12.33 mmol) at room temp. The resulting mixture was stirred for 17 hr at room temp. The 10 mixture was diluted with EtOAc and washed with 10% K3PO 4 . The aqueous layer was extracted with CH 2 C1 2 . The extract was dried over Na 2
SO
4 and concentrated in vacuo. The residue was chromatographed on silica-gel with n-hexane:EtOAc (5:1 to 2:1, v/v) as eluent to give 1.53g (64%) 4-nitrobenzyl carbothioamide as a pale yellow crystalline material. IR (KBr) 1529, 1446, 1326, 1315, 858cm~'; 'H-NMR (400 MHz, CDCl 3 ) 8 4.15 (s, 2H), 7.51 (d, J= 8.3 Hz, 2H), 8.24 (d, J= 15 8.8 Hz, 2H); MS (FAB) m/z 197 (M+1); Anal. Calcd for C 8 HgN 2 0 2 S: C, 48.97; H, 4.11; N, 14.28; S; 16.34. Found: C, 48.69; H, 4.06; N, 14.07; S; 16.10. To a stirred solution of 4-nitrobenzylcarbothioamide (502.0 mg, 2.558 mmol) in EtOH (5 mL) was added 1,3-dichloro-2-propanone (649.6 mg, 5.16 mmol) and the mixture was heated under reflux for 1 hr. The mixture was concentrated and the residue was diluted with CHC1 3 . The 20 solution was washed with 1N NaOH, brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the residue was chromatographed on silica-gel with n-hexane:EtOAc (4:1, v/v) as eluent to afford 495.2 mg (72 %) 4-[2-(4-nitrobenzyl)thiazolyl]methyl chloride as a pale yellow oil. 'H-NMR (400 MHz, CDCI 3 ) 8 4.43 (s, 2H), 4.68 (s, 2H), 7.23 (s, 1H), 7.49 (d, J = 8.8 Hz, 2H), 8.20 (d, J = 8.8 Hz, 2H11); MS (FAB) m/z 269 (M++1). 25 To a stirred solution of vanillic acid ethyl ester (308.0 mg, 1.570 mmol) and MeONa (89 mg, 1.570 mmol) in MeOH (6.5 mL) was added a solution of 4-[2-(4-nitrobenzyl)thiazolyl] methyl chloride (211.0 mg, 0.785 mmol) in MeOH (1.4 mL) at 0 0 C. The resulting mixture was stirred at room temp for 16 hr, and heated under reflux for 1 day. The solvent was removed under a reduced pressure and the residue was extracted with CHC1 3 . The extract was washed with H 2 0, brine, and 30 dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the residue was 164 WO 01/00206 PCT/US00/18079 chromatographed on silica-gel with n-hexane:EtOAc (2:1, v/v) as eluent to afford 201.7 mg (60 %) ethyl 3-methoxy-4-[2-(4-nitrobenzyl)-4-thiazolyl]methoxybenzoate as a pale yellow oil. 'H-NMR (400 MHz, CDCl 3 ) 8 1.39 (t, J= 7.3 Hz, 3H), 3.93 (s, 3H), 4.36 (q, J= 7.3 Hz, 2H), 4.44 (s, 2H), 5.31 (s, 2H), 6.97 (d, J = 8.3 Hz, 1H), 7.28 (s, 1H), 7.48 (d, J= 8.8 Hz, 2H), 7.57 (d, J= 2.0 Hz, 5 1H), 7.64 (dd, J= 2.0, 8.3 Hz, 1H), 8.20 (d, J= 8.8 Hz, 2H); MS (FAB) m/z 429 (M+1). A stirred solution of ethyl 3-methoxy-4-[2-(4-aminobenzyl)-4-thiazolyl methoxybenzoate (201.7 mg, 0.471 mmol) and 5% Pd/C (40 mg) in EtOH (8 mL) was hydrogenated at 1 atm for 24 hr. The mixture was filtered and the filtrate was concentrated. The residue was chromatographed on silica-gel with n-hexane:EtOAc (1:1, v/v) as eluent to afford 87.8 mg (47%) ethyl 3-methoxy-4 10 [2-(4-aminobenzyl)-4-thiazolyl]methoxybenzoate as a yellowish crystalline powder. 'H-NMR (400 MHz, CDCl 3 ) 8 1.38 (t, J= 7.3 Hz, 3H), 3.93 (s, 3H), 4.21 (s, 2H11), 4.35 (q, J= 7.3 Hz, 2H), 5.30 (s, 2H), 6.66 (dd, J= 2.0, 6.4 Hz, 2H), 6.97 (d,J= 8.3 Hz, 1H), 7.09 (d,J= 8.3 Hz, 2H), 7.18 (s, 1H), 7.56 (d, J= 2.0 Hz, 1H), 7.64 (dd, J= 2.0, 8.3 Hz, 1H); MS (FAB) m/z 399 (M+1). To a solution of ethyl 3-methoxy-4-[2-(4-aminobenzyl)-4-thiazolyl]methoxybenzoate 15 (87.8 mg, 0.220 mmol) in THF (2.0 mL) was added triethylamine (30.5 ul, 0.220 mmol) and o tolyl isocyanate (30 pl), and the reaction mixture was stirred at room temp for 21 hr. The reaction mixture was poured into ice-water and the resulting precipitates filtered off. The filtrate was extracted with CHC13, washed with H 2 0, and brine. The extract was dried over Na 2
SO
4 . The solvent was removed under a reduced pressure to afford 110.4 mg (94 %) ethyl 3-methoxy-4-[2-[4 20 [N'-(2-methylphenyl) ureido]benzyl]-4-thiazolyl] methoxybenzoate as a pale yellow crystalline powder. 1H-NMR (400 MHz, CDCl 3 ) 8 1.38 (t, J= 7.3 Hz, 3H), 2.28 (s, 3H), 3.92 (s, 3H), 4.28 (s, 2H), 4.35 (q, J= 7.3 Hz, 2H), 5.29 (s, 2H), 6.20(s, 1H), 6.47 (s, 1H), 6.97 (d, J= 8.8 Hz, 1H), 7.20 (s, 1H), 7.24 (s, 2H), 7.27 (s, 2H), 7.33 (d, J= 8.3 Hz, 2H), 7.49 (d, J= 7.3 Hz, 1H), 7.56 (d, J= 2.0 Hz, 1H), 7.63 (dd, J= 2.0, 8.3 Hz, 1H); MS (FAB) m/z 532 (M+1). 25 To a stirred solution of ethyl 3-methoxy-4-[2-[4-[N'-(2-methylphenyl)ureido] benzyl]-4 thiazolyl]methoxybenzoate in THF (1.6 mL) and H 2 0 (0.4 mL) was added LiOH (6.0 mg, 0.249 mmol), and the mixture was stirred at room temp for 1 hr, and heated under reflux for 8 hr. The mixture was concentrated and diluted with CHC 3 . The solution was made basic by the addition of 1N NaOH. The aqueous extract was acidified by the addition of 1N HCI and extracted with CHC 3 . 30 The extract was washed with brine and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the obtained crude solid was recrystallized from n-hexane-EtOAc-EtOH to 165 WO 01/00206 PCT/US00/18079 afford 59.6 mg (57%) 41 as a white crystalline powder. mp 243 - 245; IR (KBr) 3282, 1685, 1637, 1600, 1554, 1516, 1278, 1234, 763, 748cm-'; 'H-NMR (400 MHz, DMSO-d 6 ) 8 2.27 (s, 3H), 3.83 (s, 3H), 4.30 (s, 2H), 5.21 (s, 2H), 6.97 (t, J= 7.3 Hz, 1H), 7.15 -7.24 (m, 3H), 7.29 (d,J= 8.8 Hz, 2H), 7.47 (d, J= 8.3 Hz, 2H), 7.50 (s, 1H), 7.58 (d, J= 8.3 Hz, 1H), 7.62 (s, 1H), 7.86 (d, J= 5 7.8 Hz, 1l), 7.97 (s, 1H), 9.09 (s, 1H),12.70 (br s, 1H); MS (FAB) m/z 504 (M*+1); Anal. Calcd for C 27
H
25
N
3 0 5 S-1/4H 2 0: C, 63.83; H, 5.06; N, 8.27. Found: C, 63.74; H, 4.99; N, 8.10. Example 38 4-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-methyl-2-pyrrffolidinyl] methoxy] 3-nitrobenzoic acid / H H >. ? 'COOH 10 2 N COOH 42 To a stirred solution of the N-tert-butoxycarbonylproline (6.00g, 0.0279mol) in MeOH:benzene (1:4, v/v) was added dropwise 2.0 M-n-hexane solution of TMSCHN, (16.7mL, 0.0334mol) at room temp. After the resulting solution was stirred for lhr at room temp, the mixture was evaporated off in vacuo to afford 6.39g (100%) N-tert-butoxycarbonylproline methyl 15 ester as a yellow syrup. 'H-NMR (CDCl 3 ) 8 1.41 (s, 9H), 1.85-1.98 (m, 4H), 2.21-2.28 (m, 2H), 3.72 (s, 3H), 4.29 (m, 1H). To a stirred solution of diisopropylanmine (2.02mL, 0.0144mol) in THF (30mL) was added dropwise 1.59 M n-hexane solution of n-BuLi (9.06mL, 0.0144mol) at minus 78 0 C for over 5 min. The resulting solution was stirred for 20 min at minus 78 0 C. To the solution was added dropwise 20 N-tert-butoxycarbonylproline methyl ester (3.00g, 0.013 Immol) in THF (30mL) at minus 78 0 C for over 5 min. The resulting solution was stirred for 10 min at minus 78 0 C. To the solution was added dropwise Mel (0.900miL, 0.0144mol) at minus 78 0 C. The resulting solution was stirred for 30 min at minus 78 0 C. The solution was quenched by the addition of sat. NH 4 C1. The resulting mixture was extracted with CHC1 3 . The extract was washed with water, dried over Na 2
SO
4 , and 25 evaporated in vacuo to afford 3.20g (q.y.) N-tert-butoxycarbonyl-2-methylproline methyl ester as a yellow syrup. 'H-NMR (CDCl 3 ) 8 1.33 (s, 9H), 1.38 (s, 3H), 1.72-2.20 (m, 4H) 3.27-3.59 (m, 2H) 3.63 (d, J=6.3Hz, 3H). To a stirred solution of N-tert-butoxycarbonyl-2-methylproline methyl ester (3.20g, 0.0131lmol) in THF (20mL) was added 1N NaOH (15.7mL) at room temp. After the resulting 166 WO 01/00206 PCT/US00/18079 mixture was stirred for 24 hr, the mixture was diluted with water and washed with EtOAc. The separated aqueous layer was acidified by the addition of IN HCI, and extracted with EtOAc. The extract was dried over Na 2
SO
4 and evaporated in vacuo to afford 1.71g(57%) N-tert butoxycarbonyl-2-methylproline as a yellow syrup. 'H-NMR (CDCl 3 ) 8 1.42 (s, 9H), 1.48 (s, 3H11), 5 1.88-2.31 (min, 4H), 3.34-3.57 (min, 2H), 9.35 (br s, 1H) To a stirred solution of N-tert-butoxycarbonyl-2-methylproline (1.10g, 4.80mmol) in THF (20mL) was added dropwise BH 3 -SMe 2 (0.546mL, 5.76mmol) at room temp. After the resulting mixture was stirred for 6 hr at 80 0 C, the mixture was evaporated in vacuo. The residue was diluted with MeOH, washed with n-hexane (3x), and evaporated in vacuo to afford 0.648g (60%) 10 N-tert-butoxycarbonyl-2-hydroxymethyl-2-methylpyrrolidine as a yellow syrup. 'H-NMR (CDCl 3 ) 8 1.47 (s, 9H), 1.76-2.05 (min, 4H1), 3.28-3.48 (min, 2H), 3.66 (min, 2H, d). To a stirred solution of N-tert-butoxycarbonyl-2-hydroxymethyl-2-methylpyrrolidine (0.648g, 3.01mmol), methyl 4-hydroxy-3-nitrobenzoate (0.593g, 3.01mmol), and Ph 3 P (0.868g, 3.31 mmol) in THF (10mL) was added dropwise DIAD (95%) (0.686mL, 3.31 mmol) at 0oC. After 15 the resulting mixture was stirred for 24hr at 80oC, the mixture was evaporated in vacuo. The residue was diluted with CH 2 C1 2 (5mL) and added TFA (5mL). After the resulting mixture was stirred for 2hr at room temp, the mixture was evaporated in vacuo. The residue was diluted with 0.5N HCI and extracted with CHC1 3 . The aqueous layer was neutralized with sat. NaHCO 3 , and extracted with CHCl 3 . The extract was dried over Na 2
SO
4 and evaporated in vacuo to afford 20 0.188g (21%) methyl 3-nitro-4-(2-methyl-2-pyrrolidinylmethoxy) benzoate as a yellow syrup. A mixture of methyl 3-nitro-4-(2-methyl-2-pyrrolidinylmethoxy)benzoate (0.188g, 0.920mmol), 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (0.289g, 0.920mmol), HOBt (0.149g, 1.10mmol), DMAP (11.2mg, 0.0920mmol) and EDC (0.21 1g, 1.10mmol) in DMF (5mL) was stirred for 14hr at room temp. EtOAC was added to the mixture and the solution was 25 washed successively with 0.5 N HCI, sat. NaHCO 3 , and brine. The organic layer was dried over Na 2
SO
4 and evaporated in vacuo to afford 0.489g (q.y.) methyl 4-[[1-[3-methoxy-4-[N'-(2 methylphenyl)ureido] phenylacetyl]-2-methyl-2-pyrrolidinyl]methoxy]-3-nitrobenzoate as a yellow crystalline material. 'H-NMR (CDC1 3 ) 8 1.26 (d, J=5.9Hz), 1.85-4.50 (mn, 10H), 2.30 (s, 3H), 3.67 (s, 2H), 3.92 (s, 3H), 6.36 (s, 2H), 6.75-7.52 (min, 7H1), 8.02 (d, J=7.8Hz, 1H), 8.15 (d, J=8.8Hz, 30 1H1), 8.47 (s, 1H). 167 WO 01/00206 PCT/US00/18079 A stirred mixture of 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2 methyl-2-pyrrolidinyl]methoxy]-3-nitrobenzoate (0.489g, 0.828mmol) in MeOH (5mL) and IN NaOH (1.24mL) was heated under reflux for 2hr After cooling, the mixture was diluted with water and extracted with CHCl 3 . The aqueous layer was acidified with iN HC1, and extracted with 5 CHCl 3 . The extract was dried over Na 2
SO
4 and evaporated in vacuo to afford 0.366g (99%) 42 as a yellow crystalline material. Example 39 4-[4-hydroxy-l-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinylmethoxy] 3- methoxybenzoic acid .,OH 10 COOH 43 A stirred mixture of 4-[4-benzyloxy-1l-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-2-pyrrolidinylmethoxy]-3-methoxybenzoate (440 mg, 0.645 mmol) and 5% Pd/C (400 mg) in AcOH:EtOH (1:1, v/v,100 mL) was hydrogenated at 1 atm for 5 hr. The mixture was filtered to remove the catalyst and the filtrate was concentrated in vacuo. The residue was 15 chromatographed on silica-gel with CHCl 3 :EtOH (10:1, v/v) as eluent to give 90 mg (24%) ethyl 4 [4-hydroxy- 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-2-pyrrolidinylmethoxy]-3 methoxybenzoate as a pale yellow oil. 'H-NMR (CDCl 3 ) 8 1.39 (3 H, t, J 7.3 Hz), 2.04-2.37 (total 5 H, min), 3.44-4.70 (16 H, series of min), 6.63 (1 H, s), 6.70-6.80 (2 H, min), 6.84 (1 H, d, J= 8.3 Hz), 7.11 (1 H, t, J= 7.8 Hz), 7.20-7.24 (3 H, min), 7.45 (1 H, d, J= 2.0 Hz), 7.59 (2 H, dd, J= 8.3, 20 2.0 Hz), 8.01 (1 H, d, J =7.8 Hz). A stirred mixture of ethyl 4-[4-hydroxy-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-2-pyrrolidinylmethoxy]-3-methoxybenzoate (90 mg, 0.152 mmol) in 0.25 N NaOH (5 mL, 1.25 mmol) and THF (5 mL) was heated under reflux overnight. The mixture was poured into ice-1 N HC1 (200 mL). The precipitate was collected with suction and recrystallized from 25 CHC13-MeOH-n-hexane to give 40 ming (47%) 43 as a colorless amorphous solid. 'H-NMR (DMSO d 6 ) 8 1.92-2.11 (2 H, min), 2.24 (3 H, s), 3.31-5.07 (14 H, series of m), 6.73 (1 H, d, J= 8.3 Hz), 6.84 (1 H, s), 6.93 (1 H, t, J= 7.8 Hz), 7.01-7.17 (3 H, min), 7.44 (1 H, s), 7.52 (1 H, d, J= 8.8 Hz), 7.79 (1 H, d, J= 8.3 Hz), 7.99 (1 H, d, J= 7.8 Hz), 8.46 (1 H, s), 8.55 (1 H, s), 12.67 (1 H, br s); MS (FAB) m/z 564 (M+1). 168 WO 01/00206 PCT/US00/18079 Example 40 (2S,4R)-3-amino-4-[4-hydroxy-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2 pyrrolidinyl]methoxybenzoic acid 9H ON 1 H2 N N' 0 ' H H OOH 44 5 To a stirred solution of (2S,4R)-4-benzyloxy-1-(tert-butoxycarbonyl)-2-prolinol (891 mg, 2.9 mmol), methyl 4-hydroxy-3-nitrobenzoate (572 mg, 2.9 mmol),and PPh 3 (839 mg, 3.2 mmol) in THF (6 mL) was added DIAD (630 mL, 3.2 mmol) and the mixture was heated under reflux overnight. After removal of the solvent, the residue was chromatographed on silica-gel with n hexane:EtOAc (1:1) and toluene:EtOAc (10:1, v/v) as eluent to give 700 mg (50%) methyl 10 (2S,4R)-4-[4-benzyloxy-l-(tert-butoxycarbonyl)-2-pyrrolidinyljmethoxy-3-nitrobenzoate as a pale yellow oil. To a stirred solution of methyl (2S, 4R)-4-[4-benzyloxy-l-(tert-butoxycarbonyl)-2 pyrrolidinyl] methoxy-3-nitrobenzoate (681 mg, 1.4 mmol) in CH 2 C1 2 (2 mL) was added TFA (2 mL), and the resulting mixture was stirred for 2 hr. After the reaction mixture was concentrated, 15 the residue was made basic by the addition of sat. NaHCO 3 and extracted with CHCl 3 . The extract was washed with H20, dried over MgSO 4 , and evaporated to give 511 mg (95%) methyl (2S,4R)-4 [4-benzyloxy-2-pyrrolidinyl]methoxy-3-nitrobenzoate as a yellow oil. A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (409 mg, 1.3 mmol), methyl (2S,4R)-4- (benzyloxy-2-pyrrolidinyl)methoxy-3-nitrobenzoate (502 mg, 1.3 20 mmol), EDC (383 mg, 2 mmol), and DMAP (159 mg, 1.3 mmol) in DMF (20 mL) was stirred for 3 days. The mixture was poured into 1 N HC1 and the resulting precipitate was collected with suction. The residue was dissolved in CHC1 3 and dried over MgSO 4 . After removal of solvent, the residue was chromatographed on silica-gel with CHC13:MeOH (200:1, v/v) as eluent to give 680 mg (91%) methyl (2S,4R)-4-[4-benzyloxy-l-[3-methoxy-4-[N'-(2-methylphenyl)ureido] 25 phenylacetyl]-2-pyrrolidinyl]methoxy-3-nitrobenzoate as a white amorphous solid. A solution of methyl (2S,4R)-4- [4-benzyloxy-1l-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenylacetyl]-2-pyrrolidinyl]methoxy-3-nitrobenzoate (676 mg, 0.99 mmol) and 5% Pd-C 169 WO 01/00206 PCT/US00/18079 (1 g) in EtOH:AcOH (1:1, v/v, 30 mL) was hydrogenated at 1 atm for 6 hr. The mixture was filtered and the filtrate was evaporated to give an oil, which was made basic by the addition of sat. NaHCO 3 . The mixture was extracted with EtOAc. The extract was washed with brine, dried over MgSO 4 ,.and evaporated. The residue was recrystallized from CHC1 3 -EtOH-n-hexane as eluent to 5 give 120 mg (22%) 44 as a pale yellow crystalline powder. MS (FAB) m/z 549 (M+1) Example 41 4-[[4-fluoro-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl] methoxy] 3-methoxybenzoic acid F H H OOH 45 10 A stirred mixture of ethyl 4-[4-benzyloxy-1-(tert-butoxycarbonyl)-2-pyrrolidinyl]methoxy 3-methoxybenzoate (1.189 g, 2.449 mmol) and 5% Pd-C (240 mg) in EtOH (10 mL) was hydrogenated overnight at room temp. The mixture was filtered to remove the catalyst and the filtrate was concentrated in vacuo to give ethyl 4-[1-(tert-butoxycarbonyl)-4-hydroxy-2 pyrrolidinyl]methoxy-3-methoxybenzoate (735.3 mg, 76%) as a pale yellow oil. To a stirred cold 15 (minus 78 0 C) solution of DAST (0.491 mL, 3.718 mmol) in CH 2 C1 2 (7.4 mL) was added dropwise a solution of this compound in CH 2 C1 2 (2mL), and the resulting mixture was stirred overnight. The mixture was quenched with water and extracted with CHC1 3 . The extract was washed with brine and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the residue was chromatographed on silica-gel with n-hexane:EtOAc (3:1, v/v) as eluent to afford 418.7 mg 20 (57%) ethyl 4-[1-(tert-butoxycarbonyl)-4-fluoro-2-pyrrolidinyl]methoxy-3-methoxybenzoate as an oil. 1 H-NMR (400 MHz, CDC1 3 ) 5 1.39 (t, J= 7.3 Hz, 3H), 1.49 (s, 9H), 2.16 (br m, 1H), 2.58 (dd, J= 15.6, 19.0 Hz, 1H), 3.60 - 3.75 (min, 2H), 3.91 (s, 3H), 3.97 (t, J= 9.3 Hz, 1H), 4.35 (q, J = 7.3 Hz, 2H), 4.33 - 4.53 (min, 2H), 5.25 (d, J = 52.7 Hz, 1H), 7.04 (dd, J= 7.8, 56.2 Hz, 1H), 7.55 (s, 1H), 7.65 (br s, 1H); MS (FAB) m/z 398 (M +I). 25 To a stirred solution of ethyl 4-[1-(tert-butoxycarbonyl)-4-fluoro-2-pyrrolidinyl]methoxy 3-methoxybenzoate (482.2 mg, 1.213 mmol) in CH 2 C1 2 (10.0 mL) was added TFA (1.9 mL) at 0oC, and the mixture was stirred at room temp for 2 hr. The solvent was removed under a reduced pressure and the residue was made basic by the addition of IN NaOH and extracted with CHC1 3 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated under a reduced pressure 170 WO 01/00206 PCT/US00/18079 to afford 348.7 mg (97%) ethyl 4-(4-fluoro-2-pyrrolidinyl)methoxy-3-methoxybenzoate as a brownish oil. 'H-NMR (400 MHz, CDCl 3 ) 8 1.39 (t, J= 6.8 Hz, 3H), 1.97 (ddt, J= 1.5, 5.4, 14.7 Hz,1H), 2.27 (dddd, J= 5.9, 8.8, 14.7, 32.7 Hz,1H), 3.02 (ddd, J= 3.9, 13.1, 35.2 Hz,IH), 3.36 (dd, J= 12.7, 21.5 Hz,1H), 3.65 (min, 1H), 3.90 (s, 3H), 4.09 (m, 1H), 4.35 (q, J= 6.8 Hz, 2H), 5 5.17, 5.31 (br m each, 1H), 6.90 (d,J= 8.3 Hz, 1H), 7.75 (d,J= 2.0 Hz, 1H), 7.65 (dd, J= 2.0, 8.3 Hz, 1H11); MS (FAB) m/z 298 (M+1). A mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureido ]phenylacetate (404.0 mg, 0.840 mmol), ethyl 4-(4-fluoro-2-pyrrolidinyl)methoxy-3-methoxybenzoate (250.0 mg, 0.840 mmol), Et 3 N (141,l, 1.009 mmol) in DMF (4.0 mL) was stirred for 1 hr at room temp. The 10 mixture was diluted with EtOAc, washed with water, brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the residue was chromatographed on silica-gel with n hexane:EtOAc (1:3, v/v) to afford 502 mg (q.y.) of ethyl 4-[[4-fluoro-1-[3-methoxy-4-[N'-(2 methylphenyl)ureido] phenylacetyl]-2-pyrrolidinyl]methoxy]-3-methoxybenzoate as a yellow oil. To a stirred solution of this compound in THF (8.0 mL) and 1H120 (2.0 mL) was added LiOH 15 (60.4mg, 2.520 mmol), and the mixture was stirred at room temp. overnight, and 50'C for 1 day. The mixture was diluted with CHCl 3 , and extracted with IN NaOH. The aqueous layer was acidified by the addition of IN HCI and extracted with CHCI 3 . The extract was washed with brine and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the obtained crude solid was recrystallized from EtOAc-CHCl 3 -EtOH-n-hexane to afford 294.8 mg (62%) 45 as a 20 white crystalline powder. IR (KBr) 2958, 2937, 1687, 1601, 1531, 1454, 1419, 1267, 1214, 1029cm'; 'H-NMR (400 MHz, DMSO-d 6 ) 5 1.86 - 2.09 (min, 5H), 2.06 (s, 3H), 2.25 (s, 3H), 3.47 3.67 (min, 6H), 3.76 (s, 3H), 4.05 - 4.12 (min, 2H), 4.30 - 4.31 (min, 1H11), 6.51 (s, 1H1), 6.55 (s, 1H), 6.73 - 6.95 (min, 2H1), 7.11 - 7.17 (min, 2H), 7.64 (s, 1H), 7.79 (d, J= 7.8 Hz, 1H1), 7.99 (d, J= 7.8 Hz, 1H), 8.47 (s, 1H1), 8.55 (s, 1H); MS (FAB) m/z 566 (M +1); Anal Calcd for 25 C 3 0H 32
FN
3 0 7 1/2H20: C, 62.71; H, 5.79; F, 3.31; N, 7.31. Found: C, 63.13; H, 6.17; F, 3.12; N, 7.04. Example 42 3-acetylamino-4-[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl methoxy]benzoic acid 171 WO 01/00206 PCT/US00/18079 0 N N COOH H H 46 A solution of 3-amino-4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-2 pyrrolidinylmethoxylbenzoic acid (130mg, 0.244mmol) and DMAP (2.9mg, 0.0244mmol) in pyridine (5mL) and acetic anhydride (5mL) was stirred for 2 hr at room temp. The mixture was 5 evaporated off in vacuo (exess acetic anhydride was azeotropically removed with toluene). Water was added to the residue, and extracted with CHCl 3 . The extract was dried over NaSO 4 and evaporated in vacuo. The residue was chromatographed on silica-gel with MeOH:CHCl 3 (1:15 to 1:1, v/v) as eluent to afford 29mg (21%) 46 as a white crystalline material. 'H-NMR (DMSO-d 6 ) 8 1.80-2.30 (m, 4H), 2.04 (s, 3H), 2.26 (s, 3H), 3.33(s, 3H), 3.40-4.80 (m, 7H), 6.59 (s, 1H), 6.74(d, 10 J=8.3Hz, 1H), 6.79 (d, J=8.8Hz, 1H), 7.07-7.57 (m, 6H), 7.75 (d, J=8.8Hz, 1H11), 8.07 (d, J=8.3Hz, 1H), 8.41 and 8.96 (each s, each 1H); MS (FAB) m/z 575 (M+I). Example 43 3-chloro-2-[[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl] methoxy] pyridine-5-carboxylic acid N N 15 N N COOH 47 To a stirred solution of 3-chloro-2-hydroxypyridine-5-carboxylic acid (1 g, 5.762 mmol) in benzene (16 mL) and MeOH (4 mL) was added dropwise TMSCHN 2 (3.17 mL, 6.338 mmol) at 0 0 C, and the resulting mixture was stirred overnight at room temp. The reaction mixture was quenched by the addition of AcOH and the mixture was evaporated off. The residue was 20 suspended in water and precipitate was collected. The crude solid was washed with Et 2 0, and dried under a reduced pressure to give 728.1 mg (67%) methyl 3-chloro-2-hydroxypyridine-5 carboxylate as a white crystalline powder. IR (KBr) 1655, 1282, 1245, 769cm'; 'H-NMR (400 MHz, DMSO-d6) 8 3.79 (s, 3H), 8.01 (s, 1H), 8.06 (s, 1); MS (FAB) m/z 188 (M'+1); Anal. Calcd for C 7
H
6
CINO
3 : C, 44.82; H, 3.22; Cl, 18.90; N, 7.47. Found: C, 44.74; H, 3.22; Cl, 19.00; 25 N, 7.34. To a stirred solution of methyl 3-chloro-2-hydroxypyridine-5-carboxylate (300 mg, 1.599 mmol), N-tert-butoxycalbonylprolinol (321.9 mg, 1.599 mmol), and Ph 3 P (503 mg, 1.919 mmol) in 172 WO 01/00206 PCT/US00/18079 THF (3 mL) was slowly added DIAD (378 p,1, 1.919 mmol) at room temp, and the mixture was stirred for 13 hr at 700C. The mixture was concentrated and the residue was chromatographed on silica-gel with n-hexane - EtOAc (3:1, v/v) as eluent to give 235.6 mg (40%) methyl 3-chloro-2 [[1-(tert-butoxycarbonyl)-2-pyrrolidinyl]methoxy]pyridine-5-carboxylate as a pale yellowish oil. 5 1 H-NMR (400 MHz, CDCl 3 ) 8 1.46 (s, 9H), 1.87 (m, 1H), 2.05 (br s, 3H11), 3.43 (br s, 2H), 3.92 (s, 3H), 4.17, 4.26 (br s each, 1H), 4.45 - 4.51 (m, 1H), 4.50 (s, 1H1), 8.21 (s, 1H), 8.67 (d, J= 2.0 Hz, 1H); MS (FAB) m/z 371 (M'+1). To a stirred solution of methyl 3-chloro-2-[[1-(tert-butoxycarbonyl)-2 pyrrolidinyl]methoxy] pyridine-5-carboxylate (235.6mg, 0.635 mmol) in CH 2
C
2 (5.0 mL) was 10 added TFA (1.0 mL) at 0oC, and the reaction mixture was stirred at room temp for 2 hr. The solvent was removed under a reduced pressure. The residue was made basic by the addition of IN NaOH and extracted with CHCl 3 . The extract was dried over Na 2
SO
4 , concentrated under a reduced pressure to afford 172.3 mg (q.y.) methyl 3-chloro-2-[(2-pyrrolidinyl)methoxy]pyridine-5 carboxylate as a pale yellowish oil. 'H-NMR (400 MHz, CDCl 3 ) 8 1.55 - 1.63 (m, 1H), 1.76 - 1.99 15 (m, 3H), 2.93 - 2.99 (m, 1), 3.02 - 3.08 (m, 1H), 3.57 -3.62 (m, 1H), 3.92 (s, 3H), 4.33 (dd, J = 7.3, 10.7 Hz, 1H), 4.44 (dd, J= 4.4, 10.7 Hz, 1H), 8.21 (d, J = 2.0 Hz, 1H), 8.67 (d, J= 2.0 Hz, 1H); MS (FAB) m/z 271 (M +I). The mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetate (317.0 mg, 0.660 mmol), methyl 3-chloro-2-[(2-pyrrolidinyl)methoxy] pyridine-5 20 carboxylate (172.0 mg, 0.635 mmol), Et 3 N (105 ul, 0.756 mmol) in DMF (2.0 mL) was stirred for 1 hr at room temp. The mixture was diluted with EtOAc, washed with brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure to afford methyl 3-chloro-2-[[1-[3 methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-2-pyrrolidinyl]methoxy]Jpyridine-5 carboxylate as a brownish oil. To a stirred solution of this compound in THF (6.0 mL) and H 2 0 25 (2.0 mL) was added LiOH (45.3 mg, 1.89 mmol), and the reaction mixture was stirred for 5 hr at room temp. The mixture was diluted with n-hexane and extracted with IN-NaOH. The aqueous layer was acidified by the addition of IN HCI and extracted with CHC 3 . The extract was washed with brine, dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the obtained crude solid was recrystallized from n-hexane-EtOAc-EtOH to afford 242.2 mg (70%) 47 30 as an orange crystalline powder. mp 122 - 125; IR (KBr) 3354, 1709, 1593, 1535, 1454, 1257cm'1; 'H-NMR (400 MHz, DMSO-d 6 ) 8 1.67 - 2.03 (m, 4H), 2.50 (s, 3H), 3.33 - 3.42 (m, 1H), 3.52 (m, 2H), 3.58 (d, J= 4.4 Hz, 1H), 3.83 (s, 3H), 4.27 - 4.31 (m, 2H), 4.42 - 4.47 (m, 1H), 6.73 (d, J= 173 WO 01/00206 PCT/US00/18079 7.8 Hz, 1H), 6.87 -6.95 (m, 3H), 7.11 -7.17 (m, 2H), 7.79 (d,J = 8.3 Hz, 1H), 7.99 (d,J= 8.3 Hz, 1H), 8.14 (dd, J= 2.0, 8.8 Hz, 1H), 8.46 (s, 1H), 8.56 (s, 1H), 8.69 (d, J= 2.0 Hz, 1H), 13.06 (br s, 1H); MS (FAB) m/z 553 (M++1); Anal. Calcd for C 28
H
29
CIN
4 0 6 : C, 60.81; H, 5.29; N, 10.31. Found: C, 60.98; H, 5.50; N, 9.46. 5 Example 44 2-[[1-[4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenylacetyl]-2-pyrrolidinyl ]methoxy] pyridine-5 carboxylic acid HK H (A -COOH 48 To a stirred solution of 6-hydroxynicotinic acid (2 g, 14.38 mmol) in benzene (32 mL) 10 and MeOH (8 mL) was added dropwise TMSCHN 2 (1.97 mL, 3.953 mmol) at 0oC, and the resulting mixture was stirred for 2hr at room temp. The mixture was quenched by the addition of AcOH and concentrated in vacuo. The residue was suspended in water and the solid was collected. The crude solid was washed with Et20, and dried in vacuo to give 1.566 g (71%) methyl 2 hydroxypyridine-5-carboxylate as a pale brown crystalline powder. IR (KBr) 1655, 1645, 1610, 15 1433, 1300, 1113, 777, 642cm-'; 'H-NMR (400 MHz, DMSO-d 6 ) 8 3.77 (s, 3H), 6.37 (d, J= 9.8 Hz, 1H), 7.99 (dd, J= 2.4, 9.8 Hz, 1H), 8.03 (d, J= 2.4 Hz, 1H); MS (FAB) nm/z. 154 (M*+1); Anal. Calcd for CTHTNO 3 : C, 54.90; H, 4.61; N, 9.15. Found: C, 54.89; H, 4.60; N, 9.13. To a stirred solution of methyl 2-hydroxypyridine-5-carboxylate (1.00 g, 6.529 mmol), N tert-butoxycarbonylprolinol (1.31 g, 6.529 mmol), and PhP (2.06 g, 7.836 mmol) in THF (10 mL) 20 was added DIAD (1.54 mL, 7.836 mmol) at room temp, and the resulting mixture was stirred for 13 hr at 70'C. The mixture was concentrated and the residue was chromatographed on silica-gel with n-hexane:EtOAc (3:1, v/v) as eluent to give 712.3 mg (32%) methyl 2-[[1-(tert butoxycarbonyl)-2-pyrrolidinyl] methoxy]pyridine-5-carboxylate as a pale yellow oil. 'H-NMR (400 MHz, CDCl 3 ) 6 1.47 (s, 9H), 1.85 - 1.98 (m, 4H), 3.37 (br s, 2H), 3.92 (s, 3H), 4.12 - 4.33 (br 25 m, 2H), 4.48 (brs, 1H), 6.75 (m, 1H), 8.15 (m, 1H), 8.79 (m, 1H); MS (FAB) m/z 337 (M'+1). To a stirred solution of methyl 2-[[1-(tert-butoxycarbonyl)-2 pyrrolidinyl]methyloxy]pyridine -5-carboxylate (232.3mg, 0.691 mmol) in CH 2 C1 2 (4.6 mL) was added TFA (0.9 mL) at 0oC, and the reaction mixture was stirred at room temp for 2 hr. The solvent was removed under a reduced pressure and the residue was made basic by the addition of 30 IN NaOH. The aqueous solution was extracted with CHC1 3 , washed with brine, and the dried 174 WO 01/00206 PCT/US00/18079 over Na 2
SO
4 . The solvent was evaporated under a reduced pressure to afford 146.2 mg (90%) methyl 2-(2-pyrrolidinyl)methoxypyridine-5-carboxylate as an oil. 'H-NMR (400 MHz, CDCl 3 ) 8 1.49 - 1.58 (m, 1H), 1.72 - 2.18 (m, 3H), 2.92 - 3.05 (m, 2H), 3.50 -3.57 (m, 1H), 3.91 (s, 3H), 4.23 (dd, J= 8.0, 10.7 Hz, 1H), 4.38 (dd, J= 4.4, 10.3 Hz, 1H), 6.78 (d, J= 8.8 Hz, 1H), 8.15 (dd, 5 J = 2.4, 8.8 Hz, 1H), 8.80 (d, J= 2.4 Hz, 1H); MS (FAB) m/z 237 (M+1). The mixture of pentafluorophenyl 4-[N'-(2-fluorophenyl)ureido]-3-methoxy-phenylacetate (314.8 mg, 0.650 mmol), methyl 2-[(2-pyrrolidinyl)methoxy]pyridine-5-carboxylate (146.2 mg, 0.619 mmol), Et 3 N (103 ul, 0.743 mmol) in DMF (1.5 mL) was stirred for 1 hr at room temp. The mixture was diluted with Et 2 O, washed with brine, and dried over Na 2
SO
4 . The solvent was 10 removed under a reduced pressure to afford methyl 2-[[1-[4-[N'-(2-fluorophenyl)ureido]-3 methoxyphenylacetyl]-2-pyrrolidinyl] methoxy]pyridine-5-carboxylate as a crude pale yellow oil. To a stirred solution of this compound in THF (6.0 mL) and H20 (2.0 mL) was added LiOH (44.5 mg, 1.857 mmol), and the reaction mixture was stirred for 17 hr at room temp. The mixture was diluted with n-hexane and made basic by the addition of IN NaOH. The aqueous 15 layer was acidified by IN HCI and extracted with CHCl 3 . The extract was washed with brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure and the obtained crude solid was recrystallized from n-hexane-EtOAc-EtOH to afford 202.5 mg (63%) 48 as a white crystalline powder. IR (KBr) 1602, 1537, 1456, 1265, 752cm-'; H-NMR (400 MHz, DMSO-d 6 ) 8 1.67 - 2.03 (m, 4H), 2.50 (s, 3H11), 3.33 - 3.42 (m, 1H), 3.52 (m, 2H), 3.58 (d, J= 4.4 Hz, 1H11), 3.83 20 (s, 3H), 4.27- 4.31 (m, 2H), 4.42- 4.47 (m, 1H1), 6.73 (d, J= 7.8 Hz, 1H11), 6.87- 6.95 (m, 3H), 7.11 -7.17 (m, 2H11), 7.79 (d,J= 8.3 Hz, 1H), 7.99 (d,J= 8.3 Hz, 1H), 8.14 (dd, J= 2.0, 8.8 Hz, 1H), 8.46 (s, 1H), 8.56 (s, 1H11), 8.69 (d, J = 2.0 Hz, IH), 13.06 (br s, 1H); MS (FAB) m/z 523 (M'+1); Anal. Calcd for C 27
H
27
FN
4 06.1/2H 2 0: C, 61.01; H, 5.31; N, 10.54. Found: C, 61.52; H, 5.39; N, 10.01. 25 Example 45 4-[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinylmethyl]-1-piperazinyl acetic acid 0 NJCCOOH N 'KN \ 49 To a stirred suspension of 1-benzylpiperazine (5 g, 28.4 mmol) and K 2
CO
3 (5.89 g, 42.6 30 mmol) in DMF (30 mL) was added ethyl bromoacetate (4.74 g, 28.4 mmol) at room temp. The 175 WO 01/00206 PCT/US00/18079 resulting mixture was stirred for a further 3 hr. The mixture was diluted with EtOAc (300 mL), washed with brine (2 x 100 mL), dried over MgSO 4 and evaporated. The residue was chromatographed on silica-gel with CHCI 3 :EtOH (10:1, v/v) as eluent to give 7.45 g (q.y.) ethyl 4 benzyl-1-piperazinylacetate as a yellow oil. 'H-NMR (CDC 3 ) 8 1.27 (3 H, t, J= 7.3 Hz), 2.88 5 2.96 (8 H, m), 3.20 (2 H, s), 3.52 (2 H, s), 4.18 (2 H, q, J= 7.3 Hz), 7.22-7.32 (5 H, m). A stirred solution of ethyl 4-benzyl-l1-piperazinylacetate (2.00 g, 7.62 mmol) and 5% Pd/C (2 g) in AcOH:EtOH (1:1, 40 mL) was hydrogenated at 1 atm for 8 hr. The mixture was filtered and the filtrate was concentrated in vacuo. The residue was made basic by the addition of saturated NaHCO 3 and extracted with CHC1 3 (2 x 200 mL). The combined extracts were dried over 10 K 2
CO
3 and evaporated to give 1.16g (88%) ethyl 1-piperazinylacetate as a yellow oil. 'H-NMR (CDCl 3 ) 8 1.26-1.30 (3 H, m), 1.67(1 H, br s), 2.55(4 H, m), 2.92-2.96(4 H, m), 3.19-3.20(2 H, m), 4.16-4.22(2 H, m). To a stirred solution of N-Boc-L-prolinol (1.00 g, 5.02 mmol) and ethyl 1-piperazinyl acetate (864 mg, 5.02 mmol) in MeOH:AcOH (10:1, v/v, 11 mL) was added NaBH 3 CN (664 mg, 15 10.0 mmol) at room temp. After being stirred overnight, the mixture was poured into ice water (100 mL) and made basic by the addition of NaHCO 3 . The mixture was extracted with CHCl 3 (2 x 200 mL). The combined extracts were dried over Na 2
CO
3 and evaporated. The residue was chromatographed on silica-gel with CHCl 3 :EtOH (10:1, v/v) as eluent to give 1.20 g (67%) ethyl 4 [1-(tert-butoxy carbonyl)-2-pyrrolidinylmethyl]-1-piperazinylacetate as a colorless oil. 'H-NMR 20 (CDCl 3 ) 8 1.27 (3 H, t, J= 7.3 Hz), 1.46-1.47 (9 H, m), 1.79-3.96 (total 19 H, series of m), 4.19 (2 H, q, J= 7.3 Hz). A mixture of ethyl 4-[ 1-(tert-butoxycarbonyl)-2-pyrrolidinylmethyl]- 1-piperazinylacetate (1.20 g, 3.38 mmol) in TFA (5 mL) and CH 2 C1 2 (5 mL) was stirred overnight. After removal of the solvent, the residue was made basic by the addition of sat. NaHCO 3 . The mixture was 25 extracted with CHC1 3 (2 x 200 mL). The combined extracts were dried over Na 2
CO
3 and evaporated to give 386 mg (45%) ethyl 4-(2-pyrrolidinylmethyl)-1-piperazinylacetate as a yellow oil. MS (FAB) 256 (M++1). To a stirred solution of ethyl 4-(2-pyrrolidinylmethyl)-1-piperazinylacetate (380 mg, 1.49 mmol) and 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (468 mg, 1.49 mmol) in 30 DMF (10 mL) was added EDC-HCI (428 mg, 2.24 mmol), HOBt, and DMAP (cat.). After being stirred overnight, the mixture was diluted with EtOAc (300 mL), washed with brine (2 x 200 mL), 176 WO 01/00206 PCT/US00/18079 and dried over MgSO 4 . After removal of the solvent, the residue was chromatographed on silica gel with CHCl 3 :EtOH (9:1, v/v) as eluent to give 257 mg (31%) ethyl 4-[1-[3-methoxy-4-[N'-(2 methylphenyl) ureido] phenylacetyl]-2-pyrrolidinylmethyl]-l-piperazinylacetate as a yellow foam. 'H-NMR (CDCl 3 ) 5 1.24-1.29 (3 H, min), 1.69-4.24 (total 29 H, series of min), 6.41 (1 H, min), 6.81 (2 5 H, min), 7.13-7.26 (4 H, min), 7.52 (1 H, d, J= 7.3 Hz), 8.04 (1 H, d, J = 8.3 Hz). To a stirred solution of ethyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-2-pyrrolidinylmethyl]-1-piperazinylacetate (250 mg, 0.453 mmol) in THF (4 mL) was added 0.25 N NaOH (3.6 mL). The resulting mixture was stirred overnight. The mixture was adjusted to pH 7.5 by the addition of 1 N HCI and extracted with CHCl 3 :MeOH (4:1, 3 x 100 mL). 10 The combined extracts were dried over MgSO 4 and evaporated. The crude solid was recrystallized from CHCl 3 -MeOH-n-hexane to give 40 mg (17%) 49 as a colorless crystalline powder. mp 160 170 oC; 'H-NMR (DMSO-d 6 ) 8 1.74-4.08 (total 27 H, series of min), 6.73 (1 H, d, J= 7.8 Hz), 6.87 (1 H, s), 6.93 (1 H, t, J= 7.8 Hz), 7.11-7.17 (2 H, min), 7.79 (1 H, d, J= 7.8 Hz), 8.00 (1 H, dd, J= 7.8, 2.4 Hz), 8.47 (1 H, s), 8.56 (1 H, s); MS (FAB) 524 (M+1); Anal. Calcd for 15 C 28
H
37 N0 5 -HCl-H 2 0: C, 58.17; H, 6.97; N, 12.11. Found: C, 58.26; H, 7.26; N, 11.53. Example 46 N N J OH H H H 50 To a suspension of 4-aminophenylacetic acid (10 g, 66 mmol) in 1:1 CH 2 C1 2 :acetone (100 mnL) was added o-tolyisocyanate (8.8 g, 66 imol). The mixture was heated to reflux for 4 hr at 20 which time a white precipitate had formed. The precipitate was filtered and the solid washed generously with 1:1 CH 2 Cl 2 :acetone. The solid was recrystallized with hot methanol and dried under vacuum to yield 14.1 g (75% yield) of the desired 4-(o-tolylureido)phenylacetic acid 50. Example 47 NINI H H O OH 51 0 51 25 To a suspension of 2-amino-4-thiazoleacetic acid (4 g, 25 mmol) in 1:1 CH 2 Cl:acetone (100 mL) was added o-tolylisocyanate (3.5 g, 26 mmol). The mixture was heated to reflux for 8 hr at which time a yellow precipitate had formed. the precipitate was filtered and the solid washed generously with 1:1 CH 2 C1 2 :acetone. The solid was recrystallized with hot methanol and dried under vaccum to yield 4.8 g (66% yield) of the desired 2-(o-tolylureido)-4-thiazoleacetic acid 51. 177 WO 01/00206 PCT/US00/18079 Example 48 4-[[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinylmethylamino]benzoic acid 52 5 A stirred mixture of methyl 4-aminobenzoate (1.52 g, 10.04mmol) and 1-tert-butoxy carbonyl prolinal (3.00 g, 15.06 mmol) in toluene (30 mnL) was heated under reflux for 3 hr. After cooling to room temp, the solvent was evaporated in vacuo. The solid was dissolved in MeOH (27 mL) and AcOH (3 mL), then NaBH 3 CN (1.33 g, 20.08mmol) was added to the mixture, and the resulting mixture was stirred overnight at room temp. The reaction mixture was quenched with 10 water, and the solvent was removed under a reduced pressure. Water was added to the residue, and extracted with EtOAc. The extract was washed with H20, brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure, and the residue was chromatographed on silica-gel with n-hexane - EtOAc (3:1, v/v) as eluent to afford 2.17g (65%) 4-[1-(tert-butoxycarbonyl)-2 pyrrolidinylmethylamino] benzoate as a pale yellow oil. 'H-NMR (400 MHz, CDCl 3 ) 8 1.48 (s, 15 9H11), 1.51 - 2.09 (m, 4H), 3.05 - 3.07 and 3.43 - 3.48 (br m, 1H), 3.18 (br s, 1H), 3.36 (br s, 2H), 3.84 (s, 1H),4.06 - 4.08, 4.20 - 4.24 (br m each, 1H11), 6.49 - 6.65 (m, 2H), 7.84 (d, J = 8.3 Hz, 2H); MS (FAB) m/z 335 (M +1). To a stirred solution of methyl 4-[1-(tert-butoxycarbonyl)-2-pyrrolidinylmethylamino] benzoate (2.17 g, 6.490 mmol) in CH 2 C1 2 (44 mL) was added TFA (8.7 mL) at 0oC, and the 20 resulting mixture was stirred overnight at room temp. The solvent was removed under a reduced pressure and the residue was treated with IN NaOH. The mixture was extracted with CH 2 Cl 2 . The extract was washed with brine, dried over Na 2
SO
4 , and the solvent was concentrated under a reduced pressure to afford 1.34 g (88%) methyl 4-(2-pyrrolidinylmethylamino)benzoate as a brown oil, which is used to the subsequent reaction without further purification. 25 The mixture of the above methyl 4-(2-pyrrolidinylmethylamino)benzoate (397.8 mg, 1.69 mmol), 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (587.1 mg, 1.87mmol), EDC (HC1) (486 mg, 2.54 mmol), HOBt (23 mg, 0.17 mmol), and DMAP (21 mg, 0.17 mmol) in DMF (4 mL) was stirred overnight at room temp. The mixture was diluted with EtOAc, washed with brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure. The residue 30 was chromatographed on silica-gel with CHCl 3 - MeOH (50:1, v/v) as eluent to afford 882mg 178 WO 01/00206 PCT/US00/18079 (98%) methyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl methylamino]benzoate as a brown amorphous solid, which is used to the subsequent reaction without further purification. To a stirred solution of the above methyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] 5 phenylacetyl]-2-pyrrolidinylmethylamino]benzoate (882 mg, 1.662 mmol) in THF (18 mL) and MeOH (5.0 mL) was added IN NaOH (5.0 mL, 5.000 mmol), and the mixture was heated under reflux for 3 days. The mixture was concentrated. The residue was treated with IN HCI and extracted with CH 2 C1 2 . The extract was washed with brine, dried over Na 2
SO
4 , and evaporated in vacuo. The solid was recrystallized from n-hexane - diisopropyl ether - CHC1 3 - MeOH to afford 10 180.5 mg (21%) 52 as a pale yellow amorphous solid. IR (KBr) 1604, 1535, 1511, 1454, 1255, 1224, 1174cm-'; 'H-NMR (400 MHz, DMSO-d 6 ) 8 1.79 - 1.99 (br m, 4H), 2.25 (s, 3H), 2.90 - 2.94 (min, 1H), 3.35 - 3.62 (min, 6H), 3.87 (s, 3H), 4.12 - 4.15 (br s, 1H), 6.63 - 6.78 (min, 4H), 6.89 - 6.95 and 7.11 - 7.17 (m each, 3H), 7.65 (d,J= 8.3 Hz, 2H), 7.80 (d, J = 8.3 Hz, 1H), 8.02 (d,J= 8.3 Hz, 1H), 8.47 (s, 1H), 8.57 (s, 1H), 12.0 (br s, 1H); MS (FAB) m/z 517 (M+1); Anal. Calcd for 15 C 2 9H 32
N
4 Os 5 1H 2 0: C, 65.15; H, 6.41; N, 10.48. Found: C, 65.45; H, 6.33; N, 10.02. Example 49 4-[N-[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinylmethyl]-N methylamino]benzoic acid H H OMe COOH 53 20 To a mixture of methyl 4-[N-(2-pyrrolidinyl)methylamino]benzoate (600 mg, 1.794 mmol), 37%-formaldehyde (1.79 mL, 23.32 mmol), and NaBH 3 CN (368 mg, 5.561 mmol) in
CH
3 CN (6.0 mL) was added dropwise AcOH (0.205 mL, 3.588 mmol), and the resulting mixture was stirred for 2 hr at room temp. The reaction mixture was quenched by the addition of sat. NaHCO 3 , and extracted with EtOAc. The extract was washed with brine and dried over Na 2
SO
4 . 25 The solvent was removed under a reduced pressure, and the residue was chromatographed on silica-gel with n-hexane - EtOAc (3:1, v/v) as eluent to afford 645 mng (100%) methyl 4-[N-[1 (tert-butoxycarbonyl)-2-pyrrolidinylmethyl]-N-methylamino]benzoate as a colorless oil. 'H-NMR (400 MHz, CDC1 3 ) 8 1.50 (s, 9H), 1.76 - 1.91 (min, 4H), 3.07 (s, 3H), 3.15 - 3.43 (min, 3H), 3.67 3.71 (min, 1H), 3.85 (s, 3H), 4.11 -4.17 (min, 1H), 4.37 (s, 1H), 6.75 (d,J= 8.3 Hz, 2H), 7.89 (d,J= 30 8.8 Hz, 2H); MS (FAB) m/z 349 (M +I). 179 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[N-[1-(tert-butoxycarbonyl)-2-pyrrolidinylmethyl]-N methylamino]benzoate (645 mg, 1.794 mmol) in CH 2 C1 2 (6.5 mL) was added TFA (1.3 mL) at 0OC, and the mixture was stirred overnight at room temp. The solvent was removed under a reduced pressure and the residue was treated with IN NaOH solution. The mixture was extracted with 5 CH 2 Cl 2 . The extract was washed with brine, dried over Na 2
SO
4 , and the solvent was concentrated under a reduced pressure to afford 363.2 mg (82%) of methyl 4-[N-(2-pyrrolidinyl)methyl-N methyl]aminobenzoate as a yellowish oil, which is used to the subsequent reaction without further purification. The mixture of methyl 4-[N-(2-pyrrolidinyl)methyl-N-methyl]amninobenzoate (191.8 mg, 10 0.772 mmol), 3-methoxy-4-(N'-2-methylphenylureido)phenylacetic acid (258.1 mg, 0.811 mmol), EDC (hydrochloride) (221.9 mg, 1.158 mmol), HOBt (10.0 mg, 0.077 mmol), and DMAP (9.4 mg, 0.077 mmol) in DMF (2.0 mL) was stirred for 3 hr at room temp. The reaction mixture was diluted with Et20, washed with brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure to afford 482.5 mg methyl 4-[N-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] 15 phenylacetyl]-2-pyrrolidinylmethyl]-N-methylamino]benzoate as a white amorphous powder, which is used to the subsequent reaction without further purification. To a stirred solution of methyl 4-[N-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl acetyl]-2-pyrrolidinylmethyl]-N-methylamino]benzoate in THF (5.0 mL) was added IN NaOH (6.2 mL, 6.2 mmol), and the mixture was heated under reflux for 3 days. The reaction mixture was 20 concentrated in vacuo. The residue was neutralized with IN HC1, and extracted with CH 2 C1 2 . The extract was washed with sat. NH 4 C1, brine, dried over Na 2
SO
4 , and evaporated in vacuo. The crude solid was recrystallized from n-hexane-CHCl 3 -MeOH-isopropylether to afford 102.8 mg (25%, 2steps) 53 as a pale yellow amorphous solid. mp 144 - 146; IR (KBr) 3325, 1600, 1529, 1454, 1284, 1257, 1184cm-'; 'H-NMR (400 MHz, DMSO-d 6 ) 8 1.73 - 1.91 (m, 3H11), 2.03 -2.11 (m, 25 1H), 3.03 (s, 3H), 3.16 (dd, J= 9.3, 14.2 Hz, 1H), 3.37 - 3.60 (m, 4H), 3.76 - 3.80 (m, 1H14), 3.86 (s, 3H), 4.25 (br s, 1H), 6.75 (dd, J= 1.5, 8.3 Hz, 1H), 6.86 (d, J= 1.5 Hz, 1H), 6.90 (d, J= 8.8 Hz, 2H), 6.95 - 7.01 (m, 1H), 7.12 (t, J= 7.8 Hz, 1H1), 7.20 - 7.25 (m, 1H1), 7.73 (d, J= 8.8 Hz, 2H), 8.01 (d, J= 7.8 Hz, 1H14), 8.16 - 8.20 (m, 1H), 8.73 (s, 11), 9.19 (d, J= 2.0 Hz, 1H), 12.0 (br s, 1H); MS (FAB) m/z 535 (M+1); Anal Calcd for C 2
H
31
FN
4 Os 5 -1/2H20: C, 64.08; H, 5.93; N, 30 10.31; F, 3.49. Found: C, 64.17; H, 5.84; N, 10.06; F, 3.26. Example 50 4-[N-[ -[-4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenylacetyl]-2-pyrrffolidinylmethyl]-N 180 WO 01/00206 PCT/US00/18079 methylamino]-3-nitrobenzoic acid F OMe 0 2 N COOH 54 54 To a mixture of methyl 4-fluoro-3-nitrobenzoate (1.58 g, 4.666 mmol) and [l-(tert butoxycarbonyl)-2-pyrrolidinyl]methylamine (500 mg, 2.333 nmmol) in DMF (8.0 mL) was added 5 K 2 COz (967 mg, 6.999 mmol), the resulting mixture was stirred for 3 hr at room temp. The reaction mixture was diluted with EtOAc, washed with water, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure, and the residue was chromatographed on silica-gel with n hexane - EtOAc (3:1, v/v) as eluent to afford 834.9 mg (91%) of methyl 4-[N-[1-(tert butoxycarbonyl)-2-pyrrolidinylmethyl]-N-methyl]amino-3-nitrobenzoate as a pale yellow oil, 10 which is used to the subsequent reaction without further purification. To a ice-cooling solution of the above oil in CH 2
C
2 (8.3 mL) was added TFA (1.7 mL), and the reaction mixture was stirred overnight at room temp. The solvent was removed under a reduced pressure. The residue was treated with IN-NaOH and extracted with CHC1 3 . The extract was washed with brine, dried over Na 2
SO
4 , and evaporated under a reduced pressure to afford 15 553.6 mg (90%) methyl 4-[N-(2-pyrrolidinylmethyl)-N-methyl]amino-3-nitrobenzoate as a pale yellow oil. 'H-NMR (400 MHz, CDCl 3 ) 8 1.31 - 1.40 (min, 1H), 1.74 - 2.05 (mn, 4H), 2.73 - 2.79 (inm, 1H), 2.81 - 2.99 (min, 1H), 2.94 (s, 3H), 3.29 -3.55 (min, 2H), 3.89 (s, 3H), 7.14 (d, J= 9.3 Hz, 111), 7.98 (dd, J= 2.0, 8.8 Hz, 1H), 8.42 (d, J= 2.0 Hz, 1H); MS (FAB) m/z 294 (M 4 +1). A mixture of 3-methoxy-4-[N'-(2-fluorophenyl)ureido]phenylacetic acid (630.0 mg, 1.979 20 mmol), methyl 3-nitro-4-[N-(2-pyrrolidinyl)methyl-N-methylamino]benzoate (553.0 mg, 1.885 mmol), EDC(Hydrochloride) (542.0 mg, 2.827 mmol), HOBt (25.5 mg, 0.189 mmol), and DMAP (23.1 mg, 0.189 mmol) in DMF (5.0 mL) was stirred at room temp. for 2 hr. The mixture was diluted with Et 2 0, washed with brine, and dried over Na 2
SO
4 . The solvent was removed under a reduced pressure, and the residue was chromatographed on silica-gel with CHCI 3 - MeOH (30:1, 25 v/v) as eluent to afford 1.18 g (100%) methyl 4-[N-[1-[3-methoxy-4-[N'-(2-fluorophenyl)ureido] phenylacetyl]-2-pyrrolidinyl methyl]-N-methylamino]-3-nitrobenzoate as a yellow foam, which is used to the subsequent reaction without further purification. To a stirred solution of the above methyl 4-[N-[1-[3-methoxy-4-[N'-(2-fluorophenyl) ureido]phenylacetyl]-2-pyrrolidinylmethyl]-N-methylamino]-3-nitrobenzoate (2.50 mg, 0.421 181 WO 01/00206 PCT/US00/18079 mmol) in THF (3.0 mL) was added IN NaOH (1.5 mL, 1.500 mmol), and the mixture was heated under reflux overnight. After cooling, the mixture was concentrated to a small volume. The residue was treated with IN HCI, and extracted with CHCl 3 . The extract was washed with brine, dried over Na 2
SO
4 , and evaporated in vacuo. The crude solid was recrystallized from n-hexane 5 diethyl ether - CHCl 3 - MeOH to afford 194.9 mg (80%) of 54 as a yellow amorphous solid. IR (KBr) 1685, 1610, 1529, 1454, 1284, 1259, 1228 cm'; 'H-NMR (400 MHz, DMSO-d) 8 1.63 1.91 (min, 3H), 2.04 - 2.07 (br s, 1H), 2.60 (br s, 11), 2.80 (s, 1H), 2.99 (s, 2H), 3.05 - 3.10 (mn, 1H), 3.32 - 3.58 (min, 3H), 3.76 - 3.81 (mn, 1H), 3.81 (s, 3H), 4.25 (br s, 1H), 6.68 (t, J= 3.9 Hz, 1H), 6.71 (d, J= 8.8 Hz, 1H), 6.81 - 6.96 (min, 1H), 7.07 (t, J = 7.3 Hz, 1H), 7.17 (dd, J= 7.8, 9.8 Hz, 10 1H), 7.48 (t, J= 7.8 Hz, 1H), 7.85 (d, J= 8.8 Hz, 1H), 7.97 (t, J= 8.8 Hz, 1H), 8.11 - 8.20 (inm, 2H1), 8.68 (s, 1H), 9.14 (s, 1H), 12.8 (br s, 1H); MS (FAB) m/z 580 (M +1); Anal. Calcd for
C
2 9
H
3
FN
5 071/41-120: C, 59.63; H, 5.26; N, 11.99; F, 3.25.Found:C,59.68;H,5.34;N, 11.80; F, 3.21. Example 51 3-amino-4-[N-methyl-[ 1 -[4-[N'-(2-fluorophenyl)ureido]-3-methoxy-phenylacetyl]-2-pyrrolidinyl 15 methyl]-N-methylamino]benzoic acid F OMe H 2 N COOH 55 A stirred solution of methyl 4-[N-[1-[4-[N'-(2-fluorophenyl)ureido]-3-methoxy phenylacetyl]-2-pyrrolidinylmethyl]-N-methylamino]-3-nitrobenzoate (901.0 mg, 1.518 mmol) in MeOH (18.0 mL) was hydrogenated over 5%/Pd-C (1.35 g) at 45psi overnight. The mixture was 20 filtered and the filtrate was concentrated in vacuo. The residue was made basic with IN NaOH solution and extracted with CHCI 3 . The extract was washed with brine, dried over Na 2
SO
4 , and the solvent was removed under a reduced pressure. The residue was chromatographed on silica-gel with CHCl 3 - MeOH (24:1, v/v) as eluent to afford 283.7 mg (48%) methyl 3-amino-4-[N-[1-[4 [N'-(2-fluorophenyl)ureido
]
-3-methoxy-phenylacetyl]-2-pyrrolidinylmethyl]-N-methylamino] 25 benzoate as a brownish amorphous solid, which was used to the subsequent reaction without further purification. To a stirred solution of the above compound in THF (3.0 mL) was added IN NaOH solution (1.5 mL, 1.500 mmol), and the mixture was refluxed overnight. The mixture was concentrated, treated with IN HC1, and extracted with CHC1 3 . The extract was washed with brine, 30 dried over Na 2
SO
4 , and evaporated in vacuo. The solid was recrystallized from n-hexane - diethyl ether - CHCl 3 - MeOH to afford 179.8 ming (65%) 55 as a white amorphous solid. IR (KBr) 1614, 182 WO 01/00206 PCT/US00/18079 1601, 1537, 1454, 1228, 1219, 1184 cmr 1 ; 1 H-NMR (400 MHz, DMSO-d.) 8 1.60 - 2.20 (m, 4H), 2.61 - 2.68 (m, 1H), 2.89 (s, 3H), 3.13 - 3.18 (m, 1H), 3.40 - 3.61 (m, 4H), 3.85 (s, 3H), 4.01 (br m, 1H), 4.93 (br s, 2H), 6.50 -7.31 (m, 8H), 8.01 (dd, J= 2.9, 8.3 Hz, 1H), 8.18 (t,J= 8.3 Hz, 1H), 8.71 (s, 1H), 9.17(d, J= 1.5 Hz, 1H), 12.3 (br s, 1H); MS (FAB) m/z 550 (M+1); Anal. Calcd 5 for C 29
H
32
FN
5 0,-1/4H 2 0: C, 62.86; H, 5.91; N, 12.64; F, 3.43. Found: C, 62.71; H, 6.00; N, 12.39; F, 3.16. Example 52 4-[ 1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl]methylamino]benzoic acid NN ,cooH O 56 10 To a stirred mixture of methyl 4-[(2-pyrrolidinyl)methylamino]benzoate (220 mg, 0.94 mmol), 4 [N'-(2-methylphenyl)ureido]phenylacetic acid (285 mg, 0.94 mmol), 4-DMAP (140 mg, 1.13 mmol) and catalytic amount of HOBT in DMF (7 ml) was added EDC-HCI (220 mg, 1.13 mmol) at room temperature. The resulting mixture was stirred at room temperature for 20 hr. The mixture was pored into ice-water. The solid was collected, washed with water and air-dried. The 15 crude solid was purified by silica-gel (20 ml) column chromatography with CHCl 3 - EtOAc (3:1, v/v) to CHCl 3 - EtOH (9:1, v/v) as eluent to give methyl 4-[1-[4-[N'-(2-methylphenyl)ureido] phenyl acetyl]-2-pyrrolidinyl]methylamino]benzoate (400 mg, 85%) as a gum. 'H-NMR (CDCl 3 )8 1.75 2.05 (series of m, 4 H), 2.24 (s, 3 H), 3.18 and 3.27 (each m, each 1 H), 3.51 (m, 2 H), 3.60 (s, 2 H), 3.83 (s, 3 H), 4.52 (m, 1 H), 6.52 (m, 3 H), 6.81 (s, 1 H), 7.11-7.25 (series of m, 7 H), 7.53 (m, 20 1 H), 7.81 (d, J= 8.8 Hz, 2H). A mixture of methyl 4-[1-[4-[N-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl] methylamino]benzoate (280 mg, 0.56 mmol) in THF (3 ml) and 0.25N NaOH (6.8 ml, 1.75 mmol) was stirred for 3 hr at 60-70 oC. After cooling, the mixture was poured into ice-IN HC1 (3 ml). The solid was collected, washed with water and air-dried. The crude crystalline material was 25 recrystallized from CHC13-EtOH-IPE to give 56 (180 mg, 66%) as fine needles. MW 486.56 IR (KBr) n 3367, 3294, 1712, 1606, 1539 cmr'; 'H-NMR (CDCl 3 -DMSO-d 6 ) 8 1.80-2.05 (series of m, 4 H), 2.26 (s, 3 H), 2.94(m, 1 H), 3.38 and 3.56 (series of m, 3 H), 3.57 (s, 2 H), 4.23 (m,2 H), 6.48 (br s, 1 H), 6.69 (d, J = 8.8 Hz, 2H), 6.91 (t, J = 7 Hz, 1 H), 6.91 (m, 4 H), 7.39 (d, J = 8.3 Hz, 2H), 7.66 (d, J= 8.8 Hz, 2H), 7.80 (m, 2 H), 8.88 (s, 1 H), 11.76 (s, 1 H); MS(FAB) m/z 487 30 (M +1); Anal. Calcd for C 28
H
30
N
4 0 4 -0.75xH20: C, 67.24; H, 6.45; N, 11.20. Found: C, 67.13; H, 6.32; N, 11.01. 183 WO 01/00206 PCT/US00/18079 Example 53 methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxylphenylacetyl]-(2S)-pyrrolidinyl] methoxybenzoate r? O - COOMe I HH 6Me 57 5 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxylphenylacetyl]-(2S)-pyrrffolidinyl]methoxybenzoic acid NO N O COOH iHH Me 58 To a stirred solution of 2S-pyrrolidinemethanol (15.1 g, 149.5 mmol) in dioxane (100 ml) was added a solution of (Boc) 2 0 (32.6 g, 164.4 mmol) in dioxane (100 ml). The reaction mixture was 10 stirred at room temperature for 18 hr, and concentrated in vacuo. The residue was purified by column chromatography on silica gel with EtOAc-hexane (1:5 ,v/v) as eluent to give (1-tert butoxycarbonyl-(2S)-pyrrolidinyl)methanol (31.6 g, quant.) as a colorless oil. 'H-NMR (CDC13) 8 1.47 (s, 9H11), 1.60-2.00 (m, 3H), 3.25-3.70 (4H, m), 3.92-4.00 (m, 1H). To a stirred solution of (1-tert-butoxycarbonyl-(2S)-pyrrolidinyl)methanol (4.02 g, 20.0 15 mmol), methyl 4-hydroxybenzoate (3.04 g, 20.0 mmol) and Ph 3 P (6.28 g, 24.0 mmol) in THF (50 ml) was added DIAD (4.85 g, 24.0 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 18 hr. The mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (5:1, v/v) as eluent to give methyl 4 (1-tert-butoxycarbonyl-(2S)-pyrrolidinyl)methoxybenzoate (5.4 g, 81%) as a pale yellow oil. 'H 20 NMR (CDCl 3 ) 8 1.47 (s, 9H), 1.88-2.04 (m, 4H), 3.41 (m, 2H), 3.91 (s, 3H), 3.90-3.92 (m, 1H), 4.11-4.16 (m, 2H), 6.94 (d, J = 8.6 Hz, 2H11), 7.94 (d, J = 8.3 Hz, 2H1). To a stirred solution of methyl 4-(1-tert-butoxycarbonyl-2S-pyrrolidinyl)methoxybenzoate (2.1 g, 6.27 mmol) in CH 2 C1 2 (9.0 ml) was added TFA (6.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added 25 to the residue, and extracted with CH 2 C1 2 . The extract was washed with brine ,dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product (470 mg, 2.0 mmol), 4-[N'-(2 chlorophenyl) uredio]-3-methoxyphenylacetic acid (669 mg, 2.0 mmol), HOBt (405 mg, 3.0 mmol), and triethylamine (554 ml, 4.0 mmol) in THF (10.0 ml) and MeCN (10.0 ml) was added 184 WO 01/00206 PCT/US00/18079 EDC-HCI (576 mg, 3.0 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHC0 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (1:4, v/v) as 5 eluent to give 57 (900 mg, 82%) as a colorless oil. MW 552.02 'H-NMR (CDCI 3 ) 8 2.04-2.10 (m, 4H), 3.51-3.70 (m, 6H), 3.87 (s, 3H), 4.11-4.18 (m, 2H), 6.77-6.88 (m, 4H), 7.23-7.34 (m, 4H), 7.91-7.96 (m, 2H11), 8.17-8.19 (m, 1H). The mixture was stirred at 70 oC for 24 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HC1. The resulting solid was collected, washed with water, 10 and dried in vacuo to 58 (640 mg, 94%) as a white crystalline solid. MW 537.99 mp 126-130 oC; IR (KBr) 3324, 2938, 2877, 1604, 1533, 1249, 1166, 750 cm-'; 'H-NMR (DMSO-d) 8 1.93 2.05 (m, 4H11), 3.52-3.61 (m, 511), 3.82 (s, 3H11), 3.99-4.01 (m, 2H11), 4.18-4.20 (m, 1H), 4.29 (m, 1H11), 6.74-6.76 (d, 1H, J = 8.3 Hz), 6.87 (s, 1H), 6.99-7.04 (m, 3H), 7.25-7.29 (m, 1H), 7.41-7.43 (d, 1H, J= 8.1 Hz), 7.86-7.91 (m, 2H11), 7.95-7.97 (m, 1H1), 8.09-8.11 (d, 1H, J= 8.3 Hz), 8.87-8.92 15 (m, 1H); MS (FAB) m/z 538 (M'+1); Anal. calcd for C 28 H2 8
N
3 0 6 -0.5H 2 0: C, 61.48; H, 5.34; N, 7.68; Cl, 6.48. Found: C, 61.46; H, 5.36; N, 7.62; Cl, 6.50. For Na salt of 58 : Anal. Calcd for
C
28
H
27
N
3 0 6 -Na-1.5H 2 0: C, 57.29; H, 5.15; N, 7.16. Found: C, 57.48; H, 5.04; N, 6.99. Example 54 4-[ 1 -[4-[N'-(2-bromophenyl)ureidol-3-methoxylphenylacetyll-(2S)-pyrrolidinyl]methoxybenzoic 20 acid I N N OQ N ___, COOH Br H H OMe 59 To a stirred solution of methyl 4-(2S-pyrrolidinyl)methoxybenzoate (470 mg, 2.0 mmol), 4-[N'-(2 bromophenyl)uredio]-3-methoxyphenylacetic acid (758 mg, 2.0 mmol), HOBt (405 mg, 3.0 mmol), and triethylamine (554 ml, 3.0 mmol) in THF (10.0 ml) and MeCN (10.0 ml) was added EDC-HCI 25 (576 mg, 3.0 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (1:4, v/v) as eluent to give methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxylphenylacetyl]-(2S)-pyrrolidinyl] 30 methoxybenzoate (1.0 g, 84%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 2.04-2.10 (m, 4H), 3.52-3.54 (m, 1H1), 3.62 (s, 2H), 3.70 (s, 3H11), 3.88 (s, 3H), 4.13-4.19 (m, 2H), 6.79-6.94 (m, 4H), 7.20-7.31 185 WO 01/00206 PCT/US00/18079 (m, 1H), 7.91-8.12 (m, 2H), 8.13-8.15 (m, 1H). To a stirred solution of methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxylphenylacetyl]-(2S) pyrrolidinyl]methoxybenzoate (780 mg, 1.31 mmol) in THF (10.0 ml) and MeOH (5.0 ml) was added IN NaOH (2.0 ml, 2.0 mmol). The mixture was stirred at 70 oC for 24 hr. The mixture 5 was concentrated in vacuo, water was added thereto, and neutralized with IN HC1. The resulting solid was collected, washed with water, and dried in vacuo to give 59 (730 mg, 96%) as a white crystalline solid. MW 582.44 mp 120-125 'C; IR (KBr) 3318, 2938, 1604, 1529, 1166, 1025 cm 1; 'H-NMR (DMSO-d 6 )8 1.92-1.96 (m, 4H), 3.52-3.60 (m, 5H), 3.82 (s, 3H), 3.98-4.02 (m,1 H), 4.16-4.19 (m, 1H), 4.29 (m, 1H), 6.75 (d, J= 8.3 Hz, 1H), 6.87 (m, lH), 6.94-7.04 (m, 3H), 7.29 10 7.33 (m, 1H), 7.57-7.59 (m, 1H), 7.85-7.96 (m, 4H), 8.72 (s, 1H), 8.91 (s, 1H); MS (FAB) m/z 582 (M+1); Anal. calcd for C 28
H
28 N30Br- 1.0H 2 0: C, 56.01; H, 5.04; N, 7.00; Br, 13.31. Found: C, 56.12; H, 4.98; N, 6.96; Br, 13.57.. Example 55 3-amino-4-[1-[4-[N'-(2-hydroxyphenyl)ureido]-3-methoxyphenylacetamido]-2-pyrrolidinyl 15 methoxy]benzoic acid N 'N
NH
2 OH H H OMe OOH 60 To a stirred solution of 2-nitrophenol (10.0 g, 72.0 mmol) and K 2 COz (9.96 g, 72.0 mmol) in DMF (150 mL) was added dropwise benzyl bromide (9.40 mL, 79.2 mmol) at 0 'C. After being stirred at room temperature for 3 hr, the reaction mixture was diluted with water, which was extracted 20 with Et 2 0O. The extracts were washed with brine, dried over Na 2
SO
4 , and concentrated to dryness. Chromatography of the residue with hexane - EtOAc (2 : 1, v/v) as eluent gave 2-benzyloxy nitrobenzene (14.7 g, 89%) as a yellow oil. 1H-NMR (CDCl 3 ) 8 5.24 (s, 2H), 7.04 (t, J= 7.8 Hz, 1H), 7.12 (d, J= 7.8 Hz, 1H), 7.31 - 7.50 (m, SH), 7.51 (d, J= 1.5 Hz, 1H), 7.86 (dd, J= 7.8, 1.5 Hz, 1H). 25 To a stirred solution of 2-benzyloxynitrobenzene (9.92 g, 43.3 mmol) and NiCl 2 (20.3 g, 157mmol) in MeOH (350 mL) was added portionwise NaBH 4 (8.09 g, 214 mmol) at 0 'C. After disappearing of the starting material (monitored by TLC), the mixture was evaporated off. The black precipitate was dissolved in IN HCI, then acidic solution was alkalified by the addition of IN NaOH and extracted with EtOAc. The extracts were washed brine, dried over Na 2
SO
4 , and concentrated to 186 WO 01/00206 PCT/US00/18079 dryness. Chromatography of the residue with CHCl 3 as eluent gave 2-benzyloxy aniline (8.60 g, 100 %) as a reddish oil. 'H-NMR (CDCl 3 ) 8 3.71 (broad s ,2H), 5.06 (s, 2H), 6.68 - 6.86 (m, 4H), 7.32 - 7.44 (m, 5H); FAB-MS m/z 200 (M+1). To a solution of 2-benzyloxyaniline (1.15 g, 5.77 mmol) in benzene (60 mL) was added 5 triphosgene (1.27 g, 6.35 mmol) and Et 3 N (2.60 mL, 17.3 mmol) at 0 'C. The reaction mixture was heated under reflux for 20 hr. The resulting mixture was filtrated and washed with hexane, and the filtrate was concentrated to leave a residual oil, which was chromatographed with hexane EtOAc (4 : 1, v/v) as eluent to afford tert-butyl 4-[N'-(2-benzyloxyphenyl)ureido]-3-methoxy phenylacetate (2.38 g, 89 %) as a yellow oil. 'H-NMR (CDCl 3 ) 8 1.44 (s, 9H), 3.44 (s, 2H11), 3.78 10 (s, 3H), 5.07 (s, 2H), 6.73 (dd, J= 8.0, 1.7 Hz, 1H11), 6.78 (d, J= 1.7 Hz, 1H11), 6.90 - 6.98 (m, 3H), 7.07 (s, 1H1), 7.29 (s, 111), 7.33- 7.38 (m, 5H11), 7.91 (d,J= 8.0 Hz, 1H), 8.14 (m, 1H). To a solution of tert-butyl 4-[N'-(2-benzyloxyphenyl)ureido]-3-methoxyphenylacetate (2.35 g, 5.08 mmol) in CH 2 C1 2 (25 mL) was added TFA (25 mL) at 0 oC. After being stirred at room temperature. for 3 hr, the mixture was concentrated. The residue was dissolved in IN NaOH and 15 washed with Et20. The basic water layer was poured into ice - IN HCI and the resulting mixture was extracted with CHC1 3 - MeOH (4 : 1, v/v). The extracts were washed with brine, dried over Na 2
SO
4 , and concentrated to dryness. The residue was dissolved in isopropyl ether and hexane was added to this solution until the crystallization was completed. The solid was collected to give 4 [N'-(2-benzyloxyphenyl)ureido]-3-methoxyphenylacetic acid (1.59 g, 77 %) as a brownish solid. 20 'H-NMR (DMSO-d) 8 3.50 (s, 2H11), 3.85 (s, 3H11), 5.26 (s, 2H), 6.76 (d, J= 8.3 Hz, 1H), 6.83 6.89 (m, 2H11), 6.91 (s, 1), 7.01 (dd, J= 8.3, 2.3 Hz, 1H), 7.31 (t, J = 7.3 Hz, 1H11), 7.39 (t, J= 7.3 Hz, 2H), 7.49 (d, J= 7.3 Hz, 2H), 7.97 (d, J= 8.3 Hz, 1H1), 8.04 (d, J= 8.3 Hz, 1H), 8.80 (s, 1H), 8.86 (s, 1H), 12.24 (broad s, 1H); FAB-MS m/z 407 (M +1). To a solution of 4-[N'-(2-benzyloxyphenyl)ureido]-3-methoxyphenylacetic acid (1.12 g, 2.76 25 mmol), methyl 4-(2-pyrrolidinylmethoxy)-3-nitrobenzoate (890 mg, 2.76 mmol), HOBt (74.0 mg, 0.55 minmol), DMAP (67.0 mg, 0.55 mmol), and Et 3 N (0.58 mL, 4.13 mmol) in THF (15 mL) was added EDC-HCI (792 mg, 4.13 mmol). After being stirred at room temperature for 12 hr, the reaction mixture was diluted with water and extracted with EtOAc. The extracts were washed with brine, dried over Na 2
SO
4 , and concentrated to dryness. Chromatography of the residue with 30 EtOAc as eluent gave methyl 4-[1-[4-[N'-(2-benzyloxyphenyl)ureido]-3-methoxyphenylacetamido ]-2-pyrrolidinylmethoxy]-3-nitrobenzoate (1.52 g, 82 %) as a yellow amorphous solid. 'H-NMR 187 WO 01/00206 PCT/US00/18079
(CDCI
3 ) 8 1.91 (m, 1H1), 1.95 - 2.17 (m, 3H), 3.47 - 3.53 (m, 2H), 3.56 (s, 2H), 3.60 (m, 1H), 3.68 (s, 3H), 3.90 (s, 3H), 4.11 (d, J= 7.3 Hz, 1H), 4.45 (m, 1H), 5.08 (s, 2H), 6. 70 (dd, J= 8.3, 1.9 Hz, 1H), 6.75 (d, J = 1.9 Hz, 1H1), 6.91 - 6.99 (m, 3H), 7.16 (s, 111), 7.18 (d, J= 8.3 Hz, 1H), 7.33 -7.40 (m, 6H), 7.91 (d, J = 8.3 Hz, 1H1), 8.11 - 8.16 (m, 2H), 8.46 (s, 1H11). 5 A solution of methyl 4-[1-[4-[N'-(2-benzyloxyphenyl)ureido]-3-methoxyphenylacetamido]-2 pyrrolidinylmethoxy]-3-nitrobenzoate (1.52 g, 2.27 mmol) in MeOH (20 mL) and THF (5 mL) was hydrogenated over 5 % Pd-C (wet, 52.2%; 1.21 g) under hydrogen atmosphere (4 kg/cm 2 ) at room temperature After being stirred for 17 hr, the catalyst was filtered off and the filtrate was concentrated to dryness. Chromatography of the residue with EtOAc as eluent gave methyl 3 10 amino-4-[1-[4-[N'-(2-hydroxyphenyl)ureido]-3-methoxyphenylacetamido]-2-pyrrolidinylmethoxy] benzoate (1.12 g, 90 %) as a brownish amorphous solid. 'H-NMR (CDC13) 8 1.97 - 2.10 (m, 4H), 3.44 (s, 3H), 3.52 - 3.63 (m, 2H), 3.85 (s, 3H), 4.10 - 4.18 (m, 2H), 4.53 (m, 1H1), 6.65 - 6.67 (m, 4H), 6.93 - 7.02 (m, 3H), 7.33 (d, J= 2.2 Hz, 1H11), 7.36 (dd, J= 8.3, 2.2 Hz, 1H1), 7.60 (s, 1H1), 7.69 (d, J = 8.3 Hz, 1H), 8.08 (s, 1H), 9.47 (broad s, 1H); FAB-MS m/z 549 (M'+1). 15 To a solution of methyl 3-amino-4-[1-[4-[N'-(2-hydroxyphenyl)ureido]-3-methoxyphenyl acetamido]-2-pyrrolidinylmethoxy]benzoate (1.12 g, 2.04 mmol) in THF - MeOH (4: 1, v/v; 20 mL) was added IN NaOH (4.20 mL, 4.20 mmol). After being stirred at room temperature for 24 hr, the reaction mixture was concentrated. The residue was diluted with water and neutralized with 1N HCI at 0 'C. The mixture was extracted with CHCl 3 - MeOH (4 : 1, v/v) ,which was 20 washed with brine, dried over Na 2
SO
4 , and concentrated to dryness. Chromatography of the residue with CHCI 3 : MeOH (5: 1, v/v) as eluent gave 60 (352 mg, 32 %) as a pale yellow amorphous solid. MW 534.56 IR (KBr) 3282, 3062, 3025, 2952, 2865, 1629, 1546, 1509, 1454, 1419 cm-'; 'H-NMR (DMSO-d 6 ) 8 1.87 - 2.04 (m, 4H), 3.48 - 3.57 (m, 2H), 3.60 (s, 2H), 3.79 (s, 3H), 3.94 (dd, J= 9.5, 7.6 Hz, 1H), 4.12 (dd, J= 9.5, 3.9 Hz), 4.35 (m, 111), 4.87 (broad s, 1H), 25 6.70 -6.91 (m, 6H), 7.16 (dd, 111, J= 8.3, 2.0 Hz, 1H1), 7.26 (d, J= 2.0 Hz, 111), 7.96 (d, J= 8.3 Hz, 111), 7.97 (d, J = 8.3 Hz, 1H11), 8.80 (s, 111), 8.82 (s, 1H1); FAB-MS m/z 535 (M+1); Anal. Calcd for C 28
H
30
N
4 0 7 -4.5H 2 0: C, 55.63; H, 6.39; N, 9.10. Found: C, 55.08; H, 5.06; N, 8.69. Example 56 5-[[ -[4-[N'-(2-chlorophenyl)ureidol-3-methoxyphenylacetyl]-2-pyrrffolidinyl]methylamino] 30 pyridine-2-carboxylic acid 188 WO 01/00206 PCT/US00/18079 N 6N 1COOH IHH Me H61 5-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetyl]-2-pyrrolidinyl]methylamino] pyridine-2-carboxylic acid N N COOH N N N Me._H H Me H 6 Me 62 5 To a stirred solution of 5-(methoxycarbonyl)pyridine-2-carboxylic acid (2.5 g, 13.8 mmol) and 4 DMAP (340 mg, 2.8 mmol) in tert-BuOH (15 ml) was added Boc 2 0 (6 g, 27.6 mmol) at room temperature. After stirring for 2 hr at room temperature, ice and 0.2 N HCI (20 ml) was added to the mixture and extracted with CH 2 C1 2 . The extracts were washed with sat. NaHCO 3 , dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica gel (50ml) with CH 2 C1 2 as 10 eluent to give methyl 6-tert-butoxycarbonylnicotinate (2.92 g, 89%) as needles. IR (KBr) 2729, 1736, 1720, 1590, 1570 cm'; MS (FAB) mnz 238 (M +1); Anal. Calcd for Cl 2
H
15
NO
4 : C, 60.75; H, 6.37; N, 5.90. Found: C, 60.72; H, 6.46; N, 5.78. A mixture of methyl 6-tert-butoxycarbonylnicotinate (1.2 g, 5.06 mmol) in THF (15 ml) and 0.25 N NaOH (40 ml, 10 mol) at an ambient temperature for 0.5 hr. The mixture was poured 15 into ice-IN HCI (10 ml). The solid was collected, washed with water and air-dried. The crude solid was recrystallized from CHCl 3 -EtOH-IPE to afford 6-tert-butoxycarbonylnicotinic acid (850 mg, 76%) as needles. IR (KBr) n3095, 1728, 1705 crur'; 'H-NMR (DMAO-d) 8 1.63 (s, 9 H), 8.09 (m, 1 H), 8.17 (m, 1 H), 8.42 (dt, J= 2.4 and 8.3 Hz, 1 H), 9.21 (t, J= 2.4 and 8.8Hz, 1 H); MS (FAB) nm/z 224 (M* +1); Anal. Calcd for C 29
H
33
N
3 0 6 : C, 36.18; H,3.18; N, 3.84. Found: C, 20 36.85; H, 3.35; N, 3.79. To a stirred mixture of 6-tert-butoxycarbonylnicotinic acid (1.9 g, 8.51 mmol) and triethylamine (1.17 g, 11.49 mmol) in tert-BuOH (30 ml) and toluene (30 ml) was added a solution of diphenyl phosphoryl azide (2.93 g, 10.64 mmol) in toluene (3 ml) at room temperature. The resulting mixture was then heated at reflux for 5 hr. After cooling, ice and IN HCI (5ml) was 25 added to the mixture and extracted with toluene. The extracts were washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica gel (50ml) with toluene EtOAc (5:1, v/v) as eluent to give tert-butyl 5-tert-butoxycarbonyamino-2-pyridinecarboxylate (1.9 g, 76%) as a gum. 'H-NMR (CDCl 3 ) 8 1.53 (s, 9 H), 1.63 (s, 9 H), 6.82 (br s, 1 H), 8.01 (d, J 8.8 Hz, 1H), 8.17 (m, 1H), 8.46 (d,J= 2.4 Hz, 1 H). 189 WO 01/00206 PCT/US00/18079 To a stirred mixture of tert-butyl 5-tert-butoxycarbonyamino-2-pyridinecarboxylate (1.9 g, 6.45 mmol) in CH 2 C1 2 (20 ml) was added TFA (5 ml). The mixture was evaporated off, and the residue was dissolved in EtOH (30 ml). HCl-gas was induced to the solution with stirring at 0-10 oC for 10 min. The resulting stirred mixture was then heated at reflux for 10 hr. After cooling, 5 N 2 -gas was induced to remove of large excess of HCl-gas for 15 min. the mixture was evaporated off. The residue was alkalized by the addition of sat. NaHCO 3 , and extracted with CH 2 C1 2 . The extracts were washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica gel (30ml) with CHCl 3 -EtOH (98:2, v/v) as eluent to give ethyl 5 amino-2-pyridine carboxylate (700 mg, 65%) as a crystalline material. IR (KBr) n 3423, 3190, 10 1708, 1657, 15873338, 3296, 1691, 1641 cmr'; 'H-NMR (CDCl 3 )8 1.42 (t, J= 7.0 Hz, 3 H), 4.11 (br s, 1 H), 4.43(q, J= 7.0 Hz, 2 H), 6.99 (dd, J = 2.7 and 8.5 Hz, 1 H), 7.95 (d, J= 8.5 Hz, 1 H), 8.16 (d, J = 2.7 Hz, 1 H); MS (FAB) m/z 167 (M); Anal. Called for C 27
H,
29
CIN
4 0 6 : C, 57.47; H, 6.63; N, 16.76. Found: C, 57.27; H, 5.99; N, 16.72. A stirred mixture of ethyl 5-amino-2-pyridinecarboxylate (660 mg, 3.95 mmol) and 1 15 tert-butoxycarbonyprolinal (1.1 g, 5.33 mmol) in toluene (10 ml) was heated at reflux for 1 hr, during which time water was azeotropically removed with Dean-Stark water-trap. After cooling, the mixture was evaporated in vacuo. The residue was dissolved in MeOH-AcOH (9:1, v/v, 30ml). To the stirred solution was added NaBH 3 CN (500 mg, 7.90 mmol) at 0-5 oC. The resulting mixture was stirred for a further 12 hr at room temperature. The mixture was piured into ice-sat 20 NaHCO 3 (50 ml), and extracted with CH 2
C
2 . The extracts were washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica gel (50mi) with CHCl 3 EtOAc (98:2, v/v) as eluent to give ethyl 5-[N-[2-(1-tert-butoxycarbony)pyrrolidinyl]methylamino] pyridine-2-carboxylate (1.1 g, 70%) as a gum. 'H-NMR (CDCl 3 ) 8 1.38 (s, 9 H), 1.42 (s, 6 H), 3.93 (s, 3 H), 4.29 (s, 2 H), 4.67 (br s, 1 H), 7.15 (d, J= 8.8 Hz, 1 H), 8.18 (dd, J= 1.7 and 8.8Hz, 25 1 H), 8.52 (d, J= 1.7 Hz, 1H). A mixture of ethyl 5-[[2-(1-tert-butoxycarbony)pyrrolidinyl]methylamino]pyridine-2 carboxylate (800 mg, 2.29 mmol) in CH 2 C1 2 (17 ml) and TFA (3 ml) was stirred at room temperature for 3 hr. The mixture was evaporated, and the residue was made basic with sat. NaHCO 3 . The mixture was extracted with CH 2 C1 2 . The extracts were washed with brine, dried 30 over Na 2
SO
4 -Na 2
CO
3 , and evaporated to give ethyl 5-[(2-pyrrolidinyl)methylamino]pyridine-2 carboxylate (460 mg, 81%) as a gum. 'H-NMR (CDCl 3 ) 8 1.32 (t, J= 7Hz, 3 H), 1.58-2.10 (series of m, 4 H), 3.12-3.28 (series of m, 3 H), 3.65 (m, 1 H), 4.30 (be q, J= 7 Hz, 2 H), 6.27 (br, 190 WO 01/00206 PCT/US00/18079 1 H), 6.59 (dd, J = 2.4 and 8.5 Hz, 1 H), 7.65 (d, J = 8.5 Hz, 1 H), 7.94 (d, J = 2.4 Hz, 1H). To a stirred mixture of ethyl 5-[(2-pyrrolidinyl)methylamino]pyridine-2-carboxylate (220 mg, 0.88 mmol), 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (300 mg, 0.88 mmol), 4 DMAP (135 mg, 1.10 mmol) in DMF (7 ml) was added EDC-HCI (215 mg, 1.10 mmol) at room 5 temperature. The resulting mixture was stirred at room temperature for 20 hr. The mixture was pored into ice-water. The solid was collected, washed with water and air-dried. The crude solid was purified by silica gel (30ml) column chromatography with CHCl 3 - EtOH (98:2, v/v) as eluent and crystallized with Et 2 0 to give 5-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-2 pyrrolidinyl]methylamino]pyridine-2-carboxylate (420 mg, 84%) as fine needles. IR (KBr) 3319, 10 1703, 1628, 1585, 1529 cm'; 'H-NMR (CDCl 3 )8 1.38 (t, J= 7 Hz, 3 H), 1.73-2.17 (series of m, 4 H), 3.19 and 3.54 (each m, each 1 H), 3.63 (s, 2 H), 3.70 (s, 3 H), 4.39 (be q, J= 7 Hz, 2 H), 4.55 (min, 1 H), 6.02 (br s, 1 H), 6.78-6.84 (series of s and m, 3 H), 6.98 (dt, J = 2.4 and 8.0 Hz, 1 H), 7.16 (s, 1 H), 7.21-7.26 (series of m, 3 H), 7.34 (dd, J= 2.4 and 8.0 Hz, 1 H), 7.90 (d, J= 8.3 Hz, 1H), 7.98 (min, 2H), 8.16 (dd, J= 1.2 and 8.8 Hz, 1H); MS (FAB) m/z 566 (M +1); Anal. Calcd for 15 C 29
H
32 C1N 5 0 5 s-H 2 0: C, 59.63; H, 5.87; N, 12.37. Found: C, 60.06; H, 5.76; N, 11.95. A mixture of ethyl 5-[[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-2 pyrrolidinyl]methylamino]pyridine-2-carboxylate (300 mg, 0.53 mmol) in THF:MeOH (1:1, v/v, 16 ml) and 0.25N NaOH (11 ml, 2.75 mmol) was stirred for 3 hr at room temperature. The mixture was poured into ice-iN HCI (3 ml). The solid was collected, washed with water and air 20 dried. The crude crystalline material was collected with CH 2 C1 2 -Et 2 O to give 61 (180 mg, 63%) as an amorphous solid. MW 537.99 IR (KBr) 3319, 1701, 1620, 1585, 1533 cmr'; H-NMR (CDCl 3 ) 8 'H-NMR (DMSO-d 6 ) 8 1.80-2.05 (series of m, 4 H), 2.99 (min, 1 H), 3.50- 3.59 (series of m, 3 H), 3.60 (s, 2 H), 3.86 (s, 3 H), 4.11 (min, 1 H), 6.78 (d, J= 8.5 Hz, 1 H), 6.91 (s, 1 H), 6.94 (mn, 1 H), 7.02 (min, 1 H), 7.14 (dd, J= 2.5 and 8.5 Hz, 1 H), 7.28 (t, J= 7.0 Hz, 1 H), 7.45 (d, J= 8.0 Hz, 1 25 H), 7.78 (d,J= 8.8 Hz, 1 H), 7.97 (d,J= 8.3 Hz, 1 H), 8.08(brs, 1 H), 8.11 (mn, 1 H), 8.89 (s, 1 H), 8.94 (s, 1 H); MS (FAB) m/z 538 (M+ +1); Anal. Calcd for C 27
H
28
CIN,
5 -1.5xH 2 0: C, 57.39; H, 5.53; N, 12.39. Found: C, 57.37; H, 5.54; N, 11.74. To a stirred mixture of ethyl 5-[(2-pyrrolidinyl)methylamino]pyridine-2-carboxylate (230 mg, 0.923 mmol), 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (290 mg, 0.923 mmol), 30 4-DMAP (145 mg, 1.15 mmol) in DMF (7 ml) was added EDC-HCI (225 mg, 1.15 mmol) at room temperature. The resulting mixture was stirred at room temperature for 20 hr. The mixture was 191 WO 01/00206 PCT/US00/18079 pored into ice-water. The solid was collected, washed with water and air-dried. The crude solid was purified by silica gel (30ml) column chromatography with CHCl 3 - EtOH (98:2, v/v) as eluent to give ethyl 5-[[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl] methylamino]pyridine-2-carboxylate (400 mg, 80%) as fine needles. IR (KBr) n 3325, 1709, 5 1618, 1585, 1531 cm'; 'H-NMR (CDCl 3 ) 8 1.39 (t, J= 7 Hz, 3 H), 1.73-2.07 (series of m, 4 H), 2.28 (s, 3 H), 3.12 and 3.49 (each m, each 1 H), 3.60 (s, 2 H), 4.39 (br q, J= 7 Hz, 2 H), 4.53 (m, 1 H), 6.07 (br s, 1 H), 6.23 (br s, 1 H), 6.75-6.77 (series of s and m, 2 H), 6.82 (dd, J= 3.0 and 8.5 Hz, 1 H), 7.09-7.22 (series of m, 3 H), 7.49 (d,J= 8.0 Hz, 1 H), 7.90 (d, J = 8.5 Hz, 1H), 7.98 (d, J= 2.6 Hz, 1H), 8.06 (d, J = 8.8 Hz, 1H); MS (FAB) m/z 546 (M +1); Anal. Calcd for 10 C 30
H
35
N
5 O-1.5xH 2 0: C, 52.92; H, 6.69; N, 12.23. Found: C, 63.11; H, 6.48; N, 11.96. A mixture of ethyl 5-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2 pyrrolidinyl]methylamino]pyridine-2-carboxylate (D91-4596) (290 mg, 0.53 mmol) in THF:MeOH (1:1, v/v, 16 ml) and 0.25N NaOH (11 ml, 2.75 mmol) was stirred for 3 hr at room temperature. The mixture was poured into ice-IN HCI (3 ml). The solid was collected, washed with water and 15 air-dried. The crude crystalline material was collected with CH 2 C1 2 -Et20 to give 62 (170 mg, 62%) as an amorphous solid. MW 517.58 IR (KBr) 3283, 1701, 1618, 1529 cm'; 'H-NMR
(CDCI
3 ) 8 'H-NMR (DMSO-d 6 ) 8 1.78-2.04 (series of m, 4 H), 2.25 (s, 3 H), 2.95-3.55 (series of m, 4 H), 3.59 (s, 2 H), 3.87 (s, 3 H), 4.11 (m, 1 H), 6.75-7.24 (series of m, 7 H), 7.83-7.97 (series of m, 3 H), 8.01 (d, J= 8.3 Hz, 1 H), 8.13 (d, J= 2.6 Hz, 1 H), 8.11 (m, 1 H), 8.47 (s, 1 H), 8.57 20 (s, 1 H); MS (FAB) m/z 518 (M +1); Anal. Calcd for C 2
H
3 1 NsOs 5 0 5 -2.SxH20: C, 60.50; H, 6.39; N, 12.60. Found: C, 60.31; H, 6.28; N, 12.10. Example 57 2-[1-[[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-2-pyrrolidiny]methoxy]pyridine-5 carboxylic acid N N -T 0 25 I H Me 63 To a stirred solution of methyl 2-hydroxy-5-pyridinecarboxylate (2.0 g, 13.06 mmol), PPh 3 (4.2 g, 15.93 mmol) and 1-tert-butoxycarbony-(L)-prolinol (2.63 g, 13.06 mmol) in THF (25 ml) was added a solution of DIAD (3.3 g, 15.67 mmol) in THF (5 ml) at 0-10 oC. The resulting stirred mixture was then heated at reflux for 3 hr. After cooling, the mixture wsa evaporated in vacuo. 30 The residue was chromatographed on silica-gel (120ml) with n-hexane-EtOAc (4:1, v/v) as eluent to give methyl 2-[2-(1-tert-butoxycarbony)pyrrolidinyl]methoxypyridine-5-carboxylate (3.0 g, 192 WO 01/00206 PCT/US00/18079 68%) as a gum. 'H-NMR (CDCl 3 ) 8 1.46 (s, 9 H), 1.82-2.04 (series of m, 4 H), 3.45 (m, 2 H), 3.91 (s, 3 H), 4.10-4.32 (series of m, 2 H), 4.48 (m, 1 H), 6.32 (br, 1 H), 6.75 (d,J= 8.8 Hz, 1 H), 8.15 (dd, J= 2 and 8.8Hz, 1 H), 8.79 (d, J= 2 Hz, 1H). A mixture of methyl 2-[2-(1-tert-butoxycarbony)pyrrolidinyl]methoxypyridine-5 5 carboxylate (2.9 g, 8.62 mmol) in CH 2 Cl 2 (80 ml) and TFA (20 ml) was stirred at room temperature for 3 hr. The mxture was evaporated, and the residue was alkalized with sat. NaHCO 3 . The mixture was extracted with CH 2 Cl 2 . The extracts were washed with brine, dried over Na 2
SO
4 -Na 2
CO
3 , and evaporated to give methyl 2-(2-pyrrolidinyl)methoxypyridine-5 carboxylate (1.2 g, 59%) as a gum. 'H-NMR (CDCl 3 )8 1.58-2.050 (series of m, 4 H), 2.90-3.02 10 (series of m, 2 H), 3.87 and 3.90 (each s, 3 H), 4.23 (m, 1 H), 4.37 (m, 1 H), 6.33 (br, 1 H), 6.78 (d, J = 8.5 Hz, 1 H), 8.15 (dd, J = 2.2 and 8.8 Hz, 1 H), 8.79 (d, J= 2.2 Hz, 1H). To a stirred mixture of methyl 2-(2-pyrrolidinyl)methoxypyridine-5-carboxylate (370 mg, 1.57 mmol), 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (525 mg, 1.57 mmol), 4 DMAP (230 mg, 1.88 mmol) in DMF (10 ml) was added EDC-HCl (360 mg, 1.88 mmol) at room 15 temperature. The resulting mixture was stirred at room temperature for 20 hr. The mixture was pored into ice-water. The solid was collected, washed with water and air-dried. The crude solid was purified by silica-gel (30ml) column chromatography with CHCl 3 - EtOH (98:2, v/v) as eluent and crystallized with Et 2 0O to give methyl 2-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl acetyl]-2-pyrrolidinylmethoxy]pyridine-5-carboxylate (600 mg, 69%) as an amorphous solid. 20 'H-NMR (CDC1 3 ) 8.21 and 2.01 (each m, 4 H), 3.45-4.50 (series of s and m, 13 H which contains amide-isomers), 6.58-8.83 (series of s and m, 12 H which contains amide-isomers) A mixture of methyl 2-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-2 pyrrolidinylmethoxy]pyridine-5-carboxylate (230 mg, 0.415 mmol) in THF (1 ml) and 0.25N NaOH (4 ml, 1 mmol) was stirred for 14 hr at room temperature and for 3 hr at 60 oC. Afrter 25 cooling, the mixture was poured into ice-IN HCI (2 ml). The solid was collected, washed with water and air-dried. The crude crystalline material was purified by preparative TLC plate with CHCl 3 -EtOH (9:1, v/v) as eluent and crystallized with Et 2 0 to give 63 (150 mg, 67%) as an amorphous solid. MW 538.98 IR (KBr) 3329, 1709, 1601, 1533 cm-'; 'H-NMR (CDCl 3 ) 8 1.85 2.05 (series of m, 4 H), 3.50-3.60 (series of m, 2 H), 3.82 (s, 3 H), 3.86 (s, 2 H), 4.29 (m, 1 H), 30 4.42 (m, 1 H), 6.72-7.05 (series of m, 4 H), 7.28 (m, 1 H), 7.43 (d, J= 8Hz, 2 H), 7.95 (d, J= 8.3 Hz, 1 H), 8.09 (d, J = 8.3 Hz, 2 H), 8.64 (m, 1 H), 8.89 (s, 1 H), 8.93 (s, 1 H); MS (FAB) m/z 539 193 WO 01/00206 PCT/US00/18079 (M ); Anal. Calcd for C 27
H
2 8 C1N 4 06-1.3xH 2 0: C, 57.55; H, 5.47; N, 9.94. Found: C, 57.94; H, 5.00; N, 9.45. Example 58 5-[ -[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-2-pyrrolidinylmethoxy]pyridine-2 5 carboxylic acid N N 0 - COOH r H H 6Me 64 To a stirred mixture of methyl 5-(2-pyrrolidinyl)methoxypyridine-2-carboxylate (370 mg, 1.57 mmol), 4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (595 mg, 1.57 mmol), 4 DMAP (230 mg, 1.88 mmol) in DMF (10 ml) was added EDC-HC1 (360 mg, 1.88 mmol) at room 10 temperature. The resulting mixture was stirred at room temperature for 20 hr. The mixture was pored into ice-water. The solid was collected, washed with water and air-dried. The crude solid was purified by silica-gel (30ml) column chromatography with CHCl 3 - EtOH (98:2, v/v) as eluent and crystallized with Et20 to give methyl 5-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxypheny lacetyl]-2-pyrrolidinylmethoxy]pyridine-2-carboxylate (650 mg, 69%) as an amorphous solid. IR 15 (KBr) n 3323, 1720, 1624, 1601, 1527 cm'; 'H-NMR (CDC13) 8 1.22 and 2.00 (each m, 4 H), 3.48-4.55 (series of s and m, 13 H which contains amide-isomers), 6.93-8.82 (series of s and m, 12 H which contains amide-isomers); MS (FAB) m/z 597 (M + -1) and 599 (M +1); Anal. Called for
C
28
H
3 0 BrN 4 06-1.0xH 2 0: C, 54.55; H, 5.23; N, 9.09. Found: C, 54.13; H, 5.03; N, 9.33. A mixture of methyl 5-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-2 20 pyrrolidinylmethoxy]pyridine-2-carboxylate (300 mg, 0.5 mmol) in THF:MeOH (1:1, v/v, 2 ml) and 0.25N NaOH (4 ml, 1 mmol) was stirred for 3 hr at room temperature and for 5 hr at 60 C. Afrter cooling, the mixture was poured into ice-lN HCI (2 ml). The solid was collected, washed with water and air-dried. The crude crystalline material was purified by preparative TLC plate with CHCI 3 -EtOH (9:1, v/v) as eluent and crystallized with Et20 to give 64 (180 mg, 62%) as an 25 amorphous solid. MW 583.43 IR (KBr) n 3319, 1705, 1685, 1601, 1529 cm'; 'H-NMR (DMSO d) 8 1.82-2.05 (series of m, 4 H), 3.48-3.58 (series of m, 2 H), 3.82 (s, 3 H), 3.86 (s, 2 H), 4.42 4.55 (series of m, 3 H), 6.72-6.98 (series of m, 4 H), 7.32 (t, J= 8 Hz, 1 H), 7.60 (d, J= 8 Hz, 1 H), 7.95 (m, 2 H), 8.08 (m, 1 H), 8.63 (m, 1 H), 8.64 (m, 1 H), 8.89 (s, 1 H), 8.93 (s, 1 H); MS (FAB) m/z 583 (M ); Anal. Calcd for C 27
H
2 8 BrN 4 06-2.0xH20: C, 52.26; H, 5.20; N, 9.03. Found: 30 C, 52.72; H, 4.63; N, 8.50. 194 WO 01/00206 PCT/US00/18079 Example 59 4-[ 1-[3-[N'-(2-bromophenyl)urcido]-2-methoxy-6-pyridylacetyl]-(4S)-fluoro-(2S)-pyrrolidiny methoxy]benzoic acid N N 0- 0 -COOH rH H Me 65 5 To a stirred solution of ethyl 3-amino-2-methoxy-6-pyridylacetate (1.61 g, 7.66 mmol) in THF (10 ml) were added 2-bromophenylisocyanate (948 ml, 7.66 mmol) and Et 3 N (107 ml, 0.776 mmol). After stirring overnight, the mixture was poured into H 2 0 (100 ml) and extracted with CHCl 3 MeOH (4:1, 2 x 200 ml). The combined extracts were dried over MgSO, and evaporated. The residue was recrystallized from CHCI 3 -MeOH-hexane to give ethyl 3-[N'-(2-bromophenyl)ureidol 10 2-methoxy-6-pyridylacetate (2.91 g, 93%) as a colorless crystalline powder. mp 160-163 'C; 1
H
NMR (DMSO-d) 8 1.19 (dt, J= 7.1, 0.7 Hz, 3 H), 3.69 (s, 2 H), 3.95 (s, 3 H), 4.07-4.13 (m, 2 H), 6.90 (d, J= 7.8 Hz, 1 H), 6.99 (t, J= 7.8 Hz, 1 H), 7.33 (t, J= 7.8 Hz, 1 H), 7.61 (d, J= 7.8 Hz, 1 H), 7.96 (dd, J=7.8, 1.5 Hz, 1 H), 8.31 (d, J= 7.8 Hz, 1 H), 8.82 (s, 1 H), 9.12 (s, 1 H); MS (FAB) m/z 408 (M), 410 (M+2); Anal. Calcd for C, 7 H,BrN 3 0 4 -0.25 H 2 0: C, 49.47; H, 4.52; 9.96. 15 Found: C, 49.34; H, 4.48; N, 9.96. A mixture of ethyl 3-[N'-(2-bromophenyl)ureido]-2-methoxy-6-pyridylacetate (2.90 g, 7.10 mmol), 0.25 N NaOH (56.8 ml, 14.2 mmol), and THF (50 ml) was stirred for 5 h. The mixture was neutralized with 1 N HC1 and the resulting precipitate was collected by filtration. The residue was recrystallized from CHC13-MeOH-hexane to give 3-[N'-(2-bromophenyl)ureido]-2 20 methoxy-6-pyridylacetic acid (2.40 g, 89%) as a colorless crystalline powder. mp 195-197 'C; 1
H
NMR (DMSO-d 6 ) 8 3.59 (s, 2 H), 3.95 (s, 3 H), 6.88 (d, J= 8.1 Hz, 1 H), 6.97-7.01 (m, 1 H), 7.33 (t,J = 7.3 Hz, 1 H), 7.61 (d,J = 8.1Hz, 1 H), 7.95-7.97 (m, 1 H), 8.29 (d,J= 8.1Hz, 1 H), 8.81 (s, 1 H), 9.10 (s, 1 H), 12.35 (br s, 1 H); Anal. Calcd for C, 5
HI
4 BrN 3
O
4 : C, 47.39; H, 3.71; N, 11.05. Found: C, 47.27; H, 3.59; N, 10.86. 25 To a stirred solution of 3-[N'-(2-bromophenyl)ureido]-2-methoxy-6-pyridylacetic acid (751 mg, 1.97 mmol) and methyl (4S)-fluoro-(2S)-pyrrolidinylmethoxybenzoate (500 mg, 1.97 mmol) in DMF (10 ml) were added EDC-HCl (566 mg, 2.96 mmol), DMAP (cat.), and HOBt (cat.). After stirring overnight, the mixture was partitioned between EtOAc (200 ml) and brine (200 ml). The phases were separated. The organic phase was washed with brine (100 mnil), dried 30 over MgSO 4 , and evaporated. The resulting residue was chromatographed on silica gel with 195 WO 01/00206 PCT/US00/18079 CHCl 3 -MeOH (20:1) as eluent to give methyl 4-[1-[3-[N'-(2-bromophenyl)ureido]-2-methoxy-6 pyridylacetyl]-(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (1.16 g, 96%) as a yellow viscous solid. A mixture of methyl 4-[1-[3-[N'-(2-bromophenyl)ureido]-2-methoxy-6-pyridylacetyl] 5 (4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (1.16 g, 1.88 mmol), 0.25 N NaOH (15 ml, 3.75 mmol), and THF (15 ml) was stirred overnight. The mixture was neutralized with 1 N HCI and extracted with CHCl 3 -MeOH (4:1, 2 x 200 ml). The combined extracts were dried over MgSO 4 and evaporated. The resulting residue was chromatographed on silica gel with CHCI 3 -MeOH (40:1 to 10:1) as eluent to give 65 as a pale yellow amorphous solid. MW 601.42 'H-NMR 10 (DMSO-d) 8 2.27-2.39 (m, 2 H), 3.33-4.84 (series of m, 10 H), 5.33-5.53 (m, 1 H), 6.87-6.90 (m, 1 H), 6.99 (t, 1 H, J= 7.6 Hz), 7.08 (d, 2 H, J= 9.0 Hz), 7.34 (t, 1 H, J= 7.6 Hz), 7.61 (d, 1 H, J= 7.8 Hz), 7.88 (d, 2 H, J= 9.0 Hz), 7.97 (d, 1 H, J= 8.3 Hz), 8.28-8.32 (m, 1 H), 8.81-8.82 (m, 1 H), 9.10-9.12 (m, 1 H), 12.66 (brs, 1 H); MS (FAB) m/z 601 (M), 603 (M+2); Anal. Calcd for
C
27
H
26 BrFN 4 0 6 : C, 53.92; H, 4.36; N, 9.32. Found: C, 52.37; H, 4.62; N, 8.38. 15 Example 60 4-[(4S)-fluoro- 1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoic acid F N O COOH Me H H 66 To a stirred solution of 4-[N'-(2-methylphenyl)ureido]phenylacetic acid (337 mg, 1.18 mmol) and 20 methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (300 mg, 1.18 mmol) in DMF (10 ml) were added EDC-HCl (339 mg, 1.77 mmnol), HOBt (cat.) and DMAP (cat.). The reaction mixture was stirred overnight. The mixture was partitioned between EtOAc (200) and H20 (200 ml) and the organic phase was separated. The organic phase was washed with brine (200 ml), dried over MgSO 4 , and evaporated. The residue was chromatographed on silica gel with CHCl 3 -MeOH (50:1) 25 to give methyl 4-[(4S)-fluoro-l1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S)-pyrrolidinyl methoxy]benzoate (613 mg, quant) as a yellow viscous oil. 'H-NMR (CDCl 3 )8 2.03-2.55 (series of m, total 5 H), 3.47-4.21 (series of m, total 7 H), 4.44-4.60 (m, 3 H), 5.21 and 5.34 (m, each, total 1 H), 6.87-7.16 (m, 8 H), 7.52-7.55 (m, 3 H), 7.93 (d, J= 8.8 Hz, 2 H), 7.99 (d, J= 8.8 Hz, 1 H). To a stirred solution of methyl 4-[(4S)-fluoro-1-[4-[N'-(2-methylphenyl)ureido] 30 phenylacetyl]-(2S)-pyrrolidinylmethoxylbenzoate (613 mg, 1.18 mmol) in THF (10 ml) was added 196 WO 01/00206 PCT/US00/18079 0.25 N NaOH (9.4 ml, 2.36 mmol). The mixture was refluxed for 1 day. After cooling to rt, the mixture was poured into 1 N HCI (50 ml) and extracted with CHC13-MeOH (5:1, 2 x 200 ml). The combined extracts were dried over MgSO 4 and evporated. The residue was chromatographed on silica gel with CHCl 3 -MeOH (10:1) to give 66 (378 mg, 63%) as a colorless amorphous solid. MW 5 505.54 'H-NMR (DMSO-d 6 )8.08-2.30 (m, total 5 H), 3.47-4.63 (series of m, 7 H), 5.30-5.50 (m, 1 H), 6.94 (t, J= 7.3 Hz, 1 H), 7.02-7.17 (m, 6 H), 7.37-7.41 (m, 2 H), 7.82-7.96 (m, 4 H), 9.05 (s, 1 H); MS (FAB)m/z 506 (M+1);Anal. Calcd for C 28
H
28
FN
3 0-1.75 H 2 0: C, 62.62; H, 5.91; N, 7.82. Found: C, 62.23; H, 5.63; N, 7.18. Example 61 10 4-[(4S)-fluoro- 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S)-pyrrolidinyl methoxy]benzoic acid N N O\ COOH MeH H OMe 67 To a stirred solution of 1-(tert-butoxycarbonyl)-(4S)-fluoro-(2S)-proline (1.85 g, 7.93 minmol) in THF (15 ml) was added BH 3 -DMS (0.75 ml, 7.93 mmol) at room temperature. After being heated 15 at reflux for 5 h with stirring, the mixture was cooled to room temperature and concentrated in vacuo. The resulting residue was quenched by the addition of H 2 0 (100 ml) and extracted with CHCl 3 (2 x 200 ml). The combined extracts were washed with brine (100 ml), dried over MgSO 4 , and evaporated. The residue was chromatographed on silica gel with CHC13-EtOAc (4:1) as eluent to give 1-(tert-butoxycarbonyl)-(4S)-fluoro-(2S)-pyrrolidinylmethanol (1.76 g, quant) as a colorless 20 oil. 'H-NMR (CDCl 3 ) 8 1.48 (s, 9 H), 1.64 (m, 1 H), 1.97-2.28 (m, 2 H), 3.53-3.87 (series of m, 4 H), 4.09-4.25 (m, 1 H), 5.09 and 5.22 (m, each, total 1 H). To a stirred mixture of 1-(tert-butoxycarbonyl)-(4S)-fluoro-(2S)-pyrrolidinylmethanol (500 mg, 2.28 mmol), methyl 4-hydroxybenzoate (416 mg, 2.74 mmol), Ph 3 P (719 mg, 2.74 mmol) in THF (10 ml) was added DIAD (0.54 ml, 2.74 mmol) at room temperature. The mixture was 25 heated to reflux for 5 h with stirring. After cooling to room temperature, the mixture was concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -EtOAc as eluent (10:1 to 4:1) to give methyl 4-[1-(tert-butoxycarbonyl)-(4S)-fluoro-(2S)-pyrrolidinyl methoxy]benzoate (597 mg, 74%) as a colorless oil. 'H-NMR (CDC13) 8 1.49-1.59 (m, 9 H), 2.05 2.21 (m, 1 H), 3.56-4.43 (series of m, 8 H), 5.19 and 5.32 (m, each, total 1 H), 6.97 (m, 2 H), 7.98 30 (d, J= 8.5 Hz, 2 H). 197 WO 01/00206 PCT/US00/18079 A mixture of methyl 4-[1-(tert-butoxycarbonyl)-(4S)-fluoro-(2S)-pyrrolidinylmethoxy] benzoate (590 mg, 1.67 mmol) and TFA (5 ml) in CH 2 C1 2 (5 ml) was stirred for 3 h. After the mixture was concentrated in vacuo, the residue was made basic with sat. NaHCO 3 and extracted with CHC1 3 (2 x 200 ml). The combined extracts were dried over K 2
CO
3 and evaporated to give 5 methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (414 mg, 98%) as a yellow solid. 'H NMR (CDC 3 ) 8 1.89-2.02 (m, 1 H), 2.16-2.31 (m, 1 H), 2.98 (m, 1 H), 3.35 (m, 1 H), 3.46-3.68 (m, 1 H), 3.86 (s, 3 H), 4.00-4.08 (m, 2 H), 5.16 and 5.29 (t, each, J= 4.7 Hz, total 1 H), 6.91 (d, J =8.8 Hz, 2 H), 7.96 (d, J = 8.8 Hz, 2 H). A mixture of methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (205 mg, 0.810 10 mmol), 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (254 mg, 0.810 mmol), EDC-HCI (233 mg, 1.22 mmol), HOBt (cat.), and DMAP (cat.) in DMF (10 ml) was stirred overnight. The mixture was diluted with EtOAc (200 ml), washed with brine (2 x 100 ml), dried over MgSO 4 , and evaporated. The residue was chromatographed on silica gel with CHC13-MeOH (20:1) as eluent to give methyl 4-[(4S)-fluoro-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] 15 phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (445 mg, quant) as a light brown viscous. 'H NMR (CDC1 3 )6 2.05-2.55 (m, total 6 H), 3.55-4.13 (m, 11 H), 4.48-4.60 (m, 2 H), 5.20 and 5.33 (each m, total 1 H), 6.29 (s, 1 H), 6.79 (m, 2 H), 6.96 (d, J= 8.8 Hz, 2 H), 7.11-7.25 (m, 3 H), 7.48 (d, J= 7.6 Hz, 1 H), 7.93-8.09 (m, 4 H). A mixture of methyl 4-[(4S)-fluoro-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] 20 phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (445 mg, 1.62 mmol) and 0.25 N NaOH (15 ml, 3.75 mmol) in THF (15 ml) was stirred overnight at room temperature then for 2 h at reflux. The mixture was acidified with 1 N HCI and extracted with CHC13-MeOH (4:1, 2 x 200 ml). The combined extracts were dried over MgSO 4 and evaporated. The residue was chromtographed on silica gel with CHC1 3 -MeOH (10:1) as eluent to give 67 (260 mg, 30%) as a pale yellow 25 amorphous solid. MW 535.56 'H-NMR (DMSO-d) 8 2.25-2.51 (m, 5 H), 3.33-4.41 (series of m, 10 H), 5.30-5.50 (m, 1 H), 6.75-7.17 (m, 7 H), 7.79 (d, J= 8.1 Hz, 1 H), 7.87-8.04 (m, 3 H), 8.48 (m, 1 H), 8.58 (m, 1 H); MS (FAB) m/z 536 (M+1); Anal.Calcd for C 2 9H 30
FN
3 0 6
-H
2 0: C, 62.92; H, 5.83; N, 7.59. Found: C, 62.40; H, 5.82; N, 6.93. Example 62 30 4-[ 1-[4-[N'-(2-Bromophenyl)ureido]-3-methoxyphenylacetyl]-(4S)-fluoro-(2S)-pyrrffolidiny methoxy]benzoic acid 198 WO 01/00206 PCT/US00/18079 0 - N4 N jNj O COOH r H H 6Me 68 A mixture of methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (501 mg, 1.98 mmol), 4 [N'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (750 mg, 1.98 mmol), EDC-HCI (569 mg, 2.97 mnol), HOBt (cat.) and DMAP (cat.) in DMF (10 ml) was stirred overnight. The 5 mixture was diluted with EtOAc (300 ml), washed with brine (100 ml), dried over MgSO 4 and evaporated. The residue was chromatographed on silica gel with CHClI-EtOAc (4:1) to CHC1 3 MeOH (10:1) as eluent to give methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl acetyl]-(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (1.29 g, quant) as a brown viscous oil. 'H NMR (CDCl 3 ) 8 2.05-2.58 (m, 2 H), 3.49-4.17 (series of m, 12 H), 4.52-4.65 (m, 2 H), 6.82-7.33 10 (series of m, 8 H), 7.53 (dd, J= 8.1, 1.5 Hz, 1H), 7.95-8.02 (in, 4H), 8.14 (dd, J= 8.3, 1.7 Hz, 1H). A mixture of methyl 4-[1-[4-[N'-(2-bromophenyl)ureidol-3-methoxyphenylacetyl]-(4S) fluoro-(2S)-pyrrolidinylmethoxy]benzoate (1.29 g, 2.10 mmol) and 0.25 N NaOH (17 ml, 4.20 mmol) in THF (20 ml) was refluxed for 5 h with stirring. The mixture was poured into ice-cooled 1 N HCI (100 ml) and extracted with CHCl 3 -MeOH (4:1, 2 x 200 ml). The combined extracts were 15 dried over MgSO 4 and evaporated. The residue was chromatographed on silica gel with CHCl 3 MeOH (10:1) as eluent to give 68 (860 mg, 68%) as a colorless amorphous solid. MW 600.43 'H NMR (DMSO-d) 8 2.24-2.31 (mn, 2 H), 3.21-4.63 (series of m, 10 H), 5.31-5.51 (m, 1 H), 6.74 7.10 (m, 5 H), 7.32 (t, J= 7.8 Hz, 1 H), 7.60 (d, J= 7.8 Hz, 2 H), 7.87-7.99 (m, 4 H), 8.74-8.75 (m, 1 H), 8.92-8.94 (m, 1 H); MS (FAB) m/z 601 (M++1); Anal. Calcd for C 28
H
27 BrFN 3 0 6 -2 H20: 20 C, 52.84; H, 4.91; N, 6.60. Found: C, 52.38; H, 4.62; N, 5.99. For Na salt of 68: mp 180-182 'C; Anal. Calcd for C 28
H
27 BrFN 3 NaO6.O0.75 H 2 0: C, 52.88; H, 4.36; N, 6.61. Found: C, 52.97; H, 4.36; N, 6.61. Example 63 4-[ -[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinyl 25 methoxy]benzoic acid F N N 0O/ COOH IH H[ Me 69 A mixture of methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (205 mg, 0.810 mmol), 3 methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (254 mg, 0.810 mmol), EDC-HCI (233 199 WO 01/00206 PCT/US00/18079 mg, 1.22 mmol), HOBt (cat.) and DMAP (cat.) in DMF (10 ml) was stirred overnight. The mixture was diluted with EtOAc (200 ml), washed with brine (2 x 100 ml), dried over MgSO 4 , and evaporated. The residue was chromatographed on silica gel with CHCl 3 -MeOH (20:1) as eluent to give methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(4S)-fluoro-(2S) 5 pyrrolidinylmethoxy]benzoate (376 mg, 81%) as a yellow foam. 'H-NMR (CDCl 3 ) 8 2.07-2.56 (inm, 2 H), 3.57-4.14 (series of m, 11 H), 4.50-4.61 (min, 2 H), 5.22 and 5.35 (series of m, total 1 H), 6.80-7.33 (series of m, 9 H), 7.93-8.00 (min, 3 H), 8.16 (d, J= 8.1 Hz, 1 H). A mixture of methyl methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl] (4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (376 mg, 0.660 mmol) and 0.25 N NaOH (15 nil, 10 3.75 mmol) in THF (15 ml) was stirred overnight at room temperature then for 2 h at reflux. The mixture was acidified with 1 N HCI and extracted with CHCl 3 -MeOH (4:1, 2 x 200 mnl). The combined extracts were dried over MgSO 4 and evaporated. The residue was chromtographed on silica gel with CHCl 3 -MeOH (20:1) as eluent to give 69 (260 mg, 30%) as a pale yellow amorphous solid. MW 555.98 'H-NMR (DMSO-d6)8 2.24-2.501 (min, 2 H), 3.48-4.65 (series of m, 15 10 H), 5.30-5.50 (min, 1 H), 6.75-7.08 (min, 5 H), 7.29 (t, J= 7.3 Hz, 1 H), 7.43-7.45 (min, 1H), 7.89 7.98 (min, 2 H), 7.99 (d, J = 8.3 Hz, 1 H), 8.09 (d, J = 7.1 Hz, 1 H), 8.90-8.96 (mn, 2 H); MS (FAB) m/z 556 (M'+1); Anal.Calcd for C 2 8
H
27 CIFN30 6 -1/4H 2 0: C, 60.00; H, 4.95; N, 7.50. Found: C, 59.67; H, 5.08; N, 7.10. Example 64 20 4-[1-[4-[N'-(2-bromophenyl)ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoic acid N N O COOH r H H 70 Methyl 4-[ 1-(4-benzyloxycarbonylaminophenylacetyl)-(4S)-fluoro-(2S)-pyrrolidinylmethoxy] benzoate (300 mg, 0.576 mmol) was tsdded EtOH-THF (5:1, 30 ml) and the solution was 25 hydrogenated over 5% Pd/C (300 ml) for 12 h while stirring. The mixture was filtered to remove the catalyst. The filtrate was concentrated under a reduced pressure. The residue was chromatographed on silica gel with CHCl 3 -MeOH (20:1) as eluent to give methyl 4-[1-(4 aminophenylacetyl)-(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (200 mg, 90%) as a yellow oil. 1 H-NMR (CDCl 3 ) 8 2.01-2.56 (series of min, 2 H), 3.50-4.14 (series of m, 5 H), 4.45-4.62 (min, 2 H), 30 5.21 and 5.34 (each m, total 1 H), 6.60-6.65 (min, 2 H), 6.88 (d, J= 8.8 Hz, 0.5 H), 6.99-7.05 (inm, 3.5 H), 7.95-8.00 (min, 2 H). 200 WO 01/00206 PCT/US00/18079 methyl 4-[1-(4-aminophenylacetyl)-(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (200 mg, 0.518 mmol) was dissolved THF (10 ml). Et 3 N (108 ul, 0.776 mmol) and 2-bromophenylisocyanate (96 ul, 0.776 mmol) were added to the solution. The mixture was stirred overnight and diluted with EtOAc (200 ml). The solution was washed with brine (100 ml), dried over MgSO 4 , and the solvent 5 was removed under a reduced pressure. The residue was chromatographed on silica gel with CHCl 3 -EtOAc (4:1) to CHCl 3 -MeOH (10:1) as eluent to give methyl 4-[1-[4-[N'-(2-bromophenyl) ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (303 mg, quant) as a yellow oil. 'H-NMR (CDCl 3 ) 8 2.08-2.60 (series of m, 2 H), 3.56-4.69 (series of m, 10 H), 5.28 and 5.40 (m, each, total 1 H), 6.84-6.92 (m, 3 H), 7.03-7.10 (m, 3 H),7.14(d, J=8.1 Hz,1 H), 7.23 (t,J= 8.1 10 Hz, 1 H), 7.39-7.44 (m, 2 H), 7.89 (d,J= 8.1 Hz, 1 H), 7.98-8.03 (m, 2H), 8.09(d,J= 8.1 Hz, 1H). Methyl 4-[ 1 -[4-[N'-(2-bromophenyl)ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinyl-methoxy] benzoate (300 mg, 0.513 mmol) was dissolved THF (5 ml), and 0.25 N NaOH (4.0 ml, 1.00 mmol) was added to this solution. After being stirred for 3 days, the mixture was poured into 1 N HCI (100 ml), and extracted with CHC1 3 -MeOH (5:1, 2 x 200 ml). The combined extracts were dried 15 over MgSO 4 and the solvent was removed under a reduced pressure. The residue was chromatographed on silica gel with CHC13-MeOH (10:1) to give 70 (209 mg, 71%) as a colorless amorphous solid. MW 570.41 'H-NMR (DMSO-d 6 )8 2.24-2.51 (m, 2 H), 3.36-4.64 (series of m, 7 H), 5.31-5.50 (m, 1 H), 6.97 (t, J = 7.8 Hz, 1 H), 7.04 (d, J = 8.5 Hz, 1 H), 7.09 (d, J= 8.8 Hz, 1 H), 7.14-7.20 (m, 2 H), 7.34 (t, J= 7.8 Hz, 1 H), 7.38-7.43 (m, 2 H), 7.61 (d, J= 8.1 Hz, 1 H), 20 7.87-7.92 (m, 2 H), 8.08 (d,J= 8.1 Hz, 1 H), 8.15 (s, 1 H), 9.45-9.47 (m, 1 H), 12.66 (br s, 1 H); MS (FAB) m/z 572 (M+2), 570 (Me); Anal. Calcd for C 27
H
25 BrFN 3 0 5 -1.5 H20: C, 54.28; H, 4.72; N, 7.03. Found: C, 54.67; H, 4.51; N, 6.61. Example 65 4-[ -[4-[N'-(2-iodophenyl)ureido]-3-methoxyphenylacetyl]-(4S)-fluoro-(2S)-pyrrffolidinylmethoxy] 25 benzoic acid F N N O- COOH 71 H HiMe 71 To a stirred solution of tert-butyl 4-amino-3-methoxyphenylacetate (1.94 g, 8.16 mmol) in THF (20 ml) was added 2-iodophenylisocyanate (2.0 g, 8.16 mmol) and Et 3 N (114 ul, 0.816 mmol). After stirring overnight, the mixture was poured into 1 N HCI (200 ml). The resulting precipitate 30 was collected by filtlation and dissolved in CHC1 3 (200 ml). The solution was dried over MgSO 4 and evaporated to give tert-butyl 4-[N'-(2-iodophenyl)ureido]-3-methoxyphenylacetate (3.93 g, 201 WO 01/00206 PCT/US00/18079 quant) as a pale yellow amorphous solid. 'H-NMR (CDCl 3 )8 1.44 (s, 9 H), 3.49 (s, 2 H), 3.85 (s, 3 H), 6.78-6.88 (min, 4 H), 7.07 (s, 1 H), 7.31-7.35 (min, 1 H), 7.76 (dd, J= 7.8, 1.5 Hz, 1 H), 7.95 (d, J = 8.3 Hz, 1 H), 7.99 (dd, J= 8.3, 1.5 Hz, 1 H). MS (ESI), m/z 483 (MW+1). A stirred mixture of tert-butyl 4-[N'-(2-iodophenyl)ureido]-3-methoxyphenylacetate (3.93 5 g, 8.16 mmol) and TFA (5 ml) in CH 2 Cl 2 (5 ml) was refluxed for 3 h. After cooling to rt, the mixture was concentrated in vacuo and H 2 0 (50 ml) was added to this residue. The resulting precipitate was collected by filtration and purified by column chromatography on silica gel with CHCl 3 -MeOH (9:1) as eluent to give 4-[N'-(2-iodophenyl)ureido]-3-methoxyphenylacetic acid (2.89 g, 83%) as a pale yellow crystalline powder. 'H-NMR (DMSO-d 6 )8 3.62 (s, 2 H), 3.88 (s, 3 10 H), 6.78 (d, J= 8.3 Hz, 1 H), 6.83-6.87 (min, 1 H), 6.94 (s, 1 H), 7.32-7.36 (min, 1 H), 7.69 (dd, J= 8.3, 1.5 Hz, 1 H), 7.84 (dd, J= 8.3, 1.5 Hz, 1 H), 7.97-8.00 (min, 1 H), 8.55 (m, 1 H), 8.82 (m, 1 H), 12.26 (br s, 1 H). A mixture of 4-[N'-(2-iodophenyl)ureido]-3-methoxyphenylacetic acid (505 mg, 1.18 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (300 mg, 1.18 mmol), EDC-HCI (339 15 mg, 1.77 mmol), DMAP (catatlytic amount) and HOBt (catalytic amount) in DMF (10 ml) was stirred overnight. The mixture was diluted with EtOAc (300 ml) and washed with brine (2 x 200 ml). The solution was dried over MgSO 4 and evaporated. The resulting residue was chromatographed on silica gel with CHCl 3 -EtOAc (4:1) as eluent to give methyl 4-[1-[4-[N'-(2 iodophenyl)ureido]-3-methoxyphenylacetylJ]-(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (500 20 mg, 64%) as a colorless viscous oil. 'H-NMR (CDCl 3 ) 8 07-2.58 (m, 2 H), 3.59-4.20 (min, 11 H), 4.51-4.64 (min, 2 H), 5.24 and 5.37 (min, each, total 1 H), 6.80-6.91 (min, 5 H), 6.98 (d, J= 8.8 Hz, 2 H), 7.34 (t, J = 7.8 Hz, 1 H), 7.78 (dd, J = 7.8, 1.2 Hz, 1 H), 7.95-8.02 (m, 4 H). To a stirred solution of methyl 4-[1-[4-[N'-(2-iodophenyl)ureido]-3-methoxyphenyl acetyl]-(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (500 mg, 0.756 mmol) in THF (6 ml) was 25 added 0.25 N NaOH (6 ml). The stirring was continued overnight at room temperature then 5 h at reflux. After cooling to rt, the solution was poured into 1 N HCI (100 ml) and extracted with CHCl 3 -MeOH (4:1, 2 x 200 ml). The combined extracts were dried over MgSO 4 and evaporated. The residue was chromatographed on silica gel with CHCl 3 -MeOH (10:1) as eluent to give 71 (295 mg, 60%) as a colorless amorphous solid. MW 647.43 'H-NMR (DMSO-d 6 )8 2. 2.09-2.31 30 (min, 2 H), 3.33-4.41 (series of m, 10 H), 5.30-5.50 (min, 1 H), 6.77-6.92 (min, 3 H), 7.03-7.09 (mn, 2 H), 7.34 (t, J= 8.1 Hz, 1 H), 7.69 (dd, J= 8.3, 1.5 Hz, 1 H), 7.83-7.99 (min, 4 H), 8.54 (mn, 1 H), 202 WO 01/00206 PCT/US00/18079 8.82 (min, 1 H); MS (FAB) m/z 648 (M++1); Anal. Calcd for C 28
H
27
FIN
3 0 4 : C, 51.94; H, 4.20; N, 6.49. Found: C, 51.17; H, 4.53; N, 5.76. Example 66 4-[(4S)-fluoro-l-[4-(N '-phenylureido)phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoic acid F N N 't& 0' \ COOH 5H H 72 To a stirred solution of ethyl 4-aminophenylacetate (6.43 g, 35.9 mmol) and Et 3 N (5.50 ml, 39.5 mmol) in THF (70 ml) was added phenyl isocyanate (3.90 ml, 35.9 mmol), and the reaction mixture was stirred at room temperature for 4 days. Resulting precipitate was collected under a reduced pressure and the filtrate was washed with n-hexane to give ethyl 4-(N'-phenylureido) 10 phenylacetate (9.64 g, 90%) as a white crystalline powder. mp 153-155 0 C; 'H-NMR (CDC13)8 1.26 (t, J = 7.1 Hz, 3 H), 3.52 (s, 2 H), 4.15 (q, J= 7.1 Hz, 2 H), 6.98-7.04 (min, 1 H), 7.07-7.11 (min, 4 H), 7.18-7.25 (min, 5 H), 7.42 (s, 1 H); MS (FAB) m/z 299 (M+1); Anal. Calcd for
C
17
H
18 sN 2 0 3 : C, 68.44; H, 6.08; N, 9.39. Found: C, 68.22; H, 6.10; N, 9.36. To a stirred solution of ethyl 4-(N'-phenylureido)phenylacetate (9.64 g, 32.3 mmol) in 15 THF (80 ml) was added 0.5 N NaOH (80 ml) and the reaction mixture was heated under reflux for 5 hr. After cooled to room temperature, the mixture was poured into ice-1 N HC1. The resulting precipitate was collected under a reduced pressure and the crude solid was recrystallized from MeOH-CHCl 3 to give 4-(N'-phenylureido)phenylacetic acid (8.14 g, 93%) as a white crystalline powder. MS (FAB) m/z 271 (M +I); Anal. Calcd for C 15
HI
14
N
2 0 3 : C, 66.66; H, 5.22; N, 10.36. 20 Found: C, 66.45; H, 5.22; N, 10.30. A mixture of 4-(N'-phenylureido)phenylacetic acid (310 mg, 1.15 mmol), methyl 4-[(4S) fluoro-(2S)-pyrrolidinylmethoxy]benzoate (287 mg, 1.13 mmol), EDCHC1 (260 mg, 1.36 mmol), HOBt (185 mg, 1.37 mmol), and Et 3 N (190 ml, 1.36 mmol) in DMF (5 ml) was stirred at room temperature overnight. The mixture was diluted with H 2 0, and extracted with EtOAc. The extract 25 was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1 to 50:1, v/v) as eluent to give methyl 4 [(4S)-fluoro-1-[4-(N '-phenylureido)phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (570 mg, 99%) as a pale yellow foam. 'H-NMR (CDCl 3 ) 8 2.07-2.58 (min, 2 H), 3.55-3.56 (min, 1 H), 3.69 3.98 (series of s and m, total 6 H), 4.01-4.08 and 4.21-4.25 (each m, 1 H), 4.46-4.65 (mn, 2 H), 30 5.23-5.25 and 5.38 (each min, 1 H), 6.88-7.07 (min, 7 H), 7.15-7.20 (min, 2 H), 7.28-7.30 (mn, 2 H), 7.34 203 WO 01/00206 PCT/US00/18079 and 7.40 (each s, 1 H), 7.71 and 7.81 (each s, 1 H), 7.91-7.95 and 7.99-8.01 (each m, 2 H); MS (ESI) m/z 506 (M++1). To a stirred solution of methyl 4-[(4S)-fluoro-1-[4-(N'-phenylureido)phenylacetyl]-(2S) pyrrolidinylmethoxy]benzoate (570 mg, 1.13 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml), 5 and the reaction mixture was heated under reflux for 5 hr. After cooled to room temperature, the mixture was poured into ice-I N HCI, and the resulting precipitate was collected under a reduced pressure. The crude solid was recrystallized from MeOH-CHCl 3 -IPE to give 72 (348 mg, 63%) as a white crystalline powder. MW 491.51 mp 169-171oC; 'H-NMR (DMSO-d) 8 2.24-2.36 (m, 2 H), 3.47-4.08 (m, 5 H), 4.20-4.64 (m, 2 H), 5.31-5.50 (m, 1 H), 6.94-7.46 (series of m, total 11 H), 10 7.87-7.92 (m, 2 H), 8.64-8.67 (m, 2 H), 12.63 (br s, 1 H); MS (FAB) m/z 492 (M+1); Anal. Calcd for C 27 H2 6
FN
3 0 6 1/4H 2 0: C, 65.38; H, 5.38; N,8.47; F,3.83. Found: C,65.13; H,5.38; N,8.25; F,3.78 Example 67 4-[(4S)-fluoro-1 -[3-methyl-4-(N '-phenylurcido)phenylacetyl]-(2S)-pyrrffolidinylmethoxy]benzoic 15 acid F N NN e N - COOH 73 H H Me 73 To a stirred solution of tert-butyl 4-amino-3-methylphenylacetate (1.20 g, 5.42 mmol) and Et 3 N (830 ml, 5.95 mmol) in THF (10 ml) was added phenyl isocyanate (650 ml, 5.98 mmol) and the reaction mixture was stirred at room temperature overnight. The reaction mixture was 20 concentrated to a small volume and diluted with n-hexane. Resulting precipitate was collected under a reduced pressure and the filtrate was washed with n-hexane to give tert-butyl 3-methyl-4 (N'-phenylureido)phenylacetate (1.12 g, 61%) as a white crystalline powder. mp 143-145oC; 'H NMR (CDC13) 8 1.47 (s, 9 H), 2.09 (s, 3 H), 3.47 (s, 2 H), 6.44 (s, 1 H), 7.01-7.07 (m, 4 H), 7.16 7.27 (m, 2 H), 7.30-7.33 (m, 2 H), 7.45-7.47 (m, 1 H). 25 To a stirred solution of tert-butyl 3-methyl-4-(N'-phenylureido)phenylacetate (1.12 g, 3.29 mmol) in CH 2 C1 2 (10 ml) was added TFA (10 ml) and the reaction mixture was stirred at room temperature for 4 hr. The reaction mixture was concentrated to a small volume and poured into ice-H 2 0. The resulting precipitate was collected under a reduced pressure and the crude solid was recrystallized from MeOH-CHCl 3 to give 3-methyl-4-(N'-phenylureido)phenylacetic acid (680 mg, 30 73%) as white needles. 'H-NMR (DMSO-d( 6 ) 8 2.22 (s, 3 H), 3.46 (s, 2 H), 6.93-7.05 (m, 3 H), 204 WO 01/00206 PCT/US00/18079 7.25-7.29 (m, 2 H), 7.43-7.46 (m, 2 H), 7.72-7.74 (m, 1 H), 7.90 (s, 1 H), 8.98 (s, 1 H), 12.26 (br s, 1 H); Anal. Calcd for C 16
H
1 6
N
2 0 3 : C, 67.59; H, 5.67; N, 9.85. Found: C, 67.47; H, 5.68; N, 9.73. A mixture of 3-methyl-4-(N'-phenylureido)phenylacetic acid (301 mg, 1.06 mmol), methyl 4 [(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (268 mg, 1.06 mmol), EDCHCI (243 mg, 1.27 5 mmol), HOBt (172 mg, 1.27 mmol), and Et 3 N (180 ml, 1.29 mmol) in DMF (5 ml) was stirred at room temperature overnight. The mixture was diluted with H 2 0, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCI 3 -MeOH (100:1 to 60:1, v/v) as eluent to give methyl 4-[(4S)-fluoro- 1-[3-methyl-4-(N'-phenylureido)phenylacetyl]-(2S)-pyrrolidinylmethoxy] 10 benzoate (550 mg, q.y.) as a white foam. 'H-NMR (CDCl 3 ) 8 1.79 and 1.87 (each s, 3 H), 2.04 2.61 (m, 2 H), 3.52-3.54 (m, 1 H), 3.73-4.27 (series of s and m, total 7 H), 4.47-4.67 (m, 2 H), 5.26-5.27 and 5.40 (each m, 1 H), 6.79-6.99 (m, 6 H), 7.14-7.18 (m, 2 H), 7.27-7.31 (m, 2 H), 7.40-7.44 (m, 1 H), 7.89-8.01 (m, 3 H); MS (ESI) m/z 520 (M'+1). To a stirred solution of methyl 4-[(4S)-fluoro--[3-methyl-4-(N'-phenylureido) 15 phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (550 mg, 1.06 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml), and the reaction mixture was heated under reflux for 2 hr. After cooled to room temperature, the mixture was poured into ice-1 N HC1, and the resulting precipitate was collected under a reduced pressure. The crude solid was recrystallized from MeOH-CHCl 3 -IPE to give 73 (226 mg, 42%) as a white crystalline powder. MW 505.54 mp 130-135oC; 'H-NMR 20 (DMSO-d) 8 2.18-2.30 (series of s and m, total 5 H), 3.47-3.92 (series of m, total 5 H), 4.03-4.63 (m, 2 H), 5.31-5.50 (m, 1 H), 6.94-7.10 (m, 5 H), 7.26-7.30 (m, 2 H), 7.45-7.47 (m, 2 H), 7.70 7.75 (m, 1 H), 7.87-7.92 (m, 3 H), 8.96-8.98 (m, 1 H), 12.63 (br s, 1 H); MS (ESI) m/z 506 (M+1); Anal. Calcd for C 28
H
28
FN
3 0s.1/2H 2 0: C, 65.36; H, 5.68; N, 8.17; F, 3.69. Found: C, 65.61; H, 5.71; N, 7.84; F, 3.60. 25 Example 68 4-[(4S)-fluoro-1 -[4-[N'-(2-fluorophenyl)ureido]-3-methylphenylacetyl]-(2S)-pyrrolidinylmethoxy] benzoic acid F N N O/ COOH H H Me 74 To a stirred solution of tert-butyl 4-amino-3-methylphenylacetate (1.09 g, 4.93 mmol) and Et 3 N 30 (755 ml, 5.42 mmol) in THF (10 ml) was added 2-fluorophenyl isocyanate (610 tl, 5.44 mmol) 205 WO 01/00206 PCT/US00/18079 and the reaction mixture was stirred at room temperature overnight. The reaction mixture was concentrated to a small volume and diluted with n-hexane. Resulting precipitate was collected under a reduced pressure and the filtrate was washed with n-hexane to give tert-butyl 4-[N'-(2 fluorophenyl)ureido]-3-methylphenylacetate (1.31 g, 74%) as a white crystalline powder. mp 89 5 91 0 C; 'H-NMR (CDCI 3 ) 8 1.47 (s, 9 H), 2.06 (s, 3 H), 3.49 (s, 2 H), 6.62 (s, 1 H), 6.92-7.09 (m, 5 H), 7.21 (br s, 1 H), 7.49-7.51 (mn, 1 H), 8.10-8.15 (min, 1 H); Anal. Calcd for C 20
H
2 3
FN
2 0 3 : C, 67.02; H, 6.47; N, 7.82; F, 5.30. Found: C, 66.74; H, 6.35; N, 7.85; F, 5.69. To a stirred solution of tert-butyl 4-[N'-(2-fluorophenyl)ureido]-3-methylphenylacetate (1.25 g, 3.49 mmol) in CH 2
CI
2 (10 ml) was added TFA (10 ml) and the reaction mixture was 10 stirred at room temperature overnight. The reaction mixture was concentrated to a small volume and poured into ice-H 2 0. The resulting precipitate was collected under a reduced pressure and the crude solid was recrystallized from MeOH-CHCl 3 -IPE to give 4-[N'-(2-fluorophenyl)ureido]-3 methylphenylacetic acid (830 mg, 79%) as white needles. 'H-NMR (DMSO-d) 8 2.23 (s, 3 H), 3.47 (s, 2 H), 6.96-7.30 (min, 5 H), 7.74-7.76 (min, 1 H), 8.17-8.20 (min, 1 H), 8.33 (s, 1 H), 8.94 (s, 1 15 H), 12.27 (br s, 1 H); Anal. Calcd for C,HFN 2 0 3 : C, 63.57; H, 5.00; N, 9.27; F, 6.28. Found: C, 63.28; H, 5.00; N, 9.14; F, 6.43. A mixture of 4-[N'-(2-fluorophenyl)ureido]-3-methylphenylacetic acid (321 mg, 1.06 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (269 mg, 1.06 mmol), EDCHCI (244 mg, 1.27 mmol), HOBt (172 mg, 1.27 mmol), and Et 3 N (177 ml, 1.27 mmol) in DMF (5 ml) was 20 stirred at room temperature overnight. The mixture was diluted with H 2 0, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHC13-MeOH (100:1, v/v) as eluent to give methyl 4-[(4S)-fluoro-1-[4-[N'-(2-fluorophenyl)ureido]-3-methylphenylacetyl]-(2S)-pyrrolidinyl methoxy]benzoate (560 mg, 98%) as a white foam. 'H-NMR (CDCl 3 )8 1.78 and 1.86 (each s, 3 25 H), 2.16-2.65 (min, 2 H), 3.58-3.61 (min, 1 H), 3.74-4.15 (series of s and m, total 7 H), 4.29-4.34 and 4.46-4.49 (each m, 1 H), 4.64-4.73 (min, 1 H), 5.29-5.34 and 5.43-5.47 (each m, 1 H), 6.84-6.97 (inm, 6 H), 7.04-7.07 (min, 1 H), 7.21 (br s, 1 H), 7.55-7.59 (min, 1 H), 7.85-8.02 (min, 3 H), 8.18-8.22 (mn, 1 H); MS (ESI) m/z 538 (M+1). To a stirred solution of methyl 4-[(4S)-fluoro-1-[4-[N'-(2-fluorophenyl)ureido]-3 30 methylphenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (560 mg, 1.04 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml), and the reaction mixture was heated under reflux for 5 hr. After cooled 206 WO 01/00206 PCT/US00/18079 to room temperature, the mixture was poured into ice-1 N HCI, and the resulting precipitate was collected under a reduced pressure. The crude solid was recrystallized from MeOH-CHCl 3 -IPE to give 74 (297 mg, 42%) as a white crystalline powder. MW 523.53 mp 137-143oC; 'H-NIMR (DMSO-d 6 )8 2.20-2.31 (series of s and m, total 5 H), 3.56-3.92 (series of m, total 5 H), 4.03-4.63 5 (m, 2 H), 5.31-5.50 (m, 1 H), 6.96-7.26 (series of m, total 7 H), 7.72-7.77 (m, 1 H), 7.87-7.92 (m, 2 H), 8.17-8.22 (m, 1 H), 8.32-8.36 (m, 1 H), 8.94-8.95 (m, 1 H), 12.66 (br s, 1 H); MS (ESI) mn/z 524 (M +1); Anal. Calcd for C 28
H
27
F
2
N
3 0: C, 64.24; H, 5.20; N, 8.03; F, 7.26. Found: C, 64.44; H, 5.75; N, 7.40; F, 6.73. Example 69 10 4-[(4S)-fluoro- 1-[4-[N '-(2-trifluoromethylphenyl)ureido]phenylacetyl]-(2S)-pyrrolidinylmethoxy] benzoic acid F N N k COOH
CF
3 H H 75 To a stirred solution of ethyl 4-aminophenylacetate (1.13 g, 6.31 mmol) and Et 3 N (965 ml, 6.92 mmol) in THF (10 ml) was added 2-trifluoromethylphenyl isocyanate (953 ml, 6.31 mmol) and the 15 reaction mixture was stirred at room temperature for 2 days. Resulting precipitate was collected under a reduced pressure and the filtrate was washed with n-hexane to give ethyl 4-[N'-(2 trifluoromethylphenyl)ureido]phenylacetate (1.93 g, 84%) as white needles. mp 137-139 0 C; 'H NMR (CDCI 3 ) 8 1.25-1.29 (m, 3 H), 3.59 (s, 2 H), 4.15-4.20 (m, 2 H), 7.05 (br s, 1 H), 7.13-7.23 (m, 6 H), 7.47-7.51 (m, 1 H), 7.54-7.56 (m, 1 H), 8.01-8.03 (m, 1 H). 20 To a stirred solution of ethyl 4-[N'-(2-trifluoromethylphenyl)ureido]phenylacetate (1.93 g, 5.27 mmol) in THF (10 ml) was added 1 N NaOH (10 ml) and the reaction mixture was heated under reflux for 5 hr. After cooled to room temperature, the mixture was poured into ice-1 N HCl. The resulting precipitate was collected under a reduced pressure and the crude solid was recrystallized from MeOH-CHCl 3 -IPE to give 4-[N'-(2-trifluoromethylphenyl)ureido]phenylacetic 25 acid (910 mg, 51%) as a white crystalline powder. mp 224-225°C; 'H-NMR (DMSO-d 6 ) 8 3.50 (s, 2 H), 7.18 (d, J = 8.3 Hz, 2 H), 7.25-7.29 (m, 1 H), 7.40 (d, J = 8.3 Hz, 2 H), 7.62-7.69 (m, 2 H), 7.95-7.97 (m, 1 H), 8.06 (s, 1 H), 9.37 (s, 1 H), 12.27 (br s, 1 H); Anal. Calcd for C 16
H
3
F
3
N
2 0 3 : C, 56.81; H, 3.87; N, 8.28; F, 16.85. Found: C, 56.68; H, 3.87; N, 8.16; F, 16.89. A mixture of 4-[N'-(2-trifluoromethylphenyl)ureido]phenylacetic acid (302 mg, 0.89 mmol), 30 methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (226 mg, 0.89 mmol), EDCHC1 (205 207 WO 01/00206 PCT/US00/18079 mg, 1.07 mmol), HOBt (145 mg, 1.07 mmol), and Et 3 N (150 ml, 1.08 mmol) in DMF (5 ml) was stirred at room temperature for 3 days. The mixture was diluted with H 2 0, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1 to 60:1, v/v) as eluent 5 to give methyl 4-[(4S)-fluoro-1l-[4-[N'-(2-trifluoromethylphenyl)ureido]phenylacetyl]-(2S) pyrrolidinylmethoxy]benzoate (463 mg, 90%) as a pale yellow foam. 'H-NMR (CDCl 3 ) 8 2.09 2.60 (m, 2 H), 3.56-4.12 (series of s and m, total 8 H), 4.26-4.65 (m, 2 H), 5.26-5.29 and 5.39-5.42 (each m, total 1 H), 6.87-6.93 (m, 2 H), 6.99-7.13 (m, 5 H), 7.30-7.33 (m, 1 H), 7.44-7.53 (m, 2 H), 7.88-7.92 (m, 1 H), 7.99-8.04 (m, 2 H), 8.09-8.15 (m, 1 H); MS (ESI) m/z 574 (M++i). 10 To a stirred solution of methyl 4-[(4S)-fluoro-1-[4-[N'-(2-trifluoromethylphenyl)ureido] phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (460 mg, 0.80 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml), and the reaction mixture was heated under reflux for 5 hr. After cooled to room temperature, the mixture was poured into ice-1 N HCI, and the resulting precipitate was collected under a reduced pressure. The crude solid was recrystallized from MeOH-CHCl 3 -IPE to 15 give 75 (169 mg, 38%) as a white crystalline powder. MW 559.51 mp 130-135 0 C; 'H-NMR (DMSO-d 6 ) 8 2.24-2.30 (m, 2 H), 3.51-4.24 (series of m, total 5 H), 4.38-4.40 and 4.61 (each m, total 2 H), 5.31-5.50 (m, 1 H), 7.03-7.42 (series of m, total 7 H), 7.62-7.69 (m, 2 H), 7.87-8.07 (m, 4 H), 9.36-9.37 (m, 1 H), 12.64 (br s, 1 H); MS (ESI) m/z 560 (M+1); Anal. Calcd for
C
28
H
2
,F
4
N
3 0s: C, 60.11; H, 4.50; N, 7.51; F, 13.58. Found: C, 60.10; H, 4.85; N, 7.01; F, 12.90. 20 Example 70 4-[(4S)-fluoro-1-[3-methoxy-4-[N'-(2-trifluoromethylphenyl)ureido]phenylacetyl]-(2S)-pyrrolidinyl methoxy]benzoic acid F N N M NCOOH 76 FH HiMe 76 To a stirred solution of tert-butyl 4-amino-3-methoxyphenylacetate (1.11 g, 4.68 mmol) and Et 3 N 25 (720 ml, 5.17 mmol) in THF (10 ml) was added 2-trifluoromethylphenyl isocyanate (707 ml, 4.68 mmol) and the reaction mixture was stirred at room temperature for 2 days. The reaction mixture was concentrated to a small volume and diluted with n-hexane. Resulting precipitate was collected under a reduced pressure and washed with n-hexane to give tert-butyl 3-methoxy-4-[N'-(2 trifluoromethylphenyl)ureido]phenylacetate (1.11 g, 56%) as a white crystalline powder. mp 131 30 133oC; 'H-NMR (CDCl 3 ) 8 1.44 (s, 9 H), 3.49 (s, 2 H), 3.85 (s, 3 H), 6.83-6.88 (m, 3 H), 6.98 (br s, 1 H), 7.17-7.21 (m, 1 H), 7.52-7.59 (m, 2 H), 7.89-7.91 (m, 1 H), 8.04-8.06 (m, 1 H). 208 WO 01/00206 PCT/US00/18079 To a stirred solution of tert-butyl 3-methoxy-4-[N'-(2-trifluoromethylphenyl)ureido] phenylacetate (1.11 g, 2.62 mmol) in CH 2 C1 2 (10 ml) was added TFA (10 ml), and the reaction mixture was stirred at room temperature for 4 hr. The reaction mixture was concentrated to a small volume and poured into ice-H 2 0. The resulting precipitate was collected under a reduced 5 pressure and the crude solid was recrystallized from MeOH-CHC13-IPE to give 3-methoxy-4-[N' (2-trifluoromethylphenyl)ureido]phenylacetic acid (839 mg, 87%) as a white crystalline powder. mp 218-220 0 C; 'H-NMR (DMSO-d) 8 3.51 (s, 2 H), 3.87 (s, 3 H), 6.76-6.79 (mn, 1 H), 6.93-6.94 (mn, 1 H), 7.27-7.30 (min, 1 H), 7.61-7.69 (min, 2 H), 7.82-7.84 (mn, 1 H), 7.97-7.99 (mn, 1 H), 8.71 (s, 1 H), 8.89 (s, 1 H), 12.30 (br s, 1 H); Anal. Calcd for C 7
H,
5
F
3
N
2 0 4 : C, 55.44; H, 4.11; N, 7.61; F, 10 15.47. Found: C, 55.30; H, 4.08; N, 7.63; F, 15.13. A mixture of 3-methoxyl-4-[N'-(2-trifluoromethylphenyl)ureido]phenylacetic acid (353 mg, 0.96 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (243 mg, 0.96 mmol), EDCHCI (221 mg, 1.15 mmol), HOBt (156 mg, 1.15 mmol), and Et 3 N (160 ml,, 1.15 mmol) in DMF (5 ml) was stirred at room temperature overnight. The mixture was diluted with H 2 0, and extracted with 15 EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1 to 60:1, v/v) as eluent to give methyl 4-[(4S)-fluoro-1l-[3-methoxy-4-[N'-(2-trifluoromethylphenyl)ureido] phenylacetyl] (2S)-pyrrolidinylmethoxy]benzoate (570 mg, 98%) as a white foam. 'H-NMR (CDCl 3 ) 8 2.05-2.58 (min, 2 H), 3.56-4.21 (series of m, total 11 H), 4.05-4.64 (min, 2 H), 5.23-5.25 and 5.36-5.37 (each inm, 20 total 1 H), 6.79-6.82 (min, 2 H), 6.89-7.00 (min, 2 H), 7.16-7.20 (min, 2 H), 7.39-7.43 (mn, 1 H), 7.51 7.59 (min, 2 H), 7.93-8.02 (min, 4 H); MS (ESI) m/z 604 (M+1). To a stirred solution of methyl 4-[(4S)-fluoro-1-[3-methoxy-4-[N'-(2-trifluoromethyl phenyl)ureido]phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (570 mg, 0.94 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml), and the reaction mixture was heated under reflux for 2 hr. 25 After cooled to room temperature, the mixture was poured into ice-1 N HCI, and the resulting precipitate was collected under a reduced pressure. The crude solid was recrystallized from MeOH-CHC1 3 -IPE to give 76 (234 mg, 42%) as a white crystalline powder. MW 589.54 mp 129 132 0 C; 'H-NMR (DMSO-d 6 )8 2.23-2.29 (min, 2 H), 3.54-4.38 (series of s and min, total 8 H), 4.40 4.61 (min, 2 H), 5.30-5.36 and 5.43-5.49 (each min, total 1 H), 6.72-6.91 (mn, 2 H), 7.02-7.08 (min, 2 H), 30 7.25-7.29 (min, 1 H), 7.59-7.67 (min, 2 H), 7.81-7.99 (min, 4 H), 8.69-8.70 (mn, 1 H), 8.87-8.90 (mn, 1 H), 12.67 (br s, 1 H); MS (ESI) m/z 589 (M*+1); Anal Calcd for C 29
H
27
F
4
N
3 0 6 : C, 59.08; H, 4.62; N, 7.13. Found: C, 59.22; H, 5.10; N, 6.58. 209 WO 01/00206 PCT/US00/18079 Example 71 4-[(4S)-fluoro- 1-[3-methyl-4-[N'-(2-trifluoromethylphenyl)ureido]phenylacetyl]-(2S) pyrrolidinylmethoxy]benzoic acid F Q JN N\/-COOH
CF
3 H H e 77 5 To a stirred solution of tert-butyl 4-amino-3-methylphenylacetate (927 mg, 4.19 mmol) and Et 3 N (645 iil, 4.63 mmol) in THF (10 ml) was added 2-trifluoromethylphenyl isocyanate (633 jl, 4.19 mmol) and the reaction mixture was stirred at room temperature for 2 days. The reaction mixture was concentrated to a small volume and diluted with n-hexane. Resulting precipitate was collected under a reduced pressure and the filtrate was washed with n-hexane to give tert-butyl 3 10 methyl-4-[N'-(2-trifluoromethylphenyl)ureido]phenylacetate (1.06 g, 62%) as a white crystalline powder. mp 178-180oC; 'H-NMR (CDCl 3 ) 8 1.44 (s, 9 H), 2.25 (s, 3 H), 3.51 (s, 2 H), 6.38 (br s, 1 H), 7.12-7.18 (m, 3 H), 7.36-7.37 (m, 1 H), 7.49-7.53 (m, 2 H), 8.13-8.16 (m, 1 H). To a stirred solution of tert-butyl 3-methyl-4-[N'-(2-trifluoromethylphenyl)ureido] phenylacetate (1.06 g, 2.60 mmol) in CH 2 Cl 2 (10 ml) was added TFA (10 ml) and the reaction 15 mixture was stirred at room temperature for 4 hr. The reaction mixture was concentrated to a small volume and poured into ice-H 2 0. The resulting precipitate was collected under a reduced pressure and the crude solid was recrystallized from MeOH-CHCl 3 -IPE to give 3-methyl-4-[N'-(2 trifluoromethylphenyl)ureido]phenylacetic acid (702 mg, 77%) as a white crystalline powder. mp 262-263oC; 'H-NMR (DMSO-d 6 ) 8 2.24 (s, 3 H), 3.48 (s, 2 H), 7.03 (d, J = 8.3 Hz, 1 H), 7.08 (s, 1 20 H), 7.26-7.30 (m, 1 H), 7.61-7.69 (m, 3 H), 7.88 (d, J = 8.3 Hz, 1 H), 8.39 (s, 1 H), 8.55 (s, 1 H), 12.28 (br s, 1 H); Anal. Called for C 1 7
H,
5
F
3
N
2 0 3 : C, 57.96; H, 4.29; N, 7.95; F, 16.18. Found: C, 57.73; H, 4.23; N, 7.92; F, 16.05. A mixture of 3-methyl-4-[N'-(2-trifluoromethylphenyl)ureido]phenylacetic acid (359 mg, 1.02 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (258 mg, 1.02 mmol), EDCHCI 25 (234 mg, 1.22 mmol), HOBt (165 mg, 1.22 mmol), and Et 3 N (170 4l, 1.22 mmol) in DMF (5 ml) was stirred at room temperature overnight. The mixture was diluted with H 2 0, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1 to 60:1, v/v) as eluent to give methyl 4-[(4S)-fluoro-l1-[3-methyl-4-[N'-(2-trifluoromethylphenyl)ureido]phenylacetyl] 210 WO 01/00206 PCT/US00/18079 (2S)-pyrrolidinylmethoxy]benzoate (612 mg, q.y.) as a white foam. 1 H-NMR (CDCl 3 ) 8 1.92 and 2.00 (each s, total 3 H), 2.09-2.61 (m, 2 H), 3.56-4.29 (series of m, total 8 H), 4.45-4.48 and 4.59 4.64 (each m, total 2 H), 5.24-5.30 and 5.38-5.44 (each m, total 1 H), 6.90-7.14 (m, 5 H), 7.22 7.53 (m, 5 H), 7.90-7.92 (m, 1 H), 8.00-8.06 (m, 2 H); MS (ESI) m/z 588 (M'+1). 5 To a stirred solution of methyl 4-[(4S)-fluoro-1-[3-methyl-4-[N'-(2-trifluoromethylphenyl) ureido]phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (610 mg, 1.04 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml), and the reaction mixture was heated under reflux for 2 hr. After cooled to room temperature, the mixture was poured into ice-1 N HC1, and the resulting precipitate was collected under a reduced pressure. The crude solid was recrystallized from MeOH-CHCl 3 -IPE to 10 give 77 (186 mg, 31%) as a white crystalline powder. MW 573.54 mp 123-126oC; 'H-NMR (DMSO-d 6 ) 6 2.19-2.29 (series of s and m, total 5 H), 3.64-4.21 (series of m, total 5 H), 4.36-4.60 (m, 2 H), 5.30-5.36 and 5.43-5.49 (each m, total 1 H), 6.97-7.08 (m, 4 H), 7.24-7.28 (m, 1 H), 7.59-7.68 (m, 3 H), 7.85-7.90 (m, 3 H), 8.37-8.39 (m, 1 H), 8.54-8.55 (m, 1 H), 12.67 (br s, 1 H); MS (ESI) m/z 573 (M). 15 Example 72 4-[(4S)-fluoro- 1-[3-methyl-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S)-pyrrolidinyl] methoxybenzoic acid F NN \- COOH 78 Me H H Me 78 A mixture of 3-methyl-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (250 mg, 0.84 mmol), 20 methyl 4-[(4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (400 rmg, 1.06 mmol), EDC.HCl (242 mg, 1.26 mmol) and DMAP (154 rmg, 1.26 mmol) in DMF (5 ml) was stirred at room temperature for 21 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHClI/MeOH 25 (50/1)], and then was purified on TLC [CHCljacetone (10/1)] to give methyl 4-[(4S)-fluoro-1-[3 methyl-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S)-pyrrffolidinyl]methoxybenzoate (342 mg, 76%) as a colorless amorphous solid. IR (KBr) 3356, 2951, 1716, 1651, 1604, 1537, 1252 cm'; 1 H-NMR (CDCl 3 ) 8 2.07 (d, J= 6.6 Hz, 2H), 2.12 (s, 3H), 2.27 (m, 1H), 2.24 (s, 3H), 2.30-2.59 (m, 1H1), 3.60 (d,J= 5.3 Hz, 1H), 3.65-4.23 (m, 3H), 3.87 (s, 3H), 4.50-4.62 (m, 1H), 5.31 (d,J= 30 52.4 Hz, 1H), 6.23 (d,J = 11.2 Hz, 1H), 6.26 (d,J= 11.9 Hz, 1H), 6.87-7.27 (m, 8H), 7.54-7.65 (m, 3H), 7.94-8.01 (m, 2H); MS (FAB) m/z 534 (M+1); Anal. Calcd for C 3
H
32
FN
3 0 5 s-0.7H 2 0: C, 211 WO 01/00206 PCT/US00/18079 65.97; H, 6.16; F, 3.48; N, 7.69. Found: C, 66.04; H, 6.07; F, 3.55; N, 7.64. To a stirred solution of methyl 4-[(4S)-fluoro-1-[3-methyl-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-(2S)-pyrrolidinyl]methoxybenzoate (227 mg, 0.425 mmol) in THF (3.4 ml) was added 0.25 N NaOH (3.4 ml). After stirring at room temperature for 4 days, the mixture was 5 acidified with IN HCI and extracted with CHCl 3 -MeOH (10/1). The combined extracts were dried over Na 2
SO
4 and concentrated in vacuo. The residue was purified on preparative-TLC [CHCl 3 /MeOH (10/1)] to give 78 (190 mg, 86%) as a colorless amorphous solid. MW 519.56 IR (KBr) 3356, 2974, 1604, 1537, 1454, 1252 cmr; 'H-NMR (DMSO-d6) 8 2.24 (s, 3H), 2.26 (s, 3H), 3.60 (d, J= 3.7 Hz, 2H), 3.65-4.65 (min, 8H), 5.31-5.50 (mn, 1H), 6.92-7.18 (mn, 7H), 7.67-7.92 (inm, 10 4H), 8.22-8.32 (min, 2H); MS (FAB) m/z 520 (M+1); Anal. Calcd for C 29
H
3 0
FN
3 07-1.1H 2 0: C, 64.58; H 6.02; F,3.52; N,7.79. Found: C, 64.71; H, 5.90; F, 3.24; N, 7.51. Example 73 4-[1 -[4-[N '-(2-chlorophenyl)ureido]-3-methylphenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinylI] methoxybenzoic acid F N N N COOH 15 I H H Me 79 To a stirred mixture of tert-butyl 4-amino-3-methylphenylacetate (1.00 g, 4.52 mmol), 2 chlorophenyl isocyanate (0.55 ml, 4.52 mmol) in THF (10 ml) was added Et 3 N (0.13 ml, 0.90 mmol) at room temperature. After 6 h stirring, the reaction mixture was concentrated in vacuo. The residue was triturated by the addition of n-hexane, to give tert-butyl 4-[N'-(2-chlorophenyl) 20 ureido]-3-methylphenylacetate (1.57 g, 93%) as a pale yellow powder. mp 104-106 oC (dec.); 'H NMR (CDC13) 8 1.45 (s, 9H), 2.28 (s, 3H), 3.51 (s, 2H), 6.33 (br, 1H), 6.96 (t, J= 7.6 Hz, 1H), 7.08 (br, 1H1), 7.16-7.30 (min, 4H), 7.42 (min, 1H), 8.2 (d,J= 8.1 Hz, 1H). To a stirred solution of tert-butyl 4-[N'-(2-chlorophenyl)ureido]-3-methylphenylacetate (1.57 g, 4.19 mmol) in CH 2 Cl 2 (10 ml) was added TFA (6 ml) at room temperature. After 4 h 25 stirring, the mixture was concentrated in vacuo. The residue was triturated by the addition of water to give 4-[N'-(2-chlorophenyl)ureido]-3-methylphenylacetic acid (1.33 g, 100%) as a yellow powder. mp 243-245 oC (dec.); 'H-NMR (CDCl 3 ) 8 2.24 (s, 3H), 3.47 (s, 2H1), 6.99-7.08 (mn, 3H), 7.28 (t, J= 7.6 Hz, 1H), 7.44 (dt, J= 8.0, 2.4 Hz, 1H), 7.66 (dd, J= 8.3, 1.9 Hz, 1H), 8.13 (dd, J= 6.1, 1.7 Hz, 1H), 8.61 (d,J= 6.3 Hz, 2H); MS (ESI), m/z 319 (M +1), 321 (M+3); Anal. Calcd for 212 WO 01/00206 PCT/US00/18079
C
16
H,
5
CIN
2 0 3 -0.7TFA: C, 59.33; H, 4.65; Cl, 10.85; N, 8.57. Found: C, 59.23; H, 4.64; Cl, 10.90; N, 8.40. A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methylphenylacetic acid (252 mg, 0.79 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (200 mg, 0.79 mmol), EDC-HCI (227 5 mg, 1.20 mmol) and DMAP (147 mg, 1.20 mmol) in DMF (5 ml) was stirred at room temperature for 17 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was purified on TLC [CHCl3/acetone (10/1)], to give methyl 4 [1-[4-[N '-(2-chlorophenyl)ureido]-3-methylphenylacetyll-(4S)-fluoro-(2S)-pyrrolidinyl] 10 methoxybenzoate (390 mg, 89%) as a colorless amorphous solid. IR (KBr) 3340, 2951, 1712, 1624, 1604, 1533, 1438 cm-'; 'H-NMR (CDC 3 ) 8 1.92-2.05 (min, 3H), 2.07-2.63 (mn, 2H), 3.61 (d, 2H, J= 8.8 Hz), 3.70-4.15 (min, 5H), 4.25-4.67 (min, 2H), 5.26-5.44 (min, 1H), 6.84-8.19 (mn, 13H); MS (FAB) m/z 554 (M'+1), 556 (M'+3). To a stirred solution of methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methylphenyl 15 acetyl]-(4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (268 mg, 0.484 mmol) in THF (3.8 ml) was added 0.25 N NaOH (3.8 ml). After stirring at room temperature for 1 days, the mixture was acidified with 1 N HCI and extracted with CHCI 3 -MeOH (10/1). The combined extracts were dried over Na 2
SO
4 and concentrated in vacuo. The residue was purified on TLC [CHCl I/MeOH (10/1)] to give 79 (124 mg, 47%) as a colorless amorphous solid. MW 539.98 IR (KBr) 3346, 2976, 1709, 20 1685, 1604, 1533, 1439 cm-'; 'H-NMR (DMSO-d 6 ) 8 2.20 (s, 3H, one of isomers), 2.24 (s, 3H, one of isomers), 2.30 (min, 1H), 3.60 (s, 2H), 3.71-4.62 (min, 6H), 5.30-5.50 (mn, 1H), 7.01-7.09 (mn, 5H), 7.28 (t, J= 7.8 Hz, 1H), 7.44 (d, J= 8.1 Hz, 1H), 7.66 (t, J= 8.1 Hz, 1H), 7.87 (d, J= 7.1 Hz, 2H), 8.13 (d, J= 7.9 Hz, 1H), 8.62 (d, J= 6.1 Hz, 2H); MS (FAB) m/z 540 (M+1), 542 (M+3). For Na salt: Anal. Calcd for C 28
H
27 CIFN307-Na-0.5EtOH-1.5H 2 0: C, 56.91; H, 5.27; Cl, 5.79; F, 25 3.10; N, 6.87. Found: C, 56.60; H, 4.98; Cl, 5.88; F, 3.08; N, 6.52. Example 74 4-[ 1-[4-[N '-(2-bromophenyl)ureidoj-3-methylphenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinyl] methoxybenzoic acid F N- N \COOH rH H Me 80 30 to a stirred mixture of tert-butyl 4-amino-3-methylphenylacetate (780 mg, 3.30 mmol), 2-bromo 213 WO 01/00206 PCT/US00/18079 phenyl isocyanate (0.41 ml, 3.30 mmol) in THF (7 ml) was added Et 3 N (0.092 ml, 0.66 mmol) at room temperature. After 3 h stirring, the reaction mixture was concentrated in vacuo. The residue was triturated by the addition of n-hexane, to give tert-butyl 4-[N'-(2-bromophenyl) ureido]-3 methylphenylacetate (1.57 g, 93%) as a pale yellow powder. mp 138-145 oC (dec.); 'H-NMR 5 (CDCl 3 ) 8 1.44 (s, 9H), 2.33 (s, 3H1), 3.51 (s, 2H), 6.90 (dt, J= 9.0, 1.4 Hz, 1H), 6.98 (br, 1H), 7.18-7.31 (m, 4H), 7.39 (dd, J= 8.1, 2.9 Hz, 1H), 7.44 (d, J= 8.0 Hz, 1H), 8.22 (d, J= 8.3 Hz, 2H); Anal. Calcd for C 20
H
22 BrN 2 03-0.2H 2 0: C,56.80; H,5.58; N,6.62. Found: C,56.85; H,5.42; N, 6.62. 10 To a stirred solution of tert-butyl 4-[N'-(2-bromophenyl)ureido]-3-methylphenylacetate (1.27 g, 3.03 mmol) in CH 2
CI
2 (10 ml) was added TFA (5 ml) at room temperature. After 1 h stirring, the mixture was concentrated in vacuo. The residue was triturated by the addition of water, to give 4-[N'-(2-bromophenyl)ureido]-3-methylphenylacetic acid (1.05 g, 95%) as a pale yellow powder. mp 245-248 oC (dec.); 'H-NMR (CDC13) 8 2.24 (s, 3H), 3.48 (s, 2H), 6.96 (dt, J= 15 7.3, 1.5 Hz, 1H), 7.02 (d,J= 8.3 Hz, 1H), 7.07 (s, 1H11), 7.32 (t,J= 8.1 Hz, 1H), 7.59-7.66 (m, 2H), 8.44 (s, 1H1), 8.62 (s, 1H1); MS (ESI), m/z 363 (M*+1), 365 (M+3); Anal. Calcd for
C
16 HsBrN 2 03-0.7H 2 0: C, 51.13; H, 4.40; Br, 21.26; N, 7.45. Found: C, 50.84; H, 4.62; Br, 21.72; N, 7.18. A mixture of 4-[N'-(2-bromophenyl)ureido]-3-methylphenylacetic acid (287 mg, 0.79 mmol), 20 methyl 4-[(4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (200 mg, 0.79 mmol), EDC-HCl (228 mg, 1.20 mmol), HOBT (160 mg, 1.19 mmol) and Et 3 N (0.55 ml, 3.95 mmol) in DMF (5 ml) was stirred at room temperature for 4 days. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was purified on TLC [CHCI/acetone (10/1)], 25 to give methyl 4-[(2S,4S)-1-[4-[N'-(2-chlorophenyl)ureido]-3-methylphenylacetyl]-4-fluoro-2 pyrrolidinyl]methoxybenzoate (440 mg, 93%) as a colorless amorphous solid. 'H-NMR (CDCl 3 ) 8 1.90 andl.97 (eachs, 3H11, amide isomers), 2.05-2.62 (m, 2H), 3.58 (d, J= 8.1 Hz, 1H), 3.77 (m, 1H), 3.86 and 3.89 (eachs, 3H, amide isomers), 3.92-4.64 (m, 5H), 5.24-5.42 (m, 1H1), 6.83-7.23 (m, 6H), 7.41-7.62 (m, 4H11), 7.86-8.09 (m, 3H); MS (ESI), nm/z 598 (M +1), 600 (M+3). 30 To a stirred solution of methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methylphenyl acetyl]-(4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (440 mg, 0.74 mmol) in THF (6.0 ml), 0.25 214 WO 01/00206 PCT/US00/18079 N NaOH (6.0 ml) was added. After stirring at room temperature for 1 days, the mixture was acidified with 1 N HCI and extracted with CHCl 3 -MeOH (10/1). The combined extracts were dried over Na 2
SO
4 and concentrated in vacuo. The residue was purified on TLC [CHClMeOH (10/1)] to give 80 (229 mg, 53%) as a colorless amorphous solid. MW 584.44 IR (KBr) 3325, 2972, 1709, 5 1604, 1529, 1252 cm'; 'H-NMR (DMSO-d) 8 2.25 (s, 3H), 2.31 (min, 1H), 3.17 (s, 1H), 3.60 (d, J = 4.7 Hz, 2H), 3.83-4.67 (min, 5H), 5.31-5.51 (mn, 1H), 6.97 (t, J= 7.3 Hz, 1H), 7.02-7.09 (min, 5H), 7.33 (t, J= 8.0 Hz, 1H), 7.61 (d, J= 7.8 Hz, 1H), 7.64 (d, J= 8.3 Hz, 1H), 7.87 (d, J = 8.3 Hz, 2H), 7.90 (d, J= 8.8 Hz, 1H), 8.44-8.65 (min, 2H); MS (ESI), m/z 584 (M'+1), 586 (M+3); Anal. Calcd for C 28
H
27 BrFN 3 O0 7 0.4H 2 0: C, 56.84; H, 4.74; Br, 13.51; F, 3.21; N, 7.10. Found: C, 10 56.91; H, 4.93; Br, 13.23; F, 3.15; N, 6.88. For Na salt of 80 : Anal. Calcd for
C
28
H
27 BrFN307-ONa-1.8H 2 0: C, 52.64; H, 4.67; Br, 12.51; F, 2.97; N, 6.58. Found: C, 53.04; H, 4.67; Br, 12.95; F, 3.28; N, 6.11. Example 75 4-[ 1-[4-[N'-(2-chlorophenyl)ureido]-3-methylphenylacetyl]-(2S)-pyrrolidinyl]jmethoxybenzoic acid 1N N O COOH 15 IH H e 81 To a stirred solution of methyl 4-(1-tert-butoxycarbonyl-(2S)-pyrrolidinyl)methoxybenzoate (2.0 g, 5.9 mmol) in EtOH (10.0 ml) was added concd HCI (3.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 4.0 hr. The mixture was concentrated in vacuo. The resulting solid was collected and washed with EtOH-Et20 to give methyl 4-(2S-pyrrolidinyl)methoxybenzoate HCI 20 salt (1.4 g, 87%) as a white crystalline solid. 'H-NMR (CDCl 3 ) 8 1.90-2.25 (mn, 4H), 3.25-3.45 (min, 2H11), 3.88 (s, 3H), 3.90-4.00 (min, 1H), 4.25-4.45 (mn, 2H), 6.96 (d, J = 8.5 Hz, 2H), 7.95 (d, J 8.5 Hz, 2H11). To a stirred solution of methyl 4-[(2S)-pyrrolidinyl]methoxybenzoate HC1 salt (135 mg, 0.5 mmol), 4-[N'-(2-chlorophenyl)uredio]-3-methylphenylacetic acid (159 mg, 0.5 mmol), HOBt (68 mg, 0.5 25 mmol), and triethylamine (278 ml, 2.0 mmol) in THF (5.0 ml) and MeCN (5.0 ml) was added EDC-HCI (144 mg, 0.75 mmol) at 0 'C. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc(1:2 ,v/v) as 30 eluent to give methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methylphenylacetyl]-2-pyrrolidinyl] methoxybenzoate (220 mg, 82%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.91 and 1.97 (each s, 215 WO 01/00206 PCT/US00/18079 total 3H), 2.00-2.20 (min, 4H), 3.55-3.65 (min, 4H), 3.87 and 3.89(each s, total 3H),4.10-4.20 (m,2H), 4.51 (min, 1H), 6.86-7.04 (min, 6H), 7.20-7.53 (min, 4H), 7.89-8.01 (mn, 2H), 8.22 (d, J= 8.3 Hz, 1H). To a stirred solution of methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methylphenylacetyl]-2S pyrrolidinyl]methoxybenzoate (220 mg, 0.41 mmol) in THF (8.0 ml) and MeOH (4.0 ml) was 5 added IN NaOH (0.8ml, 0.8 mmol). The mixture was stirred at 70 'C for 24 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with iN HC1. The resulting solid was collected, washed with water, and dried in vacuo to give 81 (220 mg, quant) as a white crystalline solid. MW 521.99 mp 122-124 'C; IR (KBr) 3340, 1710, 1685, 1604, 1533, 1511, 1438 cm'; tH-NMR (DMSO-d 6 ) 8 1.81-2.11 (min, 4H), 2.18 and 2.20 (each s, total 3H), 3.45-3.80 10 (min, 4H), 3.95-4.05 (min, 1H), 4.12-4.20 (min, 1H), 4.21-4.31 (min, 1H), 6.99-7.06 (mn, 5H), 7.26-7.30 (min, 1H), 7.44 (d, J= 7.8 Hz, 1H1), 7.62-7.64 (min, 1H), 7.85-7.90 (mn, 2H), 8.13 (d, J= 6.8 Hz, 1H), 8.60-8.62 (min, 2H); MS (FAB) m/z 522 (M'+1); Anal. calcd for C 28
H
2 gN 3 O0Cl-0.2H 2 0: C, 63.99; H, 5.45; N, 7.99. Found: C, 63.90; H, 5.40; N, 7.72. Example 76 15 4-[ 1-[4-[N'-(2-bromophenyl)ureido]-3-methylphenylacetyll-(2S)-pyrrolidinyl]methoxybenzoic acid N N \ COOH rH H e 82 To a stirred solution of methyl [(2S)-pyrrolidinyl]methoxybenzoate HCI salt (135 mg, 0.5 mmol), 4-[N'-(2-bromophenyl)uredio]-3-methylphenylacetic acid (181 mg, 0.5 mmol), HOBt (68 mg, 0.5 mmol), and triethylamine (278 ml, 2.0 mmol) in THF (5.0 ml) and MeCN (5.0 ml) was added 20 EDC-HCI (144 mg, 0.75 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , then dried over Na 2 SO, , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (1/2, v/v) as eluent to give methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methylphenylacetyl]-(2S) 25 pyrrolidinyl]methoxybenzoate (290 mg, quant) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.95 and 2.01 (each s, total 3H), 2.00-2.20 (min, 4H), 3.50-3.65 (min, 4H), 3.87 and 3.89 (each s, total 3H), 4.10-4.20 (min, 2H), 4.50 (min, 1H), 6.85-7.06 (min, 6H), 7.24-7.28 (min, 1H), 7.40-7.44 (min, 3H), 7.89 8.16 (min, 2H), 8.17-8.18 (min, 1H). To a stirred solution of methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methylphenylacetyl]-2S 30 pyrrolidinyl]methoxybenzoate (290 mg, 0.5 mmol) in THF (8.0 ml) and MeOH (4.0 ml) was added 216 WO 01/00206 PCT/US00/18079 IN NaOH (1.0 ml, 1.0 mmol). The mixture was stirred at 70 oC for 24 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HCI. The resulting solid was collected, washed with water, and dried in vacuo to give 82 (240 mg, 85%) as a white crystalline solid. MW 566.44 mp 125-130 oC; IR (KBr) 3340, 1604, 1529, 1434 cm-'; 'H-NMR 5 (DMSO-d 6 ) 5 1.80-2.10 (m, 4H), 2.18 and 2.20 (each s, total 3H), 3.45-3.80 (m, 4H), 3.95-4.05 (m, 1H), 4.15-4.20 (m, 1H), 4.25-4.30 (m, 1H), 6.94-7.06 (m, 5H), 7.30-7.34 (m, 1H), 7.59-7.62 (m, 2H), 7.85-7.90 (m, 2H), 8.01 (d,J = 8.1 Hz, 1H), 8.44 (s, 1H), 8.62 (s, liH); MS (FAB) m/z 566 (M); Anal. calcd for C 28
H
28
N
3 OsBr-0.5H 2 0: C, 58.44; H, 5.08; N, 7.30. Found: C, 58.57; H, 4.99; N, 7.18. 10 Example 77 4-[ 1 -[3-methyl-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoic acid N~ NN Ol c -COOH Me H H Me 83 A mixture of 3-methyl-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (438 mg, 1.47 mmol), methyl 4-[(2S)-pyrrolidinylmethoxy]benzoate (420 mg, 1.79 mmol), EDCHCI (410 mg, 2.14 15 mmol), HOBt (228 mg, 1.69 mmol) and Et 3 N (240 ml, 1.72 mmol) in DMF (5 ml) was stirred at room temperature overnight. The mixture was diluted with H 2 0 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (50:1 to 25:1, v/v) as eluent to give methyl 4-[ 1-[3-methyl-4-[N '-(2-methylphenyl)ureidojphenylacetylJ]-(2S)-pyrrolidinylmethoxy] 20 benzoate (760 mg, quant.) as a white foam. 'H-NMR (CDCl 3 ) 8 1.89 (s, 3 H), 1.94-2.14 (m, 4 H), 2.16 (s, 3 H), 3.50-3.69 (m, 4 H), 3.87 (s, 3 H), 4.09-4.17 (m, 2 H), 4.42-4.45 (m, 1 H), 6.85-7.02 (m, 6 H), 7.10-7.16 (m, 3 H), 7.51-7.53 (m, 1 H), 7.62-7.64 (m, 1 H), 7.91-7.94 (m, 2 H); MS (FAB) m/z 516 (M+1). To a stirred solution of methyl 4-[1-[3-methyl-4-[N'-(2-methylphenyl)ureido] 25 phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (420 mg, 0.71 mmol) in THF (7 ml) was added 0.5 N NaOH (7 ml) and the reaction mixture was heated under reflux for 2 hr. After cooled to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was collected under a reduced pressure. The crude solid was purified by recrystallization from CHCl 3 IPE to give 83 (526 mg, 69%) as a white crystalline powder. MW 501.57 mp 191-193 0 C; 'H 30 NMR (DMSO-d 6 ) 8 1.87-2.10 (m, 4 H), 2.20 (s, 3 H), 2.26 (s, 3 H), 3.44-3.79 (series of m, total 4 H), 3.99-4.45 (series of m, total 3 H), 6.91-7.17 (series of m, total 7 H), 7.66-7.68 (m, 1 H), 7.80 217 WO 01/00206 PCT/US00/18079 7.90 (m, 3 H), 8.19-8.21 (m, 2 H), 12.62 (br s, 1 H); MS (FAB) m/z 502 (M'+l);Anal. Calcd for
C
2~
H
31
N
3 0 5 1/4H 2 0: C, 68.83; H, 6.27; N, 8.30. Found: C, 68.81; H, 6.17; N, 8.23. Example 78 4-[(4S)-fluoro- 1-[4-[N'-(2-methoxyphenyl)ureido]-3-methylphenylacetyl]-(2S)-pyrrolidinyl 5 methoxy]benzoic acid N N o- COOH H H Me Me 84 To a stirred solution of tert-butyl 4-amino-3-methylphenylacetate (1.36 g, 6.15 mmol) and triethylamine (170 ml, 1.23 mmol) in THF (30 ml) was added 2-methoxyphenyl isocyanate (820 ml, 6.15 mmol), and the resulting mixture was stirred for 27 h. The mixture was concentrated to a 10 small volume in vacuo, and hexane was added to the residue to give precipitate, which was collected by filtration to give tert-butyl 4-[N'-(2-methoxyphenyl)ureido]-3-methylphenylacetate (1.74 g, 76%) as a white crystalline material. mp 157-158 oC; 1 H-NMR (CDC13) 8 1.46 (s, 9 H), 2.30 (s, 3 H), 3.50 (s, 2 H), 3.76 (s, 3 H), 6.43 (s, 1 H), 6.83 (br d, J= 8.4 Hz, 1 H), 6.95 (br d, J 8.0 Hz, 1 H), 6.98-6.99 (m, 2 H), 7.13 (brs, 1 H), 7.23 (br s, 1 H), 7.48 (d, J= 8.8 Hz, 1 H), 8.14 15 (d, J= 8.4 Hz, 1 H); MS (ESI) m/z 371 (M +H). To a stirred solution of tert-butyl 4-[N'-(2-methoxyphenyl)ureido]-3-methylphenylacetate (1.32 g, 3.56 mmol) in CH 2 C1 2 (15 ml) was added trifluoroacetic acid (10 ml), and the resulting mixture was heated under reflux for 30 min. The mixture was concentrated in vacuo and added water to give precipitate which was collected by filtration. The crude solid was recrystallized from 20 EtOH/hexane to give 4-[N'-(2-methoxyphenyl)ureido]-3-methylphenylacetic acid (932 mg, 83%) as a white powder. mp 260-264 'C; 'H-NMR (CD 3 OD) 8 2.30 (s, 3 H), 3.55 (s, 2 H), 4.87 (s, 3 H), 6.87-6.92 (m, 2 H), 6.97-6.99 (m, 2 H), 7.10-7.24 (m, 2 H), 7.53-7.58 (m, 1 H), 8.04 (d, J= 7.2 Hz, 1 H); MS (ESI) m/z 314 (M). To a stirred solution of 4-[N'-(2-methoxyphenyl)ureido]-3-methylphenylacetic acid (336 mg, 1.07 25 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (271 mg, 1.07 mmol) and N,N dimethylaminopyridine (130 mg, 1.07 mmol) in DMF (10 ml) was added EDCHCI (226 mg, 1.18 mmol) at rt, and the resulting mixture was stirred for 20 h. The mixture was poured into 1N-HCl aq. and extracted with EtOAc. The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 218 WO 01/00206 PCT/US00/18079 MeOH (10 : 1) as eluent to give methyl 4-[(4S)-fluoro-1-[4-[N'-(2-methoxyphenyl)ureido]-3 methylphenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (583 mg, 99%) as a colorless amorphous solid. 'H-NMR (CDCl 3 ) mixture of rotamars, 8 2.05 and 2.12 (s, total 3 H), 2.05-2.61 (m, 2 H), 3.55-4.73 (series of m, 13 H), 4.51-4.66 (m, 2 H), 5.26-5.40 (m, 1 H), 6.72-7.01 (series of m, 8 H), 5 7.38-8.13 (series of m, 3 H); MS (ESI) m/z 550 (M+H). To a stirred solution of methyl 4-[(4S)-fluoro-1-[4-[N'-(2-methoxyphenyl)ureido]-3 methylphenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (557 mg, 1.01 mmol) in MeOH-THF (1 : 1, 10 ml) was added 1.0N-NaOH aq. (4.05 ml, 4.05 mmol) at rt, and the resulting mixture was heated at 60oC for 2 h. The mixture was poured into 1N-HCI aq. and extracted with EtOAc. The 10 organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (10 : 1) as eluent to give 84 (492 mg, 91%) as a colorless amorphous solid. MW 535.56 1 H-NMR (CD 3 OD), mixture of rotamars, 8 2.96 (s, 3 H), 2.11-2.45 (m, 2 H), 3.64-4.15 (series of m, 5 H), 3.91 (s, 3 H), 4.41-4.45 (m, 1 H), 4.52-4.61 (m, 1 H), 5.25-5.38 (m, 1 H), 6.84-7.10 (series of m, 7 H), 7.54-7.58 (m, 1 H), 15 7.93 (d, J= 8.8 Hz, 2 H), 8.02 (d, J= 8.8 Hz, 2 H); MS (ESI) m/z 536 (M +H), 538 (M +Na). Example 79 4-[(4S)-fluoro-1 -[4-[N' -(2-methoxyphenyl)ureido]phenylacetyl]-(2S)-pyrro-lidinylmethoxy] benzoic acid NN N Nn 0-oCOOH H H Me 85 20 To a stirred solution of ethyl 4-amino-3-methylphenylacetate (1.32 g, 7.37 mmol) and triethylamine (205 ml, 1.47 mmol) in THF (20 ml) was added 2-methoxyphenyl isocyanate (980 ml, 7.37 mmol), and the resulting mixture was stirred for 23 h. The mixture was concentrated to a small volume in vacuo and hexane was added to the residue to give precipitate, which was collected to give ethyl 4-[N'-(2-methoxyphenyl)ureido]phenylacetate (2.44 g, quant.) as a white 25 crystalline material. mp 107-109 oC; 'H-NMR (CDCl 3 ) 8 1.26 (t, J= 7.1 Hz, 3 H), 3.56 (s, 3 H), 3.79 (s, 3 H), 4.15 (q, J= 7.1 Hz, 2 H), 6.82-6.85 (m, 1 H), 6.91-7.00 (m, 2 H), 7.08 (s, 1 H), 7.17 (d, J= 8.5 Hz, 2 H), 7.27 (d, J = 8.6 Hz, 2 H), 7.33 (s, 1 H), 8.07-8.10 (m, 1 H); MS (ESI) m/z 329 (M*+H). To a stirred solution of ethyl 4-[N'-(2-methoxyphenyl)ureido]phenylacetate (2.22 g, 6.78 219 WO 01/00206 PCT/US00/18079 mmol) in MeOH (30 ml) was added 1.0 M-NaOH aq. (10.2 ml, 10.2 mmol), and the resulting mixture was stirred overnight. IN-HCI (aq.) was added and the mixture was concentrated in vacuo. Water was added to the residue to give precipitate, which was collected by filtration. The crude solid was recrystallized from EtOH/hexane to give 4-[N'-(2-methoxyphenyl)ureido] 5 phenylacetic acid as a white powder (1.87 g, 92%). mp 165-168 oC; 'H-NMR (CD 3 OD) 8 2.30 (s, 3 H), 3.55 (s, 2 H), 4.87 (s, 3 H), 6.87-6.92 (m, 2 H), 6.97-6.99 (m, 2 H), 7.10-7.24 (m, 2 H), 7.53 7.58 (m, 1 H), 8.04 (d, J = 7.2 Hz, 1 H); MS (ESI) nm/z 300 (M). To a stirred solution of 4-[N'-(2-methoxyphenyl)ureido]phenylacetic acid (353 mg, 1.18 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (298 mg, 1.18 mmol) and NA,N 10 dimethylaminopyridine (144 mg, 1.18 mmol) in DMF (10 ml) was added EDCHCI (226 mg, 1.18 mmol) at rt, and the resulting mixture was stirred for 22 h. The mixture was poured into 1N-HCI aq. and extracted with EtOAc. The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCI 3 MeOH (10 : 1) to give methyl 4-[(4S)-fluorol-[4-[N'-(2-methoxyphenyl)ureido]phenylacetyl]-(2S) 15 pyrrolidinylmethoxy]benzoate (594 mg, 94%) as a colorless amorphous solid. 'H-NMR (CDCl 3 ) mixture of rotamars, 8 2.05-2.58 (series of m, 2 H), 3.55-4.25 (series of m, 5 H), 3.77 (s, 3 H), 3.87-3.90 (m, 3 H), 4.50-4.63 (m, 2 H), 5.23-5.37 (m, 1 H), 6.81-6.84 (m, 1 H), 6.91-6.99 (m, 4 H), 7.09-7.12 (m, 2 H), 7.18-7.26 (m, 2 H), 7.45-7.53 (m, 2 H), 7.91-8.03 (m, 2 H), 8.10-8.12 (m, 1 H); MS (ESI) m/z 536 (M +H). 20 To a stirred solution of methyl 4-[(4S)-fluoro-1-[4-[N'-(2-methoxyphenyl)ureido] phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (568 mg, 1.06 mmol) in MeOH-THF (1 : 1, 10 ml) was added 1.ON-NaOH aq. (4.24 ml, 4.24 mmol) at rt, and the resulting mixture was heated at 60 0 C for 1 h. The mixture was poured into 1N-HCI aq. and extracted with EtOAc. The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The 25 residue was chromatographed on silica gel with CHCl 3 -MeOH (10 : 1) as eluent to give 85 (516 mg, 93%) as a colorless amorphous solid. MW 521.54 1 H-NMR (CD 3 OD), mixture of rotamars, 8 2.12-2.46 (m, 2 H), 3.65-4.19 (series of m, 5 H), 3.88 (s, 3 H), 4.42-4.45 (m, 1 H), 4.52-4.62 (m, 1 H), 5.24-5.39 (m, 1 H), 6.85-6.91 (m, 1 H), 6.94-7.03 (series of m, 4 H), 7.14-7.19 (m, 2 H), 7.35-7.40 (m, 2 H), 7.92-7.96 (m, 2 H), 8.02-8.04 (m, 1 H); MS (ESI) m/z 521 (M+H), 544 30 (M++Na+). Example 80 4-[(4S)-fluoro-1-[4-[N'-(2-methoxyphenyl)ureido]-3-methoxyphenylacetyl]-(2S)-pyrrolidiny 220 WO 01/00206 PCT/US00/18079 methoxy]benzoic acid F N NQO Oi COOH H H Cl 86 To a stirred solution of tert-butyl 4-amino-3-methoxylphenylacetate (1.41 g, 5.94 mmol) and triethylamine (165 ml, 1.19 mmol) in THF (20 ml) was added 2-methoxyphenyl isocyanate (790 5 ml, 5.94 mmol), and the resulting mixture was stirred for 4 days. The mixture was concentrated to a small volume in vacuo, and hexane was added to the residue to give precipitate, which was collected to give tert-butyl 4-[N'-(2-methoxyphenyl)ureido]-3-methoxylphenylacetate (2.06 g, 90%) as a white crystalline material. mp 132-134 oC; 'H-NMR (CDCl 3 ) 8 1.46 (s, 9 H), 3.50 (s, 2 H), 3.87 (s, 3 H), 3.88 (s, 3 H), 6.84 (s, 1 H), 6.87-6.90 (m, 2 H), 6.98-7.03 (m, 2 H), 7.12 (s, 1 H), 10 7.16 (s, 1 H), 8.06 (d, J= 8.4 Hz, 1 H), 8.13 (dd, J= 7.2, 2.0 Hz, 1 H); MS (ESI) m/z 387 (MW+H). To a stirred solution of tert-butyl 4-[N'-(2-methoxyphenyl)ureido]-3-methoxylphenyl acetate (2.01 g, 5.20 mmol) in CH 2 Cl 2 (15 ml) was added trifluoroacetic acid (10 ml), and the resulting mixture was heated under reflux for 30 min. The mixture was concentrated in vacuo. Water was added to the residue to give precipitate, which was collected by filtration. The crude 15 solid was recrystallized from EtOH/hexane to give 4-[N'-(2-methoxyphenyl)ureido]-3-methoxy phenylacetic acid as white powder (1.40 g, 82%). mp 182-185 oC; 'H-NMR (CD30D) 8 3.55 (s, 2 H), 3.88 (s, 3 H), 3.89 (s, 3 H), 6.80-6.99 (m, 5 H), 7.94 (d, J = 8.4 Hz, 1 H), 8.00 (d, J= 7.2 Hz, 1 H); MS (ESI) m/z 330 (M). To a stirred solution of 4-[N'-(2-methoxyphenyl)ureido]-3-methoxyphenylacetic acid (353 mg, 20 1.07 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (271 mg, 1.07 mmol) and N,N-dimethylaminopyridine (131 mg, 1.07 mmol) in DMF (10 ml) was added EDCHCI (224 mg, 1.18 mmol) at rt, and the resulting mixture was stirred for 14 h. The mixture was poured into 1N HCI aq. and extracted with EtOAc. The organic layer was washed with brine, drying over anhydrous Na 2 SO4, then concentrated in vacuo. The residue was chromatographed on silica gel 25 with CHCl 3 -MeOH (10 : 1) as eluent to give methyl 4-[(4S)-fluoro-1-[4-[N'-(2-methoxyphenyl) ureido]-3-methoxyphenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (372 mg, 61%) as a colorless amorphous solid. 1 H-NMR (CDCl 3 ), mixture of rotamars, 8 2.04-2.57 (series of m, 2 H), 3.58-4.18 (series of m, 5 H), 3.79 and 3.83 (s, total 3 H), 3.86 (s, 3 H), 3.87 (s, 3 H), 4.51-4.63 (m, 2 H), 5.22-5.36 (m, 1 H), 6.80-6.89 (m, 3 H), 6.94-7.03 (m, 4 H), 7.15-7.25 (m, 2 H), 7.94-8.01 (m, 2 221 WO 01/00206 PCT/US00/18079 H), 8.04-8.11 (m, 2 H); MS (ESI) m/z 566 (M++H). To a stirred solution of methyl 4-[(4S)-fluoro-1-[4-[N'-(2-methoxyphenyl)ureido]-3-methoxy phenyl-acetyl]-(2S)-pyrrolidinylmethoxy]benzoate (356 mg, 0.63 mmol) in MeOH-THF (1 : 1, 10 ml) was added 1.0N-NaOH aq. (1.88 ml, 1.88 mmol) at rt, and the resulting mixture was heated at 5 60 0 C for 2 h. The mixture was poured into 1N-HCI aq. and extracted with EtOAc. The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (10 : 1) as eluent to give 86 (335 mg, 97%) as a colorless amorphous solid. MW 551.56 'H-NMR (CD 3 OD), mixture of rotamars, 8 2.14-2.48 (m, 2 H), 3.69-4.20 (series of m, 5 H), 3.88 (s, 3 H), 3.89 (s, 3 H), 4.46-4.57 (m, 2 H), 10 5.27-5.41 (m, 1 H), 6.79-7.04 (m, 7 H), 7.90-8.02 (m, 4 H); MS (ESI) m/z 552 (M++H). Example 81 4-[1 -[4-[N '-(2,6-dichlorophenyl)ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinyl]methoxy benzoic acid F NN N H'I \ COOH 87 15 To a mixture of ethyl 4-aminophenylacetate (1.62 g, 9.04 mmol) and 2,6-dichlorophenyl isocyanate (1.70 g, 9.04 mmol) in THF (40 ml) was added Et 3 N (0.25 ml, 1.81 mmol) at room temperature. After 2 h stirring, the reaction mixture was concentrated in vacuo. The residue was triturated by the addition of n-hexane, to give ethyl 4-[N'-(2,6-dichlorophenyl)ureido] phenyl acetate (3.19 g, 96%) as a colorless powder. mp 168-170 oC (dec.); 'H-NMR (CDCl 3 ) 8 1.25 (t, J= 20 7.1 Hz, 3H), 3.56 (s, 2H), 4.14 (q, J= 7.1 Hz, 2H), 6.50 (br, 1H), 6.67 (br, 1H), 7.12-7.52 (m, 7H). To a stirred solution of ethyl 4-[N'-(2,6-dichlorophenyl)ureido]phenylacetate (3.19 g, 8.69 mmol) in THF (70 ml), 0.25 N NaOH (70 ml) was added. After stirring at room temperature for 17 h, the solvent was concentrated in vacuo. The residue was triturated by the addition of water, to give 4 [N'-(2,6-dichlorophenyl)ureido]phenylacetic acid (2.44 g, 82%) as colorless powder. mp 262-263 25 oC (dec.); 'H-NMR(DMSO-d 6 ) 8 3.48 (s, 2H), 7.14 (d,J= 8.3 Hz, 2H), 7.31 (t,J= 8.3 Hz, 1H), 7.37 (d, J= 8.3 Hz, 2H), 7.52 (d, J= 8.0 Hz, 2H), 8.18 (s, 1H), 8.90 (s, 1H), 12.22 (br, 1H); MS (ESI) m/z 339 (M++I), 341 (MW+3), 343 (MW+5). A mixture of 4-[N'-(2,6-dichlorophenyl)ureido]phenylacetic acid (268 mg, 0.79 mmol), methyl 4 222 WO 01/00206 PCT/US00/18079 [(2S,4S)-4-fluoro-2-pyrrolidinyl]methoxybenzoate (200 mg, 0.79 mmol), EDC.HCI (227 mg, 1.19 mmol), HOBT (161 mg, 1.19 mmol) and Et 3 N (0.55 ml, 3.95 mmol) in DMF (4 ml) was stirred at room temperature for 18 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the 5 extracts were concentrated in vacuo. The residue was purified on TLC [CHCI/MeOH (10/1)], to give tmethyl 4-[1-[4-[N '-(2,6-dichlorophenyl)ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinyl] methoxybenzoate (465 mg, 100%) as a colorless amorphous solid. 1 H-NMR (CDCI 3 ) 8 2.05-2.57 (min, 2H), 3.60 (d, 2H, J= 3.4 Hz), 3.64-3.84 (min, 2H), 3.88 and 3.89 (each s, 3H, amide isomers), 3.92-4.63 (min, 3H), 5.22-5.38 (min, 1H), 6.87 and 6.89 (each d, each J= 7.9 Hz, 2H, amide isomers), 10 7.01-7.17 (min, 6H), 7.28 (min, 2H), 7.36 (br, 1H11), 7.92 (d,J= 8.8 Hz, 1H), 7.79 (d, J= 8.8 Hz, 1H); MS (ESI) m/z 574 (M'+1), 576 (M*+3), 578 (MW+5). To a solution of methyl 4-[(2S,4S)- 1 -[4-[N'-(2,6-dichlorophenyl)ureido]phenylacetyl]-4-fluoro-2 pyrrolidinyl]methoxybenzoate (465 mg, 0.809 mmol) in THF (40 ml), 0.25 N NaOH (40 ml) was added. After stirring at room temperature for 11 h, the mixture was acidified with 1 N HCI and 15 extracted with CHCl 3 -MeOH (10/1). The combined extracts were dried over Na 2
SO
4 and concentrated in vacuo. The residue was purified on TLC [CHCl3/MeOH (10/1)], to give 87 (340 mg, 75%) as a colorless powder. MW 560.40 mp 168-172 oC (dec.); IR (KBr) 3340, 1711, 1685, 1604, 1240, 773 cm'; 'H-NMR (DMSO-d 6 ) 8 2.22-2.30 (min, 2H), 3.61 (d, J= 7.4 Hz, 2H), 3.70 4.75 (min, 6H), 5.30-5.49 (min, 1H), 7.02-7.18 (min, 5H), 7.28-7.41 (min, 4H), 7.52 (dd, J= 8.0, 2.9 Hz, 20 2H), 7.86 (min, 2H), 8.29 (br, 1H), 9.01 (br, 1H), 12.66 (br, 1H); MS (ESI) m/z 560 (MW+1), 562 (M+3), 564 (M+5); Anal. Calcd for C 27
H
2 4 C1 2
FN
3 0 5 -0.5H 2 0: C, 56.95; H, 4.43; Cl, 12.45; F, 3.34; N, 7.38. Found: C, 57.04; H, 4.34; Cl, 12.98; F, 3.27; N, 7.21. Example 82 4-[1 -[4-[N'-(2,6-dichlorophenyl)ureido]-3-methoxyphenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinyl] 25 methoxybenzoic acid F CI N N O 0- -OOH H H Me oK9co 88 To a mixture of tert-butyl 4-amino-3-methoxyphenylacetate (2.15 g, 9.04 mmol), 2,6 dichlorophenyl isocyanate (1.70 g, 9.04 mmol) in THF (40 ml) was added Et 3 N (0.25 ml, 9.04 imol) at room temperature. After 18 h stirring, the reaction mixture was concentrated in vacuo. 30 The residue was triturated by the addition of n-hexane, to give tert-butyl 4-[N'-(2,6 223 WO 01/00206 PCT/US00/18079 dichlorophenyl)ureido]-3-methoxyphenylacetate (2.27 g, 59%) as a colorless powder. mp 177-181 oC (dec.); 'H-NMR (CDCl 3 ) 8 1.43 (s, 9H-I), 3.74 (s, 2H), 3.83 (s, 3H), 6.34 (s, 1H), 6.81 (s, 1H11), 6.84 (d, J= 8.3 Hz, 1H), 7.06 (br, 1H11), 7.27 (t, J = 8.1 Hz, 1H), 7.39 (d, J = 8.1 Hz, 2H), 8.04 (d, J = 8.3 Hz, 1H). 5 To a stirred solution of tert-butyl 4-[N'-(2,6-dichlorophenyl)ureido]-3-methoxyphenylacetate (2.27 g, 5.34 mmol) in CH 2 C1 2 (50 ml) was added TFA (20 ml) at room temperature. After 2 h stirring, the mixture was concentrated in vacuo. The residue was triturated by the addition of water, to give 4-[N'-(2,6-dichlorophenyl)ureido]-3-methoxyphenylacetic acid (1.50 g, 76%) as a colorless powder. mp 246-249 oC (dec.); 'H-NMR (DMSO-d 6 ) 6 3.49 (s, 2H), 3.88 (s, 3H), 6.75 (d, J = 8.3 10 Hz, 1H), 6.93 (s, 1H), 7.30 (t, J= 7.8 Hz, 1H), 7.52 (d, J= 8.0 Hz, 2H), 7.97 (d, J= 8.0 Hz, 1H), 8.40 (s, 1H), 8.86 (s, 1H), 12.23 (br, 1H); MS (ESI) m/z 369 (M++1), 371 (M+3), 373 (M+5). A mixture of 4-[N'-(2,6-dichlorophenyl)ureido]-3-methoxyphenylacetic acid (288 mg, 0.78 mmol), methyl 4-[(2S,4S)-4-fluoro-2-pyrrolidinyl]methoxybenzoate (200 mg, 0.79 nmmol), EDC.HC1 (227 mg, 1.19 mmol), HOBT (161 mg, 1.19 mmol) and Et 3 N (0.55 ml, 3.95 mmol) in DMF (4 ml) was 15 stirred at room temperature for 18 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHCl 3 /MeOH(40/1)] to give methyl 4-[1-[4-[N'-(2,6-dichlorophenyl)ureido]-3-methoxyphenyl acetyl]-(4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (530 mg, 100%) as a colorless amorphous 20 solid. 'H-NMR (CDCl 3 ) 8 2.03-2.62 (min, 2H), 3.61 (d, 2H, J= 4.7 Hz), 3.62-3.66 (mn, 2H), 3.73 and 3.77 (each s, 3H, amide isomers), 3.78-3.85 (min, 1H), 3.87 and 3.88 (each s, 3H, amide isomers), 3.95-4.63 (min, 4H), 5.22-5.36 (min, 1H), 6.82 (s, 1H), 6.88 (d, J= 8.8 Hz, 1H), 6.95 (d, J= 8.8 Hz, 2H), 7.14-7.25 (min, 1H), 7.38 (d, J= 8.1 Hz, 2H), 7.94-8.10 (min, 3H); MS (ESI) m/z 604 (M'+1), 606 (M+3), 608 (M t +5). 25 To a solution of methyl 4-[1-[4-[N'-(2,6-dichlorophenyl)ureido]-3-methoxyphenylacetyl]-(4S) fluoro-(2S)-pyrrolidinyl]methoxybenzoate (530 mg, 0.78 mmol) in THF (40 ml), 0.25 N NaOH (40 ml) was added. After stirring at room temperature for 11 h, the mixture was acidified with 1 N HCI and extracted with CHCl 3 -MeOH (10/1). The combined extracts were dried over Na 2
SO
4 and concentrated in vacuo. The residue was purified on TLC [CHC13/MeOH (10/1)] to give 88 (420 30 mg, 75%) as a colorless amorphous solid. MW 590.43 mp 162-168 oC (dec.); IR (KBr) 3346, 2974, 1709, 1604, 1533, 1254 cm-'; 'H-NMR (DMSO-d6) 8 1.98-2.36 (min, 2H), 3.58 (s, 2H), 3.78 224 WO 01/00206 PCT/US00/18079 3.95 (m, 6H), 4.02-4.68 (m, 2H), 5.31-5.50 (m, 1H), 6.71-7.09 (m, 4H), 7.31 (t, J= 7.8 Hz, 1H), 7.53 (d, J = 8.1 Hz, 2H), 7.87 (d, J= 8.1 Hz, 2H), 7.88-8.00 (m, 1H), 8.30-8.40 (m, 1H), 8.89 (s, 1H); MS (ESI) nm/z 590 (M'+1), 592 (M*+3), 594 (M'+5); Anal. Calcd for C 28
H
26
C
2 FN30 6 -1.5H 2 0: C,54.47; H,4.73; F,3.08; N,6.81. Found: C,54.53; H,4.49; F, 2.93; N, 6.65. 5 Example 83 4-[(2S,4S)-I-[4-[N '-(2,6-Dichlorophenyl)ureido]-3-methylphenylacetyl]-4-fluoro-2-pyrrffolidinyl] methoxybenzoic acid F CI N N y ' -P - COOH H H Me oCco 89 To a mixture of tert-butyl 4-amino-3-methylphenylacetate (1.88 g, 8.51 mmol), 2,6-dichlorophenyl 10 isocyanate (1.60 g, 8.51 mmol) in THF (40 ml) was added Et 3 N (0.24 ml, 1.70 mmol) at room temperature. After 3 h stirring, the reaction mixture was concentrated in vacuo. The residue was triturated by the addition of n-hexane, to give tert-butyl 4-[N'-(2,6-dichlorophenyl)ureido]-3 methylphenylacetate (2.58 g, 74%) as a colorless powder. mp 243-244 oC (dec.); 'H-NMR (CDC13) 8 1.45 (s, 9H), 2.30 (s, 3H), 3.49 (s, 2H), 6.24 (s, 2H), 7.12-7.16 (m, 31), 7.35 (d,J= 8.3 15 Hz, 211), 7.51 (d, J = 7.8 Hz, li). To a stirred solution of tert-butyl 4-[N'-(2,6-dichlorophenyl)ureido]-3-methylphenylacetate (2.58 g, 6.30 mmol) in CH 2 C1 2 (50 ml) was added TFA (20 ml) at room temperature. After 2 h stirring, the mixture was concentrated in vacuo. The residue was triturated by the addition of water, to give 4 [N'-(2,6-dichlorophenyl)ureido]-3-methylphenylacetic acid (2.12 g, 95%) as a colorless powder. 20 mp 274-283 oC (dec.); 'H-NMR (DMSO-d 6 ) 8 2.24 (s, 3H), 3.46 (s, 2H), 7.00 (d, J= 8.6 Hz, 1), 7.06 (s, 1iH), 7.30 (t, J= 7.8 Hz, 1H), 7.52 (d, J= 8.3 Hz, 2H), 7.65 (d, J= 8.2 Hz, 1H), 8.12 (s, 1H), 8.50 (s, 1IH), 12.22 (br, 1H); MS (ESI) m/z 353 (M +1), 355 (M+3), 357 (M++5). A mixture of 4-[N'-(2,6-dichlorophenyl)ureido]-3-methylphenylacetic acid (181 mg, 0.51 mmol), methyl 4-[(2S,4S)-4-fluoro-2-pyrrolidinyl]methoxybenzoate (130 mg, 0.51 mmol), EDC.HCI (147 25 mg, 0.77 mmol), HOBT (104 mg, 0.77 mmol) and Et 3 N (0.35 ml, 2.55 mmol) in DMF (4 ml) was stirred at room temperature for 18 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was purified on TLC [CHC13/MeOH (20/1)], to give methyl 4-[ 1-[4-[N'-(2,6-dichlorophenyl)ureido]-3-methylphenylacetyl]-(4S)-fluoro-(2S) 225 WO 01/00206 PCT/US00/18079 pyrrolidinyl]methoxybenzoate (283 mg, 94%) as a colorless amorphous solid. IH-NMR (CDCl 3 ) 8 1.95-2.61 (m, 3H), 3.55 (br, 2H), 3.67-3.81 (m, 2H11), 3.87 (s, 6H), 3.89-4.68 (m, 2H), 5.23-5.43 (m, 1H), 6.81-7.10 (m, 6H), 7.13-7.43 (m, 3H), 7.56 (br, 1H, one of isomers), 7.73 (br, 1H, one of isomers), 7.89-8.00 (m, 2H); MS (ESI) m/z 588 (M'+1), 590 (M+3), 592 (M*+5). 5 To a solution of methyl 4-[1-[4-[N'-(2,6-dichlorophenyl)ureido]-3-methylphenylacetyl]-(4S) fluoro-(2S)-pyrrolidinyl]methoxybenzoate (283 mg, 0.48 mmol) in THF (20 ml), 0.25 N NaOH (20 ml) was added. After stirring at room temperature for 11 h, the mixture was extracted with EtOAc. The remaining aqueous layer was acidified with 1 N HCI and extracted with EtOAc. The combined extracts were dried over Na 2
SO
4 and concentrated in vacuo. The residue was on TLC 10 [CHCIMeOH (20/1)] to give 89 (450 mg, 67%) as a pale brown amorphous solid. MW 574.43 mp 174-180 oC (dec.); IR (KBr) 3330, 3288, 1711, 1685, 1604, 1512, 1242 cmr'; 'H-NMR (DMSO-d 6 ) 8 2.24 (m, 3H), 3.61 (d, 2H, J= 6.1 Hz), 3.72-4.68 (m, 7H), 5.30-5.50 (m, 1H), 6.97 7.20 (m, 4H), 7.29-7.68 (m, 5H), 7.87 (m, 2H), 8.10-8.95 (m, 1H), 12.65 (br, 1H); MS (ESI) m/z 574 (M+1), 576 (M'+3), 578 (M+5); Anal. Calcd for C 2 8
H
26 C1 2
FN
3 0s-0.5H 2 0: C, 57.64; H, 4.66; 15 Cl, 12.15, F, 3.26; N, 7.20. Found: C, 57.37; H, 4.44; Cl, 12.64; F, 3.23; N, 7.25. Example 84 4-[1-[3-chloro-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinyl] methoxybenzoic acid F N N O- COOH H H Me 90 20 To a stirred solution of 3-chlorophenylacetic acid (21.76 g, 127.6 mmol) in dichloroethane (100 ml) was added MeOH (15.6 ml, 383 mmol) and H 2
SO
4 (1 ml) at room temperature. After 20 minutes stirring, the mixture was heated at 80 oC for 2 h. The reaction mixture was poured into ice water and extracted with CHC1 3 . The combined extracts were washed with aq. NaHCO 3 and brine. After dried over Na 2
SO
4 , the extract was concentrated in vacuo to give methyl 3 25 chlorophenylacetate (25.4 g, 100%) as a colorless oil. 'H-NMR (CDC13) 8 3.60 (s, 2H), 3.70 (s, 3H), 7.15-7.26 (m, 4H). To a stirred mixture of methyl 3-chlorophenylacetate (25.4 g, 128 mmol) in H 2
SO
4 (44 ml) was added HNO 3 (5.5 ml, 138 mmol) at 0 oC. The reaction mixture was gradually raised to room temperature for 4 h. The reaction mixture was poured into ice water and extracted with EtOAc. 226 WO 01/00206 PCT/US00/18079 The combined extracts were washed with aq. NaHCO 3 and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [1 kg, n hexane/EtOAc (40/1)] to give methyl 3-chloro-4-nitrophenylacetate (11.4 g, 36%) as a yellow oil. IH-NMR (CDC1 3 ) 8 3.69 (s, 2H), 3.74 (s, 3H), 7.33 (dd, J= 8.3, 1.5 Hz, 1H), 7.49 (d, J= 1.5 Hz, 5 1H1), 7.87 (d, J= 8.3 Hz, 1H1). A mixture of methyl 3-chloro-4-nitrophenylacetate (10.9 g, 47.5 mmol), reduced iron powder (8.58 g, 153.6 mmol), AcONa-3H 2 0 (6.05 g, 44.5 mmol) and AcOH (17.6 ml) in MeOH/H 2 0 (100/400 ml) was heated at 110 oC for lh. After cooled to room temperature, the reaction mixture was filtered through Celite and the filtered cake was washed with MeOH. The combined filtrate were 10 evaporated and extracted with EtOAc. The combined extracts were washed with brine, dried over Na 2
SO
4 and concentrated in vacuo. The residue was chromatographed on silica gel [150 g, CHCl3/EtOAc (10/1)] to give methyl 4-amino-3-chlorophenylacetate (4.58 g, 48%) as a red oil. 'H-NMR (CDCl 3 ) 8 3.49 (s, 2H), 3.68 (s, 3H), 4.01 (br, 2H), 6.70 (d, J= 7.4 Hz, 1H), 6.96 (dd, J= 8.1, 2.0 Hz, 1H), 7.17 (d, J= 2.0 Hz, 1H). 15 To a mixture of methyl 4-amino-3-chlorophenylacetate (1.00 g, 5.01 mmol) and 2-methylphenyl isocyanate (0.60 ml, 5.01 mmol) in THF (20 ml) was added Et 3 N (0.14 ml, 1.00 mmol) at room temperature. After 1 day stirring, 2-methylphenyl isocyanate (0.60 ml, 5.01 mmol) was added to the reaction mixture and stirred 17 h. The reaction mixture was concentrated in vacuo. The residue was triturated by the addition of n-hexane to give methyl 3-chloro-4-[N'-(2-methylphenyl) 20 ureido]phenylacetate (1.23 g, 74%) as a colorless powder. 'H-NMR (CDCl 3 ) 8 2.34 (s, 3H14), 3.54 (s, 2H), 3.68 (s, 3H), 6.24 (br, 1H1), 6.99 (br, 1H), 7.15 (dd, J = 8.3, 2.0 Hz, 1H), 7.21-7.31 (m, 5H), 7.44 (d, J= 7.6 Hz, 1H1), 8.20 (d, J= 8.5 Hz, 1H14); MS (ESI) m/z 333 (M+1), 335 (M++3). To a stirred solution of methyl 3-chloro-4-[N'-(2-methylphenyl)ureido]phenylacetate (1.23 g, 3.70 mmol) in THF (30 ml) was added 0.25 N NaOH (30 ml). After stirring at room temperature for 14 25 h, the solvent was concentrated in vacuo. The residue was triturated by the addition of 1 N HCI and dried over 60 oC for 2 days under a reduced pressure to give 3-chloro-4-[N'-(2-methylphenyl) ureido]phenylacetic acid (1.22 g, 100%) as colorless powder. 'H-NMR (DMSO-d 6 ) 8.26 (s, 3H), 3.40 (s, 2H), 6.95 (t, J = 7.3 Hz, 1H), 7.11 (d, J= 7.6 Hz, 2H), 7.16 (d, J= 7.3 Hz, 11), 7.32 (s, 1H1), 7.76 (d, J= 8.0 Hz, 114), 7.94 (dd, J= 9.3, 1.0 Hz, 1H1), 8.72 (s, 21H); MS (ESI) m/z 319 30 (M+1), 321 (M*+3), 341 (M +Na). 227 WO 01/00206 PCT/US00/18079 A mixture of 3-chloro-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (319 mg, 1.00 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (253 mg, 1.00 mmol), EDC-HCI (288 mg, 1.50 mmol), HOBT (203 mg, 1.50 mmol) and Et 3 N (0.70 ml, 5.00 mmol) in DMF (4 ml) was stirred at room temperature for 15 h. The mixture was poured into ice water and extracted with 5 EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was purified on TLC [CHCl3/acetone (5/1)] to give methyl 4-[1-[3-chloro-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(4S)-fluoro-(2S) pyrrolidinyl]methoxybenzoate (480 mg, 87%) as a colorless amorphous solid. 'H-NMR (CDClI 3 ) 8 2.10-2.60 (m, 2H), 2.29 (s, 3H), 3.56 (d, J= 6.8 Hz, 1H), 3.71-3.84 (m, 1H), 3.87 and 3.89 (each 10 s, 3H, amide isomers), 3.91-4.20 (m, 3H), 4.49-4.60 (m, 2H), 5.32 (dt, J= 53.0, 4.2 Hz, 1H), 6.80 (br, 1H), 6.89 and 6.95 (each d, J= 8.8 Hz, 2H, amide isomers), 7.09-7.26 (m, 6H), 7.50 (d, J= 7.3 Hz, 1H), 7.94 and 8.00 (each d, J= 8.8 Hz, 2H, amide isomers), 8.10 and 8.15 (each d, J= 8.3 Hz, 1H, amide isomers); MS (FAB) mn/z 554 (M++1), 556 (M++3). To a solution of methyl 4-[1-[3-chloro-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(4S)-fluoro 15 (2S)-pyrrolidinyl]methoxybenzoate (480 mg, 0.866 mmol) in THF (30 ml) was added 0.25 N NaOH (30 ml). After stirring at room temperature for 2 days, the mixture was concentrated under a reduced pressure and acidified with 1 N HCL. The precipitates were collected, washed with water and dried under a reduced pressure to give 90 (374 mg, 80%) as a colorless powder. MW 539.98 IR (KBr) 3354, 3060, 2976, 1709, 1604, 1244 cm-'; 'H-NMR (DMSO-d 6 ) 8 2.27 (s, 3H), 2.31 (s, 20 2H), 3.66 (d, J= 7.2 Hz, 2H), 3.71-4.67 (m, 5H), 5.32-5.53 (m, 1H), 6.97 (t, J= 7.3 Hz, 1H), 7.04 7.22 (m, 5H), 7.32 and 7.35 (each d, J = 1.7 Hz, 1H, amide isomers), 7.77 (d, J 7.6 Hz, 1W), 7.87 and 7.90 (each d, J= 9.0 Hz, 2H, amide isomers), 8.01 and 8.03 (each d, J= 8.5 Hz, 1H, amide isomers), 8.57 and 8.59 (each s, 1H, amide isomers), 8.63 and 8.65 (each s, 1H, amide isomers), 12.63 (s, 1H); MS (ESI) m/z 540 (M+1), 542 (M +3). 25 Example 85 4-[1 -[3-chloro-4-[N'-(2-chlorophenyl)ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinyl]methoxy benzoic acid F N N O-Q COOH 91 To a mixture of methyl 4-amino-3-chlorophenylacetate (1.00 g, 5.01 mmol) and 2-chlorophenyl 30 isocyanate (0.60 ml, 5.01 rmmol) in THF (20 ml) was added Et 3 N (0.14 ml, 1.00 mmol) at room 228 WO 01/00206 PCT/US00/18079 temperature. After 1 day stirring, 2-chlorophenyl isocyanate (0.60 ml, 5.01 mmol) was added to the reaction mixture and stirred 17 h. The reaction mixture was concentrated in vacuo. The residue was triturated by the addition of n-hexane to give methyl 3-chloro-4-[N'-(2-chlorophenyl) ureido]phenylacetate (1.35 g, 76%) as a colorless powder. 'H-NMR (CDC 3 ) 8 3.58 (s, 3H), 3.71 5 (s, 2H), 7.04 (m, 3H), 7.18 (dd, J= 8.5, 2.0 Hz, 1H), 7.27-7.39 (m, 3H), 8.07 (m, 2H); MS (ESI) m/z 353 (M+1), 355 (M+3), 357 (M+5). To a stirred solution of methyl 3-chloro-4-[N'-(2-chlorophenyl)ureido]phenylacetate (1.35 g, 3.82 mmol) in THF (30 ml) was added 0.25 N NaOH (30 ml). After stirring at room temperature for 14 h, the solvent was concentrated in vacuo. The residue was triturated by the addition of 1 N HCI 10 and dried at 60 oC for 2 days under a reduced pressure to give 3-chloro-4-[N'-(2-chlorophenyl) ureido]phenylacetic acid (1.12 g, 86%) as colorless powder. 1 H-NMR (DMSO-de) 5 3.52 (s, 2H), 7.05 (m, 1H), 7.17 (d,J= 8.5 Hz, 1H), 7.30 (d,J= 7.6 Hz, 1H), 7.37 (s, 1H), 7.46 (dd, J= 8.0, 1.5 Hz, 1H), 7.95 (dd, J= 8.3, 1.2 Hz, 1H), 8.07 (d, J= 8.3 Hz, 1H), 9.00 (d, J= 8.0 Hz, 2H); MS (FAB) m/z 339 (M+1), 341 (M +3), 343 (M++5). 15 A mixture of 3-chloro-4-[N'-(2-chlorophenyl)ureido]phenylacetic acid (339 mg, 1.00 mmol), methyl 4-[(2S,4S)-4-fluoro-2-pyrrolidinyl]methoxybenzoate (253 mg, 1.00 mmol), EDC-HCI (288 mg, 1.50 mmol), HOBT (203 mg, 1.50 mmol) and Et 3 N (0.70 ml, 5.00 mmol) in DMF (4 ml) was stirred at room temperature for 15 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , 20 the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHC13/acetone(10/1)] to give methyl 4-[1-[3-chloro-4-[N'-(2-chlorophenyl)ureido]phenylacetyl] (4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (550 mg, 96%) as a colorless amorphous solid. 1 H-NMR (CDC 3 ) 8 2.14-2.64 (m, 2H), 3.59 (d, J= 11.2 Hz, 2H), 3.78-3.82 (m, 1H), 3.86 and 3.89 (each s, 3H, aminde isomers), 3.91-4.28 (m, 2H), 4.50-4.79 (m, 2H), 5.34 and 5.39 (each dt, J= 25 52.5, 4.4 Hz, 1H, amide isomers), 6.89-6.98 (m, 3H), 7.09-7.13 (m, 2H), 7.22 (dtJ= 7.3, 2.2 Hz, 1H), 7.29 (dd, J= 8.1, 2.0 Hz, 1H), 7.79 and 7.86 (each s, 1H1, amide isomers), 7.86-8.03 (m, 4H), 8.11 (dd, J= 8.3, 1.0 Hz, 1H); MS (FAB) m/z 574 (M+1), 576 (M+3), 578 (MW+5). To a solution of methyl 4-[1-[3-chloro-4-[N'-(2-chlorophenyl)ureido]phenylacetyl]-(4S)-fluoro (2S)-pyrrolidinyl]methoxybenzoate (550 mg, 0.957 mmol) in THF (30 ml) was added 0.25 N 30 NaOH (30 ml). After stirring at room temperature for 2 days, the mixture was concentrated under a reduced pressure and acidified with 1 N HCL. The precipitates were collected, washed with water 229 WO 01/00206 PCT/US00/18079 and dried under a reduced pressure to give 91 (437 mg, 82%) as a colorless powder. MW 560.40 IR (KBr) 3348, 3072, 2954, 1703, 1604,1529, 1439 cm'; 'H-NMR (DMSO-d 6 ) 8 2.25-2.42 (m, 2H), 3.67 (d, J = 8.3 Hz, 2H), 3.81-4.68 (m, 5H), 5.39 and 5.46 (each d, J= 54.4 Hz, 1H, amide isomers), 7.04-7.10 (m, 3H), 7.18 (d, J= 8.3 Hz, 1H), 7.31 (t, J= 8.3 Hz, 111), 7.33 and 7.37 (each 5 s, 1H, amide isomers), 7.47 (d, J= 8.1 Hz, 1H), 7.88 (dd, J= 9.0, 3.2 Hz, 2H), 7.98 (dd, J= 8.5, 3.0 Hz, 1H) m, 1H1), 8.09 (d, J= 8.3 Hz, 1H), 8.99 (d, J= 2.9 Hz, 1H1), 9.02 (s, 1H), 12.64 (s, 1H); MS (ESI) m/z 560 (M++1), 562 (M*+3), 564 (M*+5); Anal. Calcd for C 27
H
24 C1 2
FN
3 0 5 s0.2H 2 0: C, 57.50; H, 4.36; N, 7.45; Cl, 12.57; F, 3.37. Found: C, 57.72; H, 4.47; N, 7.14; Cl, 12.44; F, 3.44. Example 86 10 4-[ 1-[4-[N'-(2-bromophenyl)ureido]-3-chlorophenylacetyl]-(4S)-fluoro-(2S)-pyrrffolidinyl] methoxybenzoic acid F N N NI \ COOH r 92 To a mixture of methyl 4-amino-3-chlorophenylacetate (1.00 g, 5.01 mmol) and 2-bromophenyl isocyanate (0.62 ml, 5.01 mmol) in THF (20 ml) was added Et 3 N (0.14 ml, 1.00 mmol) at room 15 temperature. After 1 day stirring, 2-bromophenyl isocyanate (0.60 ml, 5.01 mmol) was added to the reaction mixture and stirred 24 h. The reaction mixture was concentrated in vacuo. The residue was triturated by the addition of n-hexane to give methyl 4-[N'-(2-bromophenyl)ureido]-3 chlorophenylacetate (1.34 g, 67%) as a colorless powder. 'H-NMR (CDCl 3 ) 8 3.58 (s, 3H), 3.70 (s, 2H), 6.98 (m, 3H), 7.19 (dd, J= 8.3, 1.9 Hz, 1H), 7.32 (m, 1H), 7.51 (m, 2H), 8.05 (m, 1-1); MS 20 (ESI) m/z 398 (M +1), 400 (M+3), 402 (M +5). To a stirred solution of methyl 4-[N'-(2-bromophenyl)ureido]-3-chlorophenylacetate (1.34 g, 3.37 inmnol) in THF (30 ml) was added 0.25 N NaOH (30 ml). After stirring at room temperature for 14 h, the solvent was concentrated in vacuo. The residue was triturated by the addition of 1 N HCI and dried at 60 oC for 2 days under a reduced pressure to give 4-[N'-(2-bromophenyl)ureido]-3 25 chlorophenylacetic acid (1.03 g, 80%) as colorless powder. 'H-NMR (DMSO-d 6 ) 8 3.56 (s, 2H), 7.00 (m, 111), 7.17 (dd, J = 9.0, 1.7 Hz, 1W), 7.32-7.40 (m, 2H), 7.62 (dd, J= 8.0, 1.2 Hz, 1H), 7.95 (m, 2H), 8.83 (s, 1), 9.01 (s, H), 12.41 (br, 1H); MS(FAB) m/z 385 (M +2),386(M +4),388 (M+6). 30 A mixture of 4-[N'-(2-bromophenyl)ureido]-3-chlorophenylacetic acid (384 mg, 1.00 230 WO 01/00206 PCT/US00/18079 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (253 mg, 1.00 mmol), EDC-HCI (288 mg, 1.50 mmol), HOBT (203 mg, 1.50 mmol) and Et 3 N (0.70 ml, 5.00 mmol) in DMF (4 ml) was stirred at room temperature for 15 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over 5 Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHCI 3 /acetone(10/1)] to give methyl 4-[1-[4-[N' -(2-bromophenyl)ureido]-3 chlorophenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (530 mg, 86%) as a colorless amorphous solid. 'H-NMR (CDCl 3 ) 8 2.14-2.63 (m, 2H), 3.58 (d, J= 10.0 Hz, 1H), 3.73-3.83 (m, 1H), 3.86 and 3.89 (each s, 3H, amide isomers), 3.90-4.29 (m, 3H), 4.50-4.69 (m, 2H), 5.33 10 and 5.37 (each m, 1H, amide isomers), 6.88-6.93 (m, 3H), 7.11-7.14 (m 2H), 7.26 (m, 1H), 7.46 (d, J= 8.1 Hz, 1H), 7.62-7.78 (m, 2H), 7.89 and 7.93 (each m, 2H, amide isomers), 8.01 (dd, J= 8.8, 1.7 Hz, 2H); MS (FAB) m/z 618 (M), 620 (M+2), 622 (M'+4). To a solution of methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-chlorophenylacetyl]-(4S)-fluoro (2S)-pyrrolidinyl]methoxybenzoate (530 mg, 0.856 mmol) in THF (30 ml) was added 0.25 N 15 NaOH (30 ml). After stirring at room temperature for 2 days, the mixture was concentrated under a reduced pressure and acidified with 1 N HCL. The mixture was extracted with CHClIMeOH (10/1). The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [20 g, CHCl3/acetone(10/1)-CHCl 3 /MeOH(10/1)] to give 92 (59 mg, 11%) as a colorless amorphous 20 solid. MW 604.85 IR (KBr) 3329, 3060, 2976, 1712, 1526, 1435 cm-'; 'H-NMR (DMSO-d 6 ) 2.31 (m, H), 3.48-4.68 (m, 7H), 5.32-5.53 (m, 1), 6.99-7.19 (m, 4H), 7.36 (s, 1H), 7.63 (dd, J= 6.7, 1.2 Hz, 1H), 7.86-8.18 (m, 4H), 8.83 (s, 1H), 9.02 (s, 1H), 12.67 (br, 1H); MS (ESI) m/z 604 (M++I), 606 (M++3), 608 (M+5); Anal. Calcd for C 27
H
24 BrC1FN 3 0 5 -0.5H 2 0: C, 52.83; H, 4.10; N, 6.85; Cl, 5.78; F, 3.09. Found: C, 53.24; H, 4.32; N, 6.43; Cl, 6.01; F, 3.07. 25 Example 87 4-[1-[3-chloro-4-(N '-phenylureido)phenylacetylj-(4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoic acid F N N N O-&jCOOH H HI "" 93 To a mixture of methyl 4-amino-3-chlorophenylacetate (1.31 g, 6.56 mmol) and phenyl isocyanate 30 (0.71 ml, 6.56 mmol) in THF (20 ml) was added Et 3 N (0.19 ml, 1.33 immol) at room temperature. After 15 h stirring, the reaction mixture was concentrated in vacuo. The residue was triturated by 231 WO 01/00206 PCT/US00/18079 the addition of n-hexane to give methyl 3-chloro-4-(N'-phenylureido)phenylacetate (1.79 g, 86%) as a pale brown solid. 'H-NMR (CDCl 3 ) 8 3.56 (s, 2H), 3.70 (s, 3H), 6.70 (m, 1H), 7.06 (s, 1H), 7.14-7.18 (m, 2H), 7.26 (dd, J= 7.8, 1.9 Hz, 1H), 7.33-7.38 (m, 4H), 8.14 (dd, J= 8.3, 3.0 Hz, 1H); MS (ESI) m/z 319 (M'+1), 321 (M+3). 5 To a stirred solution of methyl 3-chloro-4-(N'-phenylureido)phenylacetate (1.79 g, 5.62 mmol) in THF (30 ml) was added 0.25 N NaOH (30 ml). After stirring at room temperature for 20 h, the solvent was concentrated in vacuo. The residue was triturated by the addition of 1 N HCI and dried at 60 oC for 2 days under a reduced pressure to give 3-chloro-4-(N'-phenylureido) phenylacetic acid (1.58 g, 92%) as pale brown solid. 'H-NMR (DMSO-d 6 ) 8 3.55 (s, 2H), 6.99 (t, J 10 = 7.3 Hz, 1H), 7.17 (d, J= 8.3 Hz, 1H), 7.29 (t, J= 7.6 Hz, 2H), 7.36 (s, 1H), 7.46 (d, J= 8.0 Hz, 2H), 8.07 (d, J = 8.3 Hz, 1H), 8.28 (s, 1H), 9.37 (s, 1H), 12.37 (br, 1H). A mixture of 3-chloro-4-(N'-phenylureido)phenylacetic acid (305 mg, 1.00 mmol), methyl 4 [(2S,4S)-4-fluoro-2-pyrrolidinyl]methoxybenzoate (253 mg, 1.00 mmol), EDC-HCI (288 mg, 1.50 mmol), HOBT (203 mg, 1.50 mmol) and Et 3 N (0.70 ml, 5.00 mmol) in DMF (4 ml) was stirred at 15 room temperature for 17 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [30 g, CHCl3/acetone(20/1)] to give methyl 4-[1-[3-chloro-4-(N'-phenylureido)phenylacetyl]-(4S)-fluoro (2S)-pyrrolidinyl]methoxybenzoate (720 mg, 100%) as a colorless amorphous solid. MS (FAB) 20 m/z 540 (M+1), 542 (M+3). To a solution of methyl 4-[1-[3-chloro-4-(N'-phenylureido)phenylacetyl]-(4S)-fluoro-(2S) pyrrolidinyl]methoxybenzoate (720 mg, 1.00 mmol) in THF/MeOH (30/30 ml) was added 0.25 N NaOH (30 ml). After stirring at room temperature for 2 h, the reaction mixture was heated at 50 oC for 22 h. After removed the solvent, the resulting residue was acidified with 1 N HC1. The 25 precipitates were collected, washed with water and dried under a reduced pressure to give 93 [412 mg, 78% (2 Steps)] as a colorless powder. MW 525.96 IR (KBr) 3346, 3302, 2976, 1712, 1604, 1240 cm-'; 'H-NMIR (DMSO-d6) 8 2.25-2.31 (m, 2H), 3.66 (d, J= 7.8 Hz, 2H), 3.71-4.67 (m, 5H), 5.31-5.52 (m, 1H), 6.99 (t, J= 7.3 Hz, 1H), 7.04 and 7.07 (each d, J = 8.7 Hz, 2H, amide isomers), 7.14-7.18 (m, 1H), 7.29 (t, J = 7.3 Hz, 2H), 7.35 (d, J= 1.7 Hz, 1H), 7.46 (d, J= 7.8 Hz, 2H), 7,87 30 and 7.90 (each d, J = 9.0 Hz, 2H, amide isomers), 8.04 and 8.06 (each d, J = 8.5 Hz, 1H, amide isomers), 8.26 and 8.28 (each s, 1H, amide isomers), 9.36 (s, 1H), 12.63 (s, 1H); MS (ESI) m/z 526 232 WO 01/00206 PCT/US00/18079 (M++1), 528 (M++3); Anal. Calcd for C 27
H
25
CFN
3 0s-0.5H 2 0: C, 60.62; H, 4.90; N, 7.85; Cl, 6.63; F, 3.55. Found: C, 61.00; H, 5.19; N, 7.40; Cl, 6.66; F, 3.39. Example 88 4-[1 -[3-bromo-4-[N '-(2-methylphenyl)ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrolidiny 5 ]methoxybenzoic acid F N N O -- COOH Me H H Br 94 To a stirred solution of 3-bromophenylacetic acid (10.2 g, 47.4 mmol) in dichloroethane (50 ml) was added MeOH (5.8 ml, 142 mmol) and H 2 SO4 (0.5 ml) at room temperature. After 20 minutes stirring, the mixture was heated at 80 oC for 7 h. The reaction mixture was poured into ice water 10 and extracted with CHC1 3 . The combined extracts were washed with aq. NaHCO 3 and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo to give methyl 3-bromophenyl acetate (10.8 g, 99%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 3.60 (s, 2H), 3.71 (d, J= 1.0 Hz, 3H), 7.18 7.44 (m, 4H11). To a stirred mixture of methyl 3-chlorophenylacetate (10.8 g, 47.1 mmol) in H 2
SO
4 (15.1 ml) was 15 added HNO 3 (2.8 ml, 70.7 mmol) at 0 oC. The reaction mixture was gradually raised to room temperature for 5.5 h. The reaction mixture was poured into ice water and extracted with CHC 3 . The combined extracts were washed with aq. NaHCO 3 and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [500 g, n hexane/EtOAc (10/1)] to give methyl 3-bromo-4-nitrophenylacetate (3.69 g, 29%) as a yellow oil. 20 'H-NMR (CDCl 3 ) 8 3.68 (s, 2H), 3.73 (s, 3H), 7.38 (dd, J= 8.3, 1.2 Hz, 1H), 7.67 (d, J= 1.3 Hz, 1H1), 7.83 (d, J= 8.3 Hz, 1H1). A mixture of methyl 3-bromo-4-nitrophenylacetate (14.8 g, 53.8 mmol), reduced iron powder (9.62 g, 172 mmol), AcONa-3H 2 0 (7.32 g, 53.8 mmol) and AcOH (20.0 ml) in MeOH/H20 (150/600 ml) was heated at 90 oC for lh. After cooled to room temperature, the reaction mixture was filtered 25 through Celite and the filtered cake was washed with MeOH. The combined filtrate were evaporated and extracted with EtOAc. The extracts were washed with brine, dried over Na 2
SO
4 and concentrated in vacuo. The residue was chromatographed on silica gel [400 g, CHCllEtOAc (20/1)] to give methyl 4-amino-3-bromophenylacetate (9.01 g, 69%) as a brown oil. 'H-NMR (CDCl 3 ) 5 3.48 (s, 2H), 3.68 (s, 3H), 4.05 (br, 2H), 6.69 (d, J= 8.3 Hz, 1H), 7.00 (dd, J= 8.1, 2.0 30 Hz, 1H), 7.32 (d, J= 2.0 Hz, 1H1). 233 WO 01/00206 PCT/US00/18079 To a mixture of methyl 4-amino-3-bromophenylacetate (587 mg, 2.40 mmol) and 2-methylphenyl isocyanate (0.287 ml, 2.40 mmol) in THF (2 ml) was added Et 3 N (33 ml, 0.24 mmol) at room temperature. After 21 h stirring, the reaction mixture was concentrated in vacuo. The residue was triturated by the addition of n-hexane to give methyl 3-bromo-4-[N'-(2-methylphenyl)ureido] 5 phenylacetate (650 mg, 72%) as a pale brown powder. 'H-NMR (CDCl 3 ) 82.34 (s, 3H), 3.53 (s, 2H), 3.68 (s, 3H), 6.18 (br, 1H), 6.96 (br, 1H), 7.18-7.33 (min, 4H), 7.29 (d,J= 4.4 Hz, 1H), 7.30 (d, J= 7.3 Hz, 1H), 8.19 (d, J= 8.3 Hz, 1H); MS (ESI) m/z 377 (M), 379 (M+2). To a stirred solution of methyl 3-bromo-4-[N'-(2-methylphenyl)ureido]phenylacetate (650 mg, 1.72 mmol) in THF (10 ml) was added 0.25 N NaOH (10 ml). After stirring at room temperature 10 for 14 h, the solvent was concentrated in vacuo. The residue was triturated by the addition of 1 N HCI and dried at 60 oC for 2 days under a reduced pressure to give 3-bromo-4-[N'-(2 methylphenyl)ureido] phenylacetic acid (1.22 g, 100%) as colorless powder. 'H-NMR (DMSO-de) 8 2.26 (s, 3H), 3.32 (s, 2H), 6.93 (min, 2H), 7.10-7.17 (min, 4H), 7.76 (d,J = 8.1 Hz, 2H), 8.52 (s, 1H); MS (ESI) m/z 385 (M'+Na), 387 (M'+2+Na). 15 A mixture of 3-bromo-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (80 mg, 0.22 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (56 mg, 0.22 mmol), EDC.HCI (63 mg, 0.33 mmol), HOBT (45 mg, 0.33 mmol) and Et 3 N (0.15 ml, 1.10 mmol) in DMF (1 ml) was stirred at room temperature for 18 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the 20 extracts were concentrated in vacuo. The residue was purified on TLC [CHCllacetone (5/1)] to give methyl 4-[1-[3-bromo-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(4S)-fluoro-(2S) pyrrolidinyl ]methoxybenzoate (140 mg, 100%) as a yellow oil. 'H-NMR (CDC13) 8 2.30 (s, 3H), 2.55 (min, 1H), 3.56 (d, J= 6.4 Hz, 2H), 3.70-3.84 (min, 3H), 3.87 (s, 3H), 3.99-4.59 (min, 3H), 5.23 5.38 (min, 1H), 6.83-6.94 (min, 2H), 6.95 (d, J = 8.8 Hz, 1H), 7.07-7.26 (mn, 5H), 7.36-7.63 (mn, 2H), 25 7.94-8.15 (min, 3H); MS (ESI) m/z 598 (M +1), 600 (M +3). To a solution of methyl 4-[1 -[3-bromo-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(4S)-fluoro (2S)-pyrrolidinyl]methoxybenzoate (140 mg, 0.22 nimmol) in THF (10 ml) was added 0.25 N NaOH (10 ml). After stirring at room temperature for 14 h, the mixture was concentrated under a reduced pressure and acidified with 1 N HC1. The precipitates were collected, washed with water 30 and dried under a reduced pressure to give 94 (109 mg, 85%) as a colorless powder. MW 584.43 IR (KBr) 3313, 3060, 2976, 1687, 1604, 1525, 1244 cm-'; 'H-NMR (DMSO-d 6 ) 8 2.27 (s, 3H), 234 WO 01/00206 PCT/US00/18079 2.29 (m, 2H), 3.66 (d, J= 8.1 Hz, 2H), 3.72-4.68 (m, 5H), 5.31-5.53 (m, 1H), 6.92-6.99 (m, 1H), 7.04 and 7.07 (each d, J= 8.3 Hz, 2H, amide isomers), 7.11-7.21 (m, 3H), 7.48 and 7.51 (s, 1H, amide isomers), 7.75 and 7.79 (each d, J = 8.1 Hz, 1H, amide isomers), 7.86-7.92 (m, 3H), 8.45 and 8.47 (each s, 1H, amide isomers), 8.59 (s, 1H), 12.64 (s, 1H); MS (FAB) nm/z 584 (M'+1), 586 5 (M+3); Anal. Calcd for C 28
H
2 7 BrFN 3 Os: C, 57.54; H, 4.66; N, 7.19; Br, 13.67; F, 3.25. Found: C, 57.93; H, 4.97; N, 7.04; Br, 13.35; F, 2.89. Example 89 4-[I-[3-bromo-4-[N '-(2-chlorophenyl)ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinyl] methoxybenzoic acid F 1N N \ COOH 10 H H 6r 95 To a mixture of methyl 4-amino-3-bromophenylacetate (587 mg, 2.40 mmol) and 2-chlorophenyl isocyanate (0.29 ml, 2.40 mmol) in THF (2 ml) was added Et 3 N (33 ml, 0.24 mmol) at room temperature. After 21 h stirring, the reaction mixture was concentrated in vacuo. The residue was triturated by the addition of n-hexane to give methyl 3-bromo-4-[N'-(2-chlorophenyl)ureido] 15 phenylacetate (710 mg, 74%) as a pale brown powder. 'H-NMR (CDCl 3 ) 8 3.57 (s, 2H), 3.70 (s, 3H), 7.02-7.28 (m, 2H), 7.36 (d, J= 6.8 Hz, 1H), 7.48 (s, 1H), 8.00-8.11 (m, 2H); MS (ESI) m/z 397 (M'), 399 (M++2), 401 (M+4). To a stirred solution of methyl 3-bromo-4-[N'-(2-chlorophenyl)ureido]phenylacetate (710 mg, 1.79 mmol) in THF (10 ml) was added 0.25 N NaOH (10 ml). After stirring at room temperature for 14 20 h, the solvent was concentrated in vacuo. The residue was triturated by the addition of 1 N HCI and dried at 60 oC for 2 days under a reduced pressure to give 3-bromo-4-[N'-(2-chlorophenyl) ureido]phenylacetic acid (643 mg, 94%) as colorless powder. 'H-NMR (DMSO-d) 8 3.56 (s, 2H), 7.05 (m, 1H), 7.21 (dd, J= 8.6, 1.7 Hz, 1H), 7.29 (t, J= 7.8 Hz, 1H), 7.46 (d, J = 8.1 Hz, 1H), 7.46 (d, J= 8.1 Hz, 1H), 7.53 (d, J= 1.7 Hz, 1H), 7.83 (d, J= 8.3 Hz, 1H), 8.06 (d, J= 7.6 Hz, 25 1H), 8.86 (s, 1H), 8.89 (s, 1H), 12.40 (s, 1H); MS (ESI) m/z 382 (M+1), 384(M+3). A mixture of 3-bromo-4-[N'-(2-chlorophenyl)ureido]phenylacetic acid (384 mg, 1.00 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (253 mg, 1.00 mmol), EDC-HCl (288 mg, 1.50 mmol), HOBT (203 mg, 1.50 mmol) and Et 3 N (0.70 ml, 5.00 mmol) in DMF (4 ml) was stirred at room temperature for 18 h. The mixture was poured into ice water and extracted with 30 EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [30 g, 235 WO 01/00206 PCT/US00/18079 CHCl/acetone(10/1)] to give methyl 4-[1-[3-bromo-4-[N'-(2-chlorophenyl)ureido]phenylacetyl] (4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (640 mg, 100%) as a colorless amorphous solid. 'H-NMR (CDClI) 8 2.07-2.46 (min, 2H), 2.59 (t, J = 18.4 Hz, 1H), 3.57 (d, J = 10.5 Hz, 2H), 3.63 4.67 (min, 7H), 5.26-5.44 (min, 1H), 6.89-6.96 (min, 3H), 7.13 (d,J = 7.6 Hz, 1H), 7.1 (t,J= 7.3 Hz, 5 1H), 7.26-7.29 (min, 2H), 7.52-7.94 (min, 4H), 8.01(d, J= 8.5 Hz, 1H), 8.09 (d, J= 8.5 Hz, 1H); MS (FAB) m/z 618 (M), 620 (M+3), 622 (M++5). To a solution of methyl 4-[1-[3-bromo-4-[N'-(2-chlorophenyl)ureido]phenylacetyl]-(4S)-fluoro (2S)-pyrrolidinyl]methoxybenzoate (640 mg, 1.00 mmol) in THF (40 ml) was added 0.25 N NaOH (40 ml). After stirring at room temperature for 14 h, the mixture was concentrated under a 10 reduced pressure and acidified with 1 N HCI. The precipitates were collected, washed with water and dried under a reduced pressure to give 95 (522 mg, 86%) as a pale yellow powder. MW 604.85 IR (KBr) 3317, 3072, 1709, 1685, 1604, 1529, 1290 cm'; 'H-NMR (DMSO-d 6 ) 8 2.24-2.50 (min, 2H), 3.67 (d, J = 8.3 Hz, 2H), 3.73-4.68 (min, 5H), 5.31-5.52 (min, 1), 7.03-7.09 (min, 3H), 7.22 (dt, J= 8.3, 1.7 Hz, 1), 7.30 (d, J = 7.3 Hz, 1H), 7.46 (dd, J= 8.0, 1.4 Hz, 1H), 7.49 and 7.52 15 (each d, J= 2.0 Hz, 1H, amide isomers), 7.80-7.91 (min, 3H), 8.07 (dd, J= 8.3, 1.2 Hz, 1H), 8.85 and 8.86 (each s, 1H, amide isomers), 8.96 and 8.97 (each s, 1H, amide isomers), 12.62 (s, 1H); MS (FAB) m/z 605 (M++1), 607 (M++3), 609 (M +3), 626 (M'+1+Na); Anal. Calcd for
C
27
H
24 BrC1FN 3 0,O0.8H 2 0: C, 52.37; H, 4.17; N, 6.79; F, 3.07. Found: C, 52.63; H, 4.12; N, 6.62; F, 2.97. 20 Example 90 4-[ 1-[3-bromo-4-[N'-(2-bromophenyl)ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrffolidinyl] methoxybenzoic acid N N O / COOH Br H H Br 96 To a mixture of methyl 4-amino-3-bromophenylacetate (587 mg, 2.40 mmol) and 2-bromophenyl 25 isocyanate (0.30 ml, 2.40 mmol) in THF (2 ml) was added Et 3 N (33 ml, 0.24 mmol) at room temperature. After 4 h stirring, the reaction mixture was concentrated in vacuo. The residue was triturated by the addition of n-hexane to give methyl 3-bromo-4-[N'-(2-bromophenyl)ureido] phenylacetate (770 mg, 73%) as a pale brown powder. 'H-NMR (CDC13) 8 3.55 (s, 2H), 3.70 (s, 3H), 6.97 (dd, J= 7.3, 1.5 Hz, 1H), 7.22 (dd, J= 8.5, 2.2 Hz, 1H), 7.29-7.33 (mn, 2H), 7.48 (d, J= 30 1.0, 2.2 Hz, 1H), 7.54 (dd, J = 8.0, 1.2 Hz, 1H), 8.01 (min, 2H); MS (ESI) m/z 443 (M +1), 445 (M+3), 447 (M'+5). 236 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 3-bromo-4-[N'-(2-bromophenyl)ureido]phenylacetate (770 mg, 1.74 mmol) in THF (10 ml) was added 0.25 N NaOH (10 ml). After stirring at room temperature for 14 h, the solvent was concentrated in vacuo. The residue was triturated by the addition of 1 N HCI and dried at 60 oC for 2 days under a reduced pressure to give 3-bromo-4-[N'-(2-bromophenyl) 5 ureido]phenylacetic acid (702 mg, 94%) as colorless powder. 'H-NMR (DMSO-d) 8 3.56 (s, 2H), 6.99 (dt, J= 7.8, 1.5 Hz, 1H), 7.21 (dd, J= 8.3, 1.7 Hz, 1H), 7.33 (dt, J= 7.1, 1.5 Hz, 111), 7.53 (d,J = 1.7 Hz, 1H), 7.62 (dd, J= 8.1, 1.5 Hz, 1H), 7.82 (d,J= 8.3 Hz, 1H), 7.93 (dd, J= 8.1, 1.5 Hz, 1H), 8.82 (s, 1H), 8.86 (s, 1H), 12.39 (s, 1H); MS (ESI) m/z 428 (M+1), 430(M+3). A mixture of 3-bromo-4-[N'-(2-chlorophenyl)ureido]phenylacetic acid (428 mg, 1.00 mmol), 10 methyl 4-[(2S,4S)-4-fluoro-2-pyrrolidinyl]methoxybenzoate (253 mg, 1.00 mmol), EDC-HCI (288 mg, 1.50 mmol), HOBT (203 mg, 1.50 mmol) and Et 3 N (0.70 ml, 5.00 mmol) in DMF (4 ml) was stirred at room temperature for 18 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [30 g, 15 CHClI/acetone(10/1)] to give methyl 4-[1-[3-bromo-4-[N'-(2-bromophenyl)ureido]phenylacetyl] (4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (720 mg, 100%) as a colorless amorphous solid. 'H-NMR (CDCl 3 ) 8 2.07-2.45 (m, 2H), 2.58 (m, 1H), 3.58 (d, J= 9.0 Hz, 2H), 3.63-4.69 (m, 9H), 5.26-5.43 (m, 1H), 6.88-6.99 (m, 3H), 7.16 (d, J= 8.3 Hz, 1H), 7.23-7.32 (m, 2H), 7.46 (dd, J= 8.1, 1.5 Hz, 1H1), 7.51-8.20 (m, 5H); MS (FAB) m/z 664 (M), 666 (M +3), 668 (M+5). 20 To a solution of methyl 4-[1-[3-bromo-4-[N'-(2-bromophenyl)ureido]phenylacetyl]-(4S)-fluoro (2S)-pyrrolidinyl]methoxybenzoate (720 mg, 1.00 mmol) in THF (40 ml) was added 0.25 N NaOH (40 ml). After stirring at room temperature for 14 h, the mixture was concentrated in vacuo and acidified with 1 N HC1. The mixture was extracted with CHCl/MeOH (10/1). The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were 25 concentrated in vacuo. The residue was chromatographed on silica gel [20 g, CHCljacetone (10/1)-CHClz/MeOH(20/1)] and triturated by the addition of ether to give 96 (489 mg, 75%) as a colorless amorphous solid. MW 649.30 IR (KBr) 3450, 3313, 3070, 1709, 1684, 1525, 1435 cmur'; 'H-NMR (DMSO-d 6 ) 8 2.25-2.50 (m, 2H), 3.67 (d, J= 8.3 Hz, 2H), 3.73-4.68 (m, SH), 5.31-5.53 (m, 1H), 6.98-7.08 (m, 3H), 7.21 (d, J= 8.2 Hz, 1H), 7.34 (t, J= 8.8 Hz, 1H), 7.50 and 7.53 (each 30 s, 1H1, amide isomers), 7.62 (d, J = 8.0 Hz, 1H), 7.80-7.96 (m, 4H), 8.82 (s, 1H), 8.85 and 8.86 (each s, 1H, amide isomers), 12.63 (br, 1H); MS (FAB) m/z 650 (M +1), 652 (M +3), 654 (M++3), 672 (M+Na); Anal. Called for C 27
H
2 4 BrFN 3 O-0.9H 2 0: C, 48.73; H, 3.91; N, 6.31; F, 237 WO 01/00206 PCT/US00/18079 2.85. Found: C, 48.96; H, 3.98; N, 5.92; F, 2.77. Example 91 4-[ -[4-[N'-(2-methylphenyl)ureido]-2,3-difluorophenylacetyl]-(4S)-fluoro-(2S)-pyrrolidiny methoxy]benzoic acid. N N Q' N O - - C O O H 5 MeH H F 97 To a stirred solution of tert-butyl ethyl malonate (5.35 ml, 28.2 mmol) in DMF (150 ml) was added NaH (60% in oil, 3.38 g, 84.7 mmol) at rt. After 20 min, 2,3-difluoronitrobenzene (5 g, 28.2 mmol) in DMF (50 mL) was added dropwise via dropping funnel. Following the addition, the mixture was stirred for 3 hours at rt. The mixture was poured into ice-water and sat. NH 4 C1 10 (100 mL). The mixture was extracted with EtOAc and the combined organic layer was washed with IM HCI and brine, dried over MgSO 4 , filtered and concentrated. The residue was dissolved to dichloromethane (20 mL), and added TFA (20 mL) at rt. The mixture was refluxed for 18 h. The mixture was evaporated in vacuo, coevaporated with toluene (20 mL x 2). The residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC 15 5404-FC, linear gradient hexane-EtOAc 10:0 to 1: 1, 4 50 mmn x 300 mm, 15 mL/min) to give ethyl 2,3-difluoro-4-nitrophenylacetic acid (5.85 g, 85%) as a yellow oil. 'H-NMR (CDC1 3 ) 6 1.30 (min, 3 H), 3.78 (s, 2 H), 4.22 (min, 2 H), 7.22 (min, 1 H), 7.84 (min, 1 H); MS (FAB) m/z 246 (M'+1). To a stirred solution of ethyl 2,3-difluoro-4-nitrophenylacetate (5.85 g, 23.9 mol) in EtOH (100 mL), was added SnCl 2 (16.1 g, 71.6 mmol) at rt. The stirring was continued for 18 hours at 20 reflux. After removal of the solvent, the residue was dissolved in CHCl 3 (100 mL) and poured into ice water-4M NaOH (40 mL of 4M NaOH in 300 mL of ice-water), extracted with CHCl 3 (100 mL x 2), dried over anhydrous MgSO 4 , and concentrated under a reduced pressure. The residue was chromatographed on silica gel (middle pressure chromatography system YAMAZEN YFLC 5404, linear gradient of hexane-EtOAc from 9:1 to 7:3, 4 50 mm x 500 mm, 15 ml/min) to give 25 ethyl 4-amino-2,3-difluorophenylacetic acid (1.94 g, 38%) as a colorless oil. 'H-NMR (CDC13) 6 1.25 (t, J = 7.3 Hz, 3 H), 3.55 (d, J= 1.0 Hz, 2 H), 3.78 (brs, 2 H), 4.15 (dd, J= 7.2 Hz, 14.2 Hz, 2 H), 6.49 (dt, J= 1.8, 8.2 Hz, 1 H), 6.78 (min, 1 H); MS (FAB) m/z 216 (M'+1). To a stirred solution of ethyl 4-amino-2,3-difluorophenylacetate (323 mg, 1.5 mmol) in DMF (8 mL), were added triethylamine (0.209 ml, 1.5 mminol) and 2-methylphenyl isocyanate (0.372 ml, 238 WO 01/00206 PCT/US00/18079 3.0 mmol) at rt. The stirring was continued for 48 hour at 80 0 C. The reaction mixture was evaporated in vacuo, and the solid was suspended to n-hexane. The solid was collected through filtration. The solid was dissolved in THF-MeOH (1:1, v/v, 20 mL), and was added 4M NaOH (10 mL) at rt. The stirring was continued for 18 hours at rt. The reaction was poured into IM 5 HC1, and the resulting precipitate was collected through filtration. The solid was recrystallized with CHCl 3 -n-hexane to give 4-[(2-methylphenyl)ureido]-2,3-difluorophenylacetic acid (200 mg, 42%) as a white solid. 'H-NMR (CDC13) 8 2.30 (s, 3 H), 3.35 (s, 2 H), 6.98 (min, 1 H), 7.04 (min, 1 H), 7.18 (d, J= 7.3 Hz, 2 H), 7.69 (d, J= 8.1 Hz, 1 H), 7.90 (min, 1 H); MS (FAB) nm/z 321 (M*+1). To a stirred solution of methyl 4-(4-S-4-fluoro-2-pyrrolidinyl)methoxy benzoate (63 mg, 0.25 10 mmol) and 4-[N'-(2-methylphenyl)ureido]-2,3-difluorophenylacetic acid (82 mg, 0.25 mmol) in DMF (5 mL), were added EDC-HCI (72 mg, 0.38 mmol), HOBt (69 mg, 0.48 mmol), and DMAP (cat.), and the stirring was continued overnight at rt. The mixture was diluted with EtOAc (50 mL), washed with IM NaOH, IM HCI, and brine, dried over anhydrous MgSO4, and concentrated under a reduced pressure. The residue was dissolved in THF-MeOH-HO 2 0 (21 mL, 1:1:1, v/v/v) 15 and the stirring was continued for 6 h at rt. The mixture was poured into 1M HCI and extracted with CHC1 3 -MeOH (9:1, v/v). The combined organic phase was dried over anhydrous MgSO4, and concentrated under a reduced pressure. The residue was purified with TLC (Whatman, PLK 5F, CHC1IMeOH, 20:1, v/v) to give 97 (69 mg, 51%) as a white powder. MW 541.52 IR (KBr) 3340, 1604, 1540, 1251, 1168, 754 cm-'; 'H-NMR (DMSO-d 6 ) 8 2.25 (s, 3 H), 2.32 (min, 2 H), 3.68 20 4.40 (min, 7 H), 5.32-5.55 (min, 1 H), 6.98 (min, 2 H), 7.05 (d, J = 8.8 Hz, 2 H), 7.83 (d, J = 8.8 Hz, 2 H), 7.82-7.92 (min, 2 H), 8.40 (s, 1 H), 9.14 (s, 1 H); MS (ESI) m/z 564 (M*+Na); Anal. Calcd for
C
28
H
26
F
3
N
3 0s-2.0H 2 0: C, 58.23; H, 5.24; N, 7.28. Found: C, 58.07, H, 4.84; N, 7.03. Example 92 4-[1 -[4-[N'-(2-methylphenyl)ureido]-2,5-difluorophenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinyl 25 methoxy]benzoic acid F NN \/COOH Me H H 98 To a stirred solution of di-tert-butyl ethyl malonate (6.32 ml, 28.2 mmol) in DMF (150 ml), was added NaH (60% in oil, 3.38 g, 84.7 mmol) at rt. After 20 min, 2,5-difluoronitrobenzene (5 g, 28.2 mmol) in DMF (50 mL) was added dropwise via dropping funnel. Following the addition, 30 the mixture was stirred for 3 hours at rt. The mixture was poured into ice-water and sat. NH 4 C1 239 WO 01/00206 PCT/US00/18079 (100 mL). The mixture was extracted with EtOAc and the combined organic layer was washed with IM HCI and brine, dried over MgSO 4 , filtered and concentrated. The residue was dissolved to dichloromethane (20 mL), and added TFA (20 mL) at rt. The mixture was refluxed for 18 h. The mixture was evaporated in vacuo, coevaporated with toluene (20 mL x 2). The residue was 5 dissolved in MeOH (150 mL), and added conc. H 2
SO
4 (5 mL). The mixture was refluxed for 18 h. The mixture was diluted with EtOAc (300 mL), washed with water, IM HCI, and brine, dried over anhydrous MgSO 4 , and concentrated under a reduced pressure. The residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC-5404-FC, linear gradient hexane-EtOAc 10:0 to 1: 1, 4 50 mm x 300 mm, 15 mL/min) to give ethyl 2,5-difluoro-4 10 nitrophenylacetic acid (6.53 g, 90%) as a yellow oil. 'H-NMR (CDCI 3 ) 5 3.75 (s, 2 H), 3.76 (s, 3 H), 7.29 (dd, J = 5.8 Hz, 10.5 Hz, 1 H), 7.81 (dd, J= 6.0 Hz, 8.4 Hz, 1 H); MS (ESI) m/z 232 (M+1). To a stirred solution of ethyl 2,5-difluoro-4-nitrophenylacetate (5.88 g, 25.4 mol) in EtOH (100 mL), was added SnCl 2 (17.2 g, 76.3 mmol) at rt. The stirring was continued for 18 hours at 15 reflux. After removal of the solvent, the residue was dissolved in CHC1 3 (100 mL) and poured into ice water-4M NaOH (40 mL of 4M NaOH in 300 mL of ice-water), extracted with CHC13 (100 mL x 2), dried over anhydrous MgSO 4 , and concentrated under a reduced pressure. The residue was chromatographed on silica gel (middle pressure chromatography system YAMAZEN YFLC 5404, linear gradient of hexane-EtOAc from 9:1 to 7:3, 4 50 mm x 500 mm, 15 ml/min) to give 20 ethyl 4-amino-2,5-difluorophenylacetic acid (2.85 g, 52%) as a colorless oil. ' H-NMR (CDCI 3 ) 6 1.28 (t, J= 7.3 Hz, 3 H), 3.51 (s, 2 H), 3.78 (brs, 2 H), 4.15 (dd, J= 7.2 Hz, 14.2 Hz, 2 H), 6.47 (dd, J= 7.5, 10.4 Hz, 1 H), 6.88 (dd, J= 6.7, 11.0 Hz, 1 H); MS (FAB) m/z 216 (M'+1). To a stirred solution of ethyl 4-amino-2,5-difluorophenylacetate (323 mg, 1.5 mmol) in DMF (8 mL), were added triethylamine (0.209 ml, 1.5 mmol) and 2-methylphenyl isocyanate (0.372 ml, 25 3.0 mmol) at rt. The stirring was continued for 48 hour at 80 oC. The reaction mixture was evaporated in vacuo, and the solid was suspended to n-hexane. The solid was collected through filtration. The solid was dissolved in THF-MeOH (1:1, v/v, 20 mL), and was added 4M NaOH (10 mL) at rt. The stirring was continued for 18 hours at rt. The reaction was poured into IM HC1, and the resulting precipitate was collected through filtration. The solid was recrystallized 30 with CHCl 3 -n-hexane to give 4-[(2-methylphenyl)ureido]-2,5-difluorophenylacetic acid (214 mg, 46%) as a white solid. 'H-NMR (CDCl 3 ) 8 2.30 (s, 3 H), 3.35 (m, 2 H), 7.02 (m, 2 H), 7.18 (d, J= 240 WO 01/00206 PCT/US00/18079 7.3 Hz, 2 H), 7.69 (d, J = 7.8 Hz, 1 H), 8.03 (min, 1 H); MS (FAB) m/z 321 (M+1). To a stirred solution of methyl 4-[(4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (63 mg, 0.25 mmol) and 2,5-difluoro-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (82 mg, 0.25 mmol) in DMF (5 mL) was added EDC-HCI (72 mg, 0.38 mmol), HOBt (69 mg, 0.48 mmol), and DMAP 5 (cat.), and the stirring was continued overnight at rt. The mixture was diluted with EtOAc (50 mL), washed with 1M NaOH, 1M HC1, and brine, dried over anhydrous MgSO4, and concentrated under a reduced pressure. The residue was dissolved in THF-MeOH-H 2 0 (21 mL, 1:1:1, v/v/v) and the stirring was continued for 6 h at rt. The mixture was poured into IM HCI and extracted with CHCl 3 -MeOH (9:1, v/v). The combined organic phase was dried over anhydrous MgSO4, 10 and concentrated under a reduced pressure. The residue was purified with TLC (Whatman, PLK 5F, CHC13/MeOH, 20:1, v/v) to give 98 (69 mg, 51%) as a white powder. MW 541.52 IR-ATR: 3351, 1604, 1537, 1167, 754 (cm-'); 'H-NMR (DMSO) 8 2.25 (s, 3 H), 2.32 (min, 2 H), 3.68-4.70 (min, 7 H), 5.32-5.55 (min, 1 H), 6.97 (t, J= 7.6 Hz, 1 H), 7.06 (d, J= 8.5 Hz, 2 H), 7.20 (mn, 3 H), 7.87 (d, J= 8.8 Hz, 2 H), 7.83-8.04 (min, 2 H), 8.45 (s, 1 H), 9.18 (s, 1 H); MS (ESI) m/z 564 15 (M+Na); Anal. Calcd for C 28
H
26
F
3
N
3 Os-l.75 H20: C, 58.69; H, 5.19; N, 7.33. Found: C, 58.54, H, 4.85; N, 6.98. Example 93 4-[1-[4-[N '-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-4-fluoro-2-pyrrolidinyl]methylamino benzoic acid F N N OOH 20 HH Me 99 4-[1 -[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-4-fluoro-2-pyrrolidinyl]methylamino benzoic acid F N N N 0- D?-OOH r NH Me OOH 100 To a stirred solution of methyll-tert-butoxycarbonyl-4-fluoropyrrolidine-2-carboxylate(1.2 g, 4.85 25 mmol) in MeOH (5 ml) was added IN NaOH (5 ml) and the mixture was stirred at room temperature for 1hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HC1. The mixture was extracted with EtOAc. The extract was washed with water, then dried over Na 2
SO
4 , and concentrated in vacuo to give 1-tert-butoxycarbonyl-4 241 WO 01/00206 PCT/US00/18079 fluoropyrrolidine-2-carboxylic acid (1.1 g, quant) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.47 (br s, 9H), 2.78-2.83 (br s, 3H), 4.37 (s, 2H), 6.73-6.76 (min, 3H), 7.17 (min, 1H). To a stirred solution of 1-tert-butoxycarbonyl-4-fluoropyrrolidine-2-carboxylic acid (1.1 g, 4.7 mmol) in THF (10.0 ml) was added BH 3 -THF(1.0 M solution in THF, 10.0 ml, 10.0 mmol) 5 at 0 0 C. After stirred at room temperature for 1.0 h. After cooled, the mixture was concentrated in vacuo. Water was added thereto at 0 0 C, and extracted with EtOAc. The extract was washed with water, then dried over Na 2
SO
4 , and concentrated in vacuo to give 1-tert-butoxycarbonyl-4-fluoro 2-pyrrolidinylmethanol (1.0 g, quant) as a a colorless oil. 'H-NMR (CDCl 3 ) 5 1.48 (s, 9H), 2.29 2.39 (min, 1H), 3.38-3.59 (min, 2H), 3.74-3.88 (min, 2H), 4.09-4.14 (min, 2H), 4.85 (mn, 1H), 5.03 (br s, 10 1H), 5.16 (br s, 1H). To a stirred solution of oxalyl chloride (0.28 ml, 2.3 mmol) in CH 2
CI
2 (20.0 ml) was added DMSO (0.39 ml) at -78 'C. After 5 minutes, to the mixture was added 1-tert butoxycarbonyl-4-fluoro-2-pyrrolidinylmethanol (500 mg, 2.28 mmol) in CH 2 C1 2 (5.0 ml). The mixture was stirred for 30 minutes at -78 oC, and triethylamine (1.6 ml) was added. The mixture 15 was stirred for 30 minutes at -78 oC, and stirred for 30 minutes at room temperature. Water was added to the mixture, and extracted with CH 2 C1 2 . The organic layer was dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product, methyl 4-aminobenzoate(302 mg, 2.0 mmol), and AcOH (0.13 ml) in DCE (10 ml) was added NaBH(OAc) 3 (656 mg, 3.09 mmol) at 0 20 oC. The reaction mixture was stirred at room temperature for 18 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 ClZ. The extract was washed with brine, dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with EtOAc- n-hexane (1:3, v/v) as eluent to give methyl 4-(1-tert-butoxycarbonyl-4-fluoro-2-pyrrolidinyl)methylaminobenzoate (541 mg, 77%) as 25 a pale yellow oil. 'H-NMR (CDCI 3 ) 8 1.55-1.59 (min, 1H), 2.16-2.27 (min, 1H), 2.89-3.03 (mn, 2H), 3.19-3.28 (min, 2H), 3.69-3.73 (min, 1H), 3.84 (s, 3H), 5.15 and 5.29 (each s, total 1H), 6.55-6.58 (inm, 2H), 7.84-7.86 (min, 2H). To a stirred solution of methyl 4-(1-tert-butoxycarbonyl-4-fluoro-2-pyrrolidinyl)methylamino benzoate (541 mg, 1.53 mmol) in CH 2 C1 2 (8.0 ml) was added TFA (4.0 ml) at 0 oC. The reaction 30 mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 C1 2 . The extract was washed with brine 242 WO 01/00206 PCT/US00/18079 ,dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product(151 mg, 0.6 mmol), 4-[N'-(2-chlorophenyl)uredio]-3-methoxyphenylacetic acid (201 mg, 0.6 mmol), HOBt (94 mg, 0.7 mmol), and triethylamine (167 41, 1, 1.2 mmol) in THF (10.0 ml) and MeCN (10.0 ml) 5 was added EDC-HCI (173 mg, 0.9 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc(1:2 ,v/v) as eluent to give methyl 4-[1-[4-[N' 10 (2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-4-fluoro-2-pyrrolidinyl]Jmethylaminobenzoate (320 mg, 94%) as an amorphous solid. 'H-NMR (CDCl 3 ) 8 1.80-1.95 (min, 1H), 2.42-2.58 (min, 1H), 3.20-3.51 (min, 3H11), 3.51-3.76 (min, 5H), 3.84 (s, 3H), 3.85-3.98 (min, 1H), 4.67-4.70 (min, 1H), 5.10 and 5.23 (s, each, total 1H), 5.50 (br s, 1H), 6.49-6.52 (min, 2H), 6.78-6.81 (min, 2H), 6.97-7.01 (inm, 1H), 7.14-7.18 (min, 2H), 7.24-7.36 (min, 2H), 7.80-7.82 (m, 2H11), 7.99-8.01 (min, 1H), 8.15-8.18 (inm, 15 1H). To a stirred solution of methyl 4-[ 1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl acetyl]-4-fluoro-2-pyrrolidinyl]methylaminobenzoate (320 mg, 0.56 mmol) in THF (5.0 ml) and MeOH (3.0 ml) was added IN NaOH (0.8 ml, 0.8 mmol). The mixture was stirred at 70 oC for 24 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN 20 HCL. The resulting solid was collected, washed with water, and dried in vacuo to give 99 (280 mg, 90%) as a.white crystalline solid. MW 555.00 mp 132-136 'C; IR (KBr) 3332, 2937, 1602, 1531, 1174, 752 cm'i; 'H-NMR (DMSO-d6) 5 2.00-2.40 (min, 2H), 3.50-3.90 (min, 4H), 3.75-3.85 (inm, 5H), 4.27 (min, 1H), 5.23 and 5.37 (each s, total 1H), 6.51-7.03 (min, 5H), 7.25-7.29 (min, 1H), 7.41 7.44 (min, 1H), 7.64-7.68 (min, 2H), 7.92-8.10 (min, 2H), 8.87-8.94 (min, 2H); Anal. calcd for 25 C 2 8
H
2 8
N
4 0 5 FCl-0.6H 2 0: C, 59.44; H, 5.20; N, 9.90. Found: C, 59.41; H, 5.19; N, 9.72. To a stirred solution of methyl 4-(1-tert-butoxycarbonyl-4-fluoro-2-pyrrolidinyl)methylamino benzoate (541 mg, 1.53 mmol) in CH 2 C1 2 (8.0 ml) was added TFA (4.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 C1 2 . The extract was washed with brine, 30 dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product (151 mg, 0.6 mmol), 4-[N'-(2-bromophenyl)uredio]-3-methoxyphenylacetic acid (227 mg, 0.6 mmol), HOBt (94 243 WO 01/00206 PCT/US00/18079 mg, 0.7 mmol), and triethylamine (167 41, 1.2 mmol) in THF (10.0 ml) and MeCN (10.0 ml) was added EDC-HCI (173 mg, 0.9 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then 5 dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (2:3, v/v) as eluent to give methyl 4-[1-[4-[N' (2-bromophenyl)ureido]-3-methoxyphenylacetyl]-4-fluoro-2-pyrrolidinyl]methylamino benzoate (280 mg, 76%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.80-1.98 (min, 1H), 1.42-1.58 (mn, 1H), 3.20 3.52 (min, 3H), 3.67-3.79 (min, 5H), 3.84 (s, 3H), 3.94-3.97 (min, 1H), 4.68-4.71 (min, 1H), 5.10 and 10 5.23 (each s, total 1H), 5.51 (br s, 1H), 6.50-6.52 (min, 2H1), 6.79-7.07 (min, 5H), 7.25-7.33 (min, 1H), 7.51-7.53 (min, 1H11), 7.80-7.83 (min, 2H), 7.98-8.00 (min, 1H), 8.11-8.14 (min, 1H). To a stirred solution of methyl 4-[ 1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl acetyl]-4-fluoro-2-pyrrolidinyl]methylaminobenzoate (280 ming, 0.46 mmol) in THF (8.0 ml) and MeOH (8.0 ml) was added IN NaOH (2.8 ml, 2.8 mmol). The mixture was stirred at 70 oC for 18 15 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HC1. The resulting solid was collected, washed with water, and dried in vacuo to give 100 (260 mg, 95%) as a white crystalline solid. MW 599.45 mp 131-135 'C; IR (KBr) 3332, 2935, 1602, 1529, 1174 cm-'; 'H-NMR (DMSO-d 6 ) 6 1.95-2.01 (min, 1H1), 2.20-2.35 (min, 1H), 3.10-3.20 (min, 1H), 3.50-3.70 (m,3H), 3.80-3.85 (min, 5H), 4.27 (min, 1H), 5.24 and 5.37 (each s, total 1H), 6.54-6.99 (inm, 20 5H), 7.30-7.33 (min, 1H), 7.58-7.94 (min, 3H), 7.94-7.98 (min, 2H), 8.73 (mn, 1H1), 8.93 (min, 1H1); Anal. calcd for C 28
H
28
N
4 OsBrF-0.7H 2 0: C, 54.95; H, 4.84; N, 9.15. Found: C, 54.98; H, 4.81; N, 8.93. Example 94 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(4R)-fluoro-(2S)-pyrrolidinyl methoxy]benzoic acid N N 0COOH 1 25 Me H H (Me 101 A mixture of methyl 4-[(4R)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (634 mg, 2.50 mmol), 3 methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (787 ming, 2.50 mmol), EDC'HCI (718 mg, 3.75 mmol), HOBt (cat.), DMAP (cat.) and DMF (10 ml) was stirred overnight. The mixture was diluted with EtOAc (300 ml). The solution was washed with brine (2 x 100 ml), dried over 30 MgSO 4 , and concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 244 WO 01/00206 PCT/US00/18079 EtOAc (4:1) as eluent to give methyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-(4R)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (1.37 g, quant) as a pale yellow viscous solid. 'H-NMR (CDCl 3 ) 8 2.24 (s, 3 H), 2.26-2.47 (m, 2 H), 3.46 (s, 3 H), 3.49-3.64 (m, 4 H), 3.87 (s, 3 H), 4.06 (dd, J = 9.5, 2.0 Hz, 1 H), 4.51-4.62 (m, 2 H), 5.20 and 5.33 (br s, each, 5 total 1H), 6.63 (s, 1 H), 6.72 (d, J= 8.3 Hz, 1 H), 6.77 (d, J= 9.0 Hz, 2 H), 7.05 (t, J= 7.6 Hz, 1 H), 7.16-7.20 (m, 3 H), 7.53 (s, 1 H), 7.63 (d, J= 7.8 Hz, 1 H), 7.91 (d, J= 9.0 Hz, 2 H), 8.07 (d, J= 8.1 Hz, 1 H). A mixture of methyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(4R) fluoro-(2S)-pyrrolidinylmethoxy]benzoate (1.37 g, 2.49 mmol), 0.25 N NaOH (20 ml, 5.00 mmol), 10 and THF (20 ml) was stirred for 3 days. The mixture was poured into 1 N HCI (100 ml) and extracted with CHCl 3 -MeOH (5:1, 2 x 200 ml). The combined extracts were dried over MgSO 4 and concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (20:1 to 4:1) to give 101 (930 mg, 70%) as a pale yellow amorphous solid. MW 535.56 'H-NMR (DMSO-d 6 ) 8 2.24-2.41 (m, total 5 H), 3.42-4.66 (series of m, 10 H), 5.31 and 5.44 (br s, each, 15 total 1 H), 6.71-7.16 (series of m, 7 H), 7.79 (d, J= 8.1 Hz, 1 H), 7.85-7.89 (m, 2 H), 7.98-8.00 (m, 1 H), 8.47 (s, 1 H), 8.55 (s, 1 H); MS(FAB) m/z 536 (M++1). Example 95 4-[(4S)-chloro- 1-[3-methoxy-4-[N'-(2-chlorophenyl)ureido]phenylacetyl]-(2S)-pyrrolidinyl] methoxybenzoic acid CI NN COOH 20 Cl H H Me 102 To a stirred solution of methyl 4-(trans-1-tert-butoxycarbonyl-4-hydroxy-(2S)-pyrrolidinyl) methoxybenzoate (351 mg, 1.0 mmol) and PhLP (393 mg, 1.5 mmol) in CHC1 3 (5.0 ml) was added CCl 4 (5.0ml) at room temperature. The reaction mixture was stirred at 50 0 C for 24 hr. The reaction mixture was concentrated in vacuo. The residue was purified by column chromatography 25 on silica gel with n-hexane to n-hexane-EtOAc(4:1, v/v) as eluent to give methyl 4-(cis-1-tert butoxycarbonyl-4-chloro-(2S)-pyrrolidinyl)methoxybenzoate (340 mg, 92%) as a pale yellow oil. 'H-NMR (CDCI 3 ) 8 1.48 (s, 9H), 2.38-2.65 (m, 2H), 3.50-3.60 (m, 1H), 3.88 (s, 3H), 3.89-4.05 (m, 1H), 4.26-4.41 (m, 4H), 6.95-6.97 (m, 2H), 7.98 (d, J= 8.5 Hz, 2H). To a stirred solution of methyl 4-(1-tert-butoxycarbonyl-(4S)-chloro-(2S)-pyrrolidinyl) 245 WO 01/00206 PCT/US00/18079 methoxybenzoate (369mg, 1.0 mmol) in CH 2 C1 2 (3.0 ml) was added TFA (3.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 Cl 2 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to 5 the subsequent reaction without further purification. To a stirred solution of the crude product (185 mg, 0.5 mmol), 4-[N'-(2-chlorophenyl)uredio]-3-methoxyphenylacetic acid (167 mg, 0.5 mmol), HOBt (68 mg, 0.5 mmol), and triethylamine (208ml, 1.5 mmol) in THF (8.0 ml) and MeCN (8.0 ml) was added EDC-HCI (144 mg, 0.75 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, 10 and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (1:2, v/v) as eluent to give methyl 4 [(4S)-chloro-1 -[3-methoxy-4-[N'-(2-chlorophenyl)urcido]phenylacetyl]-(2S)-pyrrolidinyllmethoxy benzoate (210 mg, 72%) as a colorless oil. 'H-NMR (CDCl 3 ) 6 3.35-3.50 (min, 1H), 3.55-3.65 (inm, 15 1H), 3.61-3.66 (min, 3H), 3.75 (s, 3H), 3.88 (s, 3H), 3.99-4.04 (min, 1H), 4.35-4.40 (min, 3H), 4.48 4.53 (min, 1H), 6.77-7.10 (min, 7H), 7.25-7.36 (min, 2H), 7.93-8.00 (min, 2H), 8.18 (d,J= 8.0 Hz, 1H). To a stirred solution of methyl 4-[(4S)-chloro-1-[3-methoxy-4-[N'-(2-chlorophenyl) ureido]phenylacetyl]-(2S)-pyrrolidinyl]methoxybenzoate(210 mg, 0.35 mmol) in THF (6.0 ml) and MeOH (3.0 ml) was added IN NaOH (0.7 ml, 0.7 mmol). The mixture was stirred at 70 oC for 18 20 hr. The mixture was concentrate in vacuo, water was added thereto, and neutralized with IN HC1. The resulting solid was collected, washed with water, and dried in vacuo to give 102 (200 mg, 98%) as a white crystalline solid. MW 572.44 mp 126-131 'C; IR (KBr) 3330, 1685, 1604, 1533, 1438 cm-1; 1 H-NMR (DMSO-d 6 ) 8 2.15-2.25 (min, 1H), 2.58-2.63 (min, 1H), 3.58-3.78 (min, 3H), 3.83 (s, 3H), 4.13-4.42 (min, 4H), 4.73 (min, 1H), 6.75-7.45 (min, 7H), 7.86-8.10 (min, 4H1), 8.90 (s, 1H), 8.95 25 (s, 1H); MS (FAB) m/z 572 (M'+1); Anal. calcd for C 28
H
27
N
3 0 6 Cl: C, 58.75; H, 4.75; N, 7.34. Found: C, 58.93; H, 4.85; N, 7.15. Example 96 4-[ 1-[4-[N'-(2-bromophenyl)ureidol-3-methoxyphenylacetyll-(4S)-chloro-(2S)-pyrroidinyl] methoxybenzoic acid CI NQI 3N N OOCOOH 30 r H H Me 103 To a stirred solution of methyl 4-[1-tert-butoxycarbonyl-(4S)-chloro-(2S)-pyrrolidinyl] 246 WO 01/00206 PCT/US00/18079 methoxybenzoate (369mg, 1.0 mmol) in CH 2 Cl 2 (3.0 ml) was added TFA (3.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 C1 2 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to 5 the subsequent reaction without further purification. To a stirred solution of the crude product (185 mg, 0.5 mmol), 4-[N'-(2-bromophenyl)uredio]-3-methoxyphenylacetic acid (190 mg, 0.5 mmol), HOBt (68 mg, 0.5 mmol), and triethylamine (208ml, 1.5 mmol) in THF (8.0 ml) and MeCN (8.0 ml) was added EDC-HCI (144 mg, 0.75 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, 10 and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (1:2 ,v/v) as eluent to give methyl 4 [1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-(4S)-chloro-(2S)-pyrrffolidinylI] methoxybenzoate (260 mg, 83%) as a colorless oil. 'H-NMR (CDC13) 5 2.32-2.50 (mn, 1H), 2.53 15 2.65 (min, 1H), 3.61-3.67 (min, 3H), 3.75 (s, 3H), 3.88 (s, 3H), 3.99-4.03 (mn, 1H), 4.35-4.40 (mn, 3H), 4.45-4.55 (min, 1H), 6.78-7.10 (min, 7H), 7.28-7.33 (min, 1H), 7.52 (d, J= 8.0 Hz, 1H), 7.94-7.99 (inm, 3H), 8.14 (d, J= 8.3 Hz, 1H11). To a stirred solution of methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl acetyl]-(4S)-chloro-(2S)-pyrrolidinyl]methoxybenzoate (260 mg, 0.4 mmol) in THF (6.0 ml) and 20 MeOH (3.0 ml) was added IN NaOH (0.8 ml, 0.8 mmol). The mixture was stirred at 70 oC for 24 hr. The mixture was concentrate in vacuo, water was added thereto, and neutralized with IN HC1. The resulting solid was collected, washed with water, and dried in vacuo to give 103 (210 mg, 83%) as a white crystalline solid. MW 616.89 mp 127-132 0 C; IR (KBr) 3330, 1685, 1604, 1529, 1434 cmr'; 'H-NMR (DMSO-d 6 ) 8 2.18-2.28 (mn, 1IH), 2.60-2.70 (min, 1H), 3.55-3.75 (min, 3H), 3.83 25 (s, 3H), 4.12-4.42 (min, 4H), 4.60-4.75 (min, 1H), 6.75-7.06 (min, 5H), 7.30-7.34 (mn, 1H), 7.60 (d, J= 7.3 Hz, 1H), 7.86-7.94 (min, 5H), 8.75 (s, 1H), 8.94 (s, 1H); MS (FAB) m/z 616 (M), 618 (MW+2); Anal. calcd for C 28
H
2 7 N30 6 C1Br: C, 54.52; H, 4.41; N, 6.81. Found: C, 54.98; H, 4.54; N, 6.66. Example 97 4-[(4R)-chloro- 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S)-pyrrolidinyl 30 methoxy]benzoic acid 247 WO 01/00206 PCT/US00/18079 N N COOH 104 rH H Me 104 4-[(4R)-chloro-1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(2S)-pyrrolidinyl methoxy]benzoic acid .... C I N N \/ COOH iH H Me 105 5 To a stirred solution of methyl 1-(tert-butoxycarbonyl)-(4S)-hydroxy-(2S)-pyrrolidinylcarboxylate (1.81 g, 7.34 mmol) in CC1 4
-CH
2 C1 2 (20 ml, 1:1, v/v) was added Ph 3 P (3.87 mmol, 14.75 mmol) and the reaction mixture was stirred at room temperature for 2 hr. To the mixture was added EtOH (5 ml) and the reaction mixture was stirred at room temperature overnight. After removal of the solvent, the residue was purified by column chromatography on silica-gel with n-hexane 10 EtOAc (3:1, v/v) as eluent to give Synthesis of methyl 1-(tert-butoxycarbonyl)-(4R)-chloro-(2S) pyrrolidinylcarboxylate (1.36 g, 70%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.42 (s, 9 H), 2.32 2.39 (min, 1 H), 2.49-2.54 (min, 1 H), 3.66-3.92 (series of s and m, total 5 H), 4.44-4.55 (min, 2 H); MS(FAB) m/z 264 (M+1). To a stirred solution of methyl 1-(tert-butoxycarbonyl)-(4R)-chloro-(2S)-pyrrolidinyl 15 carboxylate (1.35 g, 5.12 mmol) in THF (10 ml) was added 0.5 N NaOH (10 ml) and the reaction mixture was heated under reflux for 1.5 hr. After cooled to room temperature, the mixture was poured into ice- 1 N HCI and the mixture was extracted with CHCl 3 -MeOH (9:1, v/v). The extract was was washed with brine, dried over Na 2
SO
4 and evaporated to give 1 -(tert-butoxycarbonyl) (4R)-chloro-(2S)-pyrrolidinylcarboxylic acid (1.28 g, quant.) as a colorless oil. 'H-NMR (CDCl 3 ) 8 20 1.44 (s, 9 H), 2.37-2.54 (min, 2 H), 3.68-3.88 (min, 2 H), 4.42-4.45 (min, 2 H). To a stirred solution of 1-(tert-butoxycarbonyl)-(4R)-chloro-(2S)-pyrrolidinylcarboxylic acid (1.28 g, 5.13 mmol) in THF (20 ml) was added dropwise BH 3 DMS (0.60 ml, 6.33 mmol) via a syringe and the reaction mixture was stirred at room temperature for 1 hr. After removal of the solvent, the residue was dissolved in CH 2 Cl 2 . The solution was washed with H 2 0, brine, dried over 25 Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (50:1, v/v) as eluent to give 1-(tert-butoxycarbonyl)-(4R)-chloroprolinol (0.88 g, 73%) as a colorless oil. 'H-NMR (CDC1 3 ) 8 1.48 (s, 9 H), 1.98 (min, 1 H), 2.26-2.32 (min, 1 H), 3.56 248 WO 01/00206 PCT/US00/18079 3.65 (m, 2 H), 3.77 (m, 2 H), 4.24 (m, 1 H), 4.41-4.46 (m, 2 H); MS(FAB) m/z 236 (M'+1). To a cooled (0 0 C), stirred solution of methyl 4-hydroxybenzoate (560 mg, 3.68 mmol), 1 (tert-butoxycarbonyl)-(4R)-chloroprolinol (870 mg, 3.69 mmol), Ph 3 P (1.16 g, 4.42 mmol) in THF (15 ml) was added DIAD (870 ml, 4.42 mmol) and the reaction mixture was heated under reflux 5 for 10 hr. After cooled to room temperature, the mixture was evaporated. The residue was purified by column chromatography on silica-gel with n-hexane-EtOAc (5:1, v/v) as eluent to give methyl 4-[1-(tert-butoxycarbonyl)-(4R)-chloro-(2S)-pyrrolidinylmethoxy]benzoate (890 mg, 65%) as a white solid. mp 116-120oC; 'H-NMR (CDCl 3 ) 8 1.47 (s, 9 H), 2.39-2.53 (m, 2 H), 3.69-3.70 and 4.13-4.17 (m, total 3 H), 3.88 (s, 3 H), 4.30-4.41 (m, 2 H), 4.50-4.55 (m, 1 H), 6.90-6.92 (m, 2 10 H), 7.96-7.98 (m, 2 H); MS(FAB) m/z 370 (M +1); Anal Calcd for C 18
H
24
CIN
5 Os: C, 58.46; H, 6.54; Cl; 9.59; N, 3.79. Found: C, 58.35; H, 6.56; Cl, 9.75; N, 3.77. To a stirred solution of methyl 4-[1-(tert-butoxycarbonyl)-(4R)-chloro-(2S) pyrrolidinylmethoxy]benzoate (840 mg, 2.27 mmol) in CH 2 C1 2 (10 ml) was added TFA (10 ml) and the reaction mixture was stirred at room temperature overnight. The mixture was 15 concentrated in vacuo and made basic by sat. NaHCO 3 . The mixture was extracted with CHCl 3 , washed with brine, dried over Na 2
SO
4 , and evaporated to give methyl 4-[(4R)-chloro-(2S) pyrrolidinylmethoxy] benzoate (580 mg, 95%) as a white solid. mp 61-64oC; 'H-NMR (CDCl 3 ) 8 1.85 (br s, 1 H), 2.03-2.10 (m, 1 H), 2.29-2.35 (m, 1 H), 3.19-3.31 (m, 2 H), 3.88 (s, 3 H), 3.92 4.06 (m, 3 H), 4.53-4.56 (m, 1 H), 6.91 (d, J= 8.8 Hz, 2 H), 7.98 (d, J = 8.8 Hz, 2 H); MS (FAB) 20 m/z 270 (M+1). A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (385 mg, 1.22 mmol), methyl 4-[(4R)-chloro-(2S)-pyrrolidinylmethoxy]benzoate (330 mg, 1.22 mmol), EDCHCI (281 mg, 1.47 mmol), HOBt (200 mg, 1.48 mmol) and Et 3 N (205 ml, 1.47 mmol) in THF (10 ml) was stirred at room temperature overnight. The mixture was diluted with H 2 0 and extracted with 25 EtOAc. The extract was washed with brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1 to 50:1, v/v) as eluent to give methyl 4-[(4R)-chloro-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S) pyrrolidinylmethoxy]benzoate (670 mg, 97%) as a white foam. 'H-NMR (CDC13) 8 2.28 (s, 3 H), 2.33-2.57 (m, 2 H), 3.50 (s, 3 H), 3.59-3.60 (m, 2 H), 3.75-3.82 (m, 2 H), 3.88 (s, 3 H), 4.06-4.09 30 (m, 1 H), 4.51-4.63 (m, 3 H), 6.65-6.80 (m, 5 H), 7.09-7.13 (m, 1 H), 7.20-7.27 (m, 3 H), 7.56 7.58 (m, 1 H), 7.91-7.93 (m, 2 H), 8.05-8.07 (m, 1 H); MS(FAB) m/z 566 (M +1). 249 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[(4R)-chloro-1-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (480 mg, 0.85 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 2 hr. After cooled to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was 5 collected under a reduced pressure. The crude solid was dissolved in CHC13-MeOH and evaporated. The residue was washed with Et20 to give 104 (355 mg, 76%) as a white amorphous solid. MW 552.02 mp 128-132oC; 'H-NMR (DMSO-d 6 ) 8 2.25 (s, 3 H), 2.29-2.46 (min, 2 H), 3.57 3.73 (min, 2 H), 3.78 (s, 3 H), 3.81-3.99 (min, 2 H), 4.11-4.31 (min, 2 H), 4.43-4.45 and 4.64-4.67 (each m, total 1 H), 4.83-4.85 (min, 1 H), 6.71-7.17 (min, 7 H), 7.78-7.80 (min, 1 H), 7.87-7.91 (mn, 2 H), 10 7.99-8.01 (min, 1 H), 8.47 (s, 1 H), 8.56 (s, 1 H), 12.66 (br s, 1 H); MS(FAB) m/z 552 (M+1); Anal. Calcd for C 29
H
30
CN
3 0 6 3/4H 2 0: C, 61.59; H, 5.61; Cl, 6.27; N, 7.43. Found: C, 61.56; H, 5.51; Cl, 6.68; N, 7.26. A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (400 mg, 1.19 mmol), methyl 4-[(4R)-chloro-(2S)-pyrrolidinylmethoxy]benzoate (320 mg, 1.19 mmol), EDCHCI (275 15 mg, 1.43 mmol), HOBt (195 mg, 1.44 mmol) and Et 3 N (200 ml, 1.43 mmol) in THF (10 ml) was stirred at room temperature overnight. The mixture was diluted with H20 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1, v/v) as eluent to give methyl 4-[(4R)-chloro-l1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(2S) 20 pyrrolidinylmethoxy]benzoate (690 mg, 98%) as a pale yellow foam. 'H-NMR (CDCl 3 ) 8 2.35 2.41 (min, 1 H), 2.49-2.59 (min, 1 H), 3.55 (s, 3 H), 3.57-3.70 (min, 2 H), 3.73-3.86 (m, 2 H), 3.88 (s, 3 H), 4.06-4.09 (min, 1 H), 4.54-4.66 (min, 3 H), 6.67-6.81 (min, 4 H), 6.95-6.99 (min, 1 H), 7.23-7.25 (m, 1 H), 7.29-7.33 (min, 1 H), 7.47-7.49 (min, 2 H), 7.90-7.99 (min, 3 H), 8.18-8.21 (mn, 1 H); MS(FAB) m/z 586 (M+1). 25 To a stirred solution of methyl 4-[(4R)-chloro-1-[4-[N'-(2-chlorophenyl)ureido]-3 methoxyphenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (410 mg, 0.70 mmol) in TIIF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 5 hr. After cooled to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was collected under a reduced pressure. The crude solid was dissolved in CHC13-MeOH and 30 evaporated. The residue was washed with Et20 to give 105 (282 mg, 70%) as an amorphous solid. MW 572.44 mp 131-136 0 C; 'H-NMR (DMSO-d) 8 2.29-2.35 (mn, 1 H), 2.44-2.47 (mn, 1 H), 3.58 3.74 (min, 2 H), 3.78 (s, 3 H), 3.81-3.99 (min, 2 H), 4.10-4.32 (min, 2 H), 4.44-4.46 and 4.66 (each m, 250 WO 01/00206 PCT/US00/18079 total 1 H), 4.84 (m, 1 H), 6.74-7.04 (m, 5 H), 7.26-7.30 (m, 1 H), 7.43-7.45 (m, 1 H), 7.87-7.91 (m, 2 H), 7.96 (d, J = 8.3 Hz, 1 H), 8.09 (d, J = 8.3 Hz, 1 H), 8.90 (s, 1 H), 8.94 (s, 1 H); Anal. Calcd for C 28
H
27
CI
2
N
3 0 6 3/4H 2 0: C, 57.39; H, 4.90; Cl, 11.66; N, 7.17. Found: C, 57.57; H, 4.94; Cl, 11.66; N, 6.89. 5 Example 98 4-[(4S)-hydroxy-1 -[4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S)-pyrrolidinyl]Jmethoxybenzoic acid OH N N \ COOH Me H H Me 106 To a stirred solution of methyl 4-[(4S)-acetoxy-1-tert-butoxycarbonyl-(2S)-pyrrolidinyl]methoxy 10 benzoate (2.31 g, 5.87 mmol) in CH 2 C1 2 (46 ml) was added TFA (10 ml) at room temperature. After 3.5 h stirring, the mixture was concentrated in vacuo. The residue was diluted by the addition of CH 2 C1 2 and 1 N NaOH, which were extracted with CH 2 C2. The combined extracts were washed with brine, dried over Na 2
SO
4 , which was concentrated in vacuo. The residue was chromatographed on silica gel [100 g, CHCl3/MeOH(20/1)] to give tmethyl 4-[(4S)-acetoxy-(2S) 15 pyrrolidinyl]methoxybenzoate (1.89 mg, 100%) as a pale purple solid. 'H-NMR (CDCI 3 ) 8 2.10 (s, 3H), 2.14 (m, 1H), 2.65 (m, 1H), 3.52-3.63 (m, H), 3.89 (s, 3H), 4.18 (m, 1H), 4.28 (d,J= 5.9 Hz, 2H), 5.38 (m, 1H), 6.93 (d, J = 8.8 Hz, 2H), 7.99 (d, J= 8.8 Hz, 2H). A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (343 mg, 1.09 mmol), methyl 4-[(4S)-acetoxy-(2S)-pyrrolidinyl]methoxybenzoate (320 mg, 1.09 mmol), EDC-HCI (313 20 mg, 1.64 mmol), HOBT (222 mg, 1.64 mmol) and Et 3 N (0.76 ml, 5.45 mmol) in DMF (7 ml) was stirred at room temperature for 16 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHCl 3 /Aectone (5/1)], to give methyl 4-[(4S)-acetoxy-1-[3-methoxy-4-[N'-(2-methylphenyl) 25 ureido]phenylacetyl]-(2S)-pyrrolidinyl]methoxybenzoate (520 mg, 81%) as a brown amorphous solid. 'H-NMR (CDCl 3 ) 8 2.00 (s, 3H, one of isomers), 2.03 (s, 3H, one of isomers), 2.28 (m, 5H), 3.54 (s, 1H), 3.58 (s, 2H), 3.64 (s, 1H), 3.67 and 3.69 (each s, 3H, amide isomers), 3.85 (d, J = 5.4 Hz, 1H), 3.88 (s, 3H), 4.04 (t, J = 9.3 Hz, 1H), 5.27-5.34 (m, 1H), 6.51 (m, 1H), 6.76-6.89 (m, 2H), 6.94 (d, J= 8.1 Hz, 1H), 7.14 (m, 1H), 7.25 (m, 4H), 7.53 (d, J= 8.3 Hz, 1H), 7.96 (d, J 30 = 8.0 Hz, 1H), 8.00-8.10 (m, 2H); MS (ESI) nm/z 590 (M++1). 251 WO 01/00206 PCT/US00/18079 To a solution of methyl 4-[(4S)-acetoxy-1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S) pyrrolidinyl]methoxybenzoate (520 mg, 0.882 mmol) in THF (30 ml), 0.25 N NaOH (30 ml) was added. After stirring at room temperature for 2 days, the mixture was extracted with EtOAc. The aqueous layer was acidified with 1 N HCI and extracted with CHCl 3 -MeOH (10/1). The combined 5 extracts were washed with brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was crystallized by the addition of CHC1 3 , EtOH and ether to give 106 (68 mg, 14%) as a colorless powder. MW 533.57 mp 148-152 oC (dec.); IR (KBr) 3356, 2939, 1687, 1604, 1533, 1454, 1255 cm-'; 'H-NMR (DMSO-d 6 ) 8 1.95-2.09 (m, 2H), 2.25 (s, 3H), 3.59 (d, J= 5.9 Hz, 2H), 3.71 (m, 1H), 3.81 and 3.85 (each s, 3H, amide isomers), 4.13-4.47 (m, 4H), 5.19 (br, 10 1H), 6.70-7.21 (m, 7H), 7.79 (d, J= 7.9 Hz, 1H1), 7.86 (d, J= 8.8 Hz, 2H), 8.01 (d, J= 8.3 Hz, 1H), 8.47 (s, 1H), 8.57 (s, 1H); MS (ESI)m/z 533 (M'+1);Anal. Calcd for C 29
H
31
N
3 07lH 2 0O: C, 63.15; H, 6.03; N, 7.62. Found: C, 63.29; H, 5.76; N, 7.46. Example 99 4-[ -[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(4S)-hydroxy-(2S)-pyrrffolidinyl] 15 methoxybenzoic acid OH Nle N N 0 COOH 107 A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (342 mg, 1.02 mmol), methyl 4-[(2S,4S)-4-acetoxy-2-pyrrolidinyl]methoxybenzoate (300 mg, 1.02 mmol), EDC-HCI (293 mg, 1.53 mmol), HOBT (207 mg, 1.53 mmol) and Et 3 N (0.71 ml, 5.10 mmol) in DMF (6 ml) 20 was stirred at room temperature for 15 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHClIAectone (5/1)], to give methyl 4-[(4S)-acetoxy-1-[4-[N'-(2-chlorophenyl)ureido]-3 methoxyphenylacetyl]-(2S)-pyrrolidinyl]methoxybenzoate (510 mg, 82%) as a pale brown 25 amorphous solid. 'H-NMR (CDCl 3 ) 8 2.01land 2.04 (each s, 3H, amide isomers), 2.17 (m, 2H), 3.56-3.66(m, 3H), 3.61 (s, 3H), 3.88 (s, 3H), 3.89(m, 1H), 4.07 (t, J= 9.6 Hz, 1H1), 4.45 (dd, J 9.2, 3.4 Hz, 1H), 4.56 (m, 1H), 5.31-5.39 (m, 1H), 6.80-7.01 (m, 4H), 7.23 (d, J= 8.1 Hz, 4H), 7.34 (d, J = 8.1Hz, 1 H), 7.95 (d, J= 8.5 Hz, 1H1), 8.00 (m, 1H), 8.18 (d, J = 8.3 Hz, 1H); MS (ESI) m/z 610 (M'+1), 612 (M'+3). 30 To a solution of methyl 4-[(4S)-acetoxy-1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl] -(2S)-pyrrolidinyl]methoxybenzoate (510 mg, 0.836 mmol) in THF (30 ml), 0.25 N NaOH (30 ml) 252 WO 01/00206 PCT/US00/18079 was added. After stirring at room temperature for 2 days, the mixture was extracted with EtOAc. The aqueous layer was acidified with 1 N HCI and extracted with CHC13-MeOH (10/1). The combined extracts were washed with brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was crystalized by the addition of EtOH and ether to give 107 5 (22 mg, 5%) as a colorless powder. MW 553.99 mp 138-142 oC (dec.); IR (KBr) 3334, 2939, 1685, 1604, 1533, 1439, 1248 cm-'; 'H-NMR (DMSO-d 6 ) 8 1.93-2.14 (m, 2H), 3.60 (d, J= 5.7 Hz, 2H), 3.71 (m, 111), 3.81 and 3.85 (each s, 3H, amide isomers), 4.14-4.50 (m, 4H11), 5.19 (br, 1H), 6.72 and 6.76 (each m, 1H, amideisomers), 6.85 and 6.90 (each s, 1H, amide isomers), 7.00 7.08 (m, 3H), 7.28 (t, J = 7.3 Hz, 1H), 7.43 (dd, J= 8.1, 1.2 Hz, 1H), 7.86-7.95 (m, 2H), 7.97 (d,J 10 = 8.1 Hz, 1H1), 8.10 (dd, J= 8.3, 1.5 Hz, 1H), 8.90 (s, 1H11), 8.94 (s, 1H11), 12.64 (br, 1H1); MS (ESI) m/z 554 (M*+1), 556 (M'+3); Anal. Calcd for C 28
H
2 8
CIN
3 0 7 : C, 60.71; H, 5.05; Cl, 6.40; N, 7.58. Found: C, 60.47; H, 5.37; Cl, 6.31; N, 7.19. Example 100 4-[(4S)-acetoxy-1-[4-[N'-(2-bromophenyl)ureido]phenylacetyl]-(2S)-pyrrolidinyl]methoxybenzoic 15 acid OH NN COOH 10 r H H Me 108 A mixture of 4-[N'-(2-bormophenyl)ureido]-3-methoxyphenylacetic acid (387 mg, 1.02 mmol), methyl 4-[(4S)-acetoxy-(2S)-pyrrolidinyl]methoxybenzoate (300 mg, 1.02 mmol), EDC-HC (293 mg, 1.53 mmol), HOBT (207 mg, 1.53 mmol) and Et 3 N (0.71 ml, 5.10 mmol) in DMF (6 ml) was 20 stirred at room temperature for 15 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHCl3/Aectone (5/1)], to give methyl 4-[(4S)-acetoxy-1-[4-[N'-(2-bromophenyl)ureido] phenylacetyl]-(2S)-pyrrolidinyl]methoxybenzoate (510 mg, 76%) as a yellow oil. 'H-NMR 25 (CDC13) 8 2.01 and 2.04 (each s, 3H, amide isomers), 2.31 (m, 2H11), 3.54-3.68 (m, 3H11), 3.76 (s, 2H), 3.88 (s, 3H), 3.89-4.58 (m, 4H11), 5.31-5.36 (m, 1H11), 6.81-6.96 (m, 5H11), 7.19-7.32 (m, 3H), 7.51 (d, J= 8.0 Hz, 111), 7.93-8.00 (m, 3H), 8.13 (d, J= 8.3 Hz, 1H); MS (ESI) m/z 654 (M+1), 656 (M+3). To a solution of methyl 4-[(4S)-acetoxy-1-[4-[N'-(2-bromophenyl)ureido]phenylacetyl]-(2S) 30 pyrrolidinyl]methoxybenzoate (510 mg, 0.779 mmol) in THF (30 ml), 0.25 N NaOH (30 ml) was added. After stirring at room temperature for 2 days, the mixture was extracted with EtOAc. The 253 WO 01/00206 PCT/US00/18079 remaining aqueous layer was acidified with 1 N HCI and extracted with CHC1 3 -MeOH (10/1). The combined extracts were washed with brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was crystalized by the addition of EtOH and ether, to give 108 (87 mg, 19%) as a pale brown powder. MW 598.44 mp 143-151 oC (dec.); IR (KBr) 3332, 2937, 5 1685, 1604, 1529, 1529, 1435 cm'; 'H-NMR (DMSO-d 6 ) 8 1.92-2.14 (m, 2H), 3.60 (d, J= 5.9 Hz, 2H), 3.72 (m, 1H), 3.81 and 3.85 (each s, 3H, amide isomers), 4.14-4.49 (m, 4H), 5.19 (br, 1H1), 6.72 and 6.75 (each m, 1H, amide isomers), 6.85 and 6.90 (each m, 1H, amide isomers), 6.97 (t, J= 6.1 Hz, 1H), 7.06 (d, J= 8.8 Hz, 2H), 7.32 (t, J = 7.1 Hz, 1H), 7.60 (dd, J = 7.8, 1.2 Hz, 1H), 7.86 (d, J= 8.8 Hz, 2H11), 7.87-7.97 (m, 3H), 8.74 (s, 1H), 8.93 (s, 1H11), 12.60 (br, 1H); MS 10 (ESI) m/z 559 (M+1), 561 (M'+3); Anal. Calcd for C 28
H
2 8 BrN 3 07-0.1H 2 0: C, 56.03; H, 4.74; Br, 13.31; N, 7.00. Found: C, 55.80; H, 4.84; Br, 13.64; N, 6.66. Example 101 4-[ 1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(4R)-hydroxy-(2S)-pyrrolidinyl methoxy]benzoic acid .... OH 0N N N 0 \ COOH 15 I HH Me 109 To a stirred solution of methyl 4-[(4R)-acetoxy-1-(tert-butoxycarbonyl)-(2S)-pyrrolidinylmethoxy] benzoate (835 mg, 2.12 mmol) in CH 2 Cl 2 (5 ml) was added TFA (5 ml) and the reaction mixture was stirred at room temperature for 1 hr. The mixture was concentrated in vacuo and made basic by sat. NaHCO 3 . The mixture was extracted with CHCl 3 , washed with brine, dried over K 2
CO
3 20 and evaporated to give methyl 4-[(4R)-acetoxy-(2S)-pyrrolidinylmethoxy]benzoate (580 mg, 95%) as a brown oil. 'H-NMR (CDC13) 5 1.86-1.93 (m, 1 H), 2.00-2.12 (series of s and m, total 5 H), 3.03-3.29 (m, 1 H), 3.73-3.80 (m, 1 H), 3.88 (s, 3 H), 3.93-4.01 (m, 2 H), 5.27-5.30 (m, 1 H), 6.91 (d, J = 9.0 Hz, 2 H), 7.98 (d, J= 9.0 Hz, 2 H); MS (FAB) m/z 294 (M'+1). A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (365 mg, 1.09 25 mmol), methyl 4-[(4R)-acetoxy-(2S)-pyrrolidinylmethoxy]benzoate (320 mg, 1.09 mmol), EDCHCI (250 mg, 1.30 mmol), HOBt (180 mg, 1.33 mmol) and Et 3 N (182 ml, 1.31 mmol) in THF (5 ml) was stirred at room temperature for 2 days. The mixture was diluted with H20 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHC13-MeOH (50:1, v/v) 30 as eluent to give methyl 4-[(4R)-acetoxy-1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl acetyl]-(2S)-pyrrolidinylmethoxy]benzoate (500 mg, 75%) as a white foam.
'
H-NMR (CDCl 3 ) 8 254 WO 01/00206 PCT/US00/18079 2.01 (s, 3 H), 2.03-2.05 (min, 1 H), 2.20-2.26 (min, 1 H), 2.37-2.43 (min, 1 H), 3.59 (s, 2 H), 3.62 (s, 3 H), 3.66-3.87 (min, 2 H), 3.89 (s, 3 H), 4.07-4.09 (min, 1 H), 4.48-4.51 (min, 1 H), 4.59 (min, 1 H), 6.70 6.82 (min, 4 H), 6.97-7.01 (min, 1 H), 7.24-7.35 (min, 4 H), 7.92-7.98 (min, 3 H), 8.12-8.21 (mn, 1 H); MS (FAB) m/z 610 (M+ 1). 5 To a stirred solution of methyl 4-[(4R)-acetoxy-1-[4-[N'-(2-chlorophenyl)ureido]-3 methoxyphenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (500 mg, 0.82 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux overnight. After cooled to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was collected under a reduced pressure. The crude solid was purified by recrystallization from 10 CHC1 3 -IPE to give 109 (223 mg, 49%) as a white crystalline powder. MW 553.99 mp 137 142 0 C; 'H-NMR (DMSO-d 6 ) 8 1.95-2.09 (min, 2 H), 3.41-3.43 (min, 1 H), 3.57 (min, 3 H), 3.78 (s, 3 H), 4.07-4.40 (series of m, total 4 H), 5.07 (min, 1 H), 6.72-6.74 (min, 1 H), 6.85 (min, 1 H), 6.99-7.03 (min, 3 H), 7.25-7.29 (min, 1 H), 7.42-7.43 (min, 1 H), 7.85-7.87 (min, 2 H), 7.93-7.95 (mn, 1 H), 8.07 8.09 (min, 1 H), 8.88 (s, 1 H), 8.92 (s, 1 H), 12.65 (br s, 1 H); MS (FAB) m/z 554 (M++1); Anal. 15 Caled for C 28
H
28
CIN
3 0 7 1/2H 2 0: C, 59.73; H, 5.19; Cl, 6.30; N, 7.46. Found: C, 59.58; H, 5.32; Cl, 6.99; N, 7.21. Example 102 4-[(4R)-hydroxy-1-[3-methoxy-4-[N '-(2-methylphenyl)ureido]phenylacetyl]-(2S)-pyrrolidiny methoxy]benzoic acid ... OH 0N? 20N N 0/ COOH 20 Me H H Me 110 A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (320 ming, 1.02 mmol), methyl 4-[(4R)-acetoxy-(2S)-pyrrolidinylmethoxy]benzoate (300 ming, 1.02 mmol), EDCHCI (235 mg, 1.23 mmol), HOBt (166 ming, 1.23 mmol) and Et 3 N (171 ml, 1.23 mmol) in THF (5 ml) was stirred at room temperature for 2 days. The mixture was diluted with H 2 0 and extracted with 25 EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (50:1, v/v) as eluent to give methyl 4-[(4R)-acetoxy-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S) pyrrolidinylmethoxy]benzoate (420 mg, 70%) as a white foam. 'H-NMR (CDC1 3 ) 8 1.99 (s, 3 H), 2.02-2.05 (min, 1 H), 2.15-2.41 (series of s and m, total 5 H), 3.55 (s, 3 H), 3.57 (s, 2 H), 3.63-3.73 30 (min, 2 H), 3.89 (s, 3 H), 4.07-4.10 (min, 1 H), 4.45-4.48 (min, 1 H), 4.57 (mn, 1 H), 6.56 (s, 1 H), 6.66 (min, 1 H), 6.75-6.82 (min, 3 H), 7.11-7.24 (min, 4 H), 7.54-7.56 (min, 1 H), 7.92-7.94 (min, 2 H), 8.05 255 WO 01/00206 PCT/US00/18079 8.07 (m, 1 H); MS (FAB) m/z 590 (M+1). To a stirred solution of methyl 4-[(4R)-acetoxy-1-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (420 mg, 0.71 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux overnight. After 5 cooled to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was collected under a reduced pressure. The crude solid was purified by recrystallization from CHCl 3 -IPE to give 110 (182 mg, 48%) as a white crystalline powder. MW 533.57 mp 178-182oC; 'H-NMR (DMSO-d 6 ) 8 1.92-2.10 (m, 2 H), 2.23 (s, 3 H), 3.40-3.44 (m, 1 H), 3.56-3.67 (m, 3 H), 3.78 (s, 3 H), 4.05-4.39 (series of m, total 4 H), 5.06 (m, 1 H), 6.71-7.01 (m, 5 H), 7.10-7.16 (m, 2 10 H), 7.77-7.79 (m, 1 H), 7.85-7.89 (m, 2 H), 7.98-8.00 (m, 1 H), 8.45 (s, 1 H), 8.54 (s, 1 H), 12.59 (br s, 1 H); MS (FAB) m/z 534 (M+1); Anal. Calcd for C 29
H
31
N
3 0 7 1/2H 2 0: C, 64.20; H, 5.94; N, 7.74. Found: C, 64.35; H, 5.83; N, 7.68. Example 103 4-[(4S)-fluoro- 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinylmethyl]- 1 15 piperazinylacetic acid F N N COOH Me H H OMe 111 To a stirred solution ofN-(tert-butoxycarbonyl) (4S)-fluoroprolinol (1.26 g, 5.75 mmol), Et 3 N (4 ml, 28.5 mmnol) and DMSO (4.1 ml, 57.5 mmol) in CH 2
C
2 (20 ml) was added SO 3 -pyridine (2.74 g, 17.2 mmol). After 5 h stirring, the mixture was evaporated to remove CH 2
C
2 and diluted with 20 Et 2 0 (200 ml). The solution was washed with 1 N HCI (200 ml) and brine (200 ml), dried over MgSO 4 and evaporated. The resulting residue was chromatographed on silica gel with hexane EtOAc (4:1) to give N-(tert-butoxycarbonyl) (4S)-fluoroprolinal (628 mg, 50%) as a yellow oil. 'H-NMR (CDCl 3 ) 81.41-1.47 (m, 9 H), 2.02-2.48 (m, 2 H), 3.47-3.94 (m, 2 H), 4.16 and 4.29 (each d, each J= 9.8 Hz, total 1 H), 5.13 and 5.26 (each s, total 1 H). 25 To a stirred solution of N-(tert-butoxycarbonyl) (4S)-fluoroprolinal (1.44 g, 6.63 mmol), ethyl 1-piperazinylacetate (1.71 g, 9.94 mmol) and AcOH (759 ul, 13.3 mmol) in MeOH (20 ml) was added NaBH 3 CN (880 mg, 13.3 mmol). The reaction mixture was stirred overnight and evaporated. The residue was quenched with sat. NaHCO 3 (100 ml) and evaporated with CHC1 3 (2 x 200 ml). The combined extracts were dried over MgSO 4 and evaporated. The oily residue was 256 WO 01/00206 PCT/US00/18079 chromatographed on silica gel with CHC13-MeOH (20:1) to give ethyl 4-[1-(tert-butoxycarbonyl) (4S)-fluoro-2-pyrrolidinylmethyl]-1-piperazinylacetate (2.38 g, 95%) as a yellow oil. A mixture of ethyl 4-[ 1-(tert-butoxycarbonyl)-(4S)-fluoro-2-pyrrolidinylmethyl]-l piperazinylacetate (2.38 g, 6.37 mmol), TFA (5 ml) and CH 2 Cl 2 (5 ml) was stirred for 3 h. The 5 mixture was evaporated and the residue was made basic with sat. NaHCO 3 (100 ml). The mixture was extracted with CHCl 3 -MeOH (4:1, 2 x 150 ml) and the combined extracts were dried over
K
2
CO
3 and evaporated to give ethyl 4-[(4S)-fluoro-2-pyrrolidinylmethyl]-1-piperazinylacetate (1.44 g, 83%) as a brown oil. 'H-NMR (CDCI 3 ) 5 1.27 (dt, J= 7.1, 2.0 Hz, 3 H), 1.66-3.35 (series of m, 17 H), 4.18 (dq, J = 7.1, 2.0 Hz, 2 H), 5.09 and 5.22 (each m, total 1 H). 10 A mixture of ethyl 4-[(4S)-fluoro-2-pyrrolidinylmethyl]-1-piperazinylacetate (1.44 g, 5.27 mmol), 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (1.66 g, 5.27 mmol), EDC-HCl (1.52 g, 7.91 mmol), HOBt (cat.) and DMAP (cat.) in DMF (10 ml) was stirred overnight. The mixture was diluted with EtOAc-MeOH (10:1, 220 ml). The solution was washed with brine (200 ml), dried over MgSO 4 and evaporated. The residue was chromatographed on 15 silica gel with CHCl 3 -MeOH (20:1) as eluent to give ethyl 4-[(4S)-fluoro-1-[3-methoxy-4-[N'-(2 methylphenyl) ureido]phenylacetyl]-2-pyrrolidinylmethyl]-1-piperazinylacetate (2.47 g, 82%) as a yellow viscous solid. 'H-NMR (CDCl 3 ) 8 1.24-1.29 (min, 3 H), 1.92-4.36 (series of m, 7 H), 5.16 and 5.29 (each m, total 1 H), 6.43 (s, 1 H), 6.74-6.81 (mn, 2 H), 7.12-7.29 (mn, 4 H), 7.50 (d, J= 7.8 Hz, 1 H), 8.01-8.07 (min, 1 H). 20 A mixture of ethyl 4-[(4S)-fluoro-l1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2 pyrrolidinylmethyl]-1-piperazinylacetate (1.0 g, 1.76 mmol) and 0.25 N NaOH (14 ml, 3.50 mmol) in THF (15 ml) was stirred overnight. The mixture was neutralized with 1 N HCI and evaporated. The residue was purified by ion exchage resin (DIAION, HP20) with H20 to MeOH as eluent to give 111 MW 541.61 (400 mg, 40%) as a pale yellow amorphous solid. 'H-NMR (CD 3 OD) 8 25 2.00-3.95 (series of m, 24 H), 4.34-4.40 (min, 1 H), 5.23 and 5.36 (min, each, total 1 H), 6.78-6.82 (inm, 1 H), 6.92 (min, 1 H), 7.00-7.04 (min, 1 H), 7.09-7.23 (min, 4 H), 7.59 (d,J= 7.1 Hz, 1 H), 7.99-8.02 (min, 1 H); MS(FAB) m/z 542 (M+1). Example 104 4-[ -[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-4,4-difluoro-2-pyrrolidinylmethyl]-1 30 piperazinylacetic acid 257 WO 01/00206 PCT/US00/18079 N S COOH 112 N N ' IHH Me 112 To a stirred mixture of 1-(tert-butoxycarbonyl)-4,4-difluoro-2-pyrrolidinylmethanol (2.11 g, 8.89 mmol), Et 3 N (6.2 ml, 44.5 mmol), DMSO (6.3 ml, 88.9 mmol) in CH 2 C1 2 (20 ml) was added
SO
3 -pyridine (4.25 g, 26.7 mmol). After 3 h stirring, the mixture was concentrated in vacuo and 5 diluted with Et 2 0 (200 ml). The resulting mixture was washed with 1 N HCI (100 ml) and brine (100 ml), dried over MgSO 4 , and concentrated in vacuo. The residue was chromatographed on silica gel with hexane-EtOAc (4:1) as eluent to give 1-(tert-butoxycarbonyl)-4,4-difluoro-2 pyrrolidinecarbaldehyde (1.40 g, 67%) as a yellow oil. 'H-NMR (CDCl 3 ) 8 1.45-1.52 (m, 9 H), 2.49 (m, 2 H), 3.75-3.88 (m, 2 H), 4.29-4.42 (m, 1 H), 9.54 and 9.60 (s, each, total 1 H). 10 To a stirred solution of 1-(tert-butoxycarbonyl)-4,4-difluoro-2-pyrrolidinecarbaldehyde (1.40 g, 5.95 mmol) and ethyl 1-piperazinylacetate (1.02 g, 5.95 mmol) in MeOH-AcOH (12:1, 13 ml) was added NaBH 3 CN (787 mg, 11.9 mmol) at 0 0 C. After 3 days stirring, the mixture was quenched by addition of sat. NaHCO 3 (100 ml) and extracted with CHC1 3 (2 x 200 ml). The combined extracts were dried over MgSO 4 and concentrated in vacuo. The residue was chromatographed on silica 15 gel with CHCl 3 -MeOH (20:1) as eluent to give ethyl 4-[1-(tert-butoxycarbonyl)-4,4-difluoro-2 pyrrolidinylmethyl]-1-piperazinylacetate (822 mg, 35%) as a yellow oil. 'H-NMR (CDC13) 6 1.27 (t, J= 7.1 Hz, 3 H), 1.46 (m, 9 H), 1.64 (m, 2 H), 2.39-2.64 (m, 10 H), 3.19 (s, 2 H), 3.42-4.05 (series of m, 3 H), 4.18 (q, J= 7.1 Hz, 2 H). A solution of ethyl 4-[1-(tert-butoxycarbonyl)-4,4-difluoro-2-pyrrolidinylmethyl]-1 20 piperazinylacetate (820 mg, 2.09 mmol) and TFA (5 ml) in CH 2 C1 2 (5 ml) was stirred for 1 h. The mixture was concentrated in vacuo and the residue was made basic with sat. NaHCO 3 . The resulting mixture was extracted with CHCl 3 -MeOH (5:1, 2 x 200 ml). The combined extracts were dried over K 2
CO
3 and concentrated in vacuo to give ethyl 4-(4,4-difluoro-2-pyrrolidinyl methyl)-1 piperazinylacetate (493 mg, 81%) as a brown oil. 1 H-NMR (CDCl 3 ) 8 1.27 (t, J = 7.1 Hz, 3H), 25 1.91 (m, 2 H), 2.27-2.60 (m, 10 H), 3.09-3.34 (m, 4 H), 3.46-3.53 (m, 1 H), 4.19 (q, J= 7.1 Hz, 2 H). A mixture of ethyl 4-(4,4-difluoro-2-pyrrolidinylmethyl)-l-piperazinylacetate (490 mg, 1.69 mmol), 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (567 mg, 1.69 mmol), EDC.HCI (486 mg, 2.54 mmol), HOBt (cat.) and DMAP (cat.) in DMF (10 ml) was stirred 30 overnight. The mixture was diluted with EtOAc (250 ml), washed with brine (2 x 200 ml), dried over MgSO 4 , and concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -EtOAc (4:1) to CHCl 3 -MeOH (10:1) as eluent to give ethyl 4-[1-[4-[N'-(2-chlorophenyl) 258 WO 01/00206 PCT/US00/18079 ureido]-3-methoxyphenylacetyl]-4,4-difluoro-2-pyrrffolidinylmethyl]-1 -piperazinylacetate (973 mg, 95%) as a yellow viscous oil. 'H-NMR (CDCI 3 ) 5 1.25 (t, J = 7.1 Hz, 3 H), 2.31-2.68 (min, 12 H), 3.17-3.20 (min, 2 H), 3.52-3.91 (min, 4 H), 4.10-4.48 (series of m, 3 H), 6.75-6.84 (min, 2 H), 7.00 (dt, J= 7.8, 1.5 Hz, 1 H), 7.16-7.29 (min, 3 H), 7.35 (dd, J= 8.3, 1.5 Hz, 1 H), 8.00 (d,J= 8.3 Hz, 1 H), 5 8.18 (dd, J= 8.3, 1.5 Hz, 1 H). To a stirred solution of ethyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-4,4 difluoro-2-pyrrolidinylmethyl]-1-piperazinylacetate (292 mg, 0.480 mmol) in THF (4 ml) was added 0.25 N NaOH (3.8 ml, 0.960 mmol). After 2 days stirring, the mixture was neutralized with 1 N HCI and extracted with CHCl 3 -MeOH (4:1, 2 x 200 ml). The combined extracts were dried 10 over MgSO 4 and concentrated in vacuo. The residue was purified by tin layer column chromatography on silica gel with CHCl 3 -MeOH (5:1) to give 112 MW 580.02 (81.7 mg, 29%) as a pale yellow amorphous solid. MW 580.02 'H-NMR (DMSO-d6) 8 2.24-2.50 (series of m, 12 H), 3.40-4.47 (series of min, 10H), 6.76 (d,J= 8.1 Hz, 1 H), 6.88 (s, 1 H), 7.02 (t,J= 8.1 Hz, 1 H), 7.28 (t,J= 8.1 Hz, 1 H), 7.44 (d,J=8.1 Hz, 1 H), 7.97 (d,J= 8.1 Hz, 1 H), 8.08 (d,J= 8.1 Hz, 1 15 H), 8.96-8.99 (min, 2 H). MS (FAB) m/z 580 (M +1). Example 105 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(4S)-phenoxy-(2S)-pyrrolidinyl] methyl-1-piperazinylacetic acid o-O N N NN N" COOH13 Me H H Me N C0H113 20 To a stirred mixture of methyl (2S,4R)-1-tert-butoxycarbonyl-4-hydroxy-2-pyrrolidinylcarboxylate (4.69 g, 19.1 nmmol), phenol (1.98 g, 21.0 mmol) and PPh 3 (5.51 g, 21.0 mmol) in THF (80 ml) was added DIAD (4.13 ml, 21.0 mmol) at room temperature under an atmosphere of nitrogen. The mixture was stirred over night. After removal of the solvent, the resulting residue was chromatographed on silica gel [700 g, CHClJ3/EtOAc (10/1)], to give methyl (2S,4S)-1-tert 25 butoxycarbonyl-4-phenoxy-2-pyrrolidinylcarboxylate (5.31 g, 86%) as a colorless oil. 'H-NMR (CDC13) 8 1.43 (br, 9H, one of isomers), 1.48 (br, 9H, one of isomers), 2.48 (min, 1H), 3.75 (br, 3H), 4.42-4.96 (min, 2H), 6.88-7.35 (min, 5H). To a stirred solution of methyl (2S,4S)-1l-tert-butoxycarbonyl-4-phenoxy-2-pyrrolidinylcarboxylate (5.31 g, 16.5 mmol) in THF (132 ml) was added 0.25 N NaOH (132 ml, 33.0 mmol) at room 30 temperature. The resulting mixture was stirred over night. After removal of the solvent, the mixture was acidified by the addition of 1 N HCI and extracted with CHC1 3 . The combined extracts were washed with brine, dried over Na 2
SO
4 and evaporated. The residue was 259 WO 01/00206 PCT/US00/18079 recrystallized from n-hexane-CHCl 3 , to give (2S,4S)-1-tert-butoxycarbonyl-4-phenoxy-2 pyrrolidinylcarboxylic acid (2.96 g, 58%) as a white powder. 'H-NMR (DMSO-d 6 ) 8 1.36 (s, 9H), 2.16 (d, J= 13.2 Hz, 1H), 2.56 (m, 1H), 3.46 (m, 1H), 3.71 (dt, J = 12.0, 5.4 Hz, 1H11), 4.26 (dt, J 9.5, 7.1 Hz, 1H), 4.99 (m, 1H), 6.85 (m, 2H), 6.94 (t, J = 7.3 Hz, 1H), 7.28 (t, J= 7.3 Hz, 1H). 5 To a stirred solution of (2S,4S)-1-tert-butoxycarbonyl-4-phenoxy-2-pyrrolidinylcarboxylic acid (2.39 g, 7.76 mmol) in THF (50 ml) was added BH3-DMS (1.55 ml, 15.5 mmol) at 0 oC. After 10 min. stirring at the same temperature, the mixture was allowed to room temperature and then heated at 50 oC for 2 h. After cooling to room temperature, the mixture was concentrated in vacuo and quenched by the addition of water at 0 oC. The mixture was extracted with EtOAc. The 10 combined extracts were washed with brine, dried over Na 2
SO
4 and evaporated. The residue was chromatographed on silica gel [60 g ,CHCl/MeOH (50/1)] to give (2S,4S)-1-tert-butoxycarbonyl 4-phenoxy-2-pyrrolidinylmethanol (2.83 g, 100%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.47 (s, 9H), 1.95 (br, 1H1), 2.36 (m, 1H), 3.56-3.74 (m, 3H), 3.89-4.52 (m, 3H), 4.85 (br, 1H), 6.84 (dd, J = 8.8, 1.2 Hz, 2H), 6.97 (t, J= 7.2 Hz, 1H), 7.29 (t, 2H, J= 7.8 Hz). 15 To a stirred mixture of (2S,4S)-1-tert-butoxycarbonyl-4-phenoxy-2-pyrrolidinylmethanol (2.75 g, 9.37 mmol), Et 3 N (7.84 ml, 56.2 mmol), DMSO (6.66 ml, 9.37 mmol) in CH2C1 2 (30 ml) at 0 oC was added SO 3 -pyridine (4.47 g, 28.1 nimmol), then the resulting mixture was allowed to raise to room temperature. After 2.5 h stirring, the mixture was concentrated in vacuo. To the resulting mixture was added water and extracted with Et 2 O. The combined extracts were washed with 20 brine, dried over NaSO 4 and concentrated in vacuo. The residue was chromatographed on silica gel [100 g, CHCl3/acetone (5/1)] to give (2S,4S)-1-tert-butoxycarbonyl-4-phenoxy-2-pyrrolidine carbaldehyde (2.54 g, 93%) as a yellow oil. 'H-NMR (CDCl 3 ) 8 1.45 (s, 9H, one of isomers), 1.49 (s, 9H, one of isomers), 2.17 (br, 2H11), 3.65-4.31 (m, 3H), 4.91 (br, 1H1), 6.79 (d, J= 7.8 Hz, 1H), 6.77 (m, 1H), 7.28 (m, 2H), 9.66 (m, 1H). 25 To a stirred mixture of 1-tert-butoxycarbonyl-(4S)-phenoxy-(2S)-pyrrolidinecarbaldehyde (1.36 g, 4.67 mmol), ethyl 1-piperazinylacetate (1.61 g, 9.37 mmol) in THF (30 ml) was added NaBH(OAc) 3 (1.98 g, 9.34 mmol) at room temperature. After 3 h stirring, the mixture was quenched by the addition of water and extracted with EtOAc. The combined extracts were washed with aq. NaHCO 3 and brine. The organic layer was dried over Na 2
SO
4 and concentrated in vacuo. 30 The residue was chromatographed on silica gel [50 g, CHClIMeOH (10/1)], to give ethyl 4-[1 tert-butoxycarbonyl-(4S)-phenoxy-(2S)-pyrrolidinyl]methyl-l-piperazinylacetate (1.05 g, 50%) as a colorless oil. 'H-NMR (CDCI 3 ) 8 1.27 (t, J= 7.3 Hz, 3H), 1.57 (s, 9H), 2.18 (m, 1H1), 2.33-2.74 (m, 9H), 3.17 (s, 2H), 3.52-4.10 (m, 511), 4.17 (q, J= 7.3 Hz, 2H), 4.89 (br, 1H), 6.84 (d, J= 6.8 Hz, 2H), 6.95 (m, 1H1), 7.26 (m, 3H). 260 WO 01/00206 PCT/US00/18079 To a stirred solution of ethyl 4-[1-tert-butoxycarbonyl-(4S)-phenoxy-(2S)-pyrrolidinyl]methylpiperazinylacetate (1.05 g, 2.35 mmol) in CH 2
CI
2 (20 ml) was added TFA (20 ml) at room temperature. After 3 h stirring, the mixture was concentrated in vacuo, which was diluted with CHCl 3 -MeOH (10/1) and made basic by the addition of 1 N NaOH. The combined reaction 5 mixture was extracted with CHCl 3 -MeOH (10/1). The organic layer was washed with brine, dried over NaSO 4 and concentrated, to give ethyl 4-[(4S)-phenoxy-(2S)-pyrrolidinyl]methyl-1 piperazinylacetate (1.12 g, quant.) as a brown oil, which was used without further purification. 'H-NMR (CDC 3 ) 8 1.25 (tt, J= 7.1, 7.1 Hz, 3H11), 1.91 (d,J = 12.0 Hz, 1H11), 2.42-2.85 (mn, 10H), 3.22 (s, 2H), 3.50 (s, 2H), 3.54-3.82 (m, 2H), 4.15 (q,J= 7.1 Hz, 2H), 4.98 (br, 1H), 6.84 (d,J= 10 8.1 Hz, 2H), 6.76 (t, J = 7.1 Hz, 1H11), 7.26-7.31 (min, 3H11), 7.40 (br, 1H). A mixture of 4-[N'-(2-methylphenyl)ureido]-3-methoxyphenylacetic acid (337 mg, 1.07 mmol), ethyl 4-[(4S)-phenoxy-(2S)-pyrrolidinyl]methyl-1-piperazinylacetate (373 mg, 1.07 mmol), EDC-HCI (308 mg, 1.61 mmnol) and DMAP (197 mg, 1.61 mmol) in DMF (6 ml) was stirred at room temperature for 18 h. The mixture was poured into ice water and extracted with EtOAc. 15 The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHClIVMeOH (50/1)], to give ethyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] (4S)-phenoxy-(2S)-pyrrolidinyl]methyl-1-piperazinylacetate (430 mg, 62%) as a colorless amorphous solid. 'H-NMR (CDCl 3 ) 8 1.25 (t, J= 7.8 Hz, 3H11), 1.98-2.17 (min, 2H), 2.26 (s, 3H), 20 2.36-2.82 (min, 1111), 3.13 (s, 1H), 3.17 (s, 1H), 3.55 (d, J= 2.4 Hz, 1), 3.66 (d, J= 0.9 Hz, 3H11), 3.67-3.84 (min, 2H11), 3.98-4.35 (min, 3H11), 4.85-4.95 (min, 1H), 6.27-6.89 (min, 6H11), 7.08 (t, J= 7.3 Hz, 1H), 7.19 (d, J= 6.8 Hz, 1H), 7.28 (min, 2H), 7.41 (d, J= 4.9 Hz, 1), 7.55 (d, J= 7.8 Hz, 1H), 8.05 (dd, J = 7.3, 2.2 Hz, 1H). For HC1 salt: a pale brown amorphous solid. IR (KBr) 3265, 3059, 1747, 1533, 1225 cm-'; MS (FAB) m/z 644 (M'+1); Anal. Calcd for C 3 6H 45
N
5 0 6 -HCl-2.1H20: 25 C, 60.22; H, 7.05; N, 9.75. Found: C, 59.97; H, 6.72; N, 9.54. To a solution of ethyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(4S)-phenoxy (2S)-pyrrolidinyl]methyl-1-piperazinylacetate (240 mg, 0.373 mimol) in THF (3.0 ml), 0.25 N NaOH (3.0 ml) was added. After stirring at room temperature for 20 h, the mixture was neutralized with 1 N HCI and extracted with CHCl 3 -MeOH (10/1). The combined extracts were 30 dried over Na 2 SO4 and concentrated in vacuo. The residue was triturated by the addition of ether, to give 113 MW 615.72 (143 mg, 62%) as a white powder. IR (KBr) 3346, 2949, 1633, 1533, 1227 cm'; 'H-NMR (DMSO-d 6 ) 8 1.76 (min, 1H), 2.18 (br, 2H11), 2.25 (s, 3H), 2.42-2.83 (mn, 9H11), 3.17 (s, 1H), 3.20 (s, 1H), 3.38 (min, 1H11), 3.70 (s, 2H), 3.72-3.78 (min, 2H), 3.85 (s, 3H11, one of isomers), 3.87 (s, 3H, one of isomers),.3.95 (min, 1H), 4.27 (br, 1H), 5.08 (mn, 1H), 6.75 (d, J= 8.3 35 Hz, 1H), 6.93 (min, SH), 7.14 (min, 2H), 7.30 (d, J= 7.6 Hz, 1H), 7.79 (d, J= 9.0 Hz, 1H), 8.02 (inm, 1H1), 8.50 (s, 1H), 8.58 (s, 1H); MS (FAB) m/z 616(M+1); Anal. Calcd for 261 WO 01/00206 PCT/US00/18079
C
34
H
4 1 NsO 6 0.1EtOH-2H 2 0: C, 62.58; H, 7.00; N, 10.67. Found: C, 62.73; H, 6.58; N, 10.24. Example 106 4-[1 -[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(4S)-phenoxy-(2S)-pyrrolidinyl] methyl- 1-piperazinylacetic acid N N N N" 5H H Me N-COOH 114 A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (358 mg, 1.07 mmol), ethyl 4-[(4S)-phenoxy-(2S)-pyrrolidinylmethyl]-1-piperazinylacetate (373 mg, 1.07 mmol), EDC-HCI (308 mg, 1.61 mmol) and DMAP (197 mg, 1.61 mmol) inDMF (6 ml) was stirred at room temperature for 18 h. The mixture was poured into ice water and extracted with EtOAc. 10 The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHCI/MeOH (50/1)] to give ethyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl] (4S)-phenoxy-(2S)-pyrrolidinyl]methyl-1-piperazinylacetate (320 mg, 45%) as a colorless amorphous solid. 'H-NMR (CDCl 3 ) 5 1.25 (qq, J= 7.3, 7.3 Hz, 3H), 2.05-2.20 (m, 2H), 2.35-2.80 15 (m, 1H), 3.12 and 3.18 (each s, 1H, amide isomers), 3.57 (d, J= 5.4 Hz, 1H), 3.66 and 3.38 (each s, 3H, amide isomers), 3.70-3.90 (m, 2H), 4.02-4.43 (m, 3H), 4.88-4.97 (m, 1H), 6.83-6.99 (m, 6H), 7.18-7.32 (m, 3H), 7.68 (d,J= 8.3 Hz, 1H), 7.74 (s, 1H), 8.16 (dd, J= 8.3, 1.4 Hz, 1H). For HCI salt: a pale brown amorphous solid. IR (KBr) 3300, 2978, 1745, 1533, 1225 cm-'; MS (FAB) m/z 664 (M'+1), 666 (M+3); Anal. Calcd for C 3
,H
42 ClN50-HCl-2.4H 2 0: C, 56.51; H, 20 6.48; N, 9.41. Found: C, 56.51; H, 6.18; N, 9.28. To a solution of ethyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(4S)-phenoxy (2S)-pyrrolidinyl]methyl-1-piperazinylacetate (181 mg, 0.273 mmol) in THF (2.2 ml), 0.25 N NaOH (2.2 ml) was added. After stirring at room temperature for 20 h, the mixture was neutralized with 1 N HCI and extracted with CHCl 3 -MeOH (10/1). The combined extracts were 25 dried over Na 2
SO
4 and concentrated in vacuo. The residue was triturated by the addition of ether to give 114 (133 mg, 77%) as a white powder. MW 636.14 IR (KBr) 3317, 2949, 1701, 1631, 1595, 1225 cm-'; 'H-NMR (DMSO-d) 8 2.13-3.05 (m, 11H), 3.22 and 3.36 (each s, 2H, amide isomer), 3.38 (m, 1H), 3.60 (s, 2H), 3.71 (m, 1H), 3.85 (s, 3H), 3.95 (m, 1H), 4.28 (br, 1H), 5.06 (m, 1H), 6.76 (d, J = 8.3 Hz, 1H), 6.91-7.03 (m, 5H), 7.29 (m, 3H), 7.44 (d, J = 7.9 Hz, 1H), 7.97 30 (dd, J= 8.1, 4.1 Hz, 1H), 8.08 (d, J= 7.0 Hz, 1H), 8.91 (s, 1H), 8.95 (s, 1H); MS (FAB) m/z 636(M++1), 638(M+3); Anal. Calcd for C 3 3
H
38 CINsO 6 -0.2EtOH1.3H 2 0: C, 59.98; H, 6.30; N, 10.47. Found: C, 60.25; H, 6.12; N, 10.11. 262 WO 01/00206 PCT/US00/18079 Example 107 4-[1 -[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl] -(4S)-phenoxy-(2S)-pyrrolidinyl] methyl-1-piperazinylacetic acid N N NN N' r H H Me COOH 115 5 A mixture of 4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (406 mg, 1.07 mmol), ethyl 4-[(4S)-phenoxy-(2S)-pyrrolidinyl]methyl-l1-piperazinylacetate (373 mg, 1.07 mmol), EDC-HCI (308 mg, 1.61 mmol) and DMAP (197 mg, 1.61 mmol) in DMF (6 ml) was stirred at room temperature for 18 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the 10 extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHCl 3 /MeOH (50/1)] to give ethyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl] (4S)-phenoxy-(2S)-pyrrolidinyl]methyl-1-piperazinylacetate (560 mg, 74%) as a colorless amorphous solid. 'H-NMR (CDC 3 ) 8 1.25 (tt, J= 7.1, 7.1 Hz, 3H11), 2.04-2.84 (m, 13H), 3.12 and 3.18 (each s, 1, amide isomers), 3,57 (d, J= 4.4 Hz, 1H), 3.66 and 3.68 (each s, 3H, amide 15 isomers), 3.68-3.87 (m, 3H), 4.05-4.41 (m, 2H11), 4.87-4.96 (m, 1H), 6.76-6.98 (m, 6H), 7.20-7.33 (m, 3H), 7.46 (d, J= 8.1 Hz, 1H11), 7.67 and 7.71 (each s, 1H, amide isomers), 7.78 and 7.81 (each s, 1H, amide isomers), 7.95 (d, J = 8.3 Hz, 1H1), 8.09 (d, J = 8.1 Hz, 1H). For HCI salt: a pale brown amorphous solid. IR (KBr) 3384, 2978, 1745, 1340, 1120 cm-'; MS (FAB) m/z 708 (M+1), 710 (M+3); Anal. Calcd for C 35
H
42 BrN 5 06-HCl-2.5H 2 0: C, 53.20; H,6.12; N, 8.86. Found: C, 20 52.98; H, 5.79; N, 8.66. To a solution of ethyl 4-[ 1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-(4S)-phenoxy (2S)-pyrrolidinyl]methyl-1-piperazinylacetate (330 mg, 0.466 mmol) in THF (3.7 ml), 0.25 N NaOH (3.7 ml) was added. After stirring at room temperature for 20 h, the mixture was neutralized with 1 N HCI and extracted with CHCl 3 -MeOH (10/1). The combined extracts were 25 dried over Na 2 SO4 and concentrated in vacuo. The residue was triturated by the addition of ether, to give 115 (217 mg, 68%) as a white powder. MW 680.59 IR (KBr) 3315, 3095, 2941, 1631, 1529, 1435 cmi'; 'H-NMR (DMSO-d 6 ) 8 1.75 (m, 1H), 2.18 and 2.23 (each s, 2H, amide isomers), 2.30-2.78 (m, 9H), 3.15 (2, 2H11), 3.47 (m, 1H), 3.58 (s, 2H11), 3.60-3.82 (m, 3H), 3.84 (m, 3H), 3.93 (m, 1H11), 4.26 (br, 1H11), 5.08 (m, 1H1), 6.75 (d, J= 7.8 Hz, 1I), 6.98 (m, 5H), 7.30 (m, 3H), 7.60 30 (dd, J = 8.1, 2.2 Hz, 1H1), 7.94 (m, 2H11), 8.75 (s, 1IH), 8.93 (s, 1H); MS(FAB) m/z 680(M +1), 82(M +3); Anal. Calcd for C 33
H
3 BrN 5 0 6 -0.2EtOH-2H 2 0: C, 55.27; H, 6.00; N, 9.65. Found: C, 55.32; H, 5.56; N, 9.25. 263 WO 01/00206 PCT/US00/18079 Example 108 4-[(4S)-(4-carboxyphenoxy)- 1-[3-methoxy-4-[N'-(2-methylphenyl)uriedo]phenylacetyl]-(2S) pyrrolidinylmethoxy]benzoic acid OOH No-- CooH MeH H CMe 116 5 To a stirred solution of methyl 1-(tert-butoxycarbonyl)-(4R)-hydroxy-(2S)-pyrrolidinylcarboxylate (10.4 g, 0.04 mol) and imidazole (8.66 g, 0.13 mol) in DMF (40 ml) was added TBS-Cl (7.03 g, 0.05 mol) and the reaction mixture was stirred at 60oC for 3 hr. After cooled to room temperature, the mixture was diluted with brine, and extracted with Et 2 0O. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on 10 silica-gel with n-hexane-EtOAc (5:1, v/v) as eluent to give methyl 1-(tert-butoxycarbonyl)-(4R) (tert-butyldimethylsilyloxy)-(2S)-pyrrolidinylcarboxylate (15.0 g, 98%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 0.06 (s, 6 H), 0.87 (s, 9 H), 1.41 and 1.46 (each s, 9 H), 1.99-2.03 (m, 1 H), 2.16-2.18 (m, 1 H), 3.31-3.42 (m, 1 H), 3.56-3.63 (m, 1 H), 3.73 and 3.74 (each s, 3 H), 4.31-4.42 (m, 2 H); MS (ESI) m/z 360 (M +1). 15 To a stirred solution of methyl 1-(tert-butoxycarbonyl)-(4R)-(tert-butyldimethylsilyloxy)-(2S) pyrrolidinylcarboxylate (15.0 g, 0.04 mol) in THF (60 ml) was added 1 N NaOH (60 ml) and the reaction mixture was stirred at 60 0 C for 2 hr. After cooled to room temperature, the mixture was concentrated to a small volume, acidified with 1 N HC1, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated to give 1-(tert-butoxycarbonyl)-(4R) 20 (tert-butyldimethylsilyloxy)-(2S)-pyrrolidinylcarboxylic acid (12.8 g, 89%) as a colorless oil. 'H NMR (CDC1 3 ) 8 0.07 and 0.08 (each s, 6 H), 0.87 (s, 9 H), 1.49 (s, 9 H), 2.06-2.11 (m, 1 H), 2.41 2.44 (m, 1 H), 3.40-3.59 (m, 2 H), 4.36-4.50 (m, 2 H); MS (ESI) m/z 346 (M +1). To a cooled (0OC) stirred solution of 1-(tert-butoxycarbonyl)-(4R)-(tert-butyldimethylsilyloxy) (2S)-pyrrolidinylcarboxylic acid (12.8 g, 0.04 mol) in THF (150 ml) was added dropwise BH 3 DMS 25 (5.30 ml, 0.06 mol) and the reaction mixture was stirred at room temperature overnight. The reaction mixture was quenched by sat. NH 4 Cl, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with toluene-acetone (5:1, v/v) as eluent to give 1-(tert-butoxy carbonyl)-(4R)-(tert-butyldimethylsilyloxy)-(2S)-prolinol (10.5 g, 85%) as a colorless oil. 'H-NMR 30 (CDCl 3 ) 8 0.06 (s, 6 H), 0.87 (s, 9 H), 1.47 (s, 9 H), 1.58-1.63 (m, 1 H), 1.93-1.98 (m, 1 H), 3.32 264 WO 01/00206 PCT/US00/18079 3.44 (min, 2 H), 3.51-3.57 (min, 1 H), 3.67-3.71 (min, 1 H), 4.13-4.15 (min, 1 H), 4.27 (min, 1 H), 4.87 4.89 (min, 1 H). To a cooled (0OC), stirred solution of methyl 4-hydroxybenzoate (4.81 g, 0.03 mol), 1-(tert butoxycarbonyl)-(4R)-(tert-butyldimethylsilyloxy)-(2S)-prolinol (10.5 g, 0.03 mol), and Ph 3 P (9.96 5 g, 0.04 mol) in THF (160 ml) was added dropwise DIAD (7.48 ml, 0.04 mol) and the reaction mixture was heated under reflux for 7 hr. After cooled to room temperature, the mixture was evaporated. The residue was purified by column chromatography on silica-gel with n-hexane EtOAc (6:1, v/v) to give methyl 4-[1-(tert-butoxycarbonyl)-(4R)-(tert-butyldimethylsilyloxy)-(2S) pyrrolidinylmethoxy]benzoate (9.58 g, 65%) as a white solid. mp 86-88 0 C; 'H-NMR (CDCl 3 ) 8 10 0.08 (s, 6 H), 0.88 (s, 9 H), 1.46 (s, 9 H), 2.04-2.15 (min, 2 H), 3.29-3.48 (min, 2 H), 3.88 (s, 3 H), 4.06-4.30 (min, 3 H), 4.46-4.51 (min, 1 H). 6.91-6.93 (min, 2 H), 7.96-7.98 (mn, 2 H); MS (ESI) m/z 466 (M++1). To a cooled (0oC), stirred solution of methyl 4-[1-(tert-butoxycarbonyl)-(4R)-(tert-butyldimethyl silyloxy)-(2S)-pyrrolidinylmethoxy]benzoate (1.49 g, 3.20 mmol) in THF (15 ml) was added TBAF 15 (6.40 ml, 6.40 mmol, 1 M solution in THF) and the reaction mixture was stirred at room temperature for 2 hr. The mixture was diluted with EtOAc, washed with H 2 0, brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with toluene-acetone (5:1, v/v) as eluent to give methyl 4-[1-(tert-butoxycarbonyl)-(4R)-hydroxy-(2S) pyrrolidinylmethoxy]benzoate (1.05 g, 93%) as a white solid. mp 103-105oC; 'H-NMR (CDCl 3 ) 8 20 1.46 (s, 9 H), 2.11-2.28 (min, 2 H), 3.49-3.60 (min, 2 H), 3.88 (s, 3 H), 4.15-4.34 (min, 3 H), 4.53-4.57 (min, 1 H), 6.91 (d, J = 8.6 Hz, 2 H), 7.97 (d, J = 8.6 Hz, 2 H); MS (ESI) m/z 352 (M++1). To a cooled (0OC), stirred solution of methyl 4-hydroxybenzoate (0.56 g, 3.68 mmol), methyl 4-[1 (tert-butoxycarbonyl)-(4R)-hydroxy-(2S)-pyrrolidinylmethoxy]benzoate (1.30 g, 3.70 mmol), and Ph 3 P (1.16 g, 4.42 mmol) in THF (20 ml) was added dropwise DIAD (0.87 ml, 4.42 mmol) and 25 the reaction mixture was stirred at room temperature for 3 hr. The mixture was evaporated and the residue was purified by column chromatography on silica-gel with n-hexane-EtOAc (6:1, v/v) as eluent to give methyl 4-[1-(tert-butoxycarbonyl)-(4S)-(4-methoxycarbonylphenoxy)-(2S) pyrrolidinylmethoxy]benzoate (1.80 g, q.y.) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.49 (s, 9 H), 2.31-2.38 (min, 1 H), 2.45-2.49 (min, 1 H), 3.64-3.77 (min, 2 H), 3.88 (s, 6 H), 4.07-4.15 (m, 1 H), 30 4.33-4.44 (min, 2 H), 4.95-5.01 (min, 1 H), 6.85 (d, J = 8.8 Hz, 2 H), 6.94 (br s, 2 H), 7.97 (d, J = 8.8 Hz, 4 H); MS (ESI) m/z 486 (M++1). 265 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[1-(tert-butoxycarbonyl)-(4S)-(4-methoxycarbonyl phenoxy)-(2S)-pyrrolidinylmethoxy]benzoate (1.80 g, 3.71 mmol) in CH 2
C
2 (15 ml) was added TFA (15 ml) and the reaction mixture was stirred at room temperature for 1.5 hr. The mixture was concentrated in vacuo, made basic by sat. NaHCO 3 , and extracted with CHCI 3 . The extract 5 was washed with brine, dried over K 2
CO
3 , and evaporated to give methyl 4-[(4S)-(4 methoxycarbonyl phenoxy)-(2S)-pyrrolidinylmethoxy]benzoate (1.50 g, q.y.) as a pale yellow oil. 'H-NMR (CDC 3 ) 3 1.87-1.92 (m, 1 H), 2.41-2.48 (m, 1 H), 3.18-3.23 (m, 1 H), 3.34-3.37 (m, 1 H), 3.60-3.66 (m, 1 H), 3.88 (s, 3 H), 3.89 (s, 3 H), 4.04-4.13 (m, 2 H), 4.94-5.00 (m, 1 H), 6.87 6.93 (m, 4 H), 7.96-8.00 (m, 4 H); MS (ESI) m/z 386 (M++1). 10 A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (400 mg, 1.27 mmol), methyl 4-[(4S)-(4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxy]benzoate (491 mg, 1.27 mmol), EDCHC1 (293 mg, 1.53 mmol), HOBt (207 mg, 1.53 mmol), and Et 3 N (215 pl, 1.54 mmol) in THF (10 ml) was stirred at room temperature overnight. The mixture was diluted with
H
2 0, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and 15 evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (60:1 to 50:1, v/v) as eluent to give methyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)uriedo] phenylacetyl]-(4S)-(4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxy]benzoate (532 mg, 61%) as a white foam. 'H-NMR (CDCl 3 ) 5 2.26-2.49 (series ofs and m, total 5 H), 3.56-3.93 (series of s and m, total 13 H), 4.07-4.59 (series of m, total 3 H), 5.01 (m, 1 H), 6.69-6.94 (m, 7 20 H), 7.09-7.13 (m, 1 H), 7.20-7.31 (m, 3 H), 7.52-7.57 (m, 1 H), 7.92-8.00 (m, 4 H), 8.06-8.09 (m, 1 H); MS (ESI) m/z 682 (M+ 1). To a stirred solution of methyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)uriedo]phenylacetyl]-(4S) (4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxy]benzoate (532 mg, 0.78 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 5 hr. 25 After cooled to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was collected. The crude solid was recrystallized from MeOH-CHC13-Et 2 0 to give 116 (125 mg, 25%) as a pale yellow crystalline powder. MW 653.68 mp 154-159oC; 'H-NMR (DMSO-d 6 )2.24 (s, 3 H), 2.38-2.49 (m, 2 H), 3.63 (s, 2 H), 3.67-3.88 (series of s and m, total 4 H), 4.01-4.06 and 4.15-4.19 (each m, total 2 H), 4.27-4.31 and 4.38-4.42 (each m, total 2 H), 5.18 30 5.25 (m, 1 H), 6.72-6.77 (m, 1 H), 6.85-7.16 (series of m, total 8 H), 7.78-7.89 (m, 5 H), 7.99-8.02 (m, 1 H), 8.46 (s, 1 H), 8.57 (s, 1 H), 12.65 (br s, 2 H); MS (ESI) m/z 654 (M++1); Anal Calcd for
C
36
H
35
N
3 01g/2H 2 0: C, 65.25; H, 5.48; N, 6.34. Found: C, 65.29; H, 5.54; N, 6.20. 266 WO 01/00206 PCT/US00/18079 Example 109 4-[(4S)-(4-carboxyphenoxy)- 1-[4-[N'-(2-chlorophenyl)uriedo]-3-methoxyphenylacetyl]-(2S) pyrrolidinylmethoxy]benzoic acid N NQ o-C-OOH I COOHH Me 117 5 A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (420 mg, 1.25 mmol), methyl 4-[(4S)-(4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxy]benzoate (483 mg, 1.25 mmol), EDCHCI (288 mg, 1.50 mmol), HOBt (203 mg, 1.50 mmol), and Et 3 N (210 p1, 1.51 mmol) in THF (10 ml) was stirred at room temperature overnight. The mixture was diluted with
H
2 0, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and 10 evaporated. The residue was purified by column chromatography on silica-gel with CHCI 3 -MeOH (50:1, v/v) as eluent to give methyl 4-[1-[4-[N'-(2-chlorophenyl)uriedo]-3-methoxyphenylacetyl] (4S)-(4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxy]benzoate (488 mg, 55%) as a white foam. 'H-NMR (CDCl 3 ) 8 2.28-2.51 (m, 2 H), 3.62-3.94 (series of s and m, total 13 H), 4.07-4.62 (series of m, total 3 H), 4.99-5.03 (m, 1 H), 6.78-6.99 (m, 7 H), 7.23-7.34 (m, 2 H), 7.42-7.52 (m, 15 2 H), 7.92-8.01 (m, 5 H), 8.17-8.20 (m, 1 H); MS (ESI) m/z 702 (M'+1). To a stirred solution of methyl 4-[1-[4-[N'-(2-chlorophenyl)uriedo]-3-methoxyphenylacetyl]-(4S) (4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxy]benzoate (488 mg, 0.70 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 3 hr. After cooled to room temperature, the mixture was poured into ice-1 N HCI and the resulting 20 precipitate was collected. The crude solid was recrystallized from MeOH-CHC13-Et 2 0 to give 117 (137 mg, 29%) as a white crystalline powder. MW 674.10 mp 150-153oC; 'H-NMR (DMSO-d 6 ) 8 and 4.17-4.21 (each m, total 2 H), 4.30-4.34 and 4.40-4.45 (each m, total 2 H), 5.20-5.27 (m, 1 H), 6.77-6.81 (m, 1 H), 6.89-6.92 (m, 1 H), 7.01-7.08 (m, 5 H), 7.27-7.31 (m, 1 H), 7.43-7.46 (m, 1 H), 7.86-7.92 (m, 4 H), 7.97-8.00 (m, 1 H), 8.10-8.12 (m, 1 H), 8.91 (s, 1 H), 8.96 (s, 1 H), 12.65 25 (br s, 2 H); MS (ESI) m/z 674 (M+1); Anal Calcd for C 35
H
32
CIN
3 01l/4H 2 0: C, 61.95; H,4.83; N,6.19; C1,5.22. Found: C,61.77; H,4.86; N,6.13; Cl, 5.49. Example 110 4-[ -[4-[N '-(2-bromophenyl)uriedo]-3-methoxyphenylacetyl]-(4S)-(4-carboxyphenoxy)-(2S) pyrrolidinylmethoxy]benzoic acid 267 WO 01/00206 PCT/US00/18079 s0 0oH OOH N N'Q ' a COOH 18 r H H 6Me 118 A mixture of 4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (464 mg, 1.22 mmol), methyl 4-[(4S)-(4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxy]benzoate (472 mg, 1.22 mmol), EDCHC1 (282 mg, 1.47 mmol), HOBt (200 mg, 1.48 mmol), and Et 3 N (205 4d, 1.47 5 mmol) in THF (10 ml) was stirred at room temperature overnight. The mixture was diluted with H20, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (60:1 to 50:1, v/v) as eluent to give methyl 4-[1-[4-[N'-(2-bromophenyl)uriedo]-3-methoxyphenyl acetyl]-(4S)-(4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxy]benzoate (379 mg, 41%) as a 10 white foam. 'H-NMR (CDC13) 8 2.28-2.51 (mn, 2 H), 3.59-3.95 (series of s and m, total 13 H), 4.07-4.62 (series of m, total 3 H), 4.99-5.03 (mn, 1 H), 6.79-6.95 (min, 7 H), 7.27-7.36 (mn, 3 H), 7.49-7.51 (mn, 1 H), 7.93-8.01 (mn, 5 H), 8.11-8.14 (min, 1 H); MS (ESI) m/z 747 (1W+1). To a stirred solution of methyl 4-[1-[4-[N'-(2-bromophenyl)uriedo]-3-methoxyphenylacetyl]-(4S) (4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxy]benzoate (379 mg, 0.51 mmol) in THF (5 15 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 5 hr. After cooled to room temperature, the mixture was poured into ice-1 N HC1 and the resulting precipitate was collected. The crude solid was purified by preparative TLC to give 118 (51 mg, 14%) as a pale yellow amorphous solid. MW 718.55 'H-NMR (DMSO-d6) S 2.20-2.40 (min, 2 H), 3.65-3.89 (series of m, total 6 H), 4.02-4.63 (series of m, total 4 H), 5.19-5.26 (mn, 1 H), 6.74-7.06 20 (min, 7 H), 7.30-7.34 (min, 1 H), 7.59-7.61 (min, 1 H), 7.83-7.96 (min, 6 H), 8.74 (s, 1 H), 8.93 (s, 1 H); Anal. Calcd for C 3
,H
3 2 BrN 3 0 9 2H 2 0: C, 55.71; H, 4.81; N, 5.57. Found: C, 55.92; H, 4.80; N, 5.30. Example 111 4-[(4S)-(4-carboxyphenoxy)-I-[4-[N'-(2-methylphenyl)uriedo]phcnylacetyl]-(2S)-pyrrolidinyl methoxy]benzoic acid 0QCOOH N NN OOH 25 Me H H 119 A mixture of 4-[N'-(2-methylphenyl)ureido]phenylacetic acid (328 mg, 1.15 mmol), methyl 4 [(4S)-(4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxy]benzoate (444 mg, 1.15 mmol), 268 WO 01/00206 PCT/US00/18079 EDCHCI (265 mg, 1.38 mmol), HOBt (187 mg, 1.38 mmol), and Et 3 N (195 pd, 1.40 mmol) in THF (10 ml) was stirred at room temperature overnight. The mixture was diluted with 1H120, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (60:1 to 5 50:1, v/v) as eluent to give methyl 4-[(4S)-(4-methoxycarbonylphenoxy)-1-[4-[N'-(2-methyl phenyl)uriedo]phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (332 mg, 44%) as a white foam. 'H-NMR (CDC1 3 ) 8 2.13 (s, 3 H), 2.24-2.48 (min, 2 H), 3.52-3.90 (series of s and m, total 10 H), 4.05-4.58 (series of m, total 3 H), 5.01 (min, 1 H), 6.78-6.90 (min, 4 H), 6.98-7.20 (min, 8 H), 7.51-7.56 (min, 2 H), 7.90-8.00 (min, 4 H); MS (ESI) m/z 652 (M++1). 10 To a stirred solution of methyl 4-[(4S)-(4-methoxycarbonylphenoxy)-1-[4-[N'-(2-methyl phenyl)uriedo]phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (332 mg, 0.51 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 3 hr. After cooled to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was collected. The crude solid was recrystallized from MeOH-CHCI 3 -Et 2 0 to give 119 15 (118 mg, 37%) as a white crystalline powder. MW 623.65 mp 157-160oC; 'H-NMR (DMSO-d 6 ) 8 2.20-2.25 (series of s and min, total 4 H), 2.39-2.47 (min, 1 H), 3.64 (s, 2 H), 3.68-3.89 (min, 1 H), 4.02-4.08 and 4.16-4.20 (each m, total 2 H), 4.29-4.33 and 4.39-4.43 (each m, total 2 H), 5.20 5.26 (min, 1 H), 6.92-6.96 (min, 1 H), 7.02-7.08 (min, 4 H), 7.12-7.18 (min, 4 H), 7.39-7.41 (mn, 2 H), 7.84-7.92 (min, 6 H), 9.01 (s, 1 H), 12.65 (br s, 2 H); MS (ESI) m/z 624 (M++1); Anal. Calcd for 20 C 35
H
33
N
3 0 8 1H 2 0: C, 65.51; H, 5.50; N, 6.55. Found: C, 65.48; H, 5.36; N, 6.52. Example 112 4-[4-(2,4-difluorophenoxy)-1-[3-methoxy-4-[N' -(2-methylphenyl)ureido]phenylacetyl]-2 pyrrolidinylmethoxybenzoic acid F a COOH 120 N< NN 0 -7--co MeH H M e 120 25 To a stirred solution of methyl 1-(tert-butoxycarbonyl)-4-hydroxyproline (4.0 g, 16.3 mmol), PhzP (5.14 g, 19.6 mmol), and 2,4-difluorophenol (2.55 g, 19.6 mmol) in THF (50 ml) was added DIAD (3.9 ml, 19.6 mmol), and the mixture was heated at reflux for 3 h. After cooling to room temperature, the mixture was concntrated in vacuo and the residue was chromatographed on silica gel with CHCl 3 -EtOAc (4:1) to give methyl 1-(tert-butoxycarbonyl)-4-(2,4-difluorophenoxy) 30 pyrrolidine-2-carboxylate (5.82 g, quant) as yellow oil. 269 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 1-(tert-butoxycarbonyl)-4-(2,4-difluorophenoxy)pyrrolidine-2 carboxylate (5.82 g, 16.3 mmol) in THF (130 ml) was added 0.25 N NaOH (130 ml, 32.6 mmol). The resulting mixture was stirred overnight. The mixture was poured into 1 N HCI (100 ml) and extracted with CHC1 3 (2 x 200 nml). The extracts were dried over MgSO 4 and evaporated. The 5 residue was chromatographed on silica gel with CHCl 3 -EtOAc (4:1) as eluent to give 1-(tert butoxycarbonyl)-4-(2,4-difluorophenoxy)pyrrolidine-2-carboxylic acid (2.55 g, 46%) as a colorless foam. 'H-NMR (CDC1 3 ) 8 1.42-1.47 (m, 9 H), 2.29-2.74 (series of m, 2 H), 3.66-3.71 (m, 2 H), 4.46-4.51 (m, 1 H), 4.83 (m, 1 H), 6.73-6.95 (m, 3 H). To a stirred solution of 1-(tert-butoxycarbonyl)-4-(2,4-difluorophenoxy)pyrrolidine-2-carboxylic 10 acid (2.55 g, 7.43 mmol) in THF (50 ml) was added BH 3 "DMS (452 ul, 7.43 mmol). The mixture was heated at reflux overnight. After cooling to room temperature, the mixture was concentrated in vacuo and quenced by the addition of HO 2 0 (100 ml). The mixture was extracted with CHC1 3 (2 x 200 ml), dried over MgSO 4 , and evaporated. The residue was chromatographed on silica gel with CHCl 3 -EtOAc (4:1) as eluent to give 1-(tert-butoxycarbonyl)-4-(2,4-difluorophenoxy)-2 15 pyrrolidinylmethanol (1.76 g, 72%) as a colorless oil. 'H-NMR (CDC13) 5 1.45 (s, 9 H), 2.28-2.36 (m, 2 H), 3.58-4.99 (series of m, 8 H), 6.74-6.90 (m, 3 H). To a stirred solution of 1-(tert-butoxycarbonyl)-4-(2,4-difluorophenoxy)-2-pyrrolidinylmethanol (500 mg, 1.52 mmol), methyl 4-hydroxybenzoate (277 mg, 1.82 mmol), and Ph 3 P (477 mg, 1.82 mmol) in THF (10 ml) was added DIAD (358 ul, 1.82 nmmol), and the mixture was heated at reflux 20 for 5 h. After cooling to room temperature, the mixture was concentrated in vacuo and the residue was chromatographed on silica gel with CHC13-EtOAc (20:1) as eluent to give methyl 4-[1-(tert butoxycarbonyl)-4-(2,4-difluorophenoxy)-2-pyrrolidinylmethoxy]benzoate (529 mg, 75%) as a colorless oil. 1 H-NMR (CDCl 3 ) 8 1.46 (s, 9 H), 2.20-2.47 (m, 2 H), 3.64 (m, 2 H), 3.86 (s, 3 H), 4.07-4.43 (m, 3 H), 4.86 (m, 1 H), 6.74-6.87 (m, 3 H), 6.94 (d, 2 H, J = 8.5 Hz), 7.95 (d, 2 H, J= 25 8.5 Hz). To a stirred solution of methyl 4-[1-(tert-butoxycarbonyl)-4-(2,4-difluorophenoxy)-2-pyrrolidinyl methoxy]benzoate (529 mg, 1.15 mmol) in CH 2 C1 2 (5 ml) was added TFA (5 ml). The mixture was stirred overnight. The mixture was concentrated in vacuo and the residue was made basic by the addition of sat. NaHCO 3 . The mixture was extracted with CHC1 3 (2 x 100 ml). The extracts 30 were dried over K 2
CO
3 and evaporated to give methyl 4-[4-(2,4-difluorophenoxy)-2-pyrrolidinyl methoxy]benzoate (385 mg, 92%) as a yellow oil. 'H-NMR (CDCl 3 ) 8 1.89-1.95 (m, 1 H), 2.28 270 WO 01/00206 PCT/US00/18079 2.35 (m, 1 H), 3.09 (dd, J= 12.5, 4.9 Hz, 1 H), 3.33 (d, J= 12.5 Hz, 1 H), 3.60 (m, 1 H), 3.86 (s, 3 H), 4.10 (d, J= 5.6 Hz, 2 H), 4.84 (m, 1 H), 6.73-6.89 (m, 3 H), 6.91 (d, J= 8.5 Hz, 2 H), 7.96 (d, J= 8.5 Hz, 2 H). A mixture of methyl 4-[4-(2,4-difluorophenoxy)-2-pyrrolidinylmethoxy]benzoate (380 mg, 1.05 5 mmol), 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (329 mg, 1.05 mmol), EDC-HCI (302 mg, 1.58 mmol), and catalytic amount of HOBt and DMAP in DMF (10 ml) was stirred for 3 days. The mixture was diluted with EtOAc (200 ml) and washed with brine (2 x 200 ml). After removal of the solvent, residue was chromatographed on silica gel with CHCl 3 -EtOAc (4:1) to CHC13-MeOH (10:1) as eluent to give methyl 4-[4-(2,4-difluorophenoxy)-1-[3-methoxy-4 10 [N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinylmethoxybenzoate (693 mg, quant). 'H NMR (CDCl 3 ) 82.16-2.53 (m, 5 H), 3.61-4.93 (series of m, 14 H), 6.48-8.12 (series of m, 16 H). To a stirred solution of methyl 4-[4-(2,4-difluorophenoxy)-1-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenylacetyl]-2-pyrrolidinylmethoxybenzoate (693 mg, 1.05 mmol) in THF (8 ml) was added 0.25 N NaOH (8.4 ml, 2.10 mmol). The mixture was stirred overnight. The mixture was 15 poured into 1 N HCI (200 ml) and the resulting precipitate was collected with suction. The solid was chromatographed on silica gel with CHCl 3 -MeOH (50:1 to 10:1) as eluent to give 120 (323 mg, 48%) as a colorless amorphous soid. MW 645.65 'H-NMR (DMSO-d 6 ) 8 2.25 (s, 3 H), 2.35 (m, 2 H), 3.33-5.18 (series of m, 11 H), 6.75 (dd, 1 H, J= 8.3, 1.7 Hz), 6.87-7.30 (series of m, 8 H), 7.79 (d, 1 H, J= 8.3 Hz), 7.85-7.90 (m, 3 H), 8.01 (d, 1 H, J = 8.3 Hz), 8.49 (s, 1 H), 8.57 (s, 1 20 H); MS (FAB) m/z, 646 (M+1). Example 113 4-[ -[4-[N -(2-chlorophenyl)ureidol-3-methoxyphenylacetyl]-4-(6-quinolyloxy-2S-pyrrolidinylI] methoxybenzoic acid SNN ~ N 0 o-CooH i H HO O H 1 2 1 25 To a stirred solution of methyl (trans- -tert-butoxycarbonyl-4-hydroxy-2-pyrrolidinyl) methoxy benzoate (1.0 g, 3.0 mmol), 6-hydroxyquinoline (435 mg, 3.0 mmnol), and Ph 3 P (943 mg, 3.6 mmol) in THF (10 ml) was added DIAD (727 mg, 3.6 mmol) at 0 'C. The reaction mixture was stirred at room temperature for 18 hr. The mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (1:2, v/v). To a 271 WO 01/00206 PCT/US00/18079 stirred solution of the product in CH 2 C1 2 (6.0 ml) was added TFA (6.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2
CI
2 . The extract was washed with brine ,dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column 5 chromatography on silica gel with MeOH-CH 2 C1 2 (1% to 10%,v/v) as eluent to give methyl 4-[(4S) [(6-quinolyloxy-(2S)-pyrrolidinyl)]methoxybenzoate (900 mg, 82%) as a pale yellow oil. 'H-NMR (CDCl 3 ) 8 1.92-2.10 (min, 1H), 2.45-2.55 (min, 1H), 3.20-3.30 (min, 1H), 3.38-3.50 (min, 1H), 3.60-3.70 (min, 1H), 3.88 (s, 3H), 4.05-4.18 (min, 2H), 5.03 (mn, 1H), 6.91 (d,J= 8.5 Hz, 1H), 7.02 (d,J= 2.7 Hz, 1H11), 7.35-7.38 (min, 2H), 7.96 (d, J= 8.5 Hz, 1H), 8.00-8.05 (min, 2H), 8.76 (d, J= 3.2 Hz, 1H). 10 To a stirred solution of methyl 4-(4S-(6-quinolyloxy-2S-pyrrolidinyl)methoxybenzoate (300 mg, 0.79mmol), 4-[N'-(2-chlorophenyl)uredio]-3-methoxyphenylacetic acid (264 mg, 0.79 mmol), HOBt (107 mg, 0.79 mmol), and triethylamine (330 ml, 2.37 mmol) in THF (10.0 ml) and MeCN (10.0 ml) was added EDC-HCI (228 mg, 1.2 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, 15 and extracted with EtOAc. The extract was washed with sat. NaHCO 3 then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with EtOAc to EtOH-EtOAc (10%, v/v) as eluent to give methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3 methoxyphenylacetyl]-4-(6-quinolyloxy-(2S)-pyrrolidinyl]methoxybenzoate (520 mg, 95%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 2.30-2.60 (min, 3H), 3.64 (s, 2H), 3.73 (s, 3H), 3.80-3.95 (mn, 1H1), 20 3.87 (s, 3H), 4.15-4.30 (min, 1H11), 4.50-4.70 (min, 2H), 5.11 (br s, 1H11), 6.81-7.01 (mn, 6H), 7.26 -7.39 (mn, 6H), 7.93-8.03 (mn, 5H), 8.19 (d, J= 8.3 Hz, 1H), 8.80 (s, 1H). To a stirred solution of methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl acetyl]-4-(6-quinolyloxy-2S-pyrrolidinyl]methoxybenzoate (520 mg, 0.75 mmol) in THF (10.0 ml) and MeOH (5.0 ml) was added 1N NaOH (1.5 ml, 1.5 mmol). The mixture was stirred at 60 oC 25 for 18 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HC1. The resulting solid was collected, washed with water, and dried in vacuo to give 121 (450 mg, 88%) as a white crystalline solid. 681.13 mp 129-133 'C; IR (KBr) 3332, 1704, 1604, 1531, 1419, 1222, 1166 cm'; 'H-NMR (DMSO-d) 8 2.25-2.55 (mn, 2H), 3.67 (s, 2H), 3.82 (s, 3H), 3.81-3.92 (min, 1H), 4.02-4.15 (min, 2H), 4.40-4.50 (min, 2H), 5.25-5.40 (min, 1H), 5.33-7.07 (min, 5H), 30 7.26-7.49 (min, 5H), 7.83-8.23 (mn, 6H11), 8.73-8.74 (min, 1H), 8.90 (s, 1H), 8.94 (s, 1IH); MS (FAB) m/z 681 (M+1); Anal. calcd for C 37
H
33
N
4 0 7 Cl-0.5H 2 0: C, 64.39; H, 4.97; N, 8.12. Found: C, 64.22; H, 4.90; N, 7.96. 272 WO 01/00206 PCT/US00/18079 Example 114 4-[ 1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-(4S)-(6-quinolyloxy-(2S) pyrrolidinyl]methoxybenzoic acid N. N N \COOH r H H Me 122 5 To a stirred solution of methyl 4-(4S-(6-quinolyloxy-2S-pyrrolidinyl)methoxybenzoate (300 mg, 0.79mmol), 4-[N'-(2-bromophenyl)uredio]-3-methoxyphenylacetic acid (299 mg, 0.79 mmol), HOBt (107 mg, 0.79 mmol), and triethylamine (330 ml, 2.37 mmol) in THF (10.0 ml) and MeCN (10.0 ml) was added EDC.HCI (228 mg, 1.2 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, 10 and extracted with EtOAc. The extract was washed with sat. NaHCO 3 then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with EtOAc to EtOH-EtOAc (10%, v/v) as eluent to give methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3 methoxyphenylacetyl]l-(4S)-(6-quinolyloxy-(2S)-pyrrolidinyl]methoxybenzoate (530 mg, 91%) as a colorless oil. 'H-NMR (CDCI 3 ) 6 2.30-2.62 (in, 3H), 3.65 (s, 2H), 3.75 (s, 3H11), 3.80-3.95 (in, l1H), 15 3.93 (s, 3H), 4.10-4.30 (mn, 1H1), 4.50-4.70 (m, 2H), 5.11 (br s, 1H), 6.82-6.98 (in, 6H), 7.15-7.39 (in, 5H), 7.52 (d, J= 8.0 Hz, 1H1), 7.93-8.03 (in, 5H), 8.14 (d, J= 8.3 Hz, 1H), 8.80 (s, 1H1). To a stirred solution of methyl 4-[1-[4-[N'-(2-bromnophenyl)ureido]-3-mnethoxyphenyl acetyl]-4-(6-quinolyloxy-2S-pyrrolidinyl]methoxybenzoate(530 mg, 0.72 mmol) in THF (10.0 ml) and MeOH (5.0 ml) was added IN NaOH (1.4 ml, 1.4 mmninol). The mixture was stirred at 70 'C 20 for 24 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HCI. The resulting solid was collected, washed with water, and dried in vacuo to give 122 (460 mg, 88%) as a white crystalline solid. MW 725.59 mp 149-153 oC; IR (KBr) 3332, 1704, 1604, 1527, 1222, 1164 cn-'; 'H-NMR (DMSO-ds) 8 2.28-2.58 (in, 2H), 3.67 (s, 2H), 3.82 (s, 3H), 3.85-3.90 (in, 111), 4.05-4.15 (in, 2H), 4.40-4.50 (in, 2H), 5.20-5.32 (in, 1H), 6.77-7.07 (in, 5H), 25 7.31-7.61 (mn, 5H), 7.83-7.97 (in, 5H1), 8.21-8.22 (in, 1H), 8.73-8.74 (in, 2H), 8.92 (s, 1H); MS (FAB) nm/z 725 (M), 727 (M*+2); Anal. calcd for C 37
H
3
N
4 06Br-0.5H 2 0: C, 60.50; H, 4.67; N, 7.63; Br, 10.88. Found: C, 60.51; H, 4.60; N, 7.52; Br, 11.06. Example 115 4-[(2S,4S)- -[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-4-(2-naphthyloxy)-2 273 WO 01/00206 PCT/US00/18079 pyrrolidinyl]methoxybenzoic acid COOH N 0 MeH H Me 123 To a stirred mixture of methyl (2S,4R)-1-tert-butoxycarbonyl-4-hydroxy-2 pyrrolidinylcarboxylate (4.22 g, 17.2 mmol), 2-naphthol (2.73 g, 18.9 mmol) and PPh 3 (4.96 g, 5 18.9 mmol) in THF (80 ml) was added DIAD (3.72 ml, 18.9 mmol) at room temperature under an atmosphere of nitrogen. After stirring over night, the mixture was concentrated in vacuo. The residue was chromatographed on silica gel [600 g, CHCl/EtOAc (10/1)], to give methyl (2S,4S)-I tert-butoxycarbonyl-4-(2-naphthyloxy)-2-pyrrolidinylcarboxylate (5.37 g), which was used without further purification. 10 To a stirred solution of methyl (2S,4S)-1-tert-butoxycarbonyl-4-(2-napthhyloxy)-2 pyrrolidinyl carboxylate (5.37 g) in TBF (116 ml) was added 0.25 N NaOH (116 ml, 29.0 mmol) at room temperature. The resulting mixture was stirred over night. After removal of the solvent, the mixture was acidified by the addition of 1 N HCI and extracted with CHCl 3 . The combined extracts were washed with brine, dried over Na 2
SO
4 and evaporated. The residue was 15 recrystallized from n-hexane-CHC1 3 , to give (2S,4S)-1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2 pyrrolidinylcarboxylic acid [4.44 g, 85%(2 steps)] as a white powder. 'H-NMR (DMSO-d 6 ) 8 1.37 and 1.41 (s, 9H, amide isomers), 2.26 (d, J= 13.9 Hz, 1H), 2.65 (min, 1H), 3.47 (d, J= 11.5 Hz, 1H), 3.81 (min, 1H), 4.30 (min, 1H), 5.14 (min, 1H11), 7.02-7.86 (min, 7H). To a stirred solution of (2S,4S)-1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2 20 pyrrolidinylcarboxylic acid (1.12 g, 3.13 mmol) in THF (30 ml) was added BH 3 -DMS (0.63 ml, 6.3 mmol) at 0 oC. The mixture was raised to room temperature immediately and then heated at 50 oC for 1.5 h. After cooling to room temperature, the mixture was quenched by the addition of water at 0 oC and extracted with EtOAc. The combined extracts were washed with brine, dried over Na 2
SO
4 and evaporated. The residue was chromatographed on silica gel [50 g, CHCIzMeOH 25 (50/1)], to give (2S,4S)-1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2-pyrrolidinylmethanol (1.10 g, 100%) as a pale yellow oil. 'H-NMR (CDCl 3 ) 8 1.48 (s, 9H), 2.45 (mn, 1H), 3.58-4.80 (mn, 4H), 5.01 (br, 1H), 7.04-7.99 (min, 7H). To a stirred mixture of (2S,4S)-1l-tert-butoxycarbonyl-4-(2-naphthyloxy)-2-pyrrolidinyl 274 WO 01/00206 PCT/US00/18079 methanol (640 mg, 1.86 mmol), methyl 4-hydroxybenzoate (283 mg, 1.86 mmol) and PPh 3 (488 mg, 1.86 mmnol) in THF (18 ml) was added DIAD (0.37 ml, 1.86 mmol) at room temperature under an atmosphere of nitrogen. The mixture was stirred over night. After removal of the solvent, the resulting residue was chromatographed on silica gel [100 g, n-hexane/EtOAc(2/1)], to 5 give methyl 4-[(2S,4S)- 1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2-pyrrolidinyl]methoxybenzoate (830 mg, 93%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.50 (d, J= 8.3 Hz, 9H), 2.34 (m, 1H), 2.53 (d, J= 14.2 Hz, 1H), 3.72-3.85 (m, 1H), 3.86 and3.87 (s, 3H, amide isomers), 4.17 (m, 1H), 4.26 4.52 (m, 2H), 5.06 (br, 1H), 6.87 (d, J= 8.8 Hz, 1H), 6.94 (d, J= 8.8 Hz, 2H1), 7.04 (br, 2H), 7.33 (t, J = 7.3 Hz, 1H), 7.42 (t, J = 7.3 Hz, 1H), 7.64-8.02 (m, 5H). .10 To a stirred solution of methyl 4-[(2S,4S)-1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2 pyrrolidinyl]methoxybenzoate (870 mg, 1.74 mmol) in CH2C1 2 (24 ml) was added TFA (6 ml) at room temperature. The mixture was stirred over night, which was concentrated in vacuo. The residue was diluted with CH 2 C1 2 and made basic by the addition of 1 N NaOH, which was extracted with CH 2 C1 2 . The organic layer was washed with brine, dried over NaSO 4 and 15 concentrated. The residue was chromatographed on silica gel [100 g, n-hexane/EtOAc(2/1)], to give methyl 4-[(2S,4S)-4-(2-naphthyloxy)-2-pyrrolidinyl]methoxybenzoate (750 mg, 100%) as a black oil 'H-NMR (CDCl 3 ) 8 1.99 (dd, J= 14.2, 5.6 Hz, 1H), 2.48 (m, 1H1), 3.22 (dd, J= 12.2, 4.6 Hz, 1H14), 3.43 (d, J = 12.5 Hz, 1H), 3.67 (m, 1H), 3.86 and 3.87 (s, 3H, amide isomers), 4.11 (m, 2H), 5.04 (m, 1H), 6.83 (d, J= 8.5 Hz, 1H), 6.89 (d, J= 8.8 Hz, 2H), 7.07 (d, J= 2.0 Hz, 114), 20 7.12 (d, J= 9.0 Hz, 1H), 7.33 (dt, J= 8.1, 1.2 Hz, 1H), 7.44 (dt, J= 6.8, 1.2 Hz, 1H), 7.70 (d, J= 8.1 Hz, 1H), 7.75 (dd, J = 9.0, 5.1Hz, 2H), 7.90 (d,J = 8.5 Hz, 1H), 7.96 (dd,J= 6.8, 2.0 Hz, 2H). A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (333 mg, 0.106 mmol), methyl 4-[(2S,4S)-4-(2-naphthyloxy)-2-pyrrolidinyl]methoxybenzoate (400 mg, 1.06 mmol), EDC-HCI (305 mg, 1.59 mmol) and DMAP (194 mg, 1.59 mmol) in DMF (10 ml) was 25 stirred at room temperature for 3 days. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [100 g, n hexane/EtOAc(1/1)CHCl 3 /MeOH(50/1)], to give methyl 4-[(2S,4S)-1-[3-methoxy-4-[N'-(2 methylphenyl)ureido]phenylacetylJ]-4-(2-naphthyloxy)-2-pyrrolidinyl]methoxybenzoate (520 mg, 30 73%) as a pale brown amorphous. 'H-NMR (CDC13) 8 2.28 (s, 3H1), 2.29 (m, 1H1), 2.55 (d, J = 14.2 Hz, 1H), 3.60 (d, J= 3.4 Hz, 2H), 3.66 (d, J= 3.7 Hz, 3H), 3.68-4.00 (m, 5H), 4.05-4.67 (m, 3H1), 5.09 (br, 1H), 6.61 (s, 1H), 6.77 (m, 2H), 6.87 (d, J= 8.8 Hz, 1H), 6.94-7.54 (m, 8H), 7.68 275 WO 01/00206 PCT/US00/18079 8.09 (m, 8H); MS (ESI) m/z 674 (M+1). To a solution of methyl 4-[(2S,4S)-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-4-(2-naphthyloxy)-2-pyrrolidinyl]methoxybenzoate (415 mg, 0.616 mmol) in THF (4.9 ml), 0.25 N NaOH (4.9 ml) was added. After stirring at room temperature for 3 days, the 5 mixture was acidified with 1 N HCI and extracted with CHCI 3 -MeOH (10/1). The combined extracts were dried over Na 2
SO
4 and concentrated in vacuo. The residue was purified on TLC [CHCl 3 /MeOH (10/1)], to give 123 (180 mg, 44%) as a colorless amorphous. MW 659.73 IR (KBr) 3354, 2937, 1685, 1601, 1533, 1255 cm'; 'H-NMR (DMSO-d) 8 2.24 (s, 3H), 2.25-2.43 (m, 2H), 3.65 (s, 2H), 3.81 (s, 3H), 3.83 (m, 1H1), 4.05-4.70 (m, 4H), 5.21-5.33 (br, 1H1), 6.76 (d, J 10 = 7.3 Hz, 1H1), 6.86-7.35 (m, 9H), 7.44 (t, J= 7.3 Hz, 111), 7.76-7.89 (m, 6H1), 8.01 (d, J= 8.3 Hz, 1H1), 8.48 (s, 1H), 8.56 (s, 1H1); MS (FAB) m/z 660(M +1). Example 116 4-[(2S,4S)-1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-4-(2-naphtyloxy)-2 pyrrolidinyl]methoxybenzoic acid N< / COOH NN NO~CO 15 Cl H H OMe 124 A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (310 mg, 0.93 mmol), methyl 4-[(2S,4S)-4-(2-naphthyloxy)-2-pyrrolidinyl]methoxybenzoate (350 mg, 0.93 mmol), EDC-HCI (267 mg, 1.40 mmol) and DMAP (171 mg, 1.40 mmol) in DMF (10 ml) was stirred at room temperature for 3 days. The mixture was poured into ice water and extracted with 20 EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [100 g, n hexane/EtOAc(1/1)CHCl/MeOH(50/1)], to give methyl 4-[(2S,4S)-1-[4-[N'-(2-chlorophenyl) ureido]-3-methoxyphenylacetyl]-4-(2-naphthyloxy)-2-pyrrolidinyl]methoxybenzoate (450 mg, 68%) as a pale brown amorphous. 'H-NMR (CDC13) 8 2.32 (m, 1H1), 2.58 (d, J = 14.5 Hz, 111), 25 3.63 (d, J= 2.7 Hz, 1H), 3.70 (s, 3H), 3.86 (s, 3H), 3.84-3.95 (m, 2H), 4.15-4.64 (m, 4H), 5.11 (br, 1H), 6.79-7.06 (m, 7H), 7.21-7.46 (m, 7H), 7.66-7.77 (m, 3H), 7.92 (d, J= 8.8 Hz, 1H), 7.97 (m, 1H1), 8.17 (d, J= 8.4 Hz, 1H); MS (ESI) m/z 694 (M+1), 696 (M+3). To a solution of methyl 4-[(2S,4S)-1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl acetyl]-4-(2-naphthyloxy)-2-pyrrolidinyl]methoxybenzoate (381 mg, 0.535 mmol) in THF (4.3 ml), 276 WO 01/00206 PCT/US00/18079 0.25 N NaOH (4.3 ml) was added. After stirring at room temperature for 3 days, the mixture was acidified with 1 N HCI and extracted with CHCl 3 -MeOH (10/1). The combined extracts were dried over Na 2
SO
4 and concentrated in vacuo. The residue was purified on TLC [CHC13/MeOH (10/1)] to give 124 (140 mg, 39%) as a colorless amorphous. MW 680.15 IR (KBr) 3323, 2935, 1704, 5 1601, 1529, 1529, 1508 cm'; 'H-NMR (DMSO-d6) 8 2.27-2.49 (min, 2H), 3.65 (s, 2H), 3.81 (s, 3H), 3.83 (min, 1H), 4.05-4.71 (min, 4H), 5.30 (br, 1H), 6.77 (d, J= 7.8 Hz, 1H), 6.87-7.16 (min, 4H), 7.14 (dd, J= 8.8, 2.2 Hz, 1H), 7.27 (t, J = 7.3 Hz, 1H), 7.29-7.46 (min, 4H), 7.76-7.86 (min, 5H11), 7.96 (d, J = 8.3 Hz, 1H), 8.08 (dd, J= 8.3, 1.2 Hz, 1H), 8.90 (s, 1H1), 8.93 (s, 1H); MS (FAB) m/z 680 (M +1), 682 (M+3); Anal. Calcd for C 38
H
34
CIN
3 0 7 -1H 2 0: C, 65.37; H, 5.20; N, 6.02. Found: C, 10 65.43; H, 5.11; N, 5.93. Example 117 2-[(2S,4S)-1 -[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-4-(2-naphthyloxy)-2 pyrrolidinyl]methoxy-5-pyridinecarboxylic acid N - COOH N N 0 0 Me H H Me 125 15 To a stirred mixture of methyl (2S,4R)-1-tert-butoxycarbonyl-4-hydroxy-2 pyrrolidinylcarboxylate (4.22 g, 17.2 mmol), 2-naphthol (2.73 g, 18.9 mmol) and PPh 3 (4.96 g, 18.9 mmol) in THF (80 ml) was added DIAD (3.72 ml, 18.9 mmol) at room temperature under an atmosphere of nitrogen. After stirring over night, the mixture was concentrated in vacuo. The residue was chromatographed on silica gel [600 g, CHCl 3 /EtOAc (10/1)], to give methyl (2S,4S) 20 1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2-pyrrolidinylcarboxylate (5.37 g), which was used to the next reaction without further purification. To a stirred solution of methyl (2S,4S)-1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2 pyrrolidinyl carboxylate (5.37 g) in THF (116 ml) was added 0.25 N NaOH (116 ml, 29.0 mmol) at room temperature. The resulting mixture was stirred over night. After removal of the solvent, the 25 mixture was acidified by the addition of 1 N HCI and extracted with CHCl 3 . The combined extracts were washed with brine, dried over Na 2
SO
4 and evaporated. The residue was recrystallized with n-hexane-CHC 3 , to give (2S,4S)-1 -tert-butoxycarbonyl-4-(2-naphthyloxy)-2 pyrrolidinylcarboxylic acid [4.44 g, 85%(2 steps)] as a white powder. 'H-NMR (DMSO-d 6 ) 8 1.37 and 1.41 (s, 9H1, amide isomers), 2.26 (d, J= 13.9 Hz, 1H), 2.65 (min, 1H1), 3.47 (d, J= 11.5 Hz, 30 1H1), 3.81 (min, 1H), 4.30 (min, 1H), 5.14 (min, 1H11), 7.02-7.86 (mn, 7H1). 277 WO 01/00206 PCT/US00/18079 To a stirred solution of (2S,4S)-1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2 pyrrolidinylcarboxylic acid (1.12 g, 3.13 mmol) in THF (30 ml) was added BH3-DMS (0.63 ml, 6.3 mmol) at 0 oC. The mixture was raised to room temperature immediately and then heated at 50 oC for 1.5 h. After cooling to room temperature, the mixture was quenched by the addition of water at 5 0 oC and extracted with EtOAc. The combined extracts were washed with brine, dried over Na 2
SO
4 and evaporated. The residue was chromatographed on silica gel [50 g, CHC13/MeOH (50/1)], to give (2S,4S)-1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2-pyrrolidinylmethanol (1.10 g, 100%) as a pale yellow oil. 'H-NMR (CDCl 3 ) 8 1.48 (s, 9H11), 2.45 (m, 1H11), 3.58-4.80 (m, 4H), 5.01 (br, 1H), 7.04-7.99 (m, 7H). 10 To a stirred mixture of (2S,4S)-1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2 pyrrolidinylmethanol (484 mg, 1.41 mmol), methyl 2-hydroxy-5-pyridinecarboxylate (216 mg, 1.41 mmol) and PPh 3 (370 mg, 1.41 mmol) in THF (15 ml) was added DIAD (0.28 ml, 1.41 mmol) at room temperature under an atmosphere of nitrogen. The mixture was stirred over night. After removal of the solvent, the resulting residue was chromatographed on silica gel [50 g, n 15 hexane/EtOAc(2/1)], to give methyl-2-[(2S,4S)-1l-tert-butoxycarbonyl-4-(2-naphthyloxy)-2 pyrrolidinyl]methoxypyridine-5-carboxylate (170 mg, 25%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.47 (s, 9H), 2.37 (m, 1H), 2.46 (d, J= 14.2 Hz, 111), 3.71-4.00 (m, 2H), 3.89 (s, 3H), 4.30-4.56 (m, 2H11), 4.74 (dd, J= 9.8, 4.6 Hz, 1H), 5.06 (br, 1H11), 6.70 (d, J= 8.8 Hz, 2H), 7.05-7.09 (m, 2H), 7.33 (t, J= 6.9 Hz, 1H), 7.42 (t, J= 6.9 Hz, 1H11), 7.67-7.75 (m, 3H11), 8.09 (d, J= 8.8 Hz, 1H), 8.77 20 (d, J= 2.2 Hz, 1H). To a stirred solution of 5-carboxymethyl-2-[(2S,4S)-1-tert-butoxycarbonyl-4-(2 napthyloxy)-2-pyrrolidinyl]methoxypyridine (170 mg, 0.36 mmol) in CH 2 C1 2 (5 ml) was added TFA (2 ml) at room temperature. After 2 h stirring, the mixture was concentrated in vacuo, which was diluted with CH 2 Cl 2 and basified by the addition of 1 N NaOH. The combined reaction 25 mixture was extracted with CH 2
CI
2 . The organic layer was washed with brine, which were dried over NaSO 4 and concentrated. The residue was purified on TLC [CHCl 3 MeOH (10/1)], to give methyl 2-[(2S,4S)-4-(2-naphthyloxy)2-pyrrolidinyl]methoxypyridine-5-carboxylate (107 mg, 80%) as a colorless oil 'H-NMR (CDCl 3 ) 5 1.95 (m, IH), 2.27 (br, 1H), 2.46 (m, 1H), 3.19 (dd, J= 12.2, 4.9 Hz, 111), 3.41 (d, J= 12.2 Hz, 11H), 3.65 (m, 111), 3.89 (s, 311), 4.58 (m, 21H1), 5.00 (br, 30 1H), 6.77 (d, J= 8.8 Hz, 111), 7.06 (br, 111), 7.11 (dd, J= 8.8, 2.7 Hz, 111), 7.31-7.45 (m, 2H11), 7.69-7.76 (m, 3H), 8.13 (dd, J= 8.8, 2.4 Hz, IH), 8.78 (d, J= 2.2 Hz, 111). 278 WO 01/00206 PCT/US00/18079 A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (89 mg, 0.283 mmol), methyl-2-[(2S,4S)-4-(2-naphthyloxy)-2-pyrrolidinyl]methoxypyridine-5-carboxylate (107 mg, 0.78 mmol), EDC-HCI (81 mg, 0.425 mmol) and DMAP (52 mg, 0.425 mmol) in DMF (3 ml) was stirred at room temperature for 18 h. The mixture was poured into ice water and extracted 5 with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was purified on TLC [CHCI/MeOH (10/1)], to give 2-[(2S,4S)-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-4-(2 naphthyloxy)-2-pyrrolidinyl]methoxy-5-pyridinecarboxylic acid methyl ester (193 mg, 100%) as a colorless amorphous. 'H-NMR (CDCl 3 ) 8 2.27 (d, J= 3.2 Hz, 3H), 2.30 (mn, 1H), 2.49 (dd, J= 10 14.2, 2.0 Hz, 1H), 3.60 (d, J= 3.9 Hz, 1H), 3.67 (d, J= 5.9 Hz, 3H), 3.81 (s, 1H), 3.85 (s, 1H), 3.88 and 3.91 (s, 3H, amide isomers), 3.95 (min, 1H), 4.02-5.09 (min, 4H), 6.67 (d, J= 8.8 Hz, 1H), 6.73-7.13 (min, 3H), 7.20-7.45 (min, 7H), 7.53 (t, J= 7.8 Hz, 1H), 7.67-7.77 (min, 3H), 8.02-8.84 (inm, 3H). For HCI salt: a pale brown amorphous. IR (KBr) 3346, 2951, 1720, 1601, 1533, 1281 cm'; MS 15 (FAB) m/z 675 (M+1). To a solution of 2-[(2S,4S)- 1-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetyl]-4 (2-naphthyloxy)-2-pyrrolidinyl]methoxy-5-pyridinecarboxylic acid methyl ester (158 mg, 0.23 mmol) in THF (1.8 ml), 0.25 N NaOH (1.8 ml) was added. After stirring at room temperature for 22 h, the mixture was neutralized with 1 N HCI and extracted with CHC1 3 -MeOH (10/1). The 20 combined extracts were dried over Na 2
SO
4 and concentrated in vacuo. The residue was purified on TLC [CHCI/MeOH (5/1)] to give 125 (51 mg, 34%) as a colorless amorphous solid. MW 660.72 IR (KBr) 3354, 2956, 1601, 1533, 1255, 1022 cur'; 'H-NMR (DMSO-d 6 ) 8 2.24 (s, 3H), 3.30-5.32 (min, 13H), 6.72-8.82 (min, 19H); MS (FAB) m/z 661(M++1); Anal. Calcd for
C
38
H
3 6
N
4 0 7 0.5EtOH-lH 2 0: C, 66.75; H, 5.89; N, 7.98. Found: C, 66.39; H, 5.55; N, 7.66. 25 Example 118 4-[5-(R)-benzyloxymethyl-1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-(S)-pyrrolidinyl methoxy]benzoic acid N N ) COOH Me H H 126 To a stirred solution of benzyl-(S)-glycidyl ether (5.0 g, 30.5 mmol) in THF (100 ml) was 30 added allylmagnesium chloride (1.0 M in Et 2 0, 30.5 ml, 30.5 mmol) at -78 0 C, and the resulting 279 WO 01/00206 PCT/US00/18079 mixture was gradually warmed up to rt with stirring. The mixture was poured into water and concentrated in vacuo, then extracted with CHCl 3 . The organic layer was dried over anhydrous Na 2
SO
4 and concentrated in vacuo. The residue was chromatographed on silica gel with hexane EtOAc (5 : 1) as eluent to give 1-benzyloxy-2-(R)-hydroxy-5-hexene (2.18 g, 35%) as a colorless 5 oil: 'H-NMR (CDC13) 8 1.52-1.60 (m, 2 H), 2.11-2.25 (m, 2 H), 2.34 (d, J= 3.2 Hz, 1 H), 3.35 (dd, J= 9.6, 8.0 Hz, 1 H), 3.52 (dd, J= 9.6, 3.2 Hz, 1 H), 3.84-3.86 (m, 1 H), 4.57 (s, 2 H), 4.96 5.07 (series of m, 2 H), 5.78-5.88 (m, 1 H), 7.29-7.38 (m, 5 H); MS (ESI) m/z, 224 (M*+NH4). To a stirred solution of 1-benzyloxy-2-(R)-hydroxy-5-hexene (2.18 g, 10.5 mmol), triphenylphosphine (3.32 g, 12.7 mmol) and phthalimide (1.86 g, 12.7 mmol) was added 10 diisopropyl azodicarboxylate (2.62 ml, 12.7 mmol) at rt, and the resulting mixture was stirred for overnight at rt. The mixture was concentrated in vacuo and extracted with EtOAc. The organic layer was washed with water, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with hexane-EtOAc (5 : 1) as eluent to give 1 benzyloxy-2-(S)-phthalimido-5-hexene (2.95 g, 83%) as a colorless oil: 'H-NMR (CDCl 3 ) 6 1.76 15 1.84 (m, 2 H), 2.06 (dd, J= 14.4, 6.8 Hz), 2 H, 2.12-2.22 (m, 1 H), 3.69 (dd, J= 10.0, 5.6 Hz, 1 H), 4.00 (t, J = 9.6 Hz, 1 H), 4.46 (d, J = 12.0 Hz, 1 H), 4.53 (d, J= 12.0 Hz, 1 H), 4.51-4.58 (m, 1 H), 4.91-4.99 (series of m, 2 H), 5.72-5.79 (m, 1 H)7.21-7.26 (m, 5 H), 7.71-7.83 (series of m, 2 H); MS (ESI) m/z, 336 (M*+H). To a stirred solution of 1-benzyloxy-2-(S)-phthalimido-5-hexene (2.95 g, 8.80 mmol) in 20 EtOH (30 ml) was added hydrazine hydrate (80% in water, 460 ml, 11.4 mmol) at rt, and the resulting mixture was heated under reflux for 7.5 h with stirring. The solution was filtered, and the filtrate was concentrated in vacuo. The residue was poured into aq.NaHCO 3 and extracted with CHCl 3 . The organic layer was dried over anhydrous Na 2
SO
4 , then concentrated in vacuo to give 2 (S)-amino-1-benzyloxy-5-hexene (1.90 mg, quant.) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.37 25 1.55 (series of m, 4 H), 2.08-2.19 (m, 2 H), 2.99-3.03 (m, 1 H), 3.25 (dd, J= 9.2, 7.6 Hz, 1 H), 3.45 (dd, J= 9.2, 4.0 Hz, 1 H), 4.53 (s, 2 H), 4.94-5.06 (series of m, 2 H), 5.76-5.85 (m, 1 H), 7.27-7.37 (m, 5 H); MS (ESI) m/z, 206 (M++H), 247 (M+H+CH 3 CN). To a stirred solution of 2-(S)-amino-1-benzyloxy-5-hexene (1.89 g, 9.21 mmol) and triethylamine (1.28 ml, 9.21 mmol) in CH 2
CI
2 (20 ml) was added benzoyl chloride (1.07 ml, 9.21 30 mmol) at rt, and the resulting mixture was stirred for 23 h. The mixture was poured into water and extracted with CH 2 C1 2 . The organic layer was washed with water, drying over anhydrous 280 WO 01/00206 PCT/US00/18079 Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with hexane EtOAc (5 : 1) as eluent to give N-[2-(S)-(1-benzyloxy)-5-hexenyl]benzamide (2.67 g, 94%) as a colorless needles. mp 78-79 oC; 'H-NMR (CDCl 3 ) 5 1.76-1.82 (m, 2 H), 2.11-2.17 (m, 2 H), 3.59 (brs, 2 H), 4.29-4.35 (m, 1 H), 4.54 (dd, J= 19.2, 12.0 Hz, 2 H), 4.96-5.05 (series of m, 2 H), 5.78 5 5.89 (m, 1 H), 6.39 (d, J= 8.0 Hz, 1 H), 7.27-7.51 (m, 8 H), 7.74 (d, J= 7.2 Hz, 2 H); MS (ESI) m/z, 310 (M'+H). To a stirred solution of N-[2-(S)-(1-benzyloxy)-5-hexenyl]benzamide (2.41 g, 7.79 mmol) in CH 3
CN-H
2 0 (3 : 1, 40 ml) was added iodine (2.97 g, 23.4 mmol) in one portion, and the resulting mixture was stirred for 20 h. The mixture was poured into aq.Na 2
S
2 0 3 and concentrated 10 in vacuo, then extracted with CHCl 3 . The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was dissolved in CH 2 Cl 2 (30 ml) and was added di-tert-butyl dicarbonate (2.55 g, 11.7 mmol), Et 3 N (1.63 ml, 11.7 mmol) and N, N dimethyl aminopyridine (180 mg, 1.47 mmol), and the resulting mixture was stirred overnight at rt. The mixture was poured into water and extracted with CH 2 C1 2 . The organic layer was washed 15 with water, drying over Na 2
SO
4 , and concentrated in vacuo. The residue was chromatographed on silica gel with hexane-EtOAc (5 : 1) to give N-Boc-2-(S)-benzoyloxymethyl-5-(S)-benzyloxy methylpyrrolidine (1.27 g, 38%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.41 and 1.49 (s, total 9 H), 1.85-2.00 (series of m, 4 H), 3.33-4.59 (series of m, 8 H), 7.26-7.32 (m, 5 H), 7.41-7.46 (m, 2 H), 7.54-7.57 (m, 1 H), 8.02 (d, J= 7.6 Hz, 2 H); MS (ESI) m/z, 426 (M+H), 448 (M++Na ). 20 To a stirred solution of N-Boc-2-(S)-benzoyloxymethyl-5-(S)-benzyloxymethylpyrrolidine (1.23 g, 2.89 mnmol) in MeOH (30 ml) was added NaOH (1.0 M in water, 3.47 ml, 3.47 mmol) at rt, and the resulting mixture was stirred for 4 h. The mixture was neutralized with aq. 1N-HCl and concentrated in vacuo, then extracted with CHC 3 . The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on 25 silica gel with hexane-EtOAc (3 : 1) as eluent to give N-Boc-5-(S)-hydroxymethyl-2-(S) benzyloxymethyl pyrrolidine (847 mg, 91%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.41 (s, 9 H), 1.57 (brs, 1 H), 1.95-1.97 (m, 2 H), 2.05-2.18 (m, 1 H), 3.36 (t, J= 8.4 Hz, 1 H), 3.56-3.62 (m, 2 H), 3.67-3.72 (m, 2 H), 3.95 (brs, 1 H), 4.03 (brs, 1 H), 4.51 (s, 1 H), 7.28-7.37 (m, 5H); MS (FAB) m/z, 322 (MW+H). 30 To a stirred solution of N-Boc-2-(S)-hydroxymethyl-5-(S)-benzyloxymethylpyrrolidine (388 mg, 1.21 immol), triphenylphosphine (380 mg, 1.45 mmol) and methyl 4-hydroxybenzoate 281 WO 01/00206 PCT/US00/18079 (220 mg, 1.45 mmol) was added diisopropyl azodicarboxylate (200 ml, 1.45 mmol) at rt, and the resulting mixture was stirred overnight. The mixture was concentrated in vacuo and extracted with EtOAc. The organic layer was washed with water, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with hexane-EtOAc (3 : 1) 5 as eluent to give methyl 4-[2-(S)-(N-Boc-5-(S)-benzyloxymethyl)pyrrolidinylmethoxy]benzoate (462 mg, 84%) as a colorless oil: 'H-NMR (CDCl 3 ) 8 1.40 and 1.48 (s, total 9 H), 1.98-2.13 (m, 4 H), 3.83 and 3.85 (s, 3 H), 3.33-4.25 (series of m, 4 H), 4.47-4.59 (m, 2 H), 6.91-6.96 (m, 2 H), 7.26-7.34 (m, 5 H), 7.95-7.98 (m, 2 H); MS (FAB) m/z, 456 (M +H), 478 (M+Na'). To a stirred solution of methyl 4-[2-(S)-(N-Boc-5-(S)-benzyloxymethyl)pyrrolidinyl 10 methoxy] benzoate (446 mg, 0.98 mmol) in CH 2 Cl 2 (10 ml) was added trifluoroaceticacid (10 ml) at rt, and the resulting mixture was stirred for 1 h. The mixture was concentrated in vacuo and poured into aq.NaHCO 3 , then extracted with CHCI 3 . The organic layer was washed with water, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo to give methyl 4-[2-(S)-(5-(S) benzyloxymethyl) pyrrolidinylmethoxy]benzoate (363 mg, quant.) as yellowish oil. The product 15 was used for next reactions without further purification. 'H-NMR (CDCl 3 ) 8 1.43-1.65 (m, 2 H), 1.93-2.07 (m, 3 H), 3.36-3.68 (series of m, 4 H), 3.89 (s, 3 H), 3.86-3.93 (over lap, 2 H), 4.55 (s, 2 H), 6.90 (d, J = 8.4 Hz, 2 H), 7.26-7.37 (m, 5 H), 7.97 (d, J = 8.4 Hz, 2 H); MS (FAB) m/z, 356 (M'+H). To a stirred solution of methyl 4-[2-(S)-(5-(S)-benzyloxymethyl)pyrrolidinylmethoxy] 20 benzoate (115 mg, 0.32 mmol), 4-[N'-(2-methylphenyl)ureido]phenylacetic acid (92.0 mg, 0.32 mmol) and N,N-dimethylaminopyridine (52.0 mg, 0.42 mmol) in DMF (10 ml) was added EDCHCl (81.0 mg, 0.42 mmol) at rt, and the resulting mixture was stirred overnight. The reaction mixture was poured into water and extracted with CHC1 3 . The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was 25 chromatographed on silica gel with CHCl 3 -MeOH (20 : 1) as eluent to give methyl 4-[5-(R)-benzyl oxymethyl- 1-[4-[N'-(2-methylphenyl)ureido]phenylacetamido]-2-(S)-pyrrolidinylmethoxy] benzoate (169 mg, 84%) as a colorless amorphous solid. 'H-NMR (CDCl 3 ), mixture of rotamars 8 1.92-2.18 (m, 3 H), 2.24 and 2.25 (s, total 3 H), 2.20-2.31 (overlap, 1 H), 3.39-3.70 (series of m, 4 H), 3.87 and 3.89 (s, total 3 H), 4.17 and 4.18 (s, total 2 H), 4.30-4.45 (series of m, 2 H), 4.53 (s, 2 30 H), 6.43-7.13 (series of m, 9 H), 7.20-7.36 (series of m, 7 H), 7.58-7.99 (series of m, 3 H); MS (FAB) m/z, 622 (M'+H). 282 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[5-(R)-benzyloxymethyl-1-[4-[N'-(2-methylphenyl) ureido] phenylacetyl]-2-(S)-pyrrolidinylmethoxy]benzoate (156 mg, 0.24 mmol) in MeOH-THF (1 : 5, 12 ml) was added 1.0M-NaOH (1.2 ml, 1.20 mmol) at rt, and the resulting mixture was heated at 80 0 C with stirring for 7 h. The mixture was poured into IN-HCI, then extracted with CHCl 3 . 5 The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (5 : 1) as eluent to give 126 (94.0 mg, 65%) as a colorless amorphous solid. MW 607.70 'H-NMR (CD 3 OD), mixture of rotamars 6 1.85-2.35 (series of m, 4 H), 2.43-2.92 (series of m, 5 H), 2.28 (s, 3 H), 3.55-4.55 (series of m, 10 H), 6.85-7.95 (series of m, 17 H); MS (ESI) m/z, 630 (M+Na+). 10 Example 119 4-[5-(R)-benzyloxymethyl-l1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-2-(S) pyrrolidinylmethoxy]benzoic acid N N I HCH Me 127 To a stirred solution of methyl 4-[2-(S)-(5-(S)-benzyloxymethyl)pyrrolidinylmethoxy] 15 benzoate (117 mg, 0.33 mmol), 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (110 mg, 0.33 mmol) and N,N-dimethylaminopyridine (50.0 mg, 0.40 mmol) in DMF (10 ml) was added EDCHC1 (76.0 mg, 0.40 mmol) at rt, and the resulting mixture was stirred overnight. The reaction mixture was poured into water and extracted with CHCI 3 . The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was 20 chromatographed on silica gel with CHC13-MeOH (20: 1) as eluent to give methyl 4-[5-(R) benzyloxymethyl- 1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-2-(S)-pyrrolidinyl methoxy]benzoate (189 mg, 85%) as a colorless amorphous solid. 'H-NMR (CDCl 3 ), mixture of rotamars 8 1.91-2.30 (series of m, 4 H), 3.39-3.74 (series of m, 3 H), 3.73 (s, 2 H), 4.15-4.02 (m, 2 H), 4.32-4.44 (m, 1 H), 4.54 (s, 2 H), 6.71-7.02 (series of m, 5 H), 7.06 (s, 1 H), 7.17 (s, 1 H), 25 7.24-7.40 (series of m, 7 H), 7.89-8.22 (series of m, 4 H); MS (FAB) m/z, 672 (M +H). To a stirred solution of methyl 4-[5-(R)-benzyloxymethyl-l-[4-[N'-(2-chlorophenyl) ureido]-3-methoxyphenylacetyl]-2-(S)-pyrrolidinylmethoxy]benzoate (169 mg, 0.25 mmol) in MeOH-THF (2 : 5, 7 ml) was added 1.0M-NaOH (750 ml, 0.75 mmol) at rt, and the resulting mixture was heated at 80oC with stirring for 2 h. The mixture was poured into IN-HC1, then 30 extracted with CHC1 3 . The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , 283 WO 01/00206 PCT/US00/18079 then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (15 : 1) as eluent to give 127 (114 mg, 69%) as a colorless amorphous solid. MW 658.14 'H NMR (CD 3 OD), mixture of rotamars 8 1.88-2.37 (series of m, 4 H), 3.51-4.49 (series of m, 8H), 3.64 and 3.73 (s, total 3 H), 4.86 (s, 2 H), 6.85-7.95 (series of m, 16 H); MS (ESI) mnz, 658 5 (M'+H), 680 (M+Na+); Anal Calcd for C 3
H
36
CIN
3 0H 2 0: C, 63.95; H, 5.66; N, 6.21. Found: C, 63.65; H, 5.40: N, 5.95. Example 120 4-[5-(R)-benzyloxymethyl-1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-2-(S) pyrrolidinylmethoxy]benzoic acid N N - NCOOH 10 BrH H OMe 128 To a stirred solution of methyl 4-[2-(S)-(5-(S)-benzyloxymethyl)pyrrolidinylmethoxy] benzoate (119 mg, 0.34 mmol), 4-[N'-(2-bromophenyl)ureidoj-3-methoxyphenylacetic acid (127 mg, 0.34 mmol) and N,N-dimethylaminopyridine (50.0 mg, 0.40 mmol) in DMF (10 ml) was added EDCHCI (77.0 mg, 0.40 mmol) at rt, and the resulting mixture was stirred overnight. The 15 reaction mixture was poured into water and extracted with CHCl 3 . The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (20 : 1) as eluent to give methyl 4-[5-(R) benzyloxymethyl- 1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-2-(S)-pyrrolidinyl methoxy]benzoate (217 mg, 90%) as a colorless amorphous solid. 'H-NMR (CDCl 3 ), mixture of 20 rotamars 6 1.92-2.31 (series of m, 4 H), 3.39-3.73 (series of m, 3 H), 3.65 (s, 2H), 4.15-4.02 (m, 2 H), 4.32-4.44 (m, 1 H), 4.54 (s, 2 H), 6.71-6.99 (series of m, 5 H), 7.04 (s, 1 H), 7.11 (s, 1 H), 7.22-7.39 (series of m, 7 H), 7.51-8.17 (series of m, 4 H); MS (FAB) mn/z, 716 (M), 718 (M+2). To a stirred solution of methyl 4-[5-(R)-benzyloxymethyl-1-[4-[N'-(2-bromophenyl) ureido]-3-methoxyphenylacetyl]-2-(S)-pyrrolidinylmethoxy]benzoate (178 mg, 0.25 mmol) in 25 MeOH-THF (2 : 5, 7 ml) was added 1.0M-NaOH (750 ml, 0.75 mmol) at rt, and the resulting mixture was heated at 80oC with stirring for 1.5 h. The reaction mixture was poured into 1N-HCI, then extracted with CHC 3 . The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCI 3 MeOH (15 : 1) as eluent to give 128 (159 mg, 91%) as a colorless amorphous solid. MW 702.59 30 'H-NMR (CD 3 OD), mixture of rotamars 5 1.88-2.35 (series of m, 4 H), 3.51-4.49 (series of m, 8 284 WO 01/00206 PCT/US00/18079 H), 3.64 and 3.72 (s, total 3 H), 4.87 (s, 2 H), 6.65-8.05 (series of m, 16 H); MS (ESI) m/z, 702 (M), 704 (M++2); Anal. Calcd for C 36
H
3
,
6 BrN 3 07H 2 0: C, 60.00; H, 5.32; N, 5.83. Found: C, 59.66; H, 5.04: N, 5.65. Example 121 5 3-[5-(R)-benzyloxymethyl-1-[4-[N'-(2-methylphenyl)ureido]-3-methoxyphenylacetyl]-2-(S) pyrrolidinylmethoxy]benzoic acid N N \/aCOOH MeCH HOMe 129 To a stirred solution of N-Boc-2-(S)-hydroxymethyl-5-(S)-benzyloxymethylpyrrolidine (415 mg, 1.29 mmol), triphenylphosphine (410 mg, 1.55 mmol) and methyl 3-hydroxybenzoate 10 (240 mg, 1.55 mmol) was added diisopropyl azodicarboxylate (320 ml, 1.55 mmol) at rt, and the resulting mixture was stirred overnight. The mixture was concentrated in vacuo, then the residue was chromatographed on silica gel with hexane-EtOAc (3 : 1) as eluent to give methyl 3-[2-(S)-(N Boc-5-(S)-benzyloxymethyl)pyrrolidinylmethoxy]benzoate (513 mg, 87%) as a colorless oil: 'H NMR (CDCl 3 ) 8 1.40 and 1.46 (s, total 9 H), 1.95-2.20 (series of m, 4 H), 3.33-3.72 (series of m, 2 15 H), 3.82-4.00 (m, 1 H), 3.90 and 3.91 (s, total 3 H), 4.09-4.21 (m, 3 H), 4.21-4.57 (m, 2 H), 7.11 7.15 (m, 1 H), 7.26-7.37 (m, 6 H), 7.53-7.65 (m, 2 H); MS (ESI) m/z, 456 (M+H). To a stirred solution of methyl 3-[2-(S)-(N-Boc-5-(S)-benzyloxymethyl)pyrrolidinyl methoxy] benzoate (501 mg, 1.10 mmol) in CH 2 C1 2 (10 ml) was added trifluoroacetic acid (10 ml) at rt, and the resulting mixture was stirred for 1 h. The mixture was concentrated in vacuo and 20 poured into aq.NaHCO 3 , the extracted with CHC1 3 . The organic layer was washed with water, drying over anhydrous Na 2
SO
4 , and concentrated in vacuo to give methyl 3-[2-(S)-(5-(S) benzyloxymethyl) pyrrolidinyl-methoxy]benzoate (387 mg, quant.) as yellowish oil. The product was used for next reactions without further purification. 'H-NMR (CDC13) 8 1.26-1.65 (m, 2 H), 1.94-2.04 (m, 3 H), 3.37-3.52 (m, 2 H), 3.63-3.66 (m, 1 H), 3.85-3.93 (m, 1 H), 3.91 (s, 3 H), 4.55 25 (s, 2 H), 7.09-7.11 (m, 1 H), 7.27-7.54 (m, 6 H), 7.54-7.55 (m, 1H), 7.61-7.63 (m, 2 H); MS (ESI) m/z, 35 (M++H). To a stirred solution of methyl 3-[2-(S)-(5-(S)-benzyloxymethyl)pyrrolidinylmethoxy] benzoate (140 mg, 0.39 mmol), 4-[N'-(2-methylphenyl)ureido]-3-methoxyphenylacetic acid (125 mg, 0.39 mmol) and N,N-dimethylamniinopyridine (58.0 mg, 0.47 mmol) in THF (15 ml) was added 285 WO 01/00206 PCT/US00/18079 EDCHCI (90.0 mg, 0.47 mmol) at rt, and the resulting mixture was stirred overnight. The reaction mixture was poured into water and extracted with CHCI 3 . The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (10 : 1) as eluent to give methyl 3-[5-(R) 5 benzyloxymethyl- 1-[4-[N'-(2-methylphenyl)ureido]-3-methoxyphenylacetyl]-2-(S)-pyrrffolidinyl methoxy]benzoate (256 mg, quant.) as a colorless amorphous solid. 'H-NMR (CDCl 3 ), mixture of rotamars 8 1.67 (s, 3 H), 1.97-2.40 (series of m, 4 H), 3.42-3.85 (series of m, 5 H), 3.60 (s, 3 H), 3.95 and 3.97 (s, total 3 H), 4.15-4.26 (m, 2 H), 4.36-4.49 (m, 1 H), 4.59 (s, 2 H), 6.32 and 6.36 (s, total 1 H), 6.75-6.87 (series of m, 2 H), 7.20 (brs, 2 H), 7.16-7.72 (series of m, 10 H), 8.03-8.09 10 (m, 1 H); MS (ESI) m/z, 652 (M+H). To a stirred solution of methyl 3-[5-(R)-benzyloxymethyl-1-[4-[N'-(2-methylphenyl) ureido]-3-methoxyphenylacetyl]-2-(S)-pyrrolidinylmethoxy]benzoate (185 mg, 0.28 mmol) in MeOH-THF (2 : 5, 7 ml) was added 1.0M-NaOH (860 ml, 0.86 mmol) at rt, and the resulting mixture was heated at 60oC with stirring for 1 h. The reaction mixture was poured into 1N-HCI, 15 then extracted with CHC1 3 . The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCI 3 MeOH (5 : 1) as eluent to givel29 (171 mg, 94%) as a colorless amorphous solid. MW 637.72 'H-NMR (CD 3 OD), mixture of rotamars 8 1.89-2.37 (series of m, 4 H), 2,29 (s, 3 H), 3.52-4.53 (series of m, 8 H), 3.66 and 3.74 (s, total 3 H), 4.85 (s, 2 H), 6.66-7.98 (series of m, 16 H); MS 20 (ESI) m/z, 638 (M'+H), 660 (M+Na+); Anal. Calcd for C 37
H
3 9
N
3 0 7
H
2 0: C, 67.77; H, 6.30; N, 6.41. Found: C, 67.40; H, 5.95: N, 6.14. Example 122 3-[5-(R)-benzyloxymethyl-1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-2-(S) pyrrolidinylmethoxy]benzoic acid 25 COOH N N 25 1iH H Me 130 To a stirred solution of methyl 3-[2-(S)-(5-(S)-benzyloxymethyl)pyrrolidinylmethoxy] benzoate (118 mg, 0.33 mmol), 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (112 mg, 0.33 mmol) and N,N-dimethylaminopyridine (50.0 mg, 0.40 mmol) in THF (15 ml) was added EDCHCI (80.0 mg, 0.40 mmol) at rt, and the resulting mixture was stirred overnight. The 30 reaction mixture was poured into water and extracted with CHCl 3 . The organic layer was washed 286 WO 01/00206 PCT/US00/18079 with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (10 : 1) as eluent to give methyl 3-[5-(R) benzyloxymethyl-1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetamido]-2-(S) pyrrolidinylmethoxy]benzoate (226 mg, quant.) as a colorless amorphous solid. 'H-NMR (CDCI 3 ), 5 mixture of rotamars 8 1.99-2.40 (series of m, 4 H), 3.43-3.92 (series of m, 5 H), 3.67 (s, 3 H), 3.98 and 4.02 (s, total 3 H), 4.18-4.29 (m, 2 H), 4.36-4.51 (m, 1 H), 4.60 (s, 2 H), 6.75-6.92 (series of m, 2 H), 7.01-7.22 (series of m, 4 H), 7.29-7.53 (series of m, 9 H), 7.62-8.26 (series of m, 3 H); MS (ESI) m/z, 672 (M'+H). To a stirred solution of methyl 3-[5-(R)-benzyloxymethyl-1-[4-[N'-(2-chlorophenyl) 10 ureido]-3-methoxyphenylacetyl]-2-(S)-pyrrolidinylmethoxy]benzoate (169 mg, 0.25 mmol) in MeOH-THF (2 : 5, 7 ml) was added 1.0M-NaOH (760 ml, 0.76 mmol) at rt, and the resulting mixture was heated at 60 0 C with stirring for 1.5 h. The reaction mixture was poured into 1N-HC1, then extracted with CHCl 3 . The organic layer was washed with brine, dryed over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHC1 3 15 MeOH (5 : 1) as eluent to give 130 (155 mg, 94%) as a colorless amorphous solid. MW 658.14 'H-NMR (CD 3 OD), mixture of rotamars 5 1.89-2.35 (series of m, 4 H), 3.52-4.53 (series of m, 8 H), 3.68 and 3.75 (s, total 3 H), 4.85 (s, 2 H), 6.68-8.03 (series of m, 16 H); MS (ESI) m/z, 658 (M +H), 680 (M*+Na ); Anal. Calcd for C 3
,H
36 C1N 3 0H 2 0: C, 63.95; H, 5.66; N, 6.21. Found: C, 64.01; H, 5.38: N, 5.96. 20 Example 123 3-[5-(R)-benzyloxymethyl-1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-2-(S) pyrrolidinylmethoxy]benzoic acid NQI COOH N N r H H Me 131 To a stirred solution of methyl 3-[2-(S)-(5-(S)-benzyloxymethyl)pyrrolidinylmethoxy] 25 benzoate (116 mg, 0.33 mmol), 4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (124 mg, 0.33 mmol) and N,N-dimethylaminopyridine (48.0 mg, 0.39 mmol) in THF (15 ml) was added EDCHCl (75.0 mg, 0.39 mmol) at rt, and the resulting mixture was stirred overnight. The reaction mixture was poured into water and extracted with CHC1 3 . The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was 30 chromatographed on silica gel with CHCl 3 -MeOH (10 : 1) as eluent to give methyl 3-[5-(R) 287 WO 01/00206 PCT/US00/18079 benzyloxymethyl-1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-2-(S)-pyrrolidinyl methoxy]benzoate (209 mg, 89%) as a colorless amorphous solid. 'H-NMR (CDC13), mixture of rotamars 8 1.99-2.37 (series of m, 4 H), 3.43-3.91 (series of m, 5 H), 3.70 (s, 3 H), 3.93 and 3.96 (s, total 3 H), 4.19-4.28 (m, 2 H), 4.37-4.51 (m, 1 H), 4.60 (s, 2 H), 6.77-7.11 (series of m, 6 H), 5 7.28-7.74 (series of m, 10 H), 7.91-7.95 (series of m, 1 H), 8.20-8.23 (series of m, 1 H); MS (ESI) m/z 716 (M+), 718 (M'+2). To a stirred solution of methyl 3-[5-(R)-benzyloxymethyl-1-[4-[N'-(2-bromophenyl) ureido]-3-methoxyphenylacetyl]-2-(S)-pyrrolidinylmethoxy]benzoate (176 mg, 0.25 mmol) in MeOH-THF (2: 5, 7 ml) was added 1.0M-NaOH (760 ml, 0.76 mmol) at rt, and the resulting 10 mixture was heated at 60oC with stirring for 1.5 h. The reaction mixture was poured into IN-HC1, then extracted with CHCI 3 . The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 MeOH (5 : 1) as eluent to give 131 (156 mg, 88%) as a colorless amorphous solid. MW 702.59 'H-NMR (CD 3 OD), mixture of rotamars 5 1.89-2.37 (series of m, 4 H), 2,29 (s, 3 H), 3.52-4.53 15 (series of m, 8 H), 3.68 and 3.75 (s, total 3 H), 4.85 (s, 2 H), 6.67-7.95 (series of m, 16 H); MS (ESI) m/z, 702 (M+H), 704 (M*+Na'); Anal. Calcd for C 36
H
36 BrN 3 0 7
H
2 0: C, 60.00; H, 5.32; N, 5.83. Found: C, 59.65; H, 5.02: N, 5.65. Example 124 4-[(2S,4S)-4-methoxy- 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]l-2-pyrrolidinyl] 20 methoxybenzoic acid OMe ON OCCOOH N N O Me H H Me 132 To a stirred mixture of (2S,4R)-1-tert-butoxycarbonyl-2-tert-butyldiphenylsilyloxymethy 4-hydroxypyrrolidine (21.7 g, 47.6 mmol), acetic acid (3.0 ml, 52.4 mmol) and PPh 3 (12.5 g, 52.4 mmol) in THF (330 ml) was added DIAD (9.4 ml, 47.6 mmol) at room temperature under an 25 atmosphere of nitrogen. After 2 h stirring the same temperature, the mixture was heated at 50 oC for 2 h. After cooling to room temperature, the reaction mixture was concentrated in vacuo. The resulting residue was chromatographed on silica gel [1 Kg, n-hexane/EtOAc(5/1)], to give (2S,4S)-4-acetoxy-l-tert-butoxycarbonyl-2-tert-butyldiphenylsilyloxymethypyrrolidine (23.3 g, 99%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.06 (s, 9H), 1.35 and 1.43 (s, 9H, amide isomers), 30 1.92 (br, 3H), 2.20-2.45 (m, 2H), 3.31-4.07 (m, 5H), 5.17-5.30 (m, 1H), 7.36-7.44 (m, 6H), 7.65 288 WO 01/00206 PCT/US00/18079 7.71 (m, 4H). To a stirred mixture of (2S,4S)-4-acetoxy-1-tert-butoxycarbonyl-2-tert-butyldiphenyl silyloxy methypyrrolidine (23.3 g, 46.9 mmol) and acetic acid (6.0 ml, 104.8 mmol) in THF (470 ml) was added TBAF (93.8 ml, 93.8 mmol) at 0 oC. After 24 h stirring, the mixture was 5 concentrated in vacuo. The resulting residue was diluted with EtOAC and aq. NH 4 Cl and extracted with EtOAc. The combined extracts were washed with brine, which were dried over Na 2
SO
4 and concntrated in vacuo. The residue was chromatographed on silica gel [700 g, CHCl3 EtOAc (4/1)], to give (2S,4S)-4-acetoxy-l-tert-butoxycarbonyl-2-pyrrolidinemethanol (9.70 g, 8%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.47 (s, 9H), 1.63 (m, 1H1), 1.81 (m, 1H1), 2.07 (s, 3H), 10 2.34 (m, 1H), 3.42 (dd, J= 12.7, 0.9 Hz, 1H1), 3.62-3.85 (m, 3H), 4.48 (br, 1H1), 5.20 (br, 1H1). To a stirred mixture of (2S,4S)-4-acetoxy-1-tert-butoxycarbonyl-2-pyrrolidinemethanol (9.70 g, 37.4 mmol), p-hydroxybenzoic acid methyl ester (5.69 g, 37.4 mmol) and PPh 3 (10.8 g, 41.1 mmol) in THF (200 ml) was added DIAD (8.10 ml, 41.1 mmol) at room temperature. After 1.5 h stirring, the mixture was concentrated in vacuo. The resulting residue was chromatographed 15 on silica gel [700 g, CHCl 3 /EtOAc (10/1)], to give methyl 4-[(2S,4S)-4-acetoxy-l-tert butoxycarbonyl-2-pyrrolidinyl]methoxybenzoate (11.8 g, 81%) as a pale yellow oil. 'H-NMR (CDCl 3 ) 8 1.48 (s, 9H), 2.03 (s, 3H), 2.27 (m, 2H), 3.46 (m, 1H1), 3.72 (m, 1H), 3.88 (s, 3H), 3.98 (t, J= 9.0 Hz, 1H1), 4.21-4.47 (m, 2H), 5.31 (br, 1H), 6.96 (br, 2H), 7.98 (d, J= 8.8 Hz, 2H). To a stirred solution of methyl 4-[(2S,4S)4-acetoxy-1-tert-butoxycarbonyl-4-hydroxy-2 20 pyrrolidinyl]methoxybenzoate (7.43 g, 18.9 mmol) in MeOH (150 ml) was added cat. K 2 C0 3 at room temperature. After 1 day stirring, the mixture was concentrated in vacuo. The resulting residue was recrystallized by the addition of CHC13-n-hexane, to give methyl 4-[(2S,4S)-1-tert butoxycarbonyl-4-hydroxy-2-pyrrolidinyl]methoxybenzoate (5.76 g, 87%) as a colorless solid. 'H NMR (CDC1 3 ) 8 1.46 (s, 9H), 2.11 (m, 111), 2.35 (br, 1H1), 3.27-3.65 (m, 2H), 3.89 (s, 3H), 4.07 25 4.54 (m, 4H), 6.96 (d, J= 6.9 Hz, 2H), 7.99 (d, J = 6.9 Hz, 2H). To a stirred solution of methyl 4-[(2S,4S)-1-tert-butoxycarbonyl-4-hydroxy-2-pyrrolidinyl] methoxybenzoate (2.10 g, 5.98 mmol) in THF (60 ml) was added 60% oil NaH (359 mg, 8.97 mmol) at 0 oC. After 15 minutes stirring, Mel (1.20 ml, 8.97 mmol) was added to the mixture was added at same temperature, and the resulting mixture was allowed to raise to room temperature for 30 over 1 h. Then 60% oil NaH (359 mg, 8.97 mmol) and Mel (1.20 ml, 8.97 mmol) was added to 289 WO 01/00206 PCT/US00/18079 the reaction mixture at room temperature and stirred for 14 h. The reaction mixture was poured into ice water and extracted with CHC13. The combined extracts were washed with aq. NaHCO 3 and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, n-hexane/EtOAc(4/1)], to give methyl 4-[(2S,4S)-1-tert 5 butoxycarbonyl-4-methoxy-2-pyrrolidinyl]methoxybenzoate (1.32 g, 60%) as a colorless oil. 'H NMR (CDCI 3 ) 8 1.48 (s, 9H), 2.05 (m, 1H), 2.29 (d, J= 14.2 Hz, 1H), 3.30 (s, 3H), 3.36-4.38 (m, 4H), 6.76 (br, 2H), 7.97 (d, J= 8.8 Hz, 2H). To a stirred solution of methyl 4-[(2S,4S)--tert-butoxycarbonyl-4-methoxy-2 pyrrolidinyl] methoxybenzoate (2.38 g, 3.61 mmol) in CH 2 C1 2 (46 ml) was added TFA (23 ml) at 10 room temperature. After 14 h stirring, the mixture was concentrated in vacuo. The residue was diluted by the addition of CH 2 Cl 2 and 1 N NaOH, and extracted with CH 2 Cl2. The combined extracts were washed with brine, dried over Na 2
SO
4 , which was concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHCl3/MeOH (20/1)], to give methyl 4-[(2S,4S) 4-methoxy-2-pyrrolidinyl]methoxybenzoate (950 mg, 99%) as a yellow oil. 'H-NMR (CDCl 3 ) 8 15 2.16 (t,J= 5.3 Hz, 1H), 2.72 (s, 1H), 2.95 (d, J= 6.8 Hz, 1H), 3.11 (d,J= 11.0 Hz, lH), 3.26 (t,J = 1.9 Hz, 3H), 3.52 (br, 1H), 3.84 (d, J= 1.7 Hz, 3H), 3.92 (s, 1H), 4.00 (d, J= 4.1 Hz, 2H), 6.88 (m, 2H), 7.94 (m, 2H). A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (375 mg, 1.19 nummol), methyl 4-[(2S,4S)-4-methoxy-2-pyrrolidinyl]methoxybenzoate (317 mg, 1.19 mmol), 20 EDC-HCI (342 mg, 1.79 mmol), HOBT (242 mg, 1.79 mmol) and Et 3 N (0.83 ml, 5.95 mmol) in DMF (5 ml) was stirred at room temperature for 13 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHCl/Aectone (10/1)CHCl 3 /MeOH (20/1)], to give methyl 4-[(2S,4S)-4-methoxy-1-[3 25 methoxy-4-[N '-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrffolidinyl]methoxybenzoate (650 mg, 98%) as a pale brown amorphous solid 'H-NMR (CDCl 3 ) 6 2.03 (m, 1H), 2.31 (s, 3H), 2.32 (m, 1H), 3.29 (d, J= 1.0 Hz, 3H), 3.57-3.68 (m, 5H), 3.88 (d, J = 1.0 Hz, 3H), 3.99-4.06 (m, 2H), 4.46 (m, 1H), 6.19 (m, 1H), 6.80 (s, 1H), 6.81 (d, J= 9.0 Hz, 1H), 6.96-7.19 (m, 4H), 7.29 (m, 2H), 7.50 (d, J = 6.7 Hz, 1H), 7.95-8.10 (m, 3H); MS (ESI) nm/z 562 (M++1). 30 To a solution of methyl 4-[(2S,4S)-4-methoxy-1-[3-methoxy-4-[N'-(2-methylphenyl) ureido] phenylacetyl]-2-pyrrolidinyl]methoxybenzoate (650 mg, 1.16 mmol) in THF (18.5 ml), 290 WO 01/00206 PCT/US00/18079 0.25 N NaOH (18.5 ml) was added. After stirring at room temperature for 12 h, the mixture was acidified with 1 N HCI and extracted with CHCl 3 -MeOH (10/1). The combined extracts were dried over Na 2
SO
4 and concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHCI3/MeOH (20/1)] to give 132 (540 mg, 85%) as a colorless amorphous solid. MW 547.60 IR 5 (KBr) 3354, 2937, 1709, 1685, 1604, 1533, 1454 cm-'; 'H-NMR (DMSO-d 6 ) 6 2.11 (mn, 2H), 2.25 (s, 3H), 3.22 (s, 3H), 3.49-3.78 (min, 4H), 3.82 and 3.86 (s, 3H11, amide isomers), 3.87-4.52 (m, 4H), 6.71-7.17 (min, 7H11), 7.79 (d,J= 8.1 Hz, 1H), 7.86-8.03 (min, 3H), 8.45-8.57 (min, 2H), 12.64 (br, 1H); MS (ESI) m/z 548 (M +1); Anal. Calcd for C 30 oH 3 3
NO
7 INa-1.5H 2 0: C, 60.29; H, 6.07; N, 7.03. Found: C, 59.90; H, 5.59; N, 6.69. 10 Example 125 4-[(2S,4S)- 1-[4-[N '-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-4-methoxy-2-pyrrolidinyl] methoxybenzoic acid OMe e N C O O H NN 0 I HH Me 133 A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (398 mg, 1.19 15 mmol), methyl 4-[(2S,4S)-4-methoxy-2-pyrrolidinyl]methoxybenzoate (317 mg, 1.19 mmol), EDC-HCI (342 mg, 1.79 mmol), HOBT (242 mg, 1.79 mmol) and Et 3 N (0.83 ml, 5.95 mmol) in DMF (5 ml) was stirred at room temperature for 13 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica 20 gel [50 g, CHClJ3/Aectone (10/1)CHCl 3 /MeOH (20/1)], to give methyl 4-[(2S,4S)-1-[4-[N'-(2 chlorophenyl)ureido]-3-methoxyphenylacetyl]-4-methoxy-2-pyrrolidinyl]methoxybenzoate (600 mg, 87%) as a colorless amorphous solid 'H-NMR (CDCI 3 ) 8 1.99-2.06 (min, 111), 2.34 (d, J= 13.9 Hz, 1H), 3.30 (s, 3H), 3.59 (d, J= 7.4 Hz, 111), 3.62 (d, J= 3.2 Hz, 2H11), 3.83 (s, 3H), 3.88 (s, 3H), 4.00-4.18 (m, 3H11), 4.42-4.51 (min, 2H), 6.82-7.07 (min, 711), 7.28 (d, J= 8.3 Hz, 1H), 7.35 (dd, J 25 7.9, 1.5 Hz, 111), 7.94-8.00 (min, 3H),8.18(d, J= 8.3Hz, 1I); MS (ESI) m/z 582 (M+1), 584 (MW+3). To a solution of methyl 4-[(2S,4S)-1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl acetyl]-4-methoxy-2-pyrrolidinyl]methoxybenzoate (600 mg, 1.03 mmol) in THF (16 ml), 0.25 N NaOH (16 ml) was added. After stirring at room temperature for 12 h, the mixture was acidified 30 with 1 N HCI and extracted with CHCl 3 -MeOH (10/1). The combined extracts were dried over Na 2
SO
4 and concentrated in vacuo. The residue was purified on TLC [CHCl3/MeOH (10/1)] to 291 WO 01/00206 PCT/US00/18079 give 133 (495 mg, 75%) as a colorless amorphous solid. MW 568.02 IR (KBr) 3330, 3070, 2937, 1709, 1685, 1604, 1533 cm-'; 'H-NMR (DMSO-d 6 ) 8 2.11 (m, 2H11), 3.22 (s, 3H11), 3.56-3.78 (m, 4H11), 3.81 and 3.85 (s, 3H, amide isomers), 3.88-4.56 (m, 4H), 6.73 and 6.77 (d, J= 8.1 Hz, 1H, amide isomers), 6.85 and 6.91 (s, 1H, amide isomers), 7.01-7.07 (m, 3H11), 7.28 (t, J= 8.1 Hz, 1H11), 5 7.43 (d, J = 8.1 Hz, 1H11), 7.85-7.94 (m, 2H11), 7.97 (d, J = 8.6 Hz, 1H11), 8.09 (d, J = 8.3 Hz, 1H), 8.90-8.95 (m, 2H); MS (FAB) m/z 570 (M +1), 572 (M'+3). Example 126 4-[(2S,4S)- -[4-[N '-(2-bromophenyl)ureidol-3-methoxyphenylacetyl]-4-methoxy-2-pyrrolidinyl] methoxybenzoic acid OMe N ,jaCOOH NN O 10 r H H OMe 134 A mixture of 4-[N'-(2-bormophenyl)ureido]-3-methoxyphenylacetic acid (451 mg, 1.19 mmol), methyl 4-[(2S,4S)-4-methoxy-2-pyrrolidinyl]methoxybenzoate (317 mg, 1.19 mmol), EDC-HCI (342 mg, 1.79 mmol), HOBT (242 mg, 1.79 mmol) and Et 3 N (0.83 ml, 5.95 mmol) in DMF (5 ml) was stirred at room temperature for 13 h. The mixture was poured into ice water and 15 extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHCl/Aectone (10/1)], to give methyl 4-[(2S,4S)-l-[4-[N'-(2-bromophenyl)ureido]-3 methoxy phenylacetyl]-4-methoxy-2-pyrrolidinyl]methoxybenzoate (760 mg, 100%) as a yellow oil. 1H-NMR (CDC 3 ) 8 1.99-2.32 (m, 1H11), 2.34 (d, J= 13.4 Hz, 1H11), 3.30 (s, 3H), 3.59 (m, 1H), 20 3.63 (d,J= 3.2 Hz, 2H11), 3.68 (dd, J= 12.2, 5.1 Hz, 1H), 3.81 (br, 3H), 3.88 (s, 3H), 3.91-4.16 (m, 2H), 4.49-4.51 (m, 2H), 6.82-7.15 (m, 7H11), 7.31 (t, J= 8.1 Hz, 1H11), 7.52 (d,J= 8.1Hz, 1H11), 7.93 8.00 (m, 3H), 8.14 (d, J= 8.3 Hz, 1H); MS (ESI) m/z 626 (M+1), 628 (M*+3). To a solution of methyl 4-[(2S,4S)-1-[4-[N'-(2-bromophenyl)ureido]phenylacetyl]-4 methoxy-2-pyrrolidinyl]methoxybenzoate (760 mg, 1.19 mmol) in THF (19 nml), 0.25 N NaOH (19 25 ml) was added. After stirring at room temperature for 12 h, the mixture was acidified with 1 N HCI and extracted with CHCl 3 -MeOH (10/1). The combined extracts were dried over Na 2
SO
4 and concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHCl 3 /MeOH (20/1)] to give 134 (580 mg, 78%) as a colorless amorphous solid. MW 612.47 IR (KBr) 3330, 2935, 1709, 1685, 1604, 1529, 1434 cm-'; 'H-NMR (DMSO-d 6 ) 8 2.11 (m, 2H11), 3.22 (s, 3H11), 3.58 30 3.78 (m, 4H), 3.81 and 3.86 (s, 3H11, amide isomers), 3.92-4.52 (m, 4H), 6.72 (d, J= 8.6 Hz, 1H), 6.77 (d, J= 8.3 Hz, 1H), 6.85 and 6.91 (s, 1H, amide isomers), 6.97 (t, J= 7.1 Hz, 1H1), 7.02 and 292 WO 01/00206 PCT/US00/18079 7.06 (d, J= 8.6 Hz, 2H, amide isomers), 7.32 (t, J= 7.3 Hz, 1H), 7.59 (dd, J= 8.1, 1.0 Hz, 1H), 7.94 (dd, J = 8.1, 1.2 Hz, 2H), 7.95-7.98 (m, 2H), 8.74 (s, 1H), 8.94 (s, 1H), 12.63 (br, 1H); MS (FAB) m/z 612 (M-+1), 614 (M +3). Example 127 5 4-[(4R)-methoxy- 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S)-pyrrolidinyl methoxy]benzoic acid pMe N SN N \COOH 13 MeH HMe Oa OH 135 To a stirred solution of 1-(tert-butoxycarbonyl)-(4R)-methoxy-(2S)-pyrrolidinylcarboxylic acid (2.87 g, 11.7 mmol) in THF (25 ml) was added BH 3 DMS (1.66 ml, 17.5 mmol) at room 10 temperature and the reaction mixture was stirred at room temperature overnight. The mixture was evaporated and the residue was dissolved with CH 2 Cl 2 . The solution was washed with H 2 0, brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica gel with CHCl 3 -MeOH (50:1, v/v) as eluent to give 1-(tert-butoxycarbonyl)-(4R)-methoxy-(2S) prolinol (1.79 g, 66%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.47 (s, 9 H), 1.69-1.73 (m, 1 H), 15 2.12-2.17 (m, 1 H), 3.31 (s, 3 H), 3.37-3.40 (m, 1 H), 3.53-3.62 (m, 2 H), 3.68-3.73 (m, 1 H), 3.83 3.87 (m, 1 H), 4.04-4.07 (m, 1 H), 4.90-4.92 (m, 1 H); MS (FAB) m/z 232 (M+1). To a stirred solution of methyl 4-hydroxybenzoate (1.18 g, 7.76 mmol), 1-(tert-butoxy carbonyl)-(4R)-methoxy-(2S)-prolinol (1.79 g, 7.74 mmol) and Ph 3 P (2.44 g, 9.30 mmol) in THF (30 ml) was added DIAD (1.83 ml, 9.29 mmol) and the reaction mixture was heated under reflux 20 for 5 hr. After cooled to room temperature, the mixture was evaporated. The residue was filtered on silica-gel with toluene-acetone (5:1, v/v) as eluent to give the crude product. The crude product was dissolved in CH 2 C1 2 (20 ml). The solution was added TFA (20 ml) and the reaction mixture was stirred at room temperature overnight. The mixture was concentrated in vacuo and made basic by sat. NaHCO 3 . The mixture was extracted with CHCI 3 , washed with brine, dried over 25 K 2 C0 3 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (30:1 to 30:2, v/v) as eluent to give methyl 4-[(4R)-methoxy-(2S)-pyrrolidinyl methoxy]benzoate (1.67 g, 81% for 2 steps) as a reddish brown oil. 'H-NMR (CDCl 3 ) 8 1.65-1.72 (m, 1 H), 1.89 (bs, 1 H), 2.05-2.22 (m, 1 H), 2.95-3.15 (m, 2 H), 3.31 (s, 3 H), 3.69-3.76 (m, 1 H), 3.88 (s, 3 H), 3.91-4.06 (m, 3 H), 6.89-6.92 (m, 2 H), 7.96-7.98 (m, 2 H); MS (FAB) m/z 266 30 (M+1). 293 WO 01/00206 PCT/US00/18079 A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (470 mg, 1.50 mmol), methyl 4-[(4R)-methoxy-(2S)-pyrrolidinylmethoxy]benzoate (396 mg, 1.49 mmol), EDCHCI (343 mg, 1.79 mmol), HOBt (242 mg, 1.79 mmol) and Et 3 N (250 ml, 1.79 mmol) in THF (10 ml) was stirred at room temperature overnight. The mixture was diluted with HO 2 0 and 5 extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1, v/v) as eluent to give methyl 4-[(4R)-methoxy-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl acetyl]-(2S)-pyrrolidinylmethoxy]benzoate (822 mg, 98%) as a white foam. 'H-NMR (CDCl 3 ) 8 2.14-2.24 (min, 2 H), 2.27 (s, 3 H), 3.25 (s, 3 H), 3.51 (s, 3 H), 3.58-3.73 (mn, 4 H), 3.88 (s, 3 H), 10 3.98-4.09 (min, 2 H), 4.40-4.53 (min, 2 H), 6.67-7.29 (series of m, total 9 H), 7.57-7.59 (mn, 1 H), 7.91-7.93 (min, 2 H), 8.04-8.06 (m, 1 H); MS (FAB) m/z 562 (M++1). To a stirred solution of methyl 4-[(4R)-methoxy-1-[3-methoxy-4-[N'-(2-methylphenyl) ureido] phenylacetyl]-(2S)-pyrrolidinylmethoxy]benzoate (517 mg, 0.92 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 3 hr. After cooled 15 to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was collected under a reduced pressure. The crude solid was purified by recrystallization from MeOH CHCl 3 -IPE to give 135 (144 mg, 29%) as a white crystalline powder. MW 547.60 mp 112 115oC; 'H-NMR (DMSO-d 6 ) 8 2.04-2.17 (min, 2 H), 2.25 (s, 3 H), 3.21 (s, 3 H), 3.56-3.75 (min, 4 H), 3.79 (s, 3 H), 4.04-4.35 (min, 4 H), 6.73-7.17 (series of m, total 7 H), 7.79-7.81 (mn, 1 H), 7.87-7.89 20 (min, 2 H), 7.99-8.01 (mn, 1 H), 8.47 (s, 1 H), 8.55 (s, 1 H), 12.63 (bs, 1 H); MS (FAB) m/z 548 (M+1); AnaL. Calcd for C 30
H
33
N
3 0 7 l/4H 2 0: C, 65.26; H, 6.12; N, 7.61. Found: C, 65.36; H, 6.45; N, 7.24. Example 128 4-[1 -[4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenylacetyl]-(4R)-methoxy-(2S)-pyrrolidinyl 25 methoxy]benzoic acid QMe NN 0 --- COOH H H Me 136 A mixture of 4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenylacetic acid (476 mg, 1.50 mmol), methyl 4-[(4R)-methoxy-(2S)-pyrrolidinylmethoxy]benzoate (397 mg, 1.50 mmol), EDCHCI (344 rmg, 1.79 mmol), HOBt (243 mg, 1.80 mmol) and Et 3 N (250 ml, 1.79 mmol) in 30 THF (10 ml) was stirred at room temperature overnight. The mixture was diluted with H 2 0 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. 294 WO 01/00206 PCT/US00/18079 The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1, v/v) as eluent to give methyl 4-[1-[4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenylacetyl]-(4R) methoxy-(2S)-pyrrolidinylmethoxy]benzoate (806 mg, 95%) as a pale yellow foam. 'H-NMR (CDCl 3 ) 8 2.14-2.37 (m, 2 H), 3.28 (s, 3 H), 3.44 (s, 3 H), 3.48-3.74 (m, 4 H), 3.88 (s, 3 H), 4.02 5 4.15 (m, 2 H), 4.43-4.58 (m, 2 H), 6.63-7.10 (series of m, total 7 H), 7.68-7.73 (m, 1 H), 7.89-8.02 (m, 4 H), 8.16-8.20 (m, 1 H); MS (FAB) m/z 566 (MW+1). To a stirred solution of methyl 4-[1-[4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenyl acetyl]-(4R)-methoxy-(2S)-pyrrolidinylmethoxy]benzoate (491 mg, 0.87 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 3 hr. After cooled 10 to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was collected under a reduced pressure. The crude solid was purified by recrystallization from MeOH CHCl 3 -IPE to give 136 (173 mg, 36%) as a white crystalline powder. MW 551.56 mp 111-116 0 C; 1 H-NMR (DMSO-d 6 ) 8 2.08-2.17 (m, 2 H), 3.21 (s, 3 H), 3.56-3.73 (m, 4 H), 3.78 (s, 3 H), 4.04 4.33 (m, 4 H), 6.74-7.22 (series of m, total 7 H), 7.87-7.89 (m, 2 H), 7.99-8.01 (m, 1 H), 8.16-8.20 15 (m, 1 H), 8.70 (s, 1 H), 9.18 (s, 1 H), 12.64 (br s, 1 H); MS (FAB) m/z 552 (M*+1); Anal. Calcd for
C
29
H
3
FN
3
O
7 0.15H 2 0: C, 62.84; H,5.51; F,3.43; N,7.58. Found:C, 63.08; H, 5.83; F, 3.30; N, 7.15. Example 129 4-[1-[4-[N '-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(4R)-methoxy-(2S)-pyrrolidiny methoxy]benzoic acid pMe SN N 0 COOH 20 IHH Me 137 A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (460 mg, 1.37 mmol), methyl 4-[(4R)-methoxy-(2S)-pyrrolidinylmethoxy]benzoate (365 mg, 1.38 mmol), EDCHC1 (316 mg, 1.65 mmol), HOBt (223 mg, 1.65 mmol) and Et 3 N (230 ml, 1.65 mmol) in THF (10 ml) was stirred at room temperature overnight. The mixture was diluted with H20 and 25 extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1, v/v) as eluent to give methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(4R) methoxy-(2S)-pyrrolidinylmethoxy]benzoate (801 mg, q. y.) as a white foam. 'H-NMR (CDCI 3 ) 8 2.13-2.36 (m, 2 H), 3.27 (s, 3 H), 3.58 (s, 3 H), 3.61-3.73 (m, 4 H), 3.88 (s, 3 H), 4.06-4.14 (m, 2 30 H), 4.43-4.56 (m, 2 H), 6.70-6.99 (series of m, total 5 H), 7.23-7.42 (m, 4 H), 7.90-8.00 (m, 3 H), 8.17-8.20 (m, 1 H); MS (FAB) m/z 582 (M'+1). 295 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl acetyl]-(4R)-methoxy-(2S)-pyrrolidinylmethoxy]benzoate (541 mg, 0.93 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 3 hr. After cooled to room temperature, the mixture was poured into ice-1 N HC1 and the resulting precipitate was 5 collected under a reduced pressure. The crude solid was purified by recrystallization from MeOH CHC1 3 -IPE to give 137 (281 mg, 53%) as a white crystalline powder. MW 568.02 mp 116-119 0 C; 'H-NMR (DMSO-d 6 ) 8 2.08-2.17 (min, 2 H), 3.21 (s, 3 H), 3.56-3.73 (min, 4 H), 3.79 (s, 3 H), 4.04 4.33 (min, 4 H), 6.75 (d, J= 8.3 Hz, 1 H), 6.87 (s, 1 H), 7.02 (d, J = 8.3 Hz, 3 H), 7.28 (t, J =7.8Hz, 1 H), 7.44 (d, J = 7.8 Hz, 1 H), 7.87-7.89 (min, 2 H), 7.96 (d, J = 8.3 Hz, 1 H), 8.10 (d, J = 8.3 Hz, 10 1 H), 8.89 (s, 1 H), 8.93 (s, 1 H), 12.63 (br s, 1 H); MS (FAB) m/z 568 (M+ 1); Anal. Calcd for
C
29
H
30 C1N 3 0 7 l/4H 2 0: C,60.84;H,5.37;Cl,6.19;N,7.34. Found: C, 61.03; H, 5.56; Cl, 6.27; N, 7.03. Example 130 4-[ 1 -[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-(4R)-methoxy-(2S)-pyrrolidinyl methoxy]benzoic acid pMe 5N'N 0 COOH 15 rH H Me 138 A mixture of 4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (600 mg, 1.58 mmol), methyl 4-[(4R)-methoxy-(2S)-pyrrolidinylmethoxy]benzoate (420 mg, 1.58 mmol), EDCHCl (364 mg, 1.90 mmol), HOBt (214 mg, 1.58 mmol) and Et 3 N (265 mnl, 1.90 mmol) in THF (15 ml) was stirred at room temperature overnight. The mixture was diluted with H 2 0 and 20 extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHC13-MeOH (100:1, v/v) as eluent to give methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-(4R) methoxy-(2S)-pyrrolidinylmethoxy]benzoate (1.01 g, q.y.) as a pale yellow foam. 1 H-NMR (CDC13) 8 2.13-2.33 (min, 2 H), 3.27 (s, 3 H), 3.57 (s, 3 H), 3.61-3.72 (min, 4 H), 3.88 (s, 3 H), 4.05 25 4.14 (min, 2 H), 4.43-4.57 (min, 2 H), 6.70-7.00 (series of m, total 5 H), 7.29-7.52 (min, 4 H), 7.92-8.01 (m, 3 H), 8.12-8.15 (min, 1 H); MS (FAB) m/z 626 (M+1). To a stirred solution of methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl acetyl]-(4R)-methoxy-(2S)-pyrrolidinylmethoxy]benzoate (697 mg, 1.11 mmol) in THF (8 mnil) was added 0.5 N NaOH (8 ml) and the reaction mixture was heated under reflux for 2 hr. After cooled 30 to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was collected under a reduced pressure. The crude solid was purified by recrystallization from MeOH 296 WO 01/00206 PCT/US00/18079 CHC13-IPE to give 138 (252 mg, 37%) as a white crystalline powder. MW 612.47 mp 125-130oC; 'H-NMR (DMSO-d) 5 2.08-2.17 (m, 2 H), 3.21 (s, 3 H), 3.60-3.72 (m, 4 H), 3.79 (s, 3 H), 3.95 4.33 (m, 4 H), 6.75-7.08 (series of m, total 5 H), 7.31-7.34 (m, 1 H), 7.59-7.61 (m, 1 H), 7.87-7.89 (m, 2 H), 7.93-7.96 (m, 2 H), 8.73 (s, 1 H), 8.91 (s, 1 H), 12.63 (br s, 1 H); MS (FAB) m/z 612 5 (M'+1); Anal. Calcd for C 2 9
-H
3 0 BrN 3 07: C, 56.87; H, 4.94; Br, 13.05; N, 6.86. Found: C, 56.67; H, 4.97; Br, 13.07; N, 6.68. Example 131 4-[4,4-difluoro-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinylmethoxy] benzoic acid F..F NYN 0 \/-COOH 10 Me H H Me 139 To a stirred solution of N-Boc proline methyl ester (2 .0 g, 8.15 mmol) in CH 2 C1 2 were added 3 A molecular sieves (2 g) and PDC (4.60 g, 12.2 mmol). The mixture was stirred for 3 days. The mixture was filtered through a Celiete pad and the filtrate was evaporated. The residue was chromatographed on silica gel with CHCl 3 -MeOH (10:1) as eluent to give methyl 1-(tert 15 butoxycarbonyl)-4-oxopyrrolidine-2-carboxylate (1.13 g, 57%) as a colorless oil. 'H-NMR (CDC1 3 ) 8 1.46-1.48 (m, 9 H), 2.56-2.61 (m, 1 H), 2.88-3.00 (m, 1 H), 3.77 (s, 3 H), 3.82-3.88 (m, 2 H), 4.71-4.83 (m, 1 H). To a cold (-78 0 C), stirred solution of methyl 1-(tert-butoxycarbonyl)-4-oxopyrrolidine-2 carboxylate (1.13 g, 4.65 mmol) in CH 2
CI
2 (20 ml) was added methylDAST (1.1 ml, 11.6 mmol). 20 The mixture was allowed to warm to room temperature. After 15 h stirring, the mixture was poured into H20 (50 ml) and extracted with EtOAc (200 ml). The extract was washed with brine (2 x 200 ml), dried over MgSO 4 , and evaporated. The residue was chromatographed on silica gel with CHC13-EtOAc (20:1) as eluent to give methyl 1-(tert-butoxycarbonyl)-4,4-difluoropyrrolidine -2-carboxylate (885 mg, 72%) as a yellow oil. 'H-NMR (CDCl 3 ) 8 1.42 and 1.47 (s, each, total 9 25 H), 2.46 (ddd, d= 26.9 13.7, 5.1 Hz,1 IH), 2.62-2.78 (m, 1H), 3.75-3.95 (m, 5H), 4.43-4.57 (m,1H). To a stirred solution of methyl 1-(tert-butoxycarbonyl)-4,4-difluoropyrrolidine-2-carboxylate (885 mg, 3.34 mmol) in THF (25 ml) was added 0.25 N NaOH (26.7 ml, 6.67 mmol) and the stirring was continued for 1 h. The mixture was poured into 1 N HCI (100 ml) and-extracted with CHCI 3 (2 x 200 ml). The combined extracts were washed with brine (100 ml), dried over MgSO 4 , and 30 evaporated to give 1-(tert-butoxycarbonyl)-4,4-difluoropyrrolidine-2-carboxylic acid (775 mg, 297 WO 01/00206 PCT/US00/18079 92%) as a yellow crystalline solid. mp 113-117 oC; IH-NMR (CDCI 3 ) 8 1.44 and 1.49 (s, each, total 9 H), 2.53-2.80 (m, 2 H), 3.71-3.90 (m, 2 H), 4.20-4.61 (m, 1 H); MS (FAB) m/z, 252 (1±+l); Anal. Calcd for CIoH 15
F
2 0 4 : C, 47.81; H, 6.02; N, 5.58. Found: C, 48.06; H, 6.05; N, 5.45. 5 To a stirred solution ofN-(tert-butoxycarbonyl) 4,4-difluoroproline (3.00 g, 11.9 mmol) in THF (20 ml) was added BH 3 "DMS (1.1 ml, 11.9 mmol) at room temperature. The mixture was heated at reflux for 2 h. After cooling to room temperature, the mixture was concntrated in vacuo. The residue was quenched by the addition of H 2 0 (100 ml) and extracted with CHCl 3 (2 x 200 ml). The combined extracts were dried over MgSO 4 and evaporated. The residue was chromatographed 10 on silica gel with CHCl 3 -EtOAc (4:1) as eluent to give 1-(tert-butoxycarbonyl)-4,4-difluoro-2 pyrrolidinylmethanol (2.11 g, 75%) as a colorless oil. 'H-NMR (CDC13) 8 1.48 (s, 9 H), 2.04-2.55 (m, 2 H), 3.59-4.17 (m, 5 H). To a stirred mixture of 1-(tert-butoxycarbonyl)-4,4-difluoro-2-pyrrolidinylmethanol (600 mg, 2.53 mmol), methyl 4-hydroxybenzoate (462 mg, 3.03 mmol), Ph 3 P (795 mg, 3.03 mmol) in THF (10 15 ml) was added DIAD (597 ul, 3.03 mmol) at room temperature. The mixture was heated at reflux for 3 h with stirring. After cooling to room temperature, the mixture was concntrated in vacuo. The residue was chromatographed on silica gel with hexane-EtOAc (4:1) as eluent to give methyl 4-[1-(tert-butoxycarbonyl)-4,4-difluoro-2-pyrrolidinylmethoxy]benzoate (831 mg, 88%) as a colorless oil. 'H-NMR (CDC13) 8 1.48 (s, 9 H), 2.53-2.61 (m, 2 H), 3.63-4.41 (series of m, total 8 20 H), 6.94 (d, J= 8.8 Hz, 2 H), 7.99 (d, J= 8.8 Hz, 2 H). A mixture of methyl 4-[1-(tert-butoxycarbonyl)-4,4-difluoro-2-pyrrolidinylmethoxy] benzoate (830 mg, 2.23 mmol) and TFA (5 ml) in CH 2 Cl 2 (5 ml) was stirred for 3 h and concntrated in vacuo. The residue was made basic with sat. NaHCO 3 and extracted with CHC1 3 (2 x 200 ml). The combined extracts were dried over K 2 C0 3 and concntrated in vacuo to give methyl 25 4-(4,4-difluoro-2-pyrrolidinylmethoxy)benzoate (550 mg, 91%) as a pale yellow solid. 'H-NMR (CDC13) 8 2.19 (m, 1 H), 2.43 (m, 1 H), 3.19-3.41 (m, 2 H), 3.77 (m, 1 H), 3.89 (s, 3 H), 4.00-4.09 (m, 2 H), 6.92 (d, J= 9.0 Hz, 2 H), 7.99 (d, J = 9.0 Hz, 2 H); MS (FAB) m/z 272 (MW+1); Anal. Calcd for C 13
H,
15
F
2
NO
3 : C, 57.56; H, 5.57; N, 5.16. Found: C, 57.65; H, 5.67; N, 5.16. A mixture of methyl 4-(4,4-difluoro-2-pyrrolidinylmethoxy)benzoate (540 mg, 1.99 30 mmol), 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (626 mg, 1.99 mmol), 298 WO 01/00206 PCT/US00/18079 EDC-HCI (572 mg, 2.99 mmol), HOBt (cat.), and DMAP (cat.) in DMF (10 ml) was stirred overnight. The mixture was diluted with EtOAc (300 ml), washed with brine (2 x 100 ml), dried over MgSO 4 , and concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (20:1) as eluent to give methyl 4-[4,4-difluoro-1-[3-methoxy-4-[N'-(2 5 methylphenyl)ureido] phenylacetyl]-2-pyrrolidinylmethoxy]benzoate (1.00 g, 89%) as a colorless foam. 'H-NMR (CDCl 3 ) 8 2.31 (s, 3 H), 2.47-2.63 (m, 2 H), 3.52-3.97 (series of s and m, total 10 H), 4.07-4.30 (m, 2 H), 4.67-4.69 (m, 1 H), 6.45 (s, 1 H), 6.65 (d, J = 1.7 Hz, 1 H), 6.74-6.76 (m, 1 H), 6.84 (d, J= 8.8 Hz, 2 H), 7.14 (m, 2 H), 7.24 (m, 2 H), 7.52-7.54 (m, 1 H), 7.94 (d, J= 8.8 Hz, 2 H), 8.09 (d, J= 8.1 Hz, 1 H). 10 A mixture of methyl 4-[4,4-difluoro-l1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl acetyl]-2-pyrrolidinylmethoxy]benzoate (1.00 g, 1.76 mmol) and 0.25 N NaOH (14 ml, 3.50 mmol) in THF (14 ml) was stirred overnight. The mixture was acidified with 1 N HCI and extracted with CHCl 3 -MeOH (10:1, 2 x 200 ml). The combined extracts were dried over MgSO 4 and evaporated. The residue was chromatographed on silica gel with CHCl 3 -MeOH (20:1 to 10:1) as eluent to give 15 139 (658 mg, 68%) as a colorless crystalline powder. MW 553.55 mp 135-140 oC; 'H-NMR (DMSO-d 6 ) 8 2.23 (s, 3 H), 2.49-2.73 (m, 2 H), 3.36-4.55 (series of m, 10 H), 6.73 (d, J= 8.3 Hz, 1 H), 6.84 (s, 1 H), 6.93 (t, J= 7.3 Hz, 1 H), 7.00 (d, J= 8.3 Hz, 2 H), 7.10-7.16 (m, 2 H), 7.78 (d, J= 8.3 Hz, 1 H), 7.86 (d, J= 8.3 Hz, 2 H), 8.00 (d, J= 8.3 Hz, 1 H), 8.47 (s, 1 H), 8.56 (s, 1 H); MS (FAB) m/z, 554 (M+1); Anal. Calcd for C 29
H
29
F
2
N
3 0 6 -3/4H 2 0: C, 61.42; H, 5.44; N, 7.06. 20 Found: C, 61.30; H, 5.44; N, 7.06. Example 132 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-4,4-difluoro-2-pyrrolidinylmethoxy] benzoic acid F F N YN -- COOH 140 IHH Me 140 25 A mixture of methyl 4-(4,4-difluoro-2-pyrrolidinylmethoxy)benzoate (229 mg, 0.845 mmol), 4 [N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (283 mg, 0.845 mmol), EDC-HCI (243 mg, 1.27 mmol), HOBt (cat.), DMAP (cat.), and DMF (10 ml) was stirred overnight. The mixture was diluted with EtOAc (300 ml). The solution was washed with brine (2 x 100 ml), dried over MgSO 4 , and concntrated in vacuo. The residue was chromatographed on silica gel with CHC1 3 30 EtOAc (20:1 to 4:1) as eluent to give methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxy 299 WO 01/00206 PCT/US00/18079 phenylacetyl]-4,4-difluoro-2-pyrrolidinylmethoxy]benzoate (482 mg, 97%) as a colorless viscous solid. 'H-NMR (CDCl 3 ) 5 2.50-2.67 (m, 2 H), 3.54-4.71 (series of m, 13 H), 6.69 (d, J= 1.5 Hz, 1 H), 6.76 (d, J = 8.3 Hz, 1 H), 6.84 (d, J= 8.8 Hz, 2 H), 6.98 (dt, J= 7.8, 1.5 Hz, 1 H), 7.23-7.27 (m, 1 H), 7.33 (d, J= 8.3 Hz, 1 H), 7.39 (m, 2 H), 7.94 (d, J= 8.8 Hz, 2 H), 8.00 (d, J= 8.3 Hz, 1 5 H), 8.19 (dd, J= 8.3, 1.5 Hz, I H). A mixture of methyl methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl] 4,4-difluoro-2-pyrrolidinylmethoxy]benzoate (480 mg, 0.816 mmol), 0.25 N NaOH (6.5 ml, 1.65 mmol), and THF (20 ml) was stirred for 3 days. The mixture was poured into 1 N HCI (100 ml) and extracted with CHCl 3 -MeOH (5:1, 2 x 200 ml). The combined extracts were dried over 10 MgSO 4 and concntrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 MeOH (20:1 to 5:1) to give 140 (270 mg, 58%) as a pale yellow amorphous solid. MW 573.97 'H-NMR (DMSO-de) 8 2.45-2.74 (m, 2 H), 3.63-4.83 (series of m, 10 H), 6.76 (d, J= 8.3 Hz, 1 H), 6.87 (s, 1 H), 7.00-7.05 (m, 3 H), 7.26-7.30 (m, 1 H), 7.44 (dd, J=8.3, 1.2 Hz, 1 H), 7.88-7.93 (m, 2 H), 7.98 (d,J = 8.3 Hz, 1 H), 8.10 (d,J= 8.3 Hz, 1 H), 8.92 (s, 1 H), 8.96 (s, 1 H); MS (FAB) 15 m/z 574 (M +1); Anal. Calcd for C 28
H
2 6
CIF
2
N
3 0 6
-H
2 0: C, 56.81; H, 4.77; N, 7.10. Found: C, 56.75; H, 4.69; N, 6.79. Example 133 4-[(2R,3R,4S)-3,4-isopropylidenedioxy-1-[4-[N'-(2-methylphenyl)ureido]-3-methoxyphenylacetyl] 2-pyrrolidinyl]methoxybenzoic acid N N YN \/O- COOH 20 Me H H Me 141 To a solution of methyl (2S,3R,4S)-1-benzyloxycarbonyl-3,4-isopropylidenedioxy-2-pyrrolidinyl carboxylate (10.7 g, 31.9 mmol) in THF (250 ml), 0.25 N NaOH (255 ml) was added. After stirring at room temperature for 24 h, the mixture was acidified with 1 N HCI and extracted with EtOAc. The combined extracts were washed with brine, which were dried over Na 2
SO
4 and 25 concentrated in vacuo, to give (2S,3R,4S)-1-benzyloxycarbonyl-3,4-isopropylidenedioxy-2 pyrrolidinylcarboxylic acid (9.87 g, 96%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.32 (s, 3H), 1.46 (d, J= 2.7 Hz, 3H), 3.61(m, 1H), 3.82 and 3.92 (d, J= 12.7 Hz, 1H, amide isomers), 4.58 and 4.64 (s, 1H, amide isomers), 4.77 (t, J= 5.1 Hz, 1H), 4.83 and 4.89 (d, J= 5.9 Hz, 1H,amide isomers), 5.15 and 5.19 (m, 2H, amideisomers), 7.31-7.37 (m, 5H). 300 WO 01/00206 PCT/US00/18079 To a stirred solution of (2S,3R,4S)-l1-benzyloxycarbonyl-3,4-isopropylidenedioxy-2-pyrrolidinyl carboxylic acid (9.87 g, 30.7 mmol) in THF (200 ml) was added BH 3 DMS (6.14 ml, 61.4 mmol) at 0 oC. The mixture was allowed to room temperature and then heated under reflux for 2 h. After cooling to room temperature, the mixture was concentrated in vacuo and quenched by the addition 5 of water at 0 oC. The mixture was extracted with EtOAc. The combined extracts were washed with water and brine, which were dried over Na 2
SO
4 and concentrated in vacuo. The residue was chromatographed on silica gel [200 g, CHCl 3/MeOH (20/1)], to give (2R,3R,4S)-1-benzyloxy carbonyl-3,4-isopropylidenedioxy-2-pyrrolidinylmethanol (10.1 g, 100%) as a colorless oil. 'H NMR (CDCl 3 ) 8 1.31 (s, 3H), 1.45 (s, 3H), 3.56-4.74 (min, 7H), 5.14 (s, 2H), 7.34 (min, 5H). 10 To a stirred mixture of (2R,3R,4S)-1-benzyloxycarbonyl-3,4-isopropylidenedioxy-2-pyrrolidinyl methanol (312 mg, 0.64 mmol), methyl p-hydroxybenzoate (67 ml, 0.70 mmol), PPh 3 (184 mg, 0.70 mmol) in THF (7 ml) was added DIAD (138 ml, 0.70 mmol) at 0 oC under an atmosphere of nitrogen. The mixture was allowed to reach room temperature and stirred for 3 h. After removal of the solvent, the resulting residue was chromatographed on silica gel [10 g, n-hexane/EtOAc 15 (4/1)], to give methyl 4-[(2R,3R,4S)-benzyloxycarbonyl-3,4-isopropylidenedioxy-2-pyrrolidinyl] methoxybenzoate (321 mg, 83%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.01 (s, 6H), 1.03 (s, 3H11), 2.23 (min, 1H11), 2.63 (min, 1H), 3.61 (d, J= 12.5 Hz, 1H11), 3.80-4.27 (min, 4H), 4.84 (br, 1H11), 5.01 and 5.08 (ABq, J= 12.2 Hz, 1H, amide isomers), 6.75-6.87 (min, 3H11), 7.19-7.63 (min, 15H). A suspension of methyl 4-[(2R,3R,4S)-l-benzyloxycarbonyl-3,4-isopropylidenedioxy-2 20 pyrrolidinyl]methoxybenzoate (2.37 g, 5.76 mmol) and 10% Pd/C (240 mg ) in EtOH (170 ml) was stirred at room temperature under an atmosphere of hydrogen. After 1 day stirring, the catalyst and solvent were changed for 10% Pd/C (500 mg) and THF (50 ml). The suspension was stirred at room temperature under an atmosphere of hydrogen for 5 days. After removed the catalyst by filtration, the filtrates were concentrated in vacuo. The residue was chromatographed on silica gel 25 [100 g, CHC1/acetone (20/1)], to give methyl 4-[(2R,3R,4S)-3,4-isopropylidenedioxy-2 pyrrolidinyl]methoxybenzoate (930 mg, 53%) as a brown oil. 'H-NMR (CDCl 3 ) 8 1.35 (s, 3H11), 1.50 (s, 3H), 3.02 (dd, J= 13.7, 4.1 Hz, 1H), 3.13(d, J = 13.7 Hz, 1H), 3.58 (t,J= 6.3 Hz, 1H), 3.88 (s, 3H), 3.90 (dd, J= 9.3, 6.6 Hz, 1H), 4.02 (dd, J= 9.5, 3.9 Hz, 1H11), 4.74 (d, J= 5.6 Hz, 1H1), 4.79 (min, 1H11), 6.90 (d, J = 9.0 Hz, 2H11), 7.98 (d, J = 9.0 Hz, 2H). 30 A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (437 mg, 1.39 mmol), methyl 4-[(2R,3R,4S)-3,4-isopropylidenedioxy-2-pyrrolidinyl]methoxybenzoate (428 mg, 1.39 301 WO 01/00206 PCT/US00/18079 mmol), EDC-HCl (400 mg, 2.09 mmol) and DMAP (170 mg, 1.39 mmol) in DMF (12 ml) was stirred at room temperature for 20 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [70 g, 5 CHClj/acetone (10/1)], to give methyl 4-[(2R,3R,4S)-3,4-isopropylidenedioxy-1-[4-[N'-(2 methylphenyl)ureido]-3-methoxyphenylacetyl]-2-pyrrolidinyl]methoxybenzoate (840 mg, 100%) as a colorless amorphous solid IR (KBr) 3354, 2985, 2939, 1716, 1533, 1254 cm-'; 'H-NMR (CDCl 3 ) 8 1.31 (s, 3H), 1.42 (s, 3H), 2.05 (s, 3H), 3.50 (s, 3H), 3.55-3.88 (m, 4H), 3.89 (s, 3H), 4.13 (m, 1H), 4.67 (br, 1H), 4.78 (d, J= 6.1 Hz, 1H11), 4.88 (t, J= 5.6 Hz, 1H), 6.46 (s, 1H), 6.62 10 (d,J= 1.5 Hz, 1H), 6.74 (m, 3H), 7.05 (s, 1H), 7.14 (d,J= 7.3 Hz, 1H), 7.23 (m, 2H), 7.57 (d,J= 7.8 Hz, 1H), 7.91-8.08 (m, 3H); MS (ESI) m/z 604 (M'+1); Anal. Calcd for C 33
H
37
N
3 0 8 s-0.6H 2 0: C, 64.50; H, 6.27; N, 6.84. Found: C, 64.38; H, 6.18; N, 6.66. A mixture of methyl 4-[(2R,3R,4S)-3,4-isopropylidenedioxy-1-[4-[N'-(2-methylphenyl)ureido]-3 methoxyphenylacetyl]-2-pyrrolidinyl]methoxybenzoate (183 mg, 0.303 mmol) and g.HCl-MeOH 15 (6 ml) was stirred at room temperature for 17 h. The mixture was concentrated in vacuo. The residue was purified on TLC [CHCl3/MeOH (10/1)], to give methyl 4-[(2R,3R,4S)-3,4-dihydroxy 1 -[4-[N'-(2-methylphenyl)ureido]-3-methoxyphenylacetyl]-2-pyrrolidinyl]methoxybenzoate (162 mg, 95%) as a colorless amorphous solid IR (KBr) 3342, 1716, 1604, 1535, 1255 cm'; 'H-NMR (CDCl 3 ) 8 2.25 (br, 3H), 3.33-3.75 (m, 7H), 3.87 (s, 3H), 4.10 (d, J= 8.3 Hz, 1H), 4.24 (s, 2H), 20 4.37 (m, 2H), 6.62-7.94 (m, 13H); MS (ESI) m/z 564 (M+1). To a solution of methyl 4-[(2R,3R,4S)-3,4-isopropylidenedioxy-1l-[4-[N'-(2-methylphenyl)ureido] 3-methoxyphenylacetyl]-2-pyrrolidinyl]methoxybenzoate (490 mg, 0.812 mmol) in THF (9.8 ml), 0.25 N NaOH (9.8 ml) was added. After stirring at room temperature for 4 days, the mixture was acidified with 1 N HCI and extracted with CHCI 3 -MeOH (10/1). The combined extracts were dried 25 over Na 2
SO
4 and concentrated in vacuo to give 141 (445 mg, 93%) as a colorless amorphous solid. MW 689.64 IR (KBr) 3354, 2983, 2937, 1707, 1604, 1533 cmr'; 'H-NMR (DMSO-d 6 ) 6 .24 and 1.26 (s, 3H, amide isomers), 1.26 and 1.32 (s, 3H, amide isomers), 2.24 (s, 3H), 3.40 (dd, J= 14.0, 5.1 Hz, 1H), 3.60 (m, 2H), 3.71 (m, 1H), 3.76 (s, 3H), 3.82 (s, 3H), 3.92-4.96 (m, 5H), 6.74 and 6.78 (m, 1H, amide isomers), 6.83-7.16 (m, 6H), 7.79 (d, J= 8.3 Hz, 1H), 7.87 (t, J= 9.1 Hz, 30 2H), 8.01 (m, 1H), 8.49 (d, J= 3.4 Hz, 1H), 8.57 (s, 1H); MS (FAB) m/z 590(M*+1); Anal. Calcd for C 32
H
35
N
3 0O-2.3H 2 0: C, 60.90; H, 6.32; N, 6.66. Found: C, 61.00; H, 6.00; N, 6.27. 302 WO 01/00206 PCT/US00/18079 Example 134 4-[(2R,3R,4S)-3,4-dihydroxy-l-[4-[N '-(2-methylphenyl)ureido]-3-methoxyphenylacetyl]-2 pyrrolidinyl]methoxybenzoic acid ,.OH N.-OH N N 0 COOH Me H H Me 142 5 To a solution of methyl 4-[(2R,3R,4S)-3,4-dihydroxy-1-[4-[N'-(2-methylphenyl)ureido]-3 methoxyphenylacetyl]-2-pyrrolidinyl]methoxybenzoate (63 mg, 0.112 mmol) in THF (0.89 ml), 0.25 N NaOH (0.89 ml) was added. After stirring at room temperature for 3 days, the mixture was acidified with 1 N HCI and extracted with CHC1 3 -MeOH (10/1). The combined extracts were dried over Na 2
SO
4 and concentrated in vacuo to give 142 (54 mg, 88%) as a colorless amorphous solid. 10 MW 549.57 IR (KBr) 3356, 2958, 2927, 1685, 1604, 1535, 1255 cm~'; 'H-NMR (DMSO-d) 8 2.24 (s, 3H), 3.40 (m, 1H-), 3.58 (s, 2H), 3.66 (dd, J= 9.8, 6.6 Hz, 1H), 3.80 (s, 3H), 3.99-4.30 (m, 5H), 5.10 (br, 1H), 6.72 (d,J = 8.1 Hz, 1H), 6.85 (s, 1H), 6.93 (t, J= 7.3 Hz, 1H), 7.03 (d,J= 8.8 Hz, 2H), 7.14 (t, J= 8.8 Hz, 2H), 7.79 (d, J= 8.1 Hz, 2H), 7.86 (d, J= 8.8 Hz, 2H), 7.99 (d, J 8.3 Hz, 1H), 8.46 (s, 1H), 8.56 (s, 1H); MS (ESI) m/z 550(M++1); Anal. Calcd for 15 C 2
H
31
N
3 01s-0.85H 2 0: C, 61.66; H, 5.83; N, 7.44. Found: C, 62.09; H, 5.93; N, 6.95. Example 135 4-[(2R,3R,4S)-1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-3,4-isopropylidenedioxy 2-pyrrolidinyl]methoxybenzoic acid ,N N N N / COOH I HH Me 143 20 A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (487 mg, 1.45 mmol), methyl 4-[(2R,3R,4S)-3,4-isopropylidenedioxy-2-pyrrolidinyl]methoxybenzoate (447 mg, 1.45 mmol), EDC.HCl (418 mg, 2.18 mmol) and DMAP (177 mg, 1.45 mmol) in DMF (12 ml) was stirred at room temperature for 19 h. The mixture was poured into ice water and extracted with EtOAc. The combined extracts were washed with ice water and brine. After dried over Na 2
SO
4 , 25 the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [70 g, CHCl3/acetone (10/1)], to give methyl 4-[(2R,3R,4S)-1-[4-[N'-(2-chlorophenyl)ureido]-3 methoxyphenylacetyl]-3,4-isopropylidenedioxy-2-pyrrolidinyl]methoxybenzoate (850 mg, 94%) as a colorless amorphous solid IR (KBr) 3329, 2939, 1716, 1627, 1531, 1254 cm''; 'H-NMR (CDCI 3 ) 8 1.33 (s, 3H), 1.43 (s, 3H), 3.56 (s, 3H), 3.61 (s, 1H), 3.64 (s, 1H), 3.70 (m, 1H), 3.79 (d, J= 10.4 303 WO 01/00206 PCT/US00/18079 Hz, 1H11), 3.88 (s, 3H), 4.14 (dd, J= 9.8, 2.2 Hz, 1H11), 4.40 (dd, J= 9.8, 3.4 Hz, 1H), 4.67 (s, 1H), 4.80 (d, J= 6.1 Hz, 1H11), 4.90 (t, J= 4.6 Hz, 1H), 6.65 (d, J= 1.7 Hz, 1H11), 6.71-6.84 (min, 3H), 6.98 (dt, J= 7.6, 1.5 Hz, 1H), 7.27 (min, 2H), 7.33 (dd, J= 8.0, 1.2 Hz, 2H11), 7.90-8.01 (min, 3H), 8.20 (dd, J= 8.3, 1.5 Hz, 1H); MS (ESI)m/z 624 (M'+1), 626 (M*+3); Anal. Calcd for C 32
H
3 4
N
3 0-1.4H 2 0: 5 C, 59.19; H, 5.71; N, 6.47. Found: C, 58.85; H, 5.35; N, 6.21. A mixture of methyl 4-[(2R,3R,4S)-1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-3,4 isopropylidenedioxy-2-pyrrolidinyl]methoxybenzoate (177 mg, 0.284 mmol) and g.HCl-MeOH (4 ml) was stirred at room temperature for 2 days. The mixture was concentrated in vacuo. The residue was purified on TLC [CHCl 3 /MeOH (15/1)], to give methyl 4-[(2R,3R,4S)-1-[4-[N'-(2 10 chlorophenyl)ureido]-3-methoxyphenylacetyl]-3,4-isopropylidenedioxy-2-pyrrffolidinyl] methoxybenzoate (140 mg, 85%) as a colorless amorphous solid IR (KBr) 3338, 2949, 1712, 1623, 1604, 1533 cm'; 'H-NMR (CDCl 3 ) 5 2.27 (min, 1H), 2.79 (min, 1H), 3.53 (dd, J= 10.5, 5.9 Hz, 1H), 3.63 (s, 3H), 3.88 (s, 3H11), 4.21 (d, J = 7.8 Hz, 1H11), 4.31 (min, 2H), 4.43 (dd, J= 9.8, 4.4 Hz, 1H1), 4.52 (t, J = 4.6 Hz, 1H11), 6.71 (s, 1H), 6.80 (min, 3H), 6.99 (t,J= 7.3 Hz, 1H), 7.16 (s, 1H), 15 7.21(s, lH), 7.39(d, J=8.1Hz, 1H), 7.91(d, J=8.6Hz, 2H), 8.16(d,J=8.3 Hz, 1H);MS(ESI) m/z 584 (M*+1), 586 (M +3). To a solution of methyl 4-[(2R,3R,4S)- 1 -[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl] 3,4-isopropylidenedioxy-2-pyrrolidinyl]methoxybenzoate (511 mg, 0.819 mmol) in THF (9.8 ml), 0.25 N NaOH (9.8 ml) was added. After stirring at room temperature for 20 h, the mixture was 20 acidified with 1 N HCI and extracted with CHCl 3 -MeOH (10/1). The combined extracts were dried over Na 2
SO
4 and concentrated in vacuo to give 143 (504 mg, 100%) as a colorless amorphous solid. MW 610.05 IR (KBr) 3330, 2983, 2937, 1711, 1689, 1604, 1533, 1252 cm-'; 'H-NMR (DMSO-d 6 ) 8 1.26 (s, 3H1), 1.32 (s, 3H11), 3.40 (min, 1H11), 3.60 and3.61 (d, J= 2.5 Hz, 3H11, amide isomers), 3.62 (min, 1H), 3.78 and 3.83 (s, 3H11, amideisomers), 4.16 (m, 2H11), 4.42-4.98 (min, 311), 25 6.74-7.15 (min, 6H), 7.28 (t, J= 7.3 Hz, 1H11), 7.43 (d, J= 8.1Hz, 1H11), 7.78-7.97 (min, 4H), 8.08 (d, J = 8.3 Hz, 1H11), 8.89 (s, 1H), 8.92 (s, 1H11), 12.68 (br, 1H11); MS (ESI) m/z 610 (M*+1), 612 (M*+3). Example 136 4-[(2R,3R,4S)-I-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-3,4-dihydroxy-2 pyrrolidinyl]methoxybenzoic acid ,,OH C . COOH N N j-H 30 I H H CMe 144 304 WO 01/00206 PCT/US00/18079 To a solution of methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-3,4 dihydroxy-2-pyrrolidinylmethoxy]benzoate (63 mg, 0.108 mmol) in THF (0.80 ml), 0.25 N NaOH (0.80 ml) was added. After stirring at room temperature for 3 days, the mixture was acidified with 1 N HCI and extracted with CHCI 3 -MeOH (10/1). The combined extracts were dried over Na 2
SO
4 5 and concentrated in vacuo to give 144 (61 mg, 100%) as a colorless amorphous solid. MW 569.99 IR (KBr) 3338, 1687, 1604, 1533, 1255, 1169, 1036 cm-'; 'H-NMR (DMSO-d 6 ) 5 3.59 (d, J= 5.5 Hz, 2H), 3.61 (min, 1H), 3.66 (dd, J = 10.0, 7.1 Hz, 1H), 3.80 (s, 3H), 4.00-4.33 (min, 5H), 5.10 (br, 1H), 6.74 (d, J= 8.3 Hz, 1H), 6.87 (s, 1H), 7.03 (min, 3H), 7.28 (t, J = 8.3 Hz, 1H), 7.43 (d, J= 6.6 Hz, 1H), 7.87 (d, J= 8.5 Hz, 2H), 7.95 (d, J= 8.3 Hz, 1H), 8.09 (d, J = 8.3 Hz, 1H), 8.32 (s, 1H), 10 8.89 (s, 1H), 8.93 (s, 1H); MS (ESI) m/z 570 (M*+1), 572 (M+3); Anal. Calcd for
C
28
H
28
CIN
3 Os-1.4H 2 O: C, 57.19; H, 5.14; N, 7.15. Found: C, 57.52; H, 5.22; N, 6.76. Example 137 4-[ 1 -[4-[N'-(2-methylphenyl)ureido]J-3-methoxyphenylacetyl]-5-(R)-phenyl-2-(S)-pyrrolidinyl methoxy]benzoic acid N N O \/ COOH 15 Me H H OMe 145 To a stirred solution of benzy N-Boc-pyrroglutarate (8.93 g, 28.0 mmol) in THF (100 ml) was added phenyllithium (1.0 Min Et 2 0-cyclohexane, 33.5 ml, 33.5 mmol) at -78 0 C, and the resulting mixture was gradually warmed up to -40 0 C, then stirred overnight. aq.NH 4 CI was added to the reaction mixture, THF was removed in vacuo, then extracted with EtOAc. The organic layer 20 was washed with water and, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with hexane-EtOAc (4 : 1) as eluent, then recrystallized from hexane-EtOAc to give benzyl [2-(S)-(N-Boc-amino)-5-oxo-6-phenyl]pentanoate (5.02 g, 45%) as a colorless needles. mp 85-87 oC; 'H-NMR (CDCl 3 ) 6 1.43 (s, 9 H), 2.07-2.19 (min, 1 H), 2.27-2.36 (min, 1 H), 2.97-3.13 (min, 2 H), 4.44 (brs, 1 H), 5.19 (dd, J= 25.2, 12.0 Hz, 2 H), 5.19 25 (overlap, 1 H), 7.28-7.98 (series of min, 10 H); MS (ESI) m/z, 322 (M*+H). To a stirred solution of benzyl [2-(S)-(N-Boc-amino)-5-oxo-6-phenyl]pentanoate (2.20 g, 5.54 mmol) in CH 2
CI
2 (50 ml) was added trifluoroacetic acid (15 ml) at rt, and the resulting mixture was stirred for 2 h. The mixture was concentrated in vacuo and poured into aq.NaHCO 3 , then extracted with EtOAc. The organic layer was dried over anhydrous Na 2
SO
4 , then concentrated in 30 vacuo to give benzyl 5-phenyl-5-pyrroline-2-(S)-carboxylate (1.60 g, quant.) as yellowish solid. 305 WO 01/00206 PCT/US00/18079 The product was used for next reaction without further purification: 'H-NMR (CDC1 3 ) 8 2.20-2.29 (m, 1 H), 2.32-2.42 (m, 1 H), 2.96-3.05 (min, 1 H), 3.12-3.21 (min, 1 H), 4.96-5.00 (m, 1 H), 5.24 (s, 2 H), 7.31-7.49 (m, 8 H), 7.88-7.91 (m, 2 H); MS (ESI) m/z, 280 (M+H). A mixture of benzyl 5-phenyl-5-pyrroline-2-(S)-carboxylate (1.59 g, 5.69 mmol) and Pd/C (10%, 5 128 mg) in MeOH (30 ml) was stirred under H2 at rt for 28 h. The mixture was filtered and the filtrate was concentrated in vacuo. The residue was dissolved in CH 3
CN-H
2 0 (3 : 2, 25 ml), then was added di-tert-butyl dicarbonate (1.86 g, 8.54 mmol) and 1.0 M-NaOH (8.54 ml, 8.54 mmol), and the resulting mixture was stirred for 30 min. The mixture was concentrated in vacuo and poured into aq.NaHCO 3 , then extracted with EtOAc. The organic layer was washed with water, 10 saturated brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHC1 3 -MeOH (9 : 1) and recrystallized from hexane-EtOAc to give N-Boc-5-(R)-phenyl-(S)-proline (810 mg, 49%) as a colorless solid. Mp 113-117 oC; 'H NMIR (CDCl 3 ) 8 1.13 (s, 9 H), 1.43 (brs, 1 H), 1.96 (brs, 1 H), 2.09 (brs, 1 H), 2.31-2.34 (min, 1 H), 2.46 (brs, 1 H), 4.52 (brs, 1 H), 4.69 (brs, 1 H), 7.22-7.37 (min, 5 H). 15 To a stirred solution of N-Boc-5-(R)-phenyl-2-(S)-proline (1.14 g, 3.91 mmol) in THF (20 ml) was added 10M-BH 3 Me 2 S (780 ml, 7.82 mmol) at rt, and the resulting mixture was heated under reflux for 30 min. The mixture was poured into aq. 1N-HCI and extracted with EtOAc. The organic layer was dried over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHC13-MeOH (10 : 1) as eluent to give N-Boc-2-(S)-hydoxymethyl-5-(R) 20 phenylpyrrolidine (1.11 g, quant.) as a colorless oil: 'H-NMR (CDCl 3 ) 8 1.19 (brs, 9 H), 1.65 (brs, 1 H), 1.83-1.90 (min, 1 H), 1.98-2.06 (m, 1 H), 2.22-2.31 (min, 1 H), 3.75-3.86 (m, 2 H), 4.16-4.19 (min, 1 H), 4.83 (t, J= 6.8 Hz, 1 H), 4.89 (brs, 1 H), 7.19-7.31 (min, 5 H); MS (ESI) m/z, 278 (M +H). To a stirred solution of N-Boc-2-(S)-hydoxymethyl-5-(R)-phenylpyrrolidine (1.10 g, 3.97 mmol), triphenylphosphine (1.25 g, 4.76 mmol) and methyl 4-hydroxybenzoate (724 mg, 4.76 mmol) was 25 added diisopropyl azodicarboxylate (955 ml, 4.76 mmol) at rt, and the resulting mixture was stirred at 60oC for 45 min. The mixture was concentrated in vacuo, and the residue was chromatographed on silica gel with hexane-EtOAc (4 : 1) as eluent to give methyl 4-[N-Boc-5-(R) phenyl-2-(S)-pyrrolidinylmethoxy]benzoate (1.31 g, 80%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.19 and 1.47 (brs, total 9 H), 2.09-2.15 (m, 3 H), 2.33-2.37 (min, 1 H), 3.94 (s, 3 H), 4.30 (brs, 1 30 H), 4.41 (brs, 2 H), 4.77 (brs, 1 H), 7.03 (d, J= 8.8 Hz, 2 H), 7.24-7.36 (m, 5 H), 8.03-8.06 (m, 2 H); MS (ESI) m/z, 412 (M++H). 306 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[N-Boc-5-(R)-phenyl-2-(S)-pyrrolidinylmethoxy] benzoate (1.28 g, 3.11 mmol) in CH 2 C1 2 (30 ml) was added trifluoroacetic acid (10 ml) at rt, and the resulting mixture was stirred for 45 min. The mixture was concentrated in vacuo and poured into aq.NaHCO 3 , then extracted with CHCl 3 . The organic layer was washed with water, drying over 5 anhydrous Na 2
SO
4 , and concentrated in vacuo to give methyl 4-[5-(R)-phenyl-2-(S)-pirrolidinyl methoxy]benzate (363 mg, quant.) as yellowish oil. The product was used for next reactions without further purification. 'H-NMR (CDC 3 ) 8 1.71-1.83 (m, 2 H), 2.03-2.10 (m, 1 H), 2.15-2.24 (m, 1 H), 3.68-3.74 (m, 1 H), 3.89 (s, 3 H), 4.01-4.09 (m, 2 H), 4.28 (t, J = 7.2 Hz, 1 H), 6.95 (d, J = 8.8 Hz, 2 H), 7.22-7.27 (m, 1 H), 7.33 (t, J= 8.0 Hz, 2 H), 7.42 (d, J= 7.6 Hz, 2 H), 8.00 (d, J 10 8.8 Hz, 2 H); MS (ESI) m/z, 312 (M++H) 353 (M++CH 3 CN). To a stirred solution of methyl 4-[5-(R)-phenyl-2-(S)-pirrolidinylmethoxy]benzoate (135 mg, 0.43 mmol), 4-[N'-(2-methylphenyl)ureido]-3-methoxyphenylacetic acid (136 mg, 0.43 mmol) and N,N dimethylaminopyridine (52.9 mg, 0.43 mmol) in DMF (10 ml) was added EDCHCI (90.8 mg, 0.48 mmol) at rt, and the resulting mixture was stirred overnight. The reaction mixture was poured into 15 water and extracted with EtOAc. The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with hexane EtOAc (1 : 5) as eluent to give methyl 4-[1-[4-[N'-(2-methylphenyl)ureido]-3-methoxyphenyl acetyl]-5-(R)-phenyl-2-(S)-pyrrolidinylmethoxy]benzoate (271 mg, quant.) as a colorless amorphous solid. 1 H-NMR (CDCl 3 ) 8 2.00-2.18 (m, 3 H), 2.31-2.41 (m, 1 H), 2.27 (s, 3 H), 3.31 20 (s, 2 H), 3.67 (s, 3 H), 3.89 (s, 3 H), 4.35-4.48 (m, 2 H), 4.60 (brs, 1 H), 4.92 (t, J= 6.8 Hz, 1 H), 6.51 (d, J= 8.4 Hz, 1 H), 6.62 (s, 1 H), 6.96 (d, J= 8.8 Hz, 1 H), 7.11-7.40 (series of m, 8 H), 7.51 (d, J= 8.0 Hz, 1 H), 7.97-8.00 (m, 2 H); MS (ESI) m/z, 608 (M'+H). To a stirred solution of methyl 4-[1-[4-[N'-(2-methylphenyl)ureido]-3-methoxyphenylacetyl]-5 (R)-phenyl-2-(S)-pyrrolidinylmethoxy]benzoate (243 mg, 0.40 mmol) in MeOH-THF (1 : 1, 10 ml) 25 was added 1.0M-NaOH (2.4 ml, 2.40 mmol) at rt, and the resulting mixture was heated at 60 0 C with stirring for 1.5 h. The reaction mixture was poured into 1N-HC1, then extracted with CHC 3 . The organic layer was washed with brine and dried over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (10 : 1) to give 145 (224 mg, 94%) as a colorless amorphous solid. MW 593.67 'H-NMR (CD30D), mixture of rotamnars, 8 30 2.00-2.19 (m, 3 H), 2.28 and 2.30 (s, total 3 H), 2.45-2.49 (m, 1 H), 3.37 (dd, J= 39, 16Hz, 2 H), 3.77 and 3.80 (s, total 3 H), 3.92-5.18 (series of m, 4 H), 6.48-8.03 (series of m, 16 H); MS (FAB) m/z, 594 (M 4 +H). 307 WO 01/00206 PCT/US00/18079 Example 138 4-[ 1 -[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-5-(R)-phenyl-2-(S)-pyrrolidinyl methoxy]benzoic acid N N N/ COOH CI H HOMe 146 5 To a stirred solution of methyl 4-[5-(R)-phenyl-2-(S)-pirrolidinylmethoxy]benzoate (142 mg, 0.46 mmol), 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (153 mg, 0.46 mmol) and N,N dimethylaminopyridine (55.7 mg, 0.46 mmol) in DMIF (10 ml) was added EDCHCl (95.7 mg, 0.50 mmol) at rt, and the resulting mixture was stirred overnight. The reaction mixture was poured into water and extracted with EtOAc. The organic layer was washed with brine, drying over anhydrous 10 Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with hexane EtOAc (1 : 5) as eluent to give methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl acetyl]-5-(R)-phenyl-2-(S)-pyrrolidinylmethoxy]benzoate (260 mg, 90%) as a colorless amorphous solid. 'H-NMR (CDCI 3 ) 8 2.00-2.19 (m, 3 H), 2.35-2.44 (m, 1 H), 3.35 (s, 2 H), 3.76 (s, 3 H), 3.89 (s, 3 H), 4.38-4.48 (m, 2 H), 4.63 (brs, 1 H), 4.94 (t, J= 7.2 Hz, 1 H), 6.53 (d, J= 8.4 Hz, 1 H), 15 6.66 (s, 1 H), 6.96-7.01 (m, 3 H), 7.12-7.42 (series of m, 8 H), 7.87 (d, J= 8.0 Hz, 1 H), 7.98 (d, J = 8.8 Hz, 2 H), 8.17-8.19 (m, 1 H); MS (ESI) m/z, 627(M'), 628 (M'+H). To a stirred solution of methyl 4-[ 1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-5-(R) phenyl-2-(S)-pyrrolidinylmethoxy]benzoate (251 mg, 0.40 mmol) in MeOH-THF (1 : 1, 10 ml) was added 1.0M-NaOH (2.4 ml, 2.40 mmol) at rt, and the resulting mixture was heated at 60 0 C with 20 stirring for 1.5 h. The reaction mixture was poured into 1N-HCI, then extracted with CHCi 3 . The organic layer was washed with brine and dried over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (10 : 1) to give 146 (181 mg, 74%) as a colorless amorphous solid. MW 614.09 'H-NMR (CD 3 OD), mixture of rotamars, 8 1.99-2.19 (m, 3 H), 2.42-2.53 (m, 1 H), 3.38 (dd, J= 39, 15 Hz, 2 H), 3.79 and 3.80 25 (s, total 3 H), 3.94-5.19 (series of m, 4 H), 6.49-8.05 (series of m, 16 H); MS (FAB) m/z, 614 (M'+H); Anal. Calcd for C 35 1 34 C1N 3 0 6
H
2 0: C, 65.06; H, 5.62; N, 6.50. Found: C, 65.03; H, 5.75; N, 6.45. Example 139 4-[1-[4-[N' -(2-bromophenyl)ureidoj-3-methoxyphenylacetyl]-5-(R)-phenyl-2-(S)-pyrrffolidinyl 30 ethoxy]benzoic acid 308 WO 01/00206 PCT/US00/18079 N N N COOH rH H Me 147 To a stirred solution of methyl 4-[5-(R)-phenyl-2-(S)-pirrolidinylmethoxy]benzoate (146 mg, 0.47 mmol), 4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (178 mg, 0.47 mmol) and N,N dimethylaminopyridine (57.4 mg, 0.47 mmol) in DMF (10 ml) was added EDCHCI (99.0 mg, 0.52 5 mmol) at rt, and the resulting mixture was stirred overnight. The reaction mixture was poured into water and extracted with EtOAc. The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with hexane EtOAc (1 : 5) as eluent to give methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl acetyl]-5-(R)-phenyl-2-(S)-pyrrolidinylmethoxy]benzoate (288 mg, 91%) as a colorless amorphous 10 solid. 'H-NMR (CDCl 3 ) 8 2.00-2.20 (m, 3 H), 2.34-2.43 (m, 1 H), 3.35 (s, 2 H), 3.72 (s, 3 H), 3.89 (s, 3 H), 4.38-4.49 (m, 2 H), 4.62 (brs, 1 H), 4.94 (t, J = 7.2 Hz, 1H), 6.54 (d, J = 8.4 Hz, 1H), 6.67 (s, 1 H), 6.91-7.05 (m, 4 H), 7.28-7.42 (series of m, 7 H), 7.51 (d, J= 8.0 Hz, 2 H), 7.87 (d, J= 8.4 Hz, 1 H), 7.99 (d, J= 8.8 Hz, 2 H), 8.14 (d, J= 8.4 Hz, 2H); MS (ESI) m/z, 672 (M), 674 (M+2). To a stirred solution of methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-5-(R) 15 phenyl-2-(S)-pyrrolidinylmethoxy]benzoate (270 mg, 0.40 mmol) in MeOH-THF (1 : 1, 10 ml) was added 1.0M-NaOH (2.0 ml, 2.0 mmol) at rt, and the resulting mixture was heated at 60oC with stirring for 1 h. The reaction mixture was poured into 1N-HC1, then extracted with CHC1 3 . The organic layer was washed with brine and dried over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHC13-MeOH (10 : 1) to givel47 (212 20 mg, 80%) as a colorless amorphous solid. MW 658.54 'H-NMR (CD 3 OD), mixture of rotamars, 8 1.99-2.19 (m, 3 H), 2.42-2.53 (m, 1 H), 3.38 (dd, J= 39, 16 Hz, 2H), 3.79 and 3.80 (s, total 3 H), 3.94-5.19 (series of m, 4 H), 6.49-8.00 (series of m, 16 H); MS (FAB) m/z, 658 (M), 660 (M+2). Example 140 4-[ -[4-[N'-(2,6-dichlorophenyl)ureido]-3-methoxyphenylacetyll-5-(R)-phenyl-2-(S)-pyrrolidiny 25 methoxy]benzoic acid N NN 0 O- COOH IHH Me 148 To a stirred solution of methyl 4-[5-(R)-phenyl-2-(S)-pirrolidinylmethoxy]benzoate (109 mg, 0.35 309 WO 01/00206 PCT/US00/18079 mmol), 4-[N'-(2,4-dichlorophenyl)ureido]-3-methoxyphenylacetic acid (129 mg, 0.35 mmol) and N,N-dimethylaminopyridine (42.8 mg, 0.35 mmol) in DMF (10 ml) was added EDCHCI (73.4 mg, 0.39 mmol) at rt, and the resulting mixture was stirred for 6 h. The reaction mixture was poured into water and extracted with EtOAc. The organic layer was washed with brine, drying over 5 anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with hexane-EtOAc (1 : 4) as eluent to give methyl 4-[1-[4-[N'-(2,6-dichlorophenyl)ureido]-3 methoxyphenylacetyl]-5-(R)-phenyl-2-(S)-pyrrolidinylmethoxy]benzoate (208 mg, 90%) as a colorless amorphous solid. 'H-NMR (CDCl 3 ) 8 2.00-2.21 (m, 3 H), 2.33-2.39 (m, 1 H), 3.31 (s, 2 H), 3.69 (s, 3 H), 3.88 (s, 3 H), 4.34-4.45 (m, 2 H), 4.59 (brs, 1 H), 4.93 (t, J= 6.8 Hz, 1 H), 6.47 10 (d, J= 8.0 Hz, 1 H), 6.63 (s, 1 H), 6.68 (s, 1 H), 6.92-6.95 (m, 2 H), 7.12-7.41 (series of m, 9 H), 7.96-8.01 (m, 4 H); MS (FAB) m/z, 662 (M*+H). To a stirred solution of methyl 4-[1-[4-[N'-(2,6-dichlorophenyl)ureido]-3-methoxyphenylacetyl]-5 (R)-phenyl-2-(S)-pyrrolidinylmethoxy]benzoate (186 mg, 0.28 mmol) in MeOH-THF (1 : 1, 10 ml) was added 1.0M-NaOH (1.4 ml, 1.4 mmol) at rt, and the resulting mixture was heated at 60 0 C with 15 stirring for 2.5 h. The reaction mixture was poured into 1N-HC1, then extracted with CHC 3 . The organic layer was washed with brine and dried over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (10 : 1) to give 148 (166 mg, 91%) as a colorless amorphous solid. MW 648.53 'H-NMR (CDCl 3 ) 6 2.00-2.18 (m, 3 H), 2.34-2.40 (m, 1 H), 3.33 (s, 2 H), 3.68 (s, 3 H), 4.37-4.47 (m, 2 H), 4.61 (brs, 1 H), 4.94 (t, J= 20 6.8 Hz, 1 H), 6.48 (d, J = 8.0 Hz, 1 H), 6.63 (s, 1 H), 6.96 (d, J = 8.4 Hz, 3 H), 7.12-7.38 (series of m, 9 H), 7.95 (d, J= 8.0 Hz, 1 H), 8.01 (d, J= 8.8 Hz, 2 H); MS (FAB) m/z, 648 (M++H). Example 141 4-[ 1 -[4-[N'-(2-bromophenyl)ureido]-3-methylphenylacetyl]-5-(R)-phenyl-2-(S)-pyrrolidinyl methoxy]benzoic acid N N P'COOH 25 r H H Me 149 To a stirred solution of methyl 4-[5-(R)-phenyl-2-(S)-pirrolidinylmethoxy]benzoate (125 mg, 0.40 mmol), 4-[N'-(2-bromophenyl)ureido]-3-methylphenylacetic acid (146 mg, 0.40 mmol) and N,N dimethylaminopyridine (49.0 mg, 0.40 mmol) in DMF (10 ml) was added EDCHC1 (84.1 mg, 0.44 immol) at rt, and the resulting mixture was stirred for 6 h. The reaction mixture was poured into 30 water and extracted with EtOAc. The organic layer was washed with brine, drying over anhydrous 310 WO 01/00206 PCT/US00/18079 Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with hexane EtOAc (1 : 4) as eluent to give methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methylphenylacetyl] 5-(R)-phenyl-2-(S)-pyrrolidinylmethoxy]benzoate (238 mg, 90%) as a colorless amorphous solid. 'H-NMR (CDCl 3 ) 8 1.92 (s, 3 H), 2.09-2.27 (m, 3 H), 2.42-2.50 (m, 1 H), 3.22-3.41 (m, 2 H), 3.88 5 (s, 3 H), 4.39 (d, J= 4.4 Hz, 1 H), 4.64 (brs, 1 H), 5.00 (t, J= 6.8 Hz, 1 H), 6.72 (s, 1 H), 6.81 6.93 (series of m, 8 H), 7.22-7.42 (series of m, 6 H), 8.01 (d, J= 8.4 Hz, 2 H), 8.13 (d, J= 8.0 Hz, 1H); MS (FAB) m/z, 656 (M), 658 (MW+2). To a stirred solution of methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methylphenylacetyl]-5-(R) phenyl-2-(S)-pyrrolidinylmethoxy]benzoate (216 mg, 0.33 mmol) in MeOH-THF (1 : 1, 10 ml) was 10 added 1.0M-NaOH (1.7 ml, 1.7 mmol) at rt, and the resulting mixture was heated at 60oC with stirring for 2.5 h. The reaction mixture was poured into 1N-HC1, then extracted with CHC1 3 . The organic layer was washed with brine and dried over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (10 : 1) to give 149 (166 mg, 91%) as a colorless amorphous solid. MW 642.54 'H-NMR (CDC13) 8 2.01 (s, 3 H), 15 2.05-2.25 (m, 3 H), 2.43-2.48 (m, 1 H), 3.34 (dd, J= 45, 16 Hz, 2 H), 4.38-4.45 (m, 2 H), 4.66 (brs, 1 H), 4.99 (t, J= 6.8 Hz, 1 H), 6.77 (s, 1 H), 6.82-6.88 (m, 2 H), 6.94 (d, J= 8.8 Hz, 2 H), 7.15-7.55 (series of m, 10 H), 8.00 (d, J= 8.8 Hz, 2 H), 8.14 (d, J= 7.2 Hz, 1 H); MS (FAB) m/z 642 (M), 644 (M++2). Example 142 20 4-[1-[4-[N'-(2-methylphenyl)ureido]-3-methoxyphenylacetyl]-5-(R)-methyl-2-(S)-pyrrolidinyl methoxy]benzoic acid NN \/ COOH Me H H OMe 150 To a stirred solution of benzy N-Boc-pyrroglutarate (7.55 g, 23.6 mmol) in THF (100 ml) was added MeLi (1.1 M in Et 2 0, 28.4 ml, 32.4 mmol) at -78oC, and the resulting mixture was 25 gradually warmed up to rt, then stirred overnight. aq.NH 4 CI was added to the reaction mixture, THF was removed in vacuo, then extracted with EtOAc. The organic layer was washed with water and, drying over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with hexane-EtOAc (3 : 1) as eluent to give benzyl [2-(S)-(N-Boc amino)-5-oxo-6-methyl]pentanoate (5.02 g, 45%) as a colorless needles. mp 85-87 oC; 'H-NMR 30 (CDC1 3 ) 8 1.43 (s, 9 H), 1.61-2.15 (series of m, 3 H), 2.09 (s, 3H), 2.41-2.55 (m, 2 H), 4.30 (brs, 1 H), 4.70 (d, J= 5.6 Hz, 1 H), 5.12-5.21 (m, 2H), 7.29-7.37 (m, 5 H); MS (ESI) m/z, 336 (M'+H). 311 WO 01/00206 PCT/US00/18079 To a stirred solution of benzyl [2-(S)-(N-Boc-amino)-5-oxo-6-methyl]pentanoate (4.46 g, 13.3 mmol) in CH 2 Cl 2 (50 ml) was added trifluoroacetic acid (20 ml) at rt, and the resulting mixture was stirred for 1.5 h. The mixture was concentrated in vacuo, and dissolved in toluene, then evaporated to give benzyl 5-methyl-5-pyrroline-2-(S)-carboxylate trifluoroacetic acid salt (5.74 g, 5 quant.) as a crude brown oil. This compound (1.97 g, 5.94 mmol) in MeOH (30 ml) was added Pd/C (10%, 153 mg), and the resulting mixture was stirred for 3 days under H 2 atomosphere. The mixture was filtered, and the filtrate was concentrated in vacuo to give 5-methyl-5-pyrrolidine-2 (S)-carboxylic acid trifluoroacetic acid salt (956 mg, 66%) as a crude white solid. To a solution of this compound (939 mg, 3.86 mmol) and di-tert-buyl dicarbonate in MeCN-water (15 : 1, 16 ml) 10 was added 1.0M-NaOH (8.49 mmol, 8.49 ml) at rt, and the resulting mixture was stirred for lh. The resultiong mixture was evaporated and poured into aq.-1N-HC1, then extracted with CHCI/MeOH (5 : 1). The organic layer was dried over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (7: 1) to give N-Boc-5 (R)-methyl-(S)-proline (711 mg, 80%) as a colorless oil. 'H-NMR (CD 3 OD) 8 1.27 (d, J = 6.0 Hz, 15 3H11), 1.41-1.46 (min, 9H), 1.62-1.64 (m, 1H), 1.96-2.01 (min, 2H11), 2.22 (brs, 1H), 3.94 (brs, 1H11), 4.17 (brs, 1H).; MS (ESI) nm/z, 230 (M'+H). To a stirred solution of N-Boc-5-(R)-methyl-2-(S)-proline (1.03 g, 4.49 mmol) in THF (20 ml) was added O10M-BH 3 -Me 2 S (1.57 ml, 15.7 mmol) at rt, and the resulting mixture was heated under reflux for 5 h. The mixture was poured into aq. 1N-HCl and extracted with EtOAc. The 20 organic layer was dried over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with hexane-AcOEt (1 : 3) as eluent to give N-Boc-2-(S) hydoxymethyl-5-(R)-methylpyrrolidine (838 mg, 87%) as a colorless oil: 'H-NMR (CDCl 3 ) 5 1.17 (d, J= 6.0 Hz, 3H), 1.48 (s, 9H), 1.48-1.64 (m, 2H11), 1.90-2.11 (m, 2H11), 3.52-3.57 (m, 1H), 3.68 3.70 (m, 1H11), 3.94-4.13 (m, 1H). 25 To a stirred solution of N-Boc-2-(S)-hydoxymethyl-5-(R)-methylpyrrolidine (820 mg, 3.81 mmol), triphenylphosphine (1.10 g, 4.19 mmol) and methyl 4-hydroxybenzoate (580 mg, 3.81 mmol) was added diisopropyl azodicarboxylate (841 ml, 4.19 mmol) at rt, and the resulting mixture was stirred at 60oC for 1 h. The mixture was concentrated in vacuo, and the residue was chromatographed on silica gel with hexane-EtOAc (5 : 1) as eluent to give methyl 4-[N-Boc-5-(R) 30 methyl-2-(S)-pyrrolidinylmethoxy]benzoate (1.32 g, 80%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.24 (brs, 3 H1), 1.49 (s, 9 H), 1.55-1.70 (min, 2 H), 1.94-2.11 (min, 2 H), 3.88 (s, 3 H), 3.88 (overlap, 1H1), 4.06-4.20 (m, 2H11), 6.93-6.96 (min, 2H), 7.97 (d, J= 8.8 Hz, 2 H); MS (ESI) m/z, 350 (M+H). 312 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[N-Boc-5-(R)-methyl-2-(S)-pyrrolidinylmethoxy] benzoate (1.29 g, 3.70 mmol) in CH2C1 2 (30 ml) was added trifluoroacetic acid (10 ml) at rt, and the resulting mixture was stirred for 35 min. The mixture was concentrated in vacuo and poured into aq.NaHCO 3 , then extracted with CHCl 3 . The organic layer was washed with water, drying over 5 anhydrous Na 2
SO
4 , and concentrated in vacuo to give methyl 4-[5-(R)-methyl-2-(S)-pirrolidinyl methoxy]benzate (871 mg, 95%) as a colorless oil. The product was used for next reactions without further purification. 'H-NMR (CDCl 3 ) 6 1.18 (d, J= 6.4 Hz, 3 H), 1.30-1.40 (m, 1 H), 1.59-1.67 (m, 1 H), 1.87-1.97 (m, 2 H), 3.19-3.27 (m, 1 H), 3.49-3.55 (m, 1 H), 3.87 (s, 3 H), 3.89 4.05 (m, 2 H), 6.89 (d, J= 8.8 Hz, 2 H), 7.96 (d, J= 8.8 Hz, 2 H); MS (ESI) m/z, 250 (M +H) 10 To a stirred solution of methyl 4-[5-(R)-methyl-2-(S)-pirrolidinylmethoxy]benzoate (141 mg, 0.57 mmol), 4-[N'-(2-methylphenyl)ureido]-3-methoxyphenylacetic acid (178 mg, 0.57 mmol) and N,N dimethylaminopyridine (69.0 mg, 0.57 mmol) in DMF (10 ml) was added EDCHCI (120 mg, 0.62 mmol) at rt, and the resulting mixture was stirred overnight. The reaction mixture was poured into water and extracted with EtOAc. The organic layer was washed with brine, drying over anhydrous 15 Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with EtOAc as eluent to give methyl 4-[1-[4-[N'-(2-methylphenyl)ureido]-3-methoxyphenyl acetyl]-5-(R) methyl-2-(S)-pyrrolidinylmethoxy]benzoate (297 mg, 96%) as a colorless amorphous solid. 1
H
NMR (CDCl 3 ) 8 1.24-1.34 (m, 3 H), 1.93-2.18 (series of m, 4 H), 2.28 (s, 3 H), 3.65 (s, 3 H), 3.88 (s, 3 H), 3.62-3.87 (m, 3 H), 4.11-4.38 (series of m, 3 H), 6.42-8.06 (series of m, 13 H); MS (ESI) 20 m/z, 546 (M++H). To a stirred solution of methyl 4-[1-[4-[N'-(2-methylphenyl)ureido]-3-methoxyphenylacetyl]-5 (R)-methyl-2-(S)-pyrrolidinylmethoxy]benzoate (279 mg, 0.51 mmol) in MeOH-THF (1 : 1, 10 ml) was added 1.0M-NaOH (2.56 ml, 2.56 mmol) at rt, and the resulting mixture was heated at 60oC with stirring for 2 h. The reaction mixture was poured into IN-HC1, then extracted with CHC1 3 . 25 The organic layer was washed with brine and dried over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (15 : 1) to give 150 (269 mg, 99%) as a colorless amorphous solid. MW 531.60 'H-NMR (CD30D), mixture of rotamars, 8 1.28-1.35 (m, 3 H), 1.74-2.21 (series of m, 4 H), 2.28 (s, 3 H), 3.71-4.37 (series of m, 6 H), 6.76 7.99 (series of m, 11 H); MS (ESI) m/z, 532 (M+H). 30 Example 143 4-[trans-4-amino- 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S)-pyrrolidinyl] methoxybenzoic acid 313 WO 01/00206 PCT/US00/18079
NH
2 N N O-D COOH Me H H - Me 0151 To a solution of methyl 4-(trans-4-amino-1-tert-butoxycarbonyl-(2S)-pyrrolidinyl) methoxybenzoate (1.0 g, 2.86 nunol) and TEA (1.2 ml, 8.6 mmol) in CH 2
CI
2 (20.0 ml) was added trifluoroacetic anhydride (720 mg, 3.43 mmol) at 0 0 C. After stirred for 2.5 hr at room 5 temperature, water was added to the solution and extracted with CH 2 C1 2 . The extract was washed with water, then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with EtOAc-n-hexane (1:3 ,v/v) as eluent to give methyl 4 (trans-I -tert-butoxycarbonyl-4-trifluoroacetamido-(2S)-pyrrffolidinyl)methoxybenzoate (940 mg, 74%) as a colorless oil. 1H-NMR (CDCl 3 ) 8 1.46 (s, 9H), 2.02-2.18 (m, 1H), 2.41-2.52 (min, 1H11), 10 3.30-3.45 (min, 1H), 3.80-3.90 (min, 1H), 3.88 (s, 3H), 4.00-4.30 (min, 3H), 4.65-4.75 (mn, 1H), 6.50 (br s, 1H), 6.91-6.94 (min, 2H), 7.96-7.99 (min, 2H). To a stirred solution of methyl 4-(trans-1-tert-butoxycarbonyl-4-trifluoroacetamido-(2S) pyrrolidinyl)methoxybenzoate (470 mg, 1.05 mmol) in CH 2 Cl 2 (10.0 ml) was added TFA (5.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was 15 concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 C1 2 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product, 3-methoxy-4-[N'-(2-methylphenyl)uredio]phenylacetic acid (314 mg, 1.0 mmol), HOBt (162 mg, 1.2 mmol), and triethylamine (417 ml, 3.0 mmol) in THF (10.0 ml) and MeCN (10.0 ml) 20 was added EDC-HCI (288 mg, 1.5 mmol) at 0 'C. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The organic layer was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with EtOAc-n-hexane (3:1, v/v) as eluent to give methyl 4-[trans-1 25 [3 -methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-4-trifluoroacetamido-(2S)-pyrrolidinyl] methoxybenzoate (350 mg, 52%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 2.01-2.10 (min, 11-H), 2.31 (s, 3H), 2.42-2.48 (min, 1H), 3.45-3.50 (min, 1H1), 3.56-3.59 (min, SH), 3.89 (s, 3H), 4.07-4.14 (min, 2H), 4.38-4.42 (min, 1H), 4.50-4.60 (min, 11-1), 4.72-4.80 (min, 1H), 6.33 (s, 1H), 6.60-6.85 (m, 3H), 7.06 7.26 (m, 3H), 7.48-7.52 (min, 1H), 7.93-8.05 (min, 3H). 314 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[trans-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] 4-trifluoroacetamido-(2S)-pyrrolidinyl]methoxybenzoate (150 mg, 0.23 mmol) in THF (3.0 ml) and MeOH (2.0 ml) was added IN NaOH (0.70 ml, 0.70 mmol). The mixture was stirred at 60 oC for 18 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with 5 IN HC1. The resulting solid was collected, washed with water, and dried in vacuo to give 151 (100 mg, 81%) as a white crystalline solid. MW 532.59 mp 170-171 'C; IR (KBr) 3264, 2937, 1604, 1535, 1415, 1376, 1255, 1224, 1033 cm-'; 'H-NMR (DMSO-d6) 8 1.80-1.90 (m, 1H), 2.10 2.20 (m, 1H), 2.24 (s, 3H), 3.55-3.80 (m, 3H), 3.57 (s, 2H), 4.08-4.18 (m, 2H), 4.36-4.60 (m, 1IH), 6.72-7.16 (m, 7H11), 7.77-8.01 (m, 4H), 8.46 (s, 1H), 8.54 (s, 1H); MS (FAB)m/z 532 (MW+1); Anal. 10 calcd for C 2 9
H
32
N
4 0 6 -2.0H 2 0: C, 61.26; H, 6.38; N, 9.85. Found: C, 61.07; H, 6.32; N, 9.58. Example 144 methyl 4-[trans-4-amino-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S) pyrrolidinyl]methoxybenzoate HCI salt
NH
2 . ~ N N N COOMe 1 Me H HNMCOOMe 152 15 To a stirred solution of methyl 4-[trans-l-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] 4-trifluoroacetamido-(2S)-pyrrolidinyl]methoxybenzoate (200 mg, 0.31 mmol) in MeOH (4.0 mml) was added water (2.0 ml) and K 2
CO
3 (138 mg, 1.0 mml) at room temperature. After stirred for 18hr at room temperature, water was added to the mixture and extracted with CH2C1 2 . The extract was washed with water, then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was 20 purified by column chromatography on silica gel with MeOH-CH 2
CI
2 (5:95 to 15:85, v/v) as eluent. The product was dissolved in EtOH (5.0 ml), and IN HCI (in EtOH) (1.0 ml, 1.0 mmol) was added thereto. The mixture was concentrated in vacuo to give 152 (120 mg, 63%) as an amorphous solid. MW 546.61 IR (KBr) 3382, 2948, 2879, 1604, 1533, 1286, 1255, 771 cm-'; 'H-NMR (DMSO-d 6 ) 8 2.25 (s, 3H), 2.10-2.30 (m, 2H), 3.59-3.70 (m, 3H11), 3.77-3.80 (m, 8H), 4.00-4.24 (m, 25 2H), 4.47-4.67 (m, 1H), 6.70-7.16 (m, 7H11), 7.77-8.00 (m, 4H11), 8.49 (s, 1H), 8.55 (s, 1H); MS (FAB) m/z 547 (M+1);Anal. calcd for C 30
H
34
N
4 0 6 .HCI-1.4H 2 0: C, 59.24; H, 6.26; N, 9.21; Cl, 5.83 Found: C, 59.42; H, 6.42; N, 9.04; Cl, 6.11. Example 145 4-[trans-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-4-methylamino-(2S) 30 pyrrolidinyl]methoxybenzoic acid 315 WO 01/00206 PCT/US00/18079 Me'H ~5~NZ N N O COOH Me H H- Me C -a153 To a stirred solution of methyl 4-(trans-1-tert-butoxycarbonyl-4-trifluoroacetamido-(2S) pyrrolidinyl)methoxybenzoate (520 mg, 1.17 mmol) in DMF (10.0 ml) was added K 2 CO3 (321 mg, 2.33 mmol) and Mel (330 mg, 2.33 mmol) at room temperature. The reaction mixture was stirred 5 at 50 0 C for 18 hr. Water was added to the mixture and extracted with EtOAc. The organic layer was washed with water, then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with EtOAc-n-hexane (1:2 ,v/v) as eluent to give methyl 4-[trans-1-tert-butoxycarbonyl-4-(N-methyl-trifluoroacetamido)-(2S)-pyrrolidinyl] methoxybenzoate (390 mg, 73%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.46 (9H, s), 2.12-2.40 10 (min, 2H1), 2.96 and 3.05 (each s, total 3H), 3.28-3.70 (min, 2H11), 3.88 (s, 3H), 3.95-4.42 (min, 3H), 5.10 5.40 (min, 1H), 6.89-6.91 (min, 2H), 7.96-8.00 (min, 2H). To a stirred solution of methyl 4-[trans-1-tert-butoxycarbonyl-4-(N-methyl-trifluoro acetoamido)-(2S)-pyrrolidinyl]methoxybenzoate (390 mg, 0.85 mmol) in CH 2 Cl 2 (8.0 ml) was added TFA (5.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The 15 mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with
CH
2 C1 2 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product, 3-methoxy-4-[N'-(2-methylphenyl)uredio]phenylacetic acid (279 mg, 0.89 mmol), HOBt (143 mg, 1.1 mmol), and triethylamine (246 ml, 1.77 mmol) in THF (8.0 ml) 20 and MeCN (8.0 ml) was added EDC-HCl (255 mg, 1.3 mmol) at 0 C. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with EtOAc-n-hexane (4:1 ,v/v) as eluent to give methyl 4-[trans-1 25 [3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-4-(N-methyl-trifluoroacetoamido)-(2S) pyrrolidinyl]methoxybenzoate (480 mg, 82%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 2.18-2.35 (min, 2H), 2.31 (s, 3H), 2.87 and 2.97 (each s, total 3H), 3.45-3.46 (min, 3H), 3.47 (s, 3H), 3.49 (s, 2H), 3.88 (s, 3H), 4.30-4.70 (min, 2H), 5.20-5.40 (min, 1H), 6.38-6.43 (mn, 1H), 6.67-6.86 (min, 4H), 7.09-7.24 (min, 4H1), 7.51-7.54 (mn, 1H), 7.93-8.08 (min, 3H). 316 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[trans-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] 4-(N-methyl-N-trifluoroacetylamino)-(2S)-pyrrolidinyl]methoxybenzoate (240 mg, 0.37 mmol) in THF (5.0 ml) and MeOH (3.0 ml) was added IN NaOH (1.27 ml, 1.27 mmol). The mixture was stirred at 60 oC for 18 hr. The mixture was concentrated in vacuo, water was added thereto, and 5 neutralized with IN HC1. The resulting solid was collected, washed with water, and dried in vacuo to give 153 (140 mg, 70%) as a white crystalline solid. MW546.61 mp 162-164 oC; IR (KBr) 3338, 1604, 1535, 1255, 1033, 755 cm'; 'H-NMR (DMSO-d) 8 1.85-1.95 (min, 1H1), 2.10-2.20 (inm, 1H-1), 2.24 (s, 3H11), 2.34 and 2.39 (each s, total 3H), 3.41-3.71 (min, 3H), 3.58 (s, 2H), 3.80 (s, 3H), 4.05-4.20 (min, 2H), 4.36-4.60 (min, 1H1), 6.73-7.16 (min, 7H), 7.77-8.01 (min, 4H1), 8.45 (s, 1H), 8.53 (s, 10 1H); MS (FAB) mnVz 547 (M+1); Anal. calcd for C 3 0H 34 N40-2.5H 2 0: C, 60.90; H, 6.64; N, 9.47. Found: C, 61.01; H, 6.50; N, 9.31. Example 146 methyl 4-[trans- 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-4-methylamino-(2S) pyrrolidinyl]methoxybenzoate Me"H N N COOMe 15 Me H H - Me 154 To a stirred solution of methyl 4-[trans-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] 4-(N-methyltrifluoroacetamido)-(2S)-pyrrolidinyl]methoxybenzoate (240 mg, 0.36 mmol) in THF (5.0 mml) and MeOH (5.0 ml) was added water (2.0 ml) and K 2
CO
3 (138 mg, 1.0 mml) at room temperature. After stirred for 18hr at room temperature, water was added to the mixture and 20 extracted with CH 2 C1 2 . The extract was washed with water, then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with MeOH-CH 2 C1 2 (5/95 to 20/80, v/v) as eluent. The product was dissolved in EtOH (5.0 ml), and IN HCI (in EtOH) (0.71 ml, 0.71 mmol) was added thereto. The mixture was concentrated in vacuo to give 154 (180 mg, 85%) as an amorphous solid. MW 560.64 IR (KBr) 3311, 2692, 2453, 1712, 25 1604, 1533 cmr'; 'H-NMR (DMSO-d 6 ) 8 2.24 (s, 3H), 2.15-2.30 (min, 2H), 2.60 (br s, 3H1), 3.60-4.20 (min, 5H), 3.78-3.81 (min, 8H), 4.47-4.70 (min, 1), 6.71-7.16 (min, 7H), 7.77-8.00 (min, 4H), 8.48 (s, 111), 8.55 (s, 1H1), 9.21 (br s, 2H); MS (FAB) m/z 561 (M +1); Anal. calcd for C 3 zHz 6
N
4 0 6 -HCl 1.411H20: C, 59.83; H, 6.45; N, 9.00; Cl, 5.70. Found: C, 60.08; H, 6.51; N, 8.68; Cl, 5.99. Example 147 30 4-[trans-4-dimethylamino-1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S) pyrrolidinyl]methoxybenzoic acid 317 WO 01/00206 PCT/US00/18079 Me, Me N N 0 O- COOMe 155 Me H H- Me 0155 To a stirred solution of trans- 1-tert-butoxycarbonyl-(2S)-hydroxymethyl-4-hydroxypyrrolidine (2.17 g, 10.0 mmol) and imidazole (2.04 g, 30.0 mmol) in DMF (50 ml) was added TBDPS-Cl (3.03 g, 11.0 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 18 hr. 5 Water was added thereto, and extracted with EtOAc. The extract was washed with water, then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc(3:2 ,v/v) as eluent to give trans- -tert butoxycarbonyl-(2S)-(tert-butyldiphenylsilyloxy)methyl-4-hydroxypyrrolidine (1.5 g, 33%) as a white crystalline solid. 1 H-NMR (CDCl 3 ) 8 1.03 (s9H), 1.25 and 1.32 (each s, 9H1), 1.90-2.10 (inm, 10 1H), 2.30-2.40 (min, 1H), 3.40-3.80 (min, 3H), 3.95-4.15 (min, 2H), 4.45-4.55 (min, 1H), 7.37-7.39 (inm, 6H), 7.63-7.64 (min, 4H). To a stirred solution of trans-1 -tert-butoxycarbonyl-(2S)-(tert-butyldiphenylsilyloxy) methyl-4-hydroxypyrrolidine (910 mg, 2.0 mmol) and Ph 3 P (628 mg, 2.4 mmol) in THF (20 ml) was added CBr 4 (993mg, 3.0 mmol) at room temperature. The reaction mixture was stirred at 15 room temperature for 0.5 hr. n-Hexane (40ml) was added thereto. The resulting solid was filtered off, and dried in vacuo. The residue was purified by column chromatography on silica gel with n hexane to n-hexane-EtOAc (3:2, v/v) as eluent to give cis-4-bromo-1-tert-butoxycarbonyl-(2S) (tert-butyldiphenylsilyloxy)methylpyrrolidine (1.0 g, quant.) as a pale yellow oil. 'H-NMR (CDC13) 8 1.06 (s, 9H), 1.31 and 1.45 (each s, 9H), 2.63 (min, 2H), 3.49 (mn, 1H), 3.89-4.14 (mn, 5H), 20 7.35-7.42 (min, 6H), 7.64-7.66 (, 4Hm). To a stirred solution of cis-4-bromo-1-tert-butoxycarbonyl-(2S)-(tert-butyldiphenylsilyloxy) methylpyrrolidine (480 mg, 0.93 mmol) in DMF (5 ml) was added NaN 3 (241 mg, 3.70 mmol) at room temperature. The reaction mixture was stirred at 70 for 3 days. Water was added thereto, and extracted with EtOAc. The extract was washed with water, then dried over Na 2
SO
4 , and 25 concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. The solution of the crude residue in EtOH (10 ml) was hydrogenated over 10% Pd-C under an atmospheric pressure at room temperature for 4 hr. The catalyst was filtered off, and the filtrate was concentrated in vacuo to give trans-4-amino-1l-tert-butoxycarbonyl-(2S)-(tert butyldiphenylsilyloxy)methylpyrrolidine (400 mg, 95%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 318 WO 01/00206 PCT/US00/18079 1.06 (s, 9H), 1.32 and 1.45 (each s, total 9H), 2.20-2.35 (min, 1H), 3.05-3.18 (min, 1H), 3.55-4.05 (inm, 6H), 7.35-7.41 (min, 6H), 7.61-7.69 (min, 4H). To a stirred solution of trans-4-amino-l-tert-butoxycarbonyl-(2S)-(tert-butyldiphenylsilyloxy) methylpyrrolidine (400 mg, 0.88 mmol), AcOH (120 ml, 2.0 mmol), and 37% HCHO aq (500 ml) 5 in MeOH (10 ml) was added NaBH 3 CN (111 mg, 1.76 mmol) at 0 'C. The reaction mixture was stirred at room temperature for 18 hr. After concentrated in vacuo, water was added and extracted with CH 2 C1 2 . The extract was dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with MeOH-CH 2 Cl 2 (3:97, v/v) as eluent to give trans-1 -tert-butoxycarbonyl-(2S)-(tert-butyldiphenylsilyloxy)methyl-4-dimethylaminopyrrolidine 10 (330 mg, 78%) as a pale yellow oil. 'H-NMR (CDC13) 8 1.06 (s, 9H), 1.33 and 1.45 (each s, total 9H), 1.80-2.25 (min, 2H), 2.23 (br s, 6H), 2.95-4.05 (min, 6H), 7.36-7.39 (min, 6H), 7.63-7.65 (mn, 4H). To a stirred solution of trans-1-tert-butoxycarbonyl-(2S)-(tert-butyldiphenylsilyloxy)methyl-4 dimethylaminopyrrolidine (330 mg, 0.68 mmol) in THF (5 ml) was added TBAF (1.0 M solution in THF, 1.0 ml, 1.0 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 2 hr. 15 The mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with MeOH-CH 2
CI
2 (3:97 to 20:80, v/v) as eluent to give trans-1-tert-butoxycarbonyl-4 dimethylamino-(2S)-hydroxymethylpyrrolidine (180 mg, quant.) as a pale yellow oil. 'H-NMR (CDCl 3 ) 8 1.47 (s, 9H), 2.23 (s, 6H), 1.65-1.75 (m,2H),2.75-4.10 (min, 4H), 3.61 (d, J= 5.6 Hz, 2H). To a stirred solution of trans- -tert-butoxycarbonyl-4-dimethylamino-(2S)-hydroxymethyl 20 pyrrolidine (180 mg, 0.73 mmol), methyl 4-hydroxybenzoate (114 mg, 0.75 mmol), and Ph 3 P (296 mg, 1.13 mmol) in THF (10 ml) was added DIAD (227mg, 1.13 mmol) at 0 oC. The reaction mixture was stirred at 70 oC for 18 hr. The mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (1:2, v/v) to MeOH
CH
2 C1 2 (5:95, v/v) as eluent to give methyl 4-[trans- 1-tert-butoxycarbonyl-4-dimethylamino-(2S) 25 pyrrolidinyl]methoxybenzoate (180 mg, 68%) as a pale yellow oil. 'H-NMR (CDCl 3 ) 8 1.46 (s, 9H), 1.80-1.95 (min, 1H), 2.20-2.23 (min, 1H), 2.24 (s, 6H), 2.90-2.95 (min, 1H), 3.10-3.30 (min, 1H), 3.50-3.65 (min, 1H), 3.88 (s, 3H), 3.95-4.35 (min, 3H), 6.93-6.95 (min, 2H), 7.96-7.98 (min, 2H). To a stirred solution of methyl 4-(trans-l-tert-butoxycarbonyl-4-dimethylamino-(2S) pyrrolidinyl)methoxybenzoate (200 mg, 0.53 mmol) in CH 2 C1 2 (6 ml) was added TFA (3ml) at 319 WO 01/00206 PCT/US00/18079 0 0 C. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 Cl 2 . The extract was washed with brine ,dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude 5 product, 3-methoxy-4-[N'-(2-methylphenyl)uredio]phenylacetic acid (166 mg, 0.53 mmol), HOBt (71 mg, 0.53 mmol), and triethylamine (140 ml, 1.10 mmol) in THF (5 ml) and MeCN (5 ml) was added EDC.HCI (152mg, 0.79 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated 10 in vacuo. The residue was purified by column chromatography on silica gel with EtOAc to
CH
2 Cl 2 -MeOH(8:92, v/v) as eluent to give methyl 4-[trans-4-dimethylamino-1-[3-methoxy-4-[N' (2-methylphenyl) ureido]phenylacetyl]-(2S)-pyrrolidinyl]methoxybenzoate (260 mg, 86%) as a colorless oil. 'H-NMR (CDC13) 8 1.95-2.15 (min, 3H), 2.23 (s, 6H), 2.31 (s, 3H), 3.30-3.34 (mn, 1H), 3.57 (s, 2H), 3.61 (s, 3H), 3.70-3.75 ( min, 1I), 4.11-4.15 (min, 2H), 4.45-4.50 (min, 1H), 6.34 (s, 1H), 15 6.72-6.88 (min, 4H), 7.08-7.24 (min, 4H), 7.51-7.53 (min, 1H), 7.92-8.07 (min, 3H). To a stirred solution of methyl 4-[trans-4-dimethylamino-1-[3-methoxy-4-[N'-(2 methylphenyl) ureido] phenylacetyl]-2-pyrrolidinyl]methoxybenzoate (260 mg, 0.45 mmol) in THF (4.0 ml) and MeOH (2.0 ml) was added IN NaOH (0.90 ml, 0.90 mmol). The mixture was stirred at 70 oC for 24 hr. The mixture was concentrate in vacuo, water was added thereto, and 20 neutralized with IN HC1. The resulting solid was collected, washed with water, and dried in vacuo to give 155 (200 mg, 79%) as a white crystalline solid. MW 560.64 mp 145-150 oC; IR (KBr) 3355, 2948, 1698, 1604, 1533, 1454, 1417, 1255, 1226, 1166, 1035, 755 cm-'; 'H-NMR (DMSO d 6 ) 8 1.82-1.98 (min, 1H), 2.08-2.11 (min, 1H), 2.20 (s, 6H), 2.25 (s, 3H), 3.40-3.60 (min, 3H), 3.64 (s, 2H), 3.82 (s, 3H), 4.01-4.16 (min, 2H), 4.36 (min, 1H), 6.74-7.15 (min, 7H), 7.77-8.02 (min, 4H), 8.44 (s, 25 1H), 8.54 (s, 1H); MS (FAB) m/z 561 (M+1); Anal. calcd for C 3
,H
36
N
4 0 6 1.2H 2 0: C, 63.95; H, 6.65; N, 9.62. Found: C, 63.82; H, 6.72; N, 9.44. Example 148 methyl 4-[trans-4-dimethylamino- 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S) pyrrolidinyl]methoxybenzoate HCI salt 320 WO 01/00206 PCT/US00/18079 Me\ Me NYN N O-- COOMe MeH H Me HCL 156 To a stirred solution of trans-4-[4-dimethylanmino-1-[3-methoxy-4-[N'-(2-methylphenyl) ureido] phenylacetyl-(2S)-pyrrolidinyl]methoxybenzoic acid (80 mg, 0.14 mmol) in toluene (4.0 ml) and MeOH (1.0 ml) was added TMSCHN 2 (2.0 M in hexane, 100 ml, 0.20 mmol) at 0 *C. The 5 reaction mixture was stirred at room temperature for 1.5 hr. The mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with MeOH
CH
2 Cl 2 (5:95, v/v) as eluent. The product was dissolved in EtOH (5.0 ml), and IN HC1 (in EtOH) (2444 1, , 0.244mmol) was added thereto. The mixture was concentrated in vacuo to give 156 (72 mg, 88%) as an amorphous solid. MW 574.67 IR (KBr) 3345, 2950, 2586, 1712, 1604, 1511, 10 1454, 1284, 1255, 1170, 1114, 1029, 850, 771 cm'; 'H-NMR (DMSO-d) 8 2.25 (s, 3H), 2.35-2.37 (min, 2H), 2.77-2.81 (min, 6H), 3.62-3.71 (min, 2H), 3.79-3.81 (min, 8H), 3.99-4.16 (min, 3H), 4.50-4.70 (min, 1H), 6.74-7.16 (min, 7H), 7.77-8.01 (min, 4H), 8.48 (s, 1H), 8.55 (s, 1H);Anal calcd for
C
32
H
38
N
4 0 6 -1.0HCI 1.2 H 2 0: C, 60.74; H, 6.59; N, 8.85. Found: C, 61.03; H, 6.78; N, 8.33. Example 149 15 4-[cis-4-dimethylamino-1-[3-methoxy-4-[N'-(2-methylphcnyl)ureidolphenylacetyl]-(2S) pyrrolidinyl]methoxybenzoic acid Me\N-Me N: NO \/ COOH Me H H OMe 157 To a stirred solution of cis-1-tert-butoxycarbonyl-(2S)-(tert-butyldiphenylsilyloxy)methyl 4-hydroxypyrrolidine (1.82 mg, 4.0 mmol), phthalimide (647 mg, 4.4 mmol), and Ph 3 P (1.26 g, 20 4.8 mmol) in T-F (20 ml) was added DIAD (889 mg, 4.4 mmol) at 0 'C. The reaction mixture was stirred at room temperature for 18 hr. The mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (5/1, v/v) as eluent to give N-[cis-1-tert-butoxycarbonyl-(2S)-(tert-butyldiphenylsilyloxy)methyl-4 pyrrolidinyl]phthalimide (1.6 g, 69%) as an amorphous solid. 'H-NMR (CDCI 3 ) 8 1.07 (s, 9H), 25 1.30 and 1.44 (each s, total 9H), 2.27-2.37 (min, 1H), 2.94-2.96 (min, 1H), 3.81-4.09 (min, 5H), 4.72 (min, 1H), 7.37-7.38 (min, 6H), 7.67-7.74 (min, 6H), 7.84-7.86 (min, 2H). 321 WO 01/00206 PCT/US00/18079 To a stirred solution of N-[cis-l-tert-butoxycarbonyl-(2S)-(tert-butyldiphenylsilyloxy) methyl-4-pyrrolidinyl]phthalimide (1.60 g, 2.74 mmol) in EtOH (8 ml) was added
NH
2 NH2*H 2 0(206 mg, 4.11 mmol) at room temperature. The reaction mixture was stirred at 70 for 1 hr. The mixture was concentrated in vacuo. The resulting solid was filtered off, and washed 5 with CHCl 3 . The filtrate was concentrated in vacuo. The resulting solid was filtered off, and washed with CHC1 3 . The filtrate was concentrated in vacuo to give cis-4-amino-1-tert butoxycarbonyl-(2S)-(tert-butyl diphenylsilyloxy) methylpyrrolidine (1.3 g, quant) as a pale yellow oil. The crude product was used to the subsequent reaction without further purification. 'H-NMR (CDCl 3 ) 8 1.06 (s, 9H11), 1.30 and 1.45 (each s, total 9H11), 1.59 (mn, 1H11), 1.85 (mn, 1H), 2.94 (mn, 1H), 10 3.44 (min, 1H), 3.78-4.07 (min, 4H), 7.36-7.41 (min, 6H), 7.51-7.65 (min, 4H). To a stirred solution of cis-4-amino-1-tert-butoxycarbonyl-(2S)-(tert-butyldiphenyl ilyloxy) methylpyrrolidine (1.24 g, 2.74 mmol), AcOH (374.1, 5.48 mmol), and 37% HCHO aq (1.0 ml) in MeOH (20 ml) was added NaBH 3 CN (345 mg, 5.48 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 18 hr. After concentrated in vacuo, water was added 15 and extracted with CH 2 C1 2 . The extract was dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with MeOH-CH 2 Cl0 2 (3/97, v/v) as eluent to give cis- 1-tert-butoxycarbonyl-(2S)-(tert-butyldiphenylsilyloxy)methyl-4-dimethylamino pyrrolidine (1.1 g, 83%) as a pale yellow oil. 'H-NMR (CDCl 3 ) 5 1.05 (s, 9H), 1.29 and 1.45 (each s, total 9H1), 1.95-2.04 (min, 111), 2.20-2.26 (min, 1H), 2.27 (s, 6H), 2.54 (mn, 1H), 3.00-3.02 (min, 1H), 20 3.62-4.03 (min, 4H), 7.34-7.41 (mn, 6H11), 7.63-7.65 (min, 4H11). To a stirred solution of cis-1-tert-butoxycarbonyl-2-(tert-butyldiphenylsilyloxy)methyl-4-dimethyl amino pyrrolidine (1.1 g, 2.27 mmol) in THF (10 ml) was added TBAF (1.0 M solution in THF) (4.5 ml, 4.5 nmmunol) at 0 oC. The reaction mixture was stirred at room temperature for 2 hr. The mixture was concentrated in vacuo. The residue was purified by column chromatography on silica 25 gel with MeOH-CH 2 Cl 2 (3/97 to 20/80, v/v) as eluent to give cis-1-tert-butoxycarbonyl-4 dimethylamino-(2S)-hydroxymethylpyrrolidine (580 mg, quant.) as a pale yellow oil. 'H-NMR
(CDCI
3 ) 8 1.47 (s, 9H), 1.25-1.96 (min, 2H), 2.25 (s, 6H), 2.53-2.58 (mn, 1H11), 3.17-4.02 (mn, 5H). To a stirred solution of cis-1 -tert-butoxycarbonyl-4-dimethylamino-(2S)-hydroxymethylpyrrolidine (555 mg, 2.27 mmol), methyl 4-hydroxybenzoate (380 mg, 2.5 mmol), and Ph 3 P (1.07 g, 4.09 30 mmol) in THF (10 ml) was added DIAD (826 mg, 4.09 nummol) at 0 oC. The reaction mixture was 322 WO 01/00206 PCT/US00/18079 stirred at 70 oC for 18 hr. The mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (1/2, v/v)MeOH-CH 2
C
2 (5/95, v/v) as eluent to give methyl 4-(cis-1l-tert-butoxycarbonyl-4-dimethylamino-(2S)-pyrrolidinyl)methoxy benzoate (260 mg, 30%) as a pale yellow oil. 'H-NMR (CDC 3 ) 8 1.45 (s, 9H), 1.70-1.90 (mn, 1H1), 5 2.26 (s, 6H), 2.33 (min, 1H), 2.57 (min, 1H), 3.06 (min, 1H), 3.85-4.23 (mn, 4H11), 3.88 (s, 3H), 6.93 (inm, 2H), 7.95 (min, 2H). To a stirred solution of methyl 4-(cis-1-tert-butoxycarbonyl-4-dimethylanmino-(2S) pyrrolidinyl) methoxybenzoate (208 mg, 0.55 mmol) in CH 2 C1 2 (6 ml) was added TFA (3ml) at 0 0 C. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was 10 concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2
C
2 . The extract was washed with brine ,dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product, 3-methoxy-4-[N'-(2-methylphenyl)uredio]phenylacetic acid (173 mg, 0.55 mmol), HOBt (74 mg, 0.55 mmol), and triethylamine (15341, 1.1 mmol) in THF (6 ml) and MeCN (6 ml) was 15 added EDC-HCI (160 mg, 0.83 mmol) at 0 0 C. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with EtOAc-CH 2
CI
2 MeOH (5/95, v/v) as eluent to give methyl 4-[cis-4-dimethylamino-1-[3-methoxy-4-[N'-(2 20 methylphenyl) ureido]phenylacetyl]-(2S)-pyrrolidinyl]methoxybenzoate (270 img, 47%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.95-2.04 (min, 1H), 2.25 (s, 6H), 2.32 (s, 3H), 2.61 (mn, 1H), 3.21 (min, 1H1), 3.56-3.58 (min, 5H), 3.80-3.83 (min, 1H11), 3.88 (s, 3H11), 4.18-4.20 (min, 1H1), 4.41-4.45 (inm, 2H), 6.36 (s, 1H), 6.68-6.85 (min, 4H), 7.08-7.25 (min, 4H), 7.52-7.55 (min, 111), 7.91-8.07 (min, 3H11). To a stirred solution of methyl 4-[cis-4-dimethylamino-l1-[3-methoxy-4-[N'-(2 25 methylphenyl) ureido] phenylacetyl]-(2S)-pyrrolidinyl]methoxybenzoate (270 mg, 0.47 mmol) in THF (4.0 ml) and MeOH (2.0 ml) was added IN NaOH (1.0 ml, 1.0 mmol). The mixture was stirred at 70 oC for 18 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HC1. The resulting solid was collected, washed with water, and dried in vacuo to give 157 (170 mg, 65%) as a white crystalline solid. MW 560.64 mp 147-150 oC; IR (KBr) 30 3353, 2952, 1700, 1604, 1533, 1454, 1415, 1255, 1166, 1035, 755 cm'; 'H-NMR (DMSO-d6) 8 :1.83-1.84 (min, 1H1), 2.08-2.10 (min, 1H), 2.21 (br s, 6H), 2.24 (s, 3H), 3.00 (min, 2H11), 3.60 (s, 2H), 3.78 (s, 3H), 3.85-4.29 (min, 4H), 6.71-7.16 (min, 7H11), 7.77-8.01 (min, 4Hm), 8.46 (s, 1H11), 8.54 (s, 1H); 323 WO 01/00206 PCT/US00/18079 MS (FAB) m/z 561 (M+H)+; Anal calcd for C 31
H
36
N
4 0 6 *2 H 2 0: C, 62.40; H, 6.76; N, 9.39. Found: C, 62.51; H, 6.60; N, 9.36. Example 150 methyl 4-[cis-4-dimethylamino-1 -[3-methoxy-4-[N'-(2-methylphenyl)ureidojphenylacetyl]-(2S) 5 pyrrolidinyl]methoxybenzoate HCI salt Me\NMe 0 NJ N NN O- COOH Me H H Me HCI 158 HCI 158 To a stirred solution of 4-[cis-4-dimethylamino-1-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenylacetyl]-(2S)-pyrrolidinyl]methoxybenzoic acid (80 mg, 0.14 mmol) in toluene (4.0 ml) and MeOH (1.0 ml) was added TMSCHN 2 (2.0 M in hexane) (10041, 0.20 mmol) at 0 oC. The 10 reaction mixture was stirred at room temperature for 1.5 hr. The mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with MeOH-CH 2 Cl 2 (5/95, v/v) as eluent. The product was dissolved in EtOH (5.0 ml), and IN HC1 (in EtOH) (2441, 0.244mmol) was added thereto. The mixture was concentrated in vacuo to give 158 (75 mg, 79%) as an amorphous solid. MW 574.67 IR (KBr) 3345, 2950, 2456, 1712, 1646, 1604, 1511, 1454, 15 1434, 1415, 1284, 1257, 1168, 1114, 1031, 771 cmr'; 'H-NMR (DMSO-d) 8 2.10-2.20 (m, 2H), 2.25 (s, 3H), 2.83 (m, 6H), 3.60-3.62 (m, 2H), 3.76-3.81 (m, 8H), 4.20-4.33 (m, 4H), 6.71-7.17 (m, 6H), 7.77-7.98 (m, 5H), 8.47 (s, 1H), 8.55 (s, 1); MS (FAB) m/z 574 (M+H)+; Anal. calcd for
C
32
H
38
N
4 0 6 * 1.0 HCI 1.3 H20: C, 60.57; H, 6.61; N, 8.83. Found: C, 60.80; H, 6.82; N, 8.44. Example 151 20 4-[trans-1-[4-[N-(2-chlorophenyl)ureido]-3-methoxylphenylacetyl]-4-dimethylamino-(2S) pyrrolidinyl]Jmethoxybenzoic acid Me\ Me ,NM N N N/ COOH C HH Me 159 To a stirred solution of methyl 4-(trans-1-tert-butoxycarbonyl-4-dimethylamino-(2S) pyrrolidinyl) methoxybenzoate (430 mg, 1.1 mmol) in CH 2 C1 2 (10.0 ml) was added TFA (5.0 ml) 25 at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 C1 2 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated in vacuo. The crude product 324 WO 01/00206 PCT/US00/18079 was used to the subsequent reaction without further purification. To a stirred solution of the crude product, 4-[AN'-(2-chlorophenyl)uredio]-3-methoxyphenylacetic acid (368 mg, 1.1 mmol), HOBt (162 mg, 1.2 mmol), and triethylamine (417 ml, 3.0 mmol) in THF (10.0 ml) and MeCN (10.0 ml) was added EDC-HC1 (288 mg, 1.1 mmol) at 0 oC. The reaction mixture was stirred at 5 room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (1:1, v/v) as eluent to give 4-[trans-1-[4-[N' (2-chlorophenyl) ureido]-3-methoxylphenylacetyl]-4-dimethylamino-(2S)-pyrrolidinyl]lmethoxy 10 benzoate (530 mg, 78%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.94-1.99 (m, 1H), 2.48 (s, 9H), 3.06-3.12 (m, 1H11), 3.33-3.38 (m, 1H11), 3.60 (s, 2H), 3.68 (s, 3H), 3.69-3.80 (m, 1H11), 3.88 (s, 3H), 4.13-4.20 (m, 2H), 4.56 (m, 1H), 6.76-7.00 (m, 5H), 7.22-7.34 (m, 3H), 7.92-8.00 (m, 3H), 8.17 8.19 (m, 1H). For HCI salt: IR (KBr) 3324, 2950, 2454, 1710, 1604, 1511, 1284 cm'; 'H-NMR (DMSO-d) 62.30-2.40 (m, 2H), 2.77-2.80 (m, 6H), 3.60-3.75 (m, 2H), 3.75-3.85 (m, 8H), 4.00 15 4.22 (m, 3H), 4.50-4.75 (m, 11-1), 6.75-7.43 (m, 7H), 7.89-8.09 (m, 4H11), 8.87 (s, 1H), 8.91 (s, 1H); MS (FAB) m/z 595 (M+H)+; Anal. calcd for C 31
H
3
N
4 0 6 Cl- 1.0HC-1.0H 2 0: C, 57.23; H, 6.04; N, 8.61; Cl, 10.90. Found: C, 57.43; H, 6.08; N, 8.38; Cl, 10.73. To a stirred solution of methyl 4-[trans-1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyl phenylacetyl]-4-dimethylamino-(2S)-pyrrolidinyl]methoxybenzoate (190 mg, 0.32 mmol) in THF 20 (3.0 ml) and MeOH (2.0 ml) was added iN NaOH (0.64 ml, 0.64 mmol). The mixture was stirred at 70 'C for 24 hr. The mixture was concentrate in vacuo, water was added thereto, and neutralized with IN HCL. The resulting solid was collected, washed with water, and dried in vacuo to give 159 (150 mg, 83%) as a white crystalline solid. MW 581.06 mp 159-161 0 C; IR (KBr) 3318, 2938, 1604, 1531, 1438, 1340 cm'; 'H-NMR (DMSO-d 6 ) 5 2.10-2.40 (m, 8H), 2.50-2.70 (m, 25 2H), 3.85-3.90 (m, 5H), 4.02-4.18 (m, 3H), 4.30-4.60 (m, 1H), 6.75-7.43 (m, 7H11), 7.86-8.09 (m, 4H), 8.86 (s, 1H), 8.91 (s, 1H); MS (FAB) m/z 581 (M+H)+; Anal. calcd for C 30
H
33
N
4 0 6 Cl- 1.2H 2 0: C, 59.79; H, 5.92; N, 9.30. Found: C, 59.69; H, 5.93; N, 9.09. Example 152 4-[cis- 1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxylphenylacetyl]-4-dimethylamino-(2S) 30 pyrrolidinyl]methoxybenzoic acid 325 WO 01/00206 PCT/US00/18079 Me\ N Me N N N COH 160 iHH Me 160 To a stirred solution of methyl 4-(cis-1-tert-butoxycarbonyl-4-dimethylamino-(2S) pyrrolidinyl) methoxybenzoate (1.2 g, 3.2 mmol) in CH 2 C1 2 (10.0 ml) was added TFA (5.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was 5 concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2
C'
2 . The extract was washed with brine ,dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product (278 mg, 1.0 mmol), 4-[N'-(2-chlorophenyl)uredio]-3-methoxyphenylacetic acid (335 mg, 1.0 mmol), HOBt (135 mg, 1.0 mmol), and triethylamine (417 ml, 3.0 mmnol) in THF 10 (4.0 ml) and MeCN (4.0 ml) was added EDC-HCI (288 mg, 1.5 mmol) at 0 'C. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc(l:1 ,v/v) as eluent to give 15 methyl 4-[cis- 1l-[4-[N'-(2-chlorophenyl)ureido]-3-methoxylphenylacetyl]-4-dimethylamino-(2S) pyrrolidinyl] methoxybenzoate (500 mg, 84%) as a colorless oil. 'H-NMR (CDCl 3 ) 6 1.98-2.50 (min, 1H), 2.26 (s, 3H), 2.25-2.40 (min, 111), 2.58-2.65 (min, 111), 3.20-3.30 (min, 1H), 3.60 (s, 2H), 3.64 (s, 3H), 3.80-3.90 (min, 1H1), 3.88 (s, 3H), 4.18-4.20 (min, 1H), 4.42-4.46 (min, 2H), 6.72-7.00 (min, 4H), 7.20-7.35 (min, 5H), 7.91-7.94 (min, 3H11), 8.18-8.21 (min, 1H). 20 To a stirred solution of methyl 4-[cis-1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyl phenylacetyl]-4-dimethylamino-(2S)-pyrrolidinyl]methoxybenzoate (250 rg, 0.42 mmol) in THF (5.0 ml) and MeOH (3.0 ml) was added IN NaOH (1.0 ml, 1.0 mmol). The mixture was stirred at 70 oC for 18 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HC1. The resulting solid was collected, washed with water, and dried in vacuo to give 160 25 (170 mg, 70%) as a white crystalline solid. MW 581.06 mp 165-167 'C; IR (KBr) 3328, 1604, 1531, 1164, 1033 cm'; 'H-NMR (DMSO-d) 6 1.80-1.90 (min, 1H), 2.20-2.50 (mn, 7H), 3.60-3.70 (min, 2H), 3.77-3.81 (min, 5H11), 4.00-4.30 (min, 4H11), 6.72-7.44 (m,7H), 7.86-8.10 (min, 4H), 8.88-8.92 (m,2H).; MS (FAB) m/z 581 (M +1); Anal calcd for C 3
H
33
N
4 0 6 Cl-1.1H 2 0: C, 59.87; H, 6.06; N, 9.31. Found: C, 59.65; H, .5.76; N, 9.09. 326 WO 01/00206 PCT/US00/18079 Example 153 4-[cis- 1 -[4-[N'-(2-chlorophenyl)ureido]-3-methoxylphenylacetyl]-4-(2-naphthalenesulfonamido) (2S)-pyrrolidinyl]lmethoxybenzoic acid 0 \ HN- \ / 0 N N O- COOH 16 iHH Me 161 5 To a stirred solution of methyl 4-(cis-1l-tert-butoxycarbonyl-4-amino-(2S)-pyrrolidinyl)methoxy benzoate (200 mg, 0.57 mmol) and TEA (317 ml, 2.3 mmol) in CHCl 3 (10.0 mnil) was added (2 naphthyl)sulfonyl chloride (155 mg, 0.68 mmol) at 0 0 C. The reaction mixture was stirred at room temperature for 18hr. The reaction mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (2:1, v/v) as eluent to give methyl 10 4-(cis-1l-tert-butoxycarbonyl-4-(2-naphthylsulfonamido)-(2S)-pyrrolidinyl) methoxybenzoate (240 mg, 78%/) as a pale yellow oil. 1 H-NMR (CDCl 3 ) 5 1.25-1.45 (br s, 9H11), 1.70-1.80 (m, 1H11), 2.20 2.40 (m, 1H), 3.20-3.50 (m, 2H1), 3.90 (s, 3H1), 3.85-4.15 (m, 3H11), 4.55-4.65 (m, 1H), 6.90-7.10 (m, 2H), 7.58-8.04 (m, 8H), 8.43(s, 1H). To a stirred solution of methyl 4-(cis-1-tert-butoxycarbonyl-4-(2-naphthylsulfonamido) 15 (2S)-pyrrolidinyl) methoxybenzoate (240 mg, 0.44 mmol) in CH 2 Cl 2 (5.0 ml) was added TFA (5.0 ml) at 0 'C. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 Cl 2 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude 20 product, 4-[N'-(2-chlorophenyl)uredio]-3-methoxyphenylacetic acid (147 mg, 0.44 mmol), HOBt (59 mg, 0.44 mmol), and triethylamine (275 ml, 1.9 mmol) in THF (6.0 ml) and MeCN (6.0 ml) was added EDC-HCI (127 mg, 0.66 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then 25 dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (1:3, v/v) as eluent to give methyl 4-[cis-1-[4 [N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-4-(2-naphthalenesulfonamido)-(2S) pyrrolidinyl]methoxybenzoate (200 mg, 65%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.75-1.80 (m, 1H), 2.25-2.40 (m, 1H11), 3.43 (s, 2H11), 3.40-3.50 (m, 1H11), 3.60 (s, 3H), 3.65-3.75 (m, 1H), 3.90 (s, 327 WO 01/00206 PCT/US00/18079 3H11), 3.85-3.92 (m, 1H), 3.95-4.00 (m, 1H), 4.30-4.40 (m, 1H), 4.65-4.75 (m, 1H1), 6.26 (d, J= 9.3 Hz, 1H), 6.50 (d, J= 8.3 Hz, 1H), 6.23 (s, 1H), 6.86 (d, J= 8.8 Hz, 2H), 6.75-7.01 (m, 1H), 7.23 7.36 (m, 3H), 7.61-7.96 (m, 9H), 8.20 (d, J= 8.1 Hz, 1H11), 8.43 (s, 1H). To a stirred solution of methyl methyl 4-[cis-1l-[4-[N'-(2-chlorophenyl)ureido]-3-methoxylphenyl 5 acetyl]-4-(2-naphthalenesulfonamide)-(2S)-pyrrolidinyl]methoxybenzoate (200 mg, 0.26 mmol) in THF (6.0 ml) and MeOH (3.0 ml) was added IN NaOH (0.5 ml, 0.5 mmol). The mixture was stirred at 70 oC for 18 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HC. The resulting solid was collected, washed with water, and dried in vacuo to give 161 (210 mg, quant) as a white crystalline solid. MW 743.22 mp 135-142 'C; IR (KBr) 10 3332, 1685, 1604, 1531, 1421, 1159 cm'; 1 H-NMR (DMSO-d 6 ) 8 1.75-1.85 (m, 1H), 2.05-2.15 (m, 1H), 3.05-3.15 (m, 1H1), 3.47 (s, 2H11), 3.60-3.80 (m, 2H), 3.73 (s, 3H), 4.05-4.20 (m, 3H11), 6.51 (d, J = 8.5 Hz, 1H), 6.74-7.04 (m, 5H), 7.27-7.31 (m, 1H), 7.43-7.45 (m, 2H), 7.66-8.17 (m, 9H), 8.46 (s, 1H1), 8.91 (d, J= 9.5 Hz, 1H); MS (FAB) m/z743 (M+1); Anal. calcd for C 38
H
3 5
N
4 0sCIS 0.5H20: C, 60.67; H, 4.82; N, 7.45; Cl, 4.26. Found: C, 60.77; H, 4.84; N, 7.21; Cl, 4.90. 15 Example 154 4-[ 1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]J-4-(2-mesitylenesulfonamido-(2S) pyrrolidinyl]methoxybenzoic acid N N C O OH- -6CO O H rH HPM 162 To a stirred solution of methyl 4-(cis-1-tert-butoxycarbonyl-4-amino-(2S)-pyrrolidinyl) 20 methoxy benzoate (180 mg, 0.51 mmol) and TEA (283 ml, 2.0 mmol) in CHCl 3 (10.0 ml) was added (2-mesitylene)sulfonyl chloride (122 mg, 0.56 mmol) at 00C. The reaction mixture was stirred at room temperature for 18hr. The reaction mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (3:1, v/v) as eluent to give methyl 4-(cis-1-tert-butoxycarbonyl-4-(2-mesitylenesulfonamido-(2S)-pyrrolidinyl) 25 methoxy benzoate (170 mg, 62%) as a pale yellow oil. 'H-NMR (CDCl 3 ) 85 1.40 (s, 9H11), 1.85-1.95 (m, 1H), 2.29 (s, 3H), 2.35-2.45 (m, 1H1), 2.62 (s, 6H11), 3.80-4.15 (mn, 3H), 3.89 (s, 3H), 3.50-3.65 (m, 1H), 6.94 (s, 2H), 6.94-7.00 (m, 2H), 7.99 (d, J= 8.8 Hz, 2H). To a stirred solution of methyl (cis-1-tert-butoxycarbonyl-4-(2-mesitylenesulfonamido)-(2S) 328 WO 01/00206 PCT/US00/18079 pyrrolidinyl)methoxybenzoate (170 mg, 0.32 mmol) in CH2C1 2 (5.0 ml) was added TFA (5.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 C1 2 . The extract was washed with brine ,dried over Na 2
SO
4 , and concentrated in vacuo. The crude product 5 was used to the subsequent reaction without further purification. To a stirred solution of the crude product, 4-[N'-(2-bromophenyl)uredio]-3-methoxyphenylacetic acid (121 mg, 0.32 mmol), HOBt (43 rmg, 0.32 mmol), and triethylamine (139 ml, 1.0 mmol) in THF (5.0 ml) and MeCN (5.0 ml) was added EDC-HCI (91 mg, 0.48 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted 10 with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc(1:4 ,v/v) as eluent to give methyl 4-[1-[4-[N' (2-bromophenyl)ureido]-3-methoxyphenylacetyl]-4-(2-mesitylenesulfonamido)-(2S)-pyrrolidinyl] methoxybenzoate (210 mg, 83%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.85-1.90 (min, 1H), 1.95 15 2.05 (min, 1H), 2.32 (s, 3H11), 2.60 (s, 6H), 3.40-3.50 (min, 3H11), 3.60-3.70 (min, 2H), 3.68 (s, 3H), 3.89 (s, 3H11), 3.96-3.99 (min, 1H11), 3.35-3.45 (min, 1H), 3.70-3.75 (min, 1H11), 6.00 (d, J= 9.5 Hz, 1H), 6.57 7.08 (min, 9H11), 7.29 -7.34 (min, 1H), 7.51-7.53 (min, 1H), 7.92-7.96 (min, 3H11), 8.15 (d, J= 6.8 Hz, 1H). To a stirred solution of methyl 4-[ 1-[3-methoxy-4-[N'-(2-bromophenyl)ureido]phenylacetyl]-4-(2 mesitylenesulfonamido-(2S)-pyrrolidinyl]methoxybenzoate (210 mg, 0.26 mmol) in THF (5.0 ml) 20 and MeOH (3.0 ml) was added IN NaOH (0.47 ml, 0.47 mmol). The mixture was stirred at 70 oC for 24 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HC1. The resulting solid was collected, washed with water, and dried in vacuo to give 162 (180 mg, 87%) as a white crystalline solid. MW 779.70 mp 130-132 'C; IR (KBr) 3332, 1689, 1604, 1529, 1155 cm'; 'H-NMR (DMSO-d 6 ) 8 1.70-1.85 (min, 1H), 2.02-2.12 (min, 1H), 2.25 (s, 3H), 25 2.52 (s, 6H), 3.05-3.12 (min, 1H), 3.48 (s, 2H11), 3.60-3.70 (min, 2H), 3.77 (s, 3H11), 3.90-4.20 (min, 3H11), 6.59 (d,J = 8.3 Hz, 1H), 6.77-6.80 (min, 1H11), 6.95-7.01 (min, 4H11), 7.31-7.35 (min, 1H), 7.60 (d,J= 8.1 Hz, 1H), 7.86-7.97 (min, 5H11), 8.72-8.76 (min, 1H11), 8.89-8.93 (min, 1H); MS (FAB) m/z 779 (M ), 781 (M'+2); Anal. calcd for C 37
H
39
N
4 OsSBr-0.5H 2 0: C, 56.35; H, 5.11; N, 7.10; Br, 10.13. Found: C, 56.39; H, 5.07; N, 6.89; Br, 10.25. 30 Example 155 4-[ 1-[4-[N-(2-bromophenyl)ureido ]-3-methoxyphenylacetyl]-4-dansylamino-(2S)-pyrrolidinyl] methoxybenzoic acid 329 WO 01/00206 PCT/US00/18079 HNfl ,k N N OO COOH rH H Me 163 To a stirred solution of methyl 4-(cis-1-tert-butoxycarbonyl-4-amino-(2S)-pyrrolidinyl)methoxy benzoate (180 mg, 0.51 mmol) and TEA (283 ml, 2.0 mmol) in CHC1 3 (10.0 ml) was added dansyl chloride (155 mg, 0.68 mmol) at 0 0 C. The reaction mixture was stirred at room temperature for 5 18hr. The reaction mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (2:1, v/v) as eluent to give methyl 4-(cis-1 tert-butoxycarbonyl-4-dansylamino-(2S)-pyrrolidinyl) methoxylbenzoate (200 mg, 73%) as a pale yellow oil. 1 H-NMR (CDCl 3 ) 6 1.34 (s, 9H), 1.50-1.60 (min, 1H), 2.15-2.25 (min, 1H), 2.87 (s, 6H), 3.15-3.22 (min, 1H), 3.35-3.45 (min, 1H), 3.48-3.52 (min, 1H), 3.80-4.10 (min, 3H), 3.91 (s, 3H), 7.01 10 7.25 (min, 4H), 7.50-7.53 (min, 1H), 8.03 (d, J= 8.7 Hz, 2H), 8.16 (d, J= 8.5Hz, 1H), 8.28 (d, J= 7.1 Hz, 1H), 8.53 (d, J= 8.5 Hz, 1H). To a stirred solution of methyl 4-(cis-l-tert-butoxycarbonyl-4-dansylamino-(2S)-pyrrolidinyl) methoxybenzoate (200 mg, 0.34 mmol) in CH 2 C1 2 (5.0 ml) was added TFA (5.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in 15 vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2
C
2 . The extract was washed with brine ,dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product, 4 [N'-(2-bromophenyl)uredio]-3-methoxyphenylacetic acid (129 mg, 0.34 mmol), HOBt (46 mg, 0.34 mmol), and triethylamine (142 ml, 1.0 mmol) in THF (5.0 ml) and MeCN (5.0 ml) was added 20 EDC-HCI (98 mg, 0.51 mnmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc(1:4 ,v/v) as eluent to give methyl 4-[1-[4-[N'-(2-bromophenyl) ureido]-3 25 methoxyphenylacetyl]-4-dansylamino-(2S)-pyrrolidinyl]methoxybenzoate (250 mg, 88%) as a colorless oil. 'H-NMR (CDC 3 ) 8 1.55-1.65 (min, 1H), 2.15-2.25 (min, 1H), 2.87 (s, 6H11), 3.20-3.35 (min, 3H), 3.50-3.55 (min, 1H), 3.67 (s, 3H), 3.78-3.81 (min, 1H), 3.88-3.93 (min, 1H), 3.91 (s, 3H), 4.28 4.31 (min, 1H), 4.65-4.70 (min, 1H), 6.35 (d,J = 9.5 Hz, 1H11), 6.54 (d,J= 8.5 Hz, 1H), 6.63 (s, 1IH), 6.90-7.13 (min, 6H), 7.22-7.31 (min, 2H), 7.50-7.56 (min, 2H), 7.88 (d, J= 8.0 Hz, 1H), 7.99 ( d, J= 330 WO 01/00206 PCT/US00/18079 9.0 Hz, 1H), 8.13-8.16 (m, 2H), 8.27 (d, J= 7.6 Hz, 1H), 8.56 (d, J= 8.3 Hz, 1H1). To a stirred solution of methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-4 dansylamino-2S-pyrrolidinyl]methoxybenzoate (250 mg, 0.29mmol) in THF (5.0 ml) and MeOH (3.0 ml) was added IN NaOH (0.52 ml, 0.52 mmol). The mixture was stirred at 70 oC for 24 hr. 5 The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HCl. The resulting solid was collected, washed with water, and dried in vacuo to give 163 (230 mg, 94%) as a green crystalline solid. MW 830.74 mp 138-141 'C; IR (KBr) 3340, 2940, 1604, 1527, 1421, 1162, 1145 cm-'; 'H-NMR (DMSO-de) 8 1.70-1.80 (m, 1H1), 1.98-2.06 (m, 1H11), 2.81 (s, 6H11), 3.00-3.10 (m, 111), 3.39-3.40 (m, 2H), 3.50-3.80 (m, 3H1), 3.76 (s, 3H), 3.90-4.15 (m, 2H), 10 6.52 (d, J = 9.0 Hz, 1H), 6.73-7.00 (m, 4H), 7.22-7.35 (m, 2H), 7.55-7.64 (m, 3H1), 7.83-8.48 (m, 8H), 8.71-8.76 (m, 1H), 8.88-8.92 (m, 1H1); MS (FAB) m/z 830 (M), 832 (M+2); Anal. calcd for
C
4 oH 4 0 N50sBrS-0.7H 2 0: C,56.97; H,.95; N,8.30;Br,9.47.Found:C,57.06; H, 4.86; N, 7.98; Br, 9.66. Example 156 4-[4-methanesulfonamido-1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-(2S) 15 pyrrolidinyl]lmethoxybenzoic acid 0 \\ Me
HN
0 N N NN O-- COOH 164 r H H Me 164 To a stirred solution of methyl 4-(cis-4-amino-1l-tert-butoxycarbonyl-(2S)-pyrrolidinyl)methoxy benzoate (180 mg, 0.51 mmol) and TEA (283 ml, 2.0 mmol) in CHC13 (10.0 ml) was added methanesulfonyl chloride (88 mng, 0.77 mmol) at 0 0 C. The reaction mixture was stirred at room 20 temperature for 18hr. The reaction mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (1:1, v/v) as eluent to give methyl 4-(cis-1-tert-butoxycarbonyl-4-methanesulfonamido-(2S)-pyrrolidinyl) methoxybenzoate (150 mg, 69%) as a pale yellow oil. 'H-NMR (CDCl 3 ) 8 1.45 (s, 9H), 1.98-2.08 (m, 1H), 2.52-2.65 (m, 1H1), 2.99 (s, 3H), 3.40-3.50 (m, 1H1), 3.55-3.80 (m, 1H1), 3.89 (s, 1H1), 4.00-4.70 (m, 4H1), 6.98-7.00 (m, 25 2H), 8.00 (d, J = 8.8 Hz, 2H1). To a stirred solution of methyl 4-(cis-1-tert-butoxycarbonyl-4-methanesulfonamido-(2S) pyrrolidinyl) methoxybenzoate (150 mg, 0.43 mmol) in CH 2 Cl 2 (5.0 ml) was added TFA (5.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was 331 WO 01/00206 PCT/US00/18079 concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 C1 2 . The extract was washed with brine ,dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product, 4-[N'-(2-bromophenyl)uredio]-3-methoxyphenylacetic acid (163 mg, 0.42 mmol), 5 HOBt (58 mg, 0.43 mmol), and triethylamine (179 ml, 1.3 mmol) in THF (5.0 ml) and MeCN (5.0 ml) was added EDC-HCI (144 mg, 0.75 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column 10 chromatography on silica gel with n-hexane-EtOAc (1:4, v/v) to EtOH-EtOAc (10%, v/v) as eluent to give methyl 4-[4-methanesulfonamido-1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl acetyl]-(2S)-pyrrolidinyl]methoxybenzoate (210 mg, 73%) as a colorless oil. 'H-NMR (CDC13) 8 1.95-2.00 (m, 1H1), 2.55-2.65 (m, 1H), 2.17 (s, 3H), 3.55-3.70 (m, 1H11), 3.60 (s, 2H), 3.67 (s, 3H), 3.85-3.90 (m, 111), 3.89 (s, 3H), 3.95-4.18 (m, 2H), 4.45-4.55 (m, 1H), 4.70-4.80 (m, 1H), 5.87 (d, 15 J= 9.3 Hz, 1H), 6.73-6.95 (m, 5H), 7.09 (s, 2H11), 7.28-7.33 (m, 1H), 7.51 (d, J= 8.0 Hz, 1H), 7.93-7.97 (m, 3H), 8.13 (d,J= 8.3 Hz, 1H1). To a stirred solution of methyl 4-[4-methanesulfonanmido-1-[4-[N'-(2-bromophenyl) ureido]-3-methoxyphenylacetyl]-(2S)-pyrrolidinyl]methoxybenzoate (210 mg, 0.3 mmol) in THF (5.0 ml) and MeOH (3.0 ml) was added IN NaOH (0.8 ml, 0.8 mmol). The mixture was stirred at 20 70 C for 24 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HCL. The resulting solid was collected, washed with water, and dried in vacuo to give 164 (170 mg, 83%) as a white crystalline solid. MW 675.55 mp 125-128 'C; IR (KBr) 3353, 1689, 1604, 1529, 1419, 1155 cm-'; 'H-NMR (DMSO-d 6 ) 8 1.88-2.00 (m, 111), 2.35-2.45 (m, 1H), 2.96 (m, 3H), 3.15-3.23 (m, 1H), 3.60 (s, 2H), 3.50-3.70 (m, 1H11), 3.78 (s, 3H), 3.80-3.90 (m, 1H11), 3.95 25 4.05 (m, 1H1), 4.10-4.30 (m, 2H), 6.71-7.03 (m, 4H1), 7.32 (m, 1H1), 7.45 (d, J= 6.8 Hz, 1H), 7.60 (d, J = 7.8 Hz, 111), 7.87-7.95 (m, 4H), 8.74 (s, 1H), 8.92 (s, 1H); MS (FAB) nm/z 675 (M), 677 (M++2); ;Anal calcd for C 29 H3N 4 OsBrS-0.6H 2 0: C, 50.75; H, 4.73; N, 8.16. Found: C, 51.04; H, 4.62; N, 7.79. Example 157 30 4-f [1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S)-octahydroindolylmethoxy] benzoic acid 332 WO 01/00206 PCT/US00/18079 N N N OC-- COOH me H H Me 165 To a stirred solution of octahydroindole-(2S)-carboxylic acid (3.00 g, 17.7 mmol) in dioxane (20 ml) was added 1 N NaOH (45 ml) and the solution was stirred at 0OC. To the mixture was added (Boc)O 2 0 (4.26 g, 19.5 mmol) in dioxane (25 ml) at 0 0 C and the reaction mixture was stirred at 5 room temperature for 1 day. The mixture was acidified with 1 N HCI and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 and evaporated to give 1-(tert-butoxy carbonyl)octahydroindole-(2S)-carboxylic acid (4.78 g, q.y.) as a colorless solid. mp 130-132 oC; 'H-NMR (CDCl 3 ) 8 1.10-1.46 (series of s and m, total 14 H), 1.65-1.76 (min, 3 H), 1.90-2.18 (min, 2 H), 2.26-2.35 (min, 1 H), 3.77-3.86 (min, 1 H), 4.22-4.34 (min, 1 H); MS (ESI) nm/z 270 (M+1). 10 To a cooled (0OC), stirred solution of 1-(tert-butoxycarbonyl)octahydroindole-(2S) carboxylic acid (1.00 g, 3.71 mmol) in TIF (10 ml) was added BH 3 DMS (530 ml, 5.59 mmol) and the reaction mixture was stirred at room temperature overnight. The mixture was quenched by
H
2 0 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHC13-MeOH 15 (50:1, v/v) as eluent to give 1-(tert-butoxycarbonyl)octahydroindole-(2S)-methanol (940 mg, 99%) as a colorless oil. 'H-NMR (CDCl 3 ) 5 1.05-1.30 (min, 4 H), 1.47 (s, 9 H), 1.49-1.74 (min, 4 H), 1.82 1.93 (min, 3 H), 2.19-2.26 (min, 1 H), 3.56-3.61 (min, 1 H), 3.70-3.75 (min, 2 H), 3.94-3.96 (min, 1 H); MS (FAB) m/z 256 (M++I). To a cooled (0OC), stirred solution of methyl 4-hydroxybenzoate (560 mg, 3.68 mmol), 1 20 (tert-butoxycarbonyl)octahydroindole-(2S)-methanol (940 mg, 3.68 mmol) and Ph 3 P (1.16 g, 4.42 mmol) in THF (20 ml) was added DIAD (870 ml, 4.42 mmol) and the reaction mixture was heated under reflux for 8 hr. After cooled to room temperature, the mixture was evaporated. The residue was purified by column chromatography on silica-gel with n-hexane-EtOAc (5:1, v/v) as eluent to give methyl 4-[1-(tert-butoxycarbonyl)-(2S)-octahydroindolylmethoxy]benzoate (1.16 g, 81%) as a 25 colorless oil. 'H-NMR (CDCl 3 ) 8 1.14-1.47 (series of s and m, total 13 H), 1.60-2.13 (series of m, total 6 H), 2.22-2.28 (min, 1 H), 3.75-3.91 (series of s and m, total 4 H), 4.06-4.18 (min, 2 H), 4.37 (min, 1 H), 6.94-6.96 (min, 2 H), 7.96-7.98 (min, 2 H); MS (FAB) m/z 390 (M++1). To a stirred solution of methyl 4-[1-(tert-butoxycarbonyl)-(2S)-octahydroindolylmethoxy] 333 WO 01/00206 PCT/US00/18079 benzoate (1.16 g, 2.98 mmol) in CH 2 C1 2 (10 ml) was added TFA (10 ml) and the reaction mixture was stirred at room temperature for 2 hr. The mixture was concentrated in vacuo, made basic by sat. NaHCO 3 , and extracted with CHCl 3 . The extract was washed with brine, dried over K 2
CO
3 , and evaporated to give methyl 4-[(2S)-octahydroindolylmethoxy]benzoate (860 mg, q.y.) as a 5 brown oil. 'H-NMR (CDCl 3 ) 8 1.23-1.78 (series of m, total 10 H), 2.00-2.09 (m, 2 H), 3.14-3.18 (m, 1 H), 3.55-3.62 (m, 1 H), 3.88 (s, 3 H), 3.96-4.06 (m, 2 H), 6.92 (d,J= 9.1 Hz, 2 H), 7.97 (d,J = 9.1 Hz, 2 H); MS (FAB) m/z 290 (M*+1). A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (298 mg, 0.95 mmol), methyl 4-[(2S)-octahydroindolylmethoxy]benzoate (274 mg, 0.95 mmol), EDCHCI (218 10 mg, 1.14 mmol), HOBt (154 mg, 1.14 mmol), Et 3 N (160 ml, 1.15 mmol) in THF (7 ml) was stirred at room temperature overnight. The mixture was diluted with H 2 0 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1 to 50:1, v/v) as eluent to give methyl 4-[ 1 -[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S)-octahydroindolyl 15 methoxy]benzoate (532 mg, 96%) as a white foam. 'H-NMR (CDCl 3 ) 8 1.13-2.05 (series of m, total 9 H), 2.14-2.24 (m, 2 H), 2.26 (s, 3 H), 3.60 (s, 2 H), 3.62 (s, 3 H), 3.80-3.85 (m, 1 H), 3.88 (s, 3 H), 4.27-4.37 (m, 3 H), 6.62 (s, 1 H), 6.74-6.76 (m, 2 H), 6.91 (d, J= 8.8 Hz, 2 H), 7.09-7.13 (m, 1 H), 7.20-7.24 (m, 3 H), 7.55 (d, J = 7.8 Hz, 1 H), 7.94 (d, J= 8.8 Hz, 2 H), 8.04 (d, J= 7.8 Hz, 1 H); MS (FAB) nm/z 586 (M+1). 20 To a stirred solution of methyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl acetyl]-(2S)-octahydroindolylmethoxy]benzoate (532 mg, 0.91 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 3 hr. After cooled to room temperature, the mixture was poured into ice-1 N HC1, and the resulting precipitate was collected. The crude solid was recrystallized from MeOH-CHCl 3 -IPE to give 165 (278 mg, 54%) as a white 25 crystalline powder. MW 571.66 mp 130-134oC; 'H-NMR (DMSO-d) 8 1.16-2.10 (series of m, total 9 H), 2.15-2.30 (series of s and m, total 4 H), 3.55-3.79 (m, 3 H), 3.81 (s, 3 H), 3.90-3.95 (m, 1 H), 4.17-4.23 (m, 2 H), 4.34-4.36 (m, 1 H), 6.72-6.74 (m, 1 H), 6.87-6.88 (m, 1 H), 6.91-6.95 (m, 1 H), 7.03 (d, J = 8.8 Hz, 2 H), 7.10-7.16 (m, 2 H), 7.78-7.80 (m, 1 H), 7.87 (d, J= 8.8 Hz, 2 H), 7.98-8.00 (m, 1 H), 8.45 (s, 1 H), 8.54 (s, 1 H), 12.61 (br s, 1 H); MS (FAB) m/z 572 (M+'+I); 30 Anal. Calcd for C 33
H
37
N
3 0 6 'l/4H 2 0: C, 68.79; H, 6.56; N, 7.29. Found: C, 68.70; H, 6.82; N, 6.97. Example 158 4-[ 1-[4-[N '-(2-chlorophenyl)ureidol-3-methoxyphenylacetyl]-(2S)-octahydroindolylmethoxy] 334 WO 01/00206 PCT/US00/18079 benzoic acid NN N COOH I H H Me 166 A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (307 mg, 0.92 mmol), methyl 4-[(2S)-octahydroindolylmethoxy]benzoate (265 mg, 0.92 mmol), EDCHCI (211 mg, 1.10 5 mmol), HOBt (148 mg, 1.10 mmol), and Et 3 N (153 ml, 1.10 mmol) in THF (7 ml) was stirred at room temperature overnight. The mixture was diluted with H 2 0 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1 to 50:1, v/v) as eluent to give methyl 4-[ 1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(2S)-octahydroindolyl 10 methoxy]benzoate (550 mg, 99%) as a white foam. 'H-NMR (CDCI 3 ) 8 1.15-2.02 (series of m, total 9 H), 2.17-2.33 (min, 2 H), 3.58 (s, 3 H), 3.62 (s, 2 H), 3.84-3.90 (series of s and m, total 4 H), 4.06-4.40 (min, 3 H), 6.71-6.74 (min, 2 H), 6.88-7.00 (min, 3 H), 7.21-7.30 (min, 2 H), 7.62 (s, 2 H), 7.91 7.95 (min, 3 H), 8.17-8.20 (min, 1 H); MS (FAB) m/z 606 (M++1). To a stirred solution of methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(2S) 15 octahydroindolylmethoxy]benzoate (550 mg, 0.91 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 5 hr. After cooled to room temperature, the mixture was poured into ice-1 N HC1, and the resulting precipitate was collected. The crude solid was recrystallized from MeOH-CHCl 3 -IPE to give 166 (286 mg, 53%) as a white crystalline powder. MW 388.29 mp 133-136 0 C; 'H-NMR (DMSO-d 6 ) 8 1.16-2.10 (series of m, total 10 H), 20 2.24-2.27 (min, 1 H), 3.55-3.75 (min, 2 H), 3.80 (s, 3 H), 3.90-3.96 (min, 1 H), 4.17-4.23 (mn, 2 H), 4.34 4.36 (min, 1 H), 6.73-6.75 (min, 1 H), 6.88 (d, J= 1.5 Hz, 1 H), 6.99-7.05 (min, 3 H), 7.25-7.30 (min, 1 H), 7.43 (dd, J= 1.5, 8.1 Hz, 1 H), 7.87-7.89 (min, 2H), 7.95 (d,J= 8.1 Hz, 1 H), 8.08 (dd, J= 1.5, 8.3 Hz, 1 H), 8.88 (s, 1 H), 8.92 (s, 1 H), 12.61 (br s, 1 H); MS (FAB) m/z 592 (M++l); Anal. Calcd for C 32
H
34
CN
3 0 6 1/4H 2 0: C,64.42; H, 5.83; N, 7.04; Cl, 5.94. Found: C,64.55; H,6.09; 25 N,6.64; C1,5.93. Example 159 4-[ 1 -[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-(2S)-octahydroindolylmethoxy] benzoic acid 335 WO 01/00206 PCT/US00/18079 SN N O-- COOH r H H Me 167 A mixture of 4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (457 mg, 1.21 mmol), methyl 4-[(2S)-octahydroindolylmethoxy]benzoate (320 mg, 1.21 mmol), EDCHCI (277 mg, 1.44 mmol), HOBt (196 mg, 1.45 mmol), and Et 3 N (200 ml, 1.43 mmol) in THF (7 ml) was 5 stirred at room temperature overnight. The mixture was diluted with H 2 0 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1, v/v) as eluent to give methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl ]-(2S)-octahydroindolyl methoxy]benzoate (423 mg, 54%) as a white foam. 'H-NMR (CDCl 3 ) 8 1.15-1.89 (series of m, 10 total 8 H), 1.96-2.02 (min, 1 H), 2.16-2.32 (m, 2 H), 3.63 (s, 2 H), 3.65 (s, 3 H), 3.82-3.86 (min, 1 H), 3.88 (s, 3 H), 4.30-4.39 (min, 3 H), 6.75-6.77 (min, 2 H), 6.88-6.93 (mn, 3 H), 7.24-7.31 (mn, 1 H), 7.37 7.50 (min, 3 H), 7.91-7.99 (min, 3 H), 8.12-8.15 (min, 1 H); MS (FAB)m/z 650 (M*+1). To a stirred solution of methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl acetyl]-(2S)-octahydroindolylmethoxy]benzoate (420 mg, 0.65 mmol) in THF (5 ml) was added 0.5 15 N NaOH (5 ml) and the reaction mixture was heated under reflux for 5 hr. After cooled to room temperature, the mixture was poured into ice-I N HCI, and the resulting precipitate was collected. The crude solid was recrystallized from MeOH-CHCl 3 -IPE to give 167 (197 mg, 48%) as a white crystalline powder. MW 636.53 mp 118-123oC; 'H-NMR (DMSO-d 6 ) 5 1.16-2.27 (series of inm, total 10 H), 3.56-3.75 (min, 3 H), 3.81 (s, 3 H), 3.90-3.96 (min, 1 H), 4.17-4.23 (min, 2 H), 4.34-4.36 20 (min, 1 H), 6.73-6.75 (min, 1 H), 6.88-7.05 (min, 4 H), 7.30-7.34 (min, 1 H), 7.59-7.61 (mn, 1 H), 7.87 7.96 (m, 4 H), 8.73 (s, 1 H), 8.91 (s, 1 H), 12.64 (br s, 1 H); MS (FAB) m/z 636 (M++1); Anal. Calcd for C 32
H
34 BrN 3 0 6 1/4H 2 0: C,59.96;H,5.42;N,6.55; Br,12.46. Found: C,60.12; H,5.86; N,6.09; Br,12.47. Example 160 25 4-[3-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl]acetyl-4-thiazolidinyl]methoxybenzoic acid N N N 0COOH Me H H Me 168 To a stirred solution of thiazolidine-4-carboxylic acid (5.0 g, 37.6 mmol) in DMF (50.0 ml) was added (Boc) 2 0 (9.8 g, 45.1 mmol) and TEA (8.0 ml). The reaction mixture was stirred at room 336 WO 01/00206 PCT/US00/18079 temperature for 18 hr. Water was added to the mixture and extracted with EtOAc. The organic layer was washed with water, then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with EtOAc-n-hexane (1:3 ,v/v) as eluent to give 3-tert-butoxycarbonylthiazolidine-4-carboxylic acid (6.5 g, 74%) as a white crystalline solid. 5 'H-NMR (CDCl 3 ) 8 1.49 (br s, 9H), 3.20-3.30 (min, 2H), 4.09-4.87 (min, 3H). To a stirred solution of 3-tert-butoxycarbonylthiazolidine-4-carboxylic acid (2.3 g, 10.0 mmol) in THF (30 ml) was added BH3-THF(1.0 M solution in THF, 20.0 ml, 20.0 mmol) at 0oC. After stirred at room temperature for 1.0 h, the reaction mixture was heated under reflux for 1.0 hr. After cooled, the mixture was concentrated in vacuo. Water was added thereto at 0oC, and 10 extracted with EtOAc. The extract was washed with water, then dried over Na 2
SO
4 , and concentrated in vacuo to give 3-tert-butoxycarbonyl-5-hydroxymethylthiazolidine (2.0 g, quant) as a a colorless oil. 'H-NMR (CDCl 3 ) 5 1.48 (s, 9H), 2.80-2.85 (mn, 1H), 3.13-3.17 (min, 1H1), 3.20-3.30 (min, 1H), 3.64-3.70 (min, 2H), 4.34 (br s, 1H1), 4.60 (br s, 1H1). To a stirred solution of 3-tert-butoxycarbonyl-5-hydroxymethylthiazolidine (1.9 g, 8.7 mmol), 15 methyl 4-hydroxybenzoate (1.3 g, 8.7 mmol), and Ph 3 P (3.2 g, 12.2 mmol) in THF (10 ml) was added DIAD (2.2 g, 10.4 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 18 hr. The mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with EtOAc- n-hexane (1:9, v/v) as eluent to give methyl 4-(3-tert butoxycarbonyl-4-thiazolidinyl)methoxybenzoate (1.6 g, 52% ) as a pale yellow oil. 'H-NMR 20 (CDCl 3 ) 8 1.49 (s, 9H), 3.11-3.19 (min, 2H), 3.88 (s, 3H), 4.04-4.31 (mn, 3H), 4.61 (mn, 2H), 6.96 (d, J = 8.8 Hz, 2H), 7.98 (d, J= 8.8 Hz, 2H). To a stirred solution of methyl 4-(3-tert-butoxycarbonyl-4-thiazolidinyl)methoxybenzoate (440 mg, 1.25 mmol) in CH 2 C1 2 (6 ml) was added TFA (3ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to 25 the residue, and extracted with CH 2 C1 2 . The extract was washed with brine ,dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product (0.6 mmol), 3-methoxy-4-[N'-(2-methyl phenyl)uredio]phenylacetic acid (188 mg, 0.6 mmol), HOBt (81 mg, 0.6 mmol), and triethylamine (280 ml, 2.0 mmol) in THF (5 ml) and MeCN (5 ml) was added EDC-HCI (173 mg, 0.9 mmol) at 30 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. 337 WO 01/00206 PCT/US00/18079 NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (1:3, v/v) as eluent to give methyl 4-[3-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenyl]acetyl-4 thiazolidinyl] methoxybenzoate (340 mg, quant) as an amorphous solid. 'H-NMR (CDCl 3 ) 8 2.31 5 (s, 3H), 3.15-3.16 (m, 2H), 3.67-3.69 (m, 5H), 3.88 (s, 3H), 4.09-4.14 (m, 2H11), 4.22-4.90 (m, 3H), 6.30 (m, 1H), 6.74-6.96 (m, 4H), 7.11-7.25 (m, 4H), 7.49-7.51 (m, 1H), 7.95-8.12 (m, 3H). To a stirred solution of methyl 4-[3-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl]acetyl-4 thiazolidinyl] methoxybenzoate (340 mg, 0.62 mmol) in THF (5.0 ml) and EtOH (3.0 ml) was added IN NaOH (0.62 ml, 0.62 mmol). The mixture was stirred at 70 oC for 18 hr. The mixture 10 was concentrated in vacuo, water was added thereto, and neutralized with IN HC1. The resulting solid was collected, washed with water, and dried in vacuo to give 168 (290 mg, 88%) as a white crystalline solid. MW 535.62 mp 125-128 'C; IR (KBr) 3357, 2937, 1604, 1533, 1419, 1253, 1166, 1033, 773 cm'; 'H-NMR (DMSO-d 6 ) 8 2.25 (s, 3H), 3.05-3.20 (m, 2H), 3.71, 3.83, and 3.85 (each s, total 5H1), 4.03-4.15 (m, 3H1), 4.52-4.76 (m, 2H1), 6.15-6.17 (m, 7H), 7.78-8.30 (m, 4H), 15 8.30 (m, 1H1), 8.56 (m, 1H); MS (FAB) m/z 536 (M+1); Anal. calcd for C 28
H
29
N
3 06S-0.5H 2 0: C, 61.75; H, 5.55; N, 7.72. Found: C, 61.72; H, 5.55; N, 7.49. Example 161 4-[3-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-4-thiazolidinyl]methoxybenzoic acid N N -Qj COOH MeH Me 169 20 To a stirred solution of methyl 4-(3-tert-butoxycarbonyl-4-thiazolidinyl)methoxybenzoate (600 mg, 1.7 mmol) in CH 2 C1 2 (6,0 ml) was added TFA (6.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 Cl 2 . The extract was washed with brine ,dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further 25 purification. To a stirred solution of the crude product, 4-[N'-(2-chlorophenyl)uredio]-3 methoxyphenylacetic acid (570 mg, 1.7 mmol), HOBt (230 mg, 1.7 mmol), and triethylamine (709 ml, 5.1 mmol) in THF (10.0 ml) and MeCN (10.0 ml) was added EDC-HCI (490 mg, 2.55 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with 30 sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc 338 WO 01/00206 PCT/US00/18079 (1:1, v/v) as eluent to give methyl 4-[3-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-4 thiazolidinyl]methoxybenzoate (900 mg, 93%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 3.15-3.18 (min, 2H), 3.70 (s, 2H), 3.78 (s, 3H), 3.86 (s, 3H), 4.09-4.93 (min, 5H), 6.80-7.01 (min, SH), 7.19-7.35 (min, 4H), 7.94-8.18 (min, 4H11). 5 To a stirred solution of methyl 4-[3-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-4 thiazolidinyl]methoxybenzoate (900 mg, 1.6 mmol) in THF (8.0 ml) and MeOH (4.0 ml) was added IN NaOH (3.1 ml, 3.1 mmol). The mixture was stirred at 70 oC for 24 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HCL. The resulting solid was collected, washed with water, and dried in vacuo to give 169 (780 mng, 89%) as a white 10 crystalline solid. MW 556.03 mp 126-129 'C; IR (KBr) 3343, 2937, 1604, 1531, 1421, 1245, 1166, 1035, 752 cmn'; 'H-NMR (DMSO-d) 8 3.06-3.24 (min, 2H), 3.72-3.85 (min, 5H), 4.02-4.27 (inm, 3H), 4.53-4.76 (min, 2H), 6.74-7.44 (min, 7H), 7.87-8.30 (min, 4H), 8.89-8.95 (mn, 2H); MS (FAB) m/z 556 (M*+1); Anal. calcd for C 72 7
N
3 0 6 CIS-0.7H 2 0: C, 56.93; H, 5.03; N, 7.38; Cl, 6.22. Found: C, 56.89; H, 4.84; N, 7.42; Cl, 6.35. 15 Example 162 4-[3-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-4-thiazolidinyl]methoxybenzoic acid -S N N 'rCOOH rH H Me 170 To a stirred solution of methyl 4-(3-tert-butoxycarbonyl-4-thiazolidinyl)methoxybenzoate (560 mg, 1.6 mmol) in CH 2
C
2 (5.0 ml) was added TFA (5.0 ml) at 0 oC. The reaction mixture was stirred 20 at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 C1 2 . The extract was washed with brine ,dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product, 4-[N'-(2-bromophenyl)uredio]-3-methoxy phenylacetic acid (599 mg, 1.6 mmol), HOBt (213 mg, 1.6 mmol), and triethylamine (659 ml, 4.7 25 mmol) in THF (10.0 ml) and MeCN (10.0 ml) was added EDC-HC1 (455 mg, 2.4 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2 SO4, and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (2:3, v/v) as eluent to 30 give methyl 4-[3-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-4-thiazolidinyl] methoxybenzoate (870 mg, 89%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 3.00-3.20 (mn, 3H), 3.70 (s, 339 WO 01/00206 PCT/US00/18079 2H11), 3.81 (s, 3H), 3.88 (s, 3H11), 4.09-4.23 (m, 1H), 4.42 (d,J = 8.5 Hz, 1H), 4.59 (d,J= 8.5 Hz, 1H1), 4.70-4.92 (m, 1H1), 6.81-7.53 (m, 9H), 7,95-8.15 (m, 4H11). To a stirred solution of methyl 4-[3-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-4 thiazolidinyl]methoxybenzoate (870 mg, 1.4 mmol) in THF (8.0 ml) and MeOH (8.0 ml) was 5 added IN NaOH (2.8 ml, 2.8 mmol). The mixture was stirred at 70 'C for 18 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HCL. The resulting solid was collected, washed with water, and dried in vacuo to give 170 (740 mg, 87%) as a white crystalline solid. MW 600.48 mp 125-133 oC; IR (KBr) 3332, 2935, 1604, 1527, 1421, 1245, 1166, 1027, 750 cm'; 'H-NMR (DMSO-d) 8 3.01-3.25 (m, 2H), 3.72-3.85 (m, 5H), 4.02-4.30 (m, 10 2H), 4.54 (d, J= 8.8 Hz, 1H), 4.74-4.87 (m, 2H), 6.76-7.07 (m, 5H), 7.30-7.34 (m, 1H1), 7.59 (d, J = 8.1 Hz, 1H1), 7.86-7.98 (m, 4H11), 8.74 (s, 1H1), 8.92-8.94 (s, 1H); MS (FAB) m/z 600 (M+1); Anal.calcd for C 27
H
2 6 N30 6 BrS-0.3H1 2 0: C, 53.52; H,4.43 N 6.94 Found: C, 53.54; H, 4.45; N, 6.80. Example 163 cis-4-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl]lmethylamino] 15 cyclohexanecarboxylic acid ?LY 0 NI( \COOH MeH HNMe C 171 To a stirred solution of 2S-pyrrolidinemethanol (2.0 g, 20.0 mmol), 3-methoxy-4-[N'-(2-methyl phenyl)uredio]phenylacetic acid (6.28 g, 20.0 mmol), HOBt (71 mg, 0.53 mmol), and triethylamine (5.5 ml, 40.0 mmol) in THF (50.0 ml) and MeCN (40.0 ml) was added EDC-HCI 20 (5.7 g, 30.0 mmol) at 0 'C. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The organic layer was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with EtOAc to MeOH- CH 2 C1 2 (1:9, v/v) as eluent to give 1-[3-methoxy-4-[N'-(2-methylphenyl) 25 ureido]phenylacetyl]-2S-pyrrolidinemethanol (7.0 g, 89%) as a white crystalline solid. 'H-NMR (CDCl 3 ) 8 1.54-1.58 (m, 111), 1.80-2.04 (m, 3H), 2.27 (s, 3H), 3.42-3.46 (m, 1H1), 3.54-3.65 (m, 2H), 3.62 (s, 2H11), 3.69 (s, 3H), 4.21-4.23 (m, 1H11), 5.04 (m, 1H1), 6.68-6.79 (m, 3H), 7.09-7.31 (m, 4H11), 7.52 (d, J = 7.8 Hz, 1H11), 8.07 (d, J= 8.0 Hz, 111). To a stirred solution of oxalyl chloride (0.3 ml, 3.3 mmol) in CH 2 C1 2 (30.0 ml) was added DMSO 30 (6.6 ml, 0.51 mmol) at -78 oC. After 5 minutes, to the mixture was added 1-[3-methoxy-4-[N'-(2 340 WO 01/00206 PCT/US00/18079 methylphenyl)ureido]phenylacetyl]-2-pyrrolidinemethanol (1.2 g, 3.0 mmol) in CH 2 C1 2 (5.0 ml). The mixture was stirred for 30 minutes at -78 oC, and triethylamine (2.1 ml, 15.0 mmol) was added. The mixture was stirred for 30 minutes at -78 oC, and stirred for 30 minutes at room temperature. Water was added to the mixture, and extracted with CH 2 Cl 2 . The extract was washed 5 with water, then dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product, benzyl cis-4-aminocyclohexanecarboxylate (769 mg, 3.3 mmol), and AcOH (0.32 ml) in DCE (10 ml) was added NaBH(OAc) 3 (1.1 g, 5.4 mmol) at 0 'C. The reaction mixture was stirred at room temperature for 18 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the 10 residue, and extracted with CH 2
C
2 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with MeOH-CH 2 C1 2 (1:9, v/v) as eluent to give benzylcis-4-[[1-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenylacetyl]-2-pyrrolidinyl]methylamino]cyclohexanecarboxylate (1.5 g, 83% ) as an amorphous solid. 'H-NMR (CDCl 3 ) 8 1.40-1.65 (min, 6H11), 1.80-1.98 (min, 6H11), 2.26 (s, 3H), 2.45 15 2.65 (min, 3H), 2.81-2.86 (min, 1H1), 3.44-3.46 (min, 2H), 3.56 (s, 2H), 3.67 (s, 3H11), 3.90-4.15 (min, 1H), 5.09 and 5.11 (each s, total 2H), 6.74-6.83 (min, 3H), 7.07-7.20 (min, 4H), 7.31-7.35 (min, 5H), 7.53 7.55 (min, 1H), 8.02-8.06 (min, 1H). To a stirred solution of benzylcis-4-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl acetyl]-2-pyrrolidinyl]methylamino]cyclohexanecarboxylate (1.5 g, 2.45 mmol) in THF (10.0 ml) 20 and MeOH (5.0 ml) was added IN NaOH (3.68 ml, 3.68 mmol). The mixture was stirred at 70 'C for 18 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HC1. The mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with MeOH-CH 2 C1 2 (1:5, v/v) as eluent to give cis-4-[[1-[3-methoxy 4-[N'-(2-methyl phenyl)ureido]phenylacetyl]-2-pyrrolidinyl]methylamino]cyclohexanecarboxylic 25 acid 171 (940 mg, 73%) as an amorphous solid. MW 522.64 IR (KBr) 3283, 2945, 2860, 1534, 1453, 1415 cm-'; 'H-NMR (DMSO-d) 8 1.38-2.00 (min, 12H), 2.45 (s, 3H11), 2.30-3.95 (min, 4H), 3.22-3.75 (min, 2H11), 3.58 (s, 2H), 3.86 (s, 3H11), 4.12 (min, 1H), 6.73-7.16 (min, 5H), 7.77-7.79 (min, 1H), 7.98-8.02 (min, 1H), 8.51-8.52 (min, 1H1), 8.57-8.59 (min, 1H); MS (FAB) m/z 523 (M+1); Anal. calcd for C, 29
H
3
,N
4 0 5 -0.5NaCl-2.2H 2 0: C, 58.89; H, 7.23; N, 9.47. Found: C, 59.21; H, 7.11; N, 9.11. 30 Example 164 methyl cis-4-[[1-[3-methoxy-4-[N '-(2-methylphenyl)ureidoJ]phenylacetyl]-2-pyrrolidinyl]methyl amino]cyclohexanecarboxylate HC1 salt 341 WO 01/00206 PCT/US00/18079 e H"N H- COOMe 172 Me H H- Me 0172 SOC1 2 was added to MeOH at 0 0 C. After stirred for 5 minutes, cis-4-[[1-[3-methoxy-4-[N'-(2 methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyI]methylamino]cyclohexanecarboxylic acid (200 mg, 0.38 mmol) was added. The mixture was stirred at room temperature for 5 hr. The mixture 5 was concentrated in vacuo. Aq. NaHCO 3 was added to the residue, and extracted with CH 2 C1 2 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with MeOH-CH 2
CI
2 (5:95 to 18:92, v/v) as eluent. The product was dissolved in EtOH (5.0 ml), and IN HCI (in EtOH) (1.0 ml, 1.0 mmol) was added thereto. The mixture was concentrated in vacuo to give 172 (160 mg, 74%) as an 10 amorphous solid. MW 536.66 IR (KBr) 3247, 2950, 2875, 1731, 1671, 1612, 1533, 1454, 1205 cm''; 'H-NMR (DMSO-d) 5 1.45-2.10 (min, 12H), 2.25 (s, 3H), 2.60-2.70 (min, 1H1), 2.90-3.20 (inm, 3H), 2.50-2.55 (min, 2H), 3.63 (min, 5H), 3.86 (s, 3H11), 4.15-4.30 (min, 111), 6.74-7.16 (min, 5H), 7.76 7.78 (min, 1H), 8.00-8.09 (min, 1H1), 8.54-8.70 (min, 2H); MS (FAB) m/z 537 (MW+1); Anal. called for
C
3
H
40 N,0-1.0HCl1.0H 2 0: C, 60.95; H, 7.33; N, 9.48; Cl, 6.00. Found: C, 60.87; H, 7.47; N, 15 8.97: Cl, 5.90. Example 165 4-[N-[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrffolidinyl]methyl]-N methylamino]cyclohexanecarboxylic acid N N COOH NN % Me H Me C Me 173 20 To a stirred solution of methyl cis-4-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] 2-pyrrolidinyl]methylamino]cyclohexanecarboxylate (300 mg, 0.55 mmol), HCHO (300 ml), and AcOH (66 mg, 1.1 mmol) in MeOH (10.0 ml) was added NaBH 3 CN (70 mg, 1.1 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 18 hr. After concentrated in vacuo, water was added and extracted with CH 2
C
2 . The extract was washed with water, then dried over 25 Na 2
SO
4 , and concentrated in vacuo. The residue was purified by TLC with MeOH-CH 2 C1 2 (3:97, v/v) as eluent to give methyl 4-[N-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2 pyrrolidinylmethyl]-N-methylamino]cyclohexanecarboxylate (160 mg, 52%) as an amorphous solid. 'H-NMR (CDCl 3 ) 6 1.30-2.20 (min, 12H), 2.27-2.37 (min, 6H), 2.50-2.60 (min, 1H1), 3.30-3.80 (min, 5H), 3.55 (s, 2H), 3.66-3.73 (min, 6H), 4.10-4.20 (min, 1H), 6.60-7.55 (min, 7H11), 8.03 (min, 1H), 8.15 30 (min, 1H1). 342 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[N-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2 pyrrolidinylmethyl]-N-methylamino]cyclohexanecarboxylate (100 mg, 0.18 mmol) in THF (5.0 ml) and MeOH (2.5 ml) was added IN NaOH (0.36 ml, 0.36 mmol). The mixture was stirred at 60 'C for 18 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with 5 IN HC1. The mixture was concentrated in vacuo. The residue was purified by TLC with MeOH
CH
2 C1 2 (1/4, v/v) as eluent to give 173 (10 mg, 10%) as an amorphous solid. MW 536.66 IR (KBr) 3440, 2954, 1697, 1533, 1454 cm-'; 'H-NMR (DMSO-d 6 ) 8 1,20-2.30 (min, 13H), 2.24 (s, 3H), 2.35-4.00 (min, 13H), 6.50-8.10 (min, 8H), 8.50 (min, 1H); MS (FAB) m/z 537 (M++1); Anal. calcd for C 30
H
4 0
N
4 0-2.0NaCl-0.8H 2 0: C, 53.94; H, 6.28; N, 8.39. Found: C, 54.08; H, 6.52; N, 8.04. 10 Example 166 4-[[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S)-pyrrolidinyl]methoxy] cyclohexanecarboxylic acid e N LN N o- COOH 174 Me H H' Me C174 A mixture of methyl 4-(1-tert-butoxycarbonyl-(2S)-pyrrolidinyl)methoxybenzoate (1.0 g, 2.9 15 mmol) and 5% Rh on alumina (500 mg) in EtOH (10.0 ml) and AcOH (1.0 ml) was hydrogenated at room temperature at 5 atm for 36 hr. The catalyst was filtered off, and the filtrate was concentrated in vacuo. The residue was purified by column chromatography on silica gel with n hexane-EtOAc (6:1, v/v) as eluent to give methyl cis-4-[(1-tert-butoxycarbonyl-(2S)-pyrrolidinyl) methoxy]cyclohexanecarboxylate (900 mg, 89%) as a pale yellow oil. 'H-NMR (CDCl 3 ) 8 1.46 (s, 20 9H), 1.46-2.00 (m, 12H), 2.34 (min, 1H), 3.20-3.55 (min, 5H), 3.67 (s, 3H), 3.84-3.92 (min, 1H). To a stirred solution of methyl cis-4-[(1-tert-butoxycarbonyl-(2S)-pyrrolidinyl)methoxy] cyclohexanecarboxylate (900 mg, 2.6 mmol) in CH2C1 2 (5.0 ml) was added TFA (5.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 Cl 2 . The extract was 25 washed with brine, dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product (200 mg, 0.83 mmol), 3-methoxy-4-[N'-(2-methylphenyl)uredio]phenylacetic acid (260 mg, 0.83 nunmmol), HOBt (135 mg, 1.0 mmol), and triethylamine (3444 1, 1.35-2.10 (min, 12H), 2.15-2.38 (inm, 1H), 2.29 (min, 3H), 3.20-3.55 (min, 5H), 3.58 (s, 2H), 3.66 (s, 3H), 3.73 (s, 3H), 4.20-4.25 (mn, 1H1), 30 6.26-6.30 (min, 1H), 6.78-6.81 (min, 2H), 7.06-7,23 (min, 3H), 7.51-7.52 (min, 1H1), 8.01-8.03 (mn, 1H1). 343 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-(2S)-pyrrolidinyl]methoxy]cyclohexanecarboxylate (460 mg, 0.86 mmol) in THF (10.0 ml) and EtOH (5.0 ml) was added iN NaOH (1.4 ml, 1.4 mmol). The mixture was stirred at 60 oC for 18 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized 5 with IN HCl. The resulting solid was collected, washed with water, and dried in vacuo to give 174 (370 mg, 83%) as a white crystalline solid. MW 523.63 mp 110-113 oC; IR (KBr) 3345, 2937, 1612, 1533, 1454 cm'; 'H-NMR (DMSO-d 6 ) 8 1.00-2.00 (m, 12H), 2.24 (s, 3H), 2.20-2.30 (m, 1H), 3.20-3.80 (m, 5H), 3.55 (s, 2H), 3.85 (s, 3H), 4.00-4.18 (m, 1H), 6.71-7.16 (m, 5H), 7.78 (d, J = 8.0 Hz, 1H), 7.90 (d, J= 8.3 Hz, 1H), 8.45 (s, 1H), 8.54 (s,1 H); MS (FAB) m/z 524 (M'+1); 10 Anal. calcd for C 29
H
3 7 NzO 6 -0.2H 2 0: C, 66.07; H, 7.15; N, 7.97. Found: C, 66.02; H, 7.14; N, 7.87. Example 167 4-[[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxylphenylacetyl]-(2S)-pyrrolidinyl]methoxy] cyclohexanecarboxylic acid N N 0 NI' COOH 175 iHH Me 175 15 To a stirred solution of methyl 4-[(1-tert-butoxycarbonyl-2-pyrrolidinyl)methoxy] cyclohexane carboxylate (900 mg, 2.6 mmol) in CH 2 C1 2 (5.0 ml) was added TFA (5.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 C1 2 . The extract was washed with brine ,dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent 20 reaction without further purification. To a stirred solution of the crude product (200 mg, 0.83 mmol), 4-[N'-(2-chlorophenyl)uredio]-3-methoxyphenylacetic acid (277 mg, 0.83 mmol), HOBt (135 mg, 1.0 mmol), and triethylamine (344 ml, 2.48 mmol) in THF (10.0 ml) and MeCN (10.0 ml) was added EDC-HCI (238 mg, 1.24 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted 25 with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (1:6, v/v) as eluent to give methyl 4-[[1-[4-[N' (2-chlorophenyl)ureido]-3-methoxylphenylacetyl]-(2S)-pyrrolidinyl]methoxy] cyclohexane carboxylate (450 mg, 97%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.35-2.15 (m, 12H), 2.25-2.40 30 (m, lH), 3.40-3.70 (m, 5H), 3.61 (s, 2H), 3.66 (s, 3H), 3.81 (s, 3H), 4.20-4.30 (m, 1), 6.81-6.99 (m, 3H), 7.17-7.34 (m, 3H), 7.92 -7.94 (m, 2W, 8.17-8.19 (m, 2H). 344 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxylphenylacetyl]-(2S) pyrrolidinyl]methoxy]cyclohexanecarboxylate (450 mg, 0.86 mmol) in THF (10.0 ml) and MeOH (5.0 ml) was added IN NaOH (1.4 ml, 1.4 mmol). The mixture was stirred at 70 oC for 24 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HCl. 5 The resulting solid was collected, washed with water, and dried in vacuo to give 175 (370 mg, 84%) as a white crystalline solid. MW 544.04 mp 111-115 oC; IR (KBr) 3330, 2938, 1704, 1594, 1533, 1438, 1199 cm-'; 'H-NMR (DMSO-d 6 ) 8 1.00-2.00 (min, 12H), 2.20-2.30 (min, 1H), 3.20-3.80 (min, 5H), 3.55 (s, 2H), 3.85 (s, 3H), 4.00-4.20 (min, liH), 6.73-6.75 (min, 1H), 6.87 (s, 1H), 6.99-7.03 (min, 1H), 7.25-7.29 (min, 1H), 7.42 (d,J= 7.1 Hz, 1H), 7.95 (d,J= 8.3 Hz, 1H), 8.08 (d,J= 8.0, 10 1H), 8.78 (s, 1H), 8.92 (s, 1H); MS (FAB) nm/z 544 (M+1); Anal. calcd for C 28
H
34
N
3 0 6 C'-0.2H 2 0: C, 61.41; H, 6.33; N, 7.67; Cl, 6.47. Found: C, 61.37; H, 6.32; N, 7.56; Cl, 6.55. Example 168 4-[ 1-[4-[N'-(2-bromophenyl)ureido]-3-methoxylphenylacetyl]-(2S)-pyrrolidinyl]methoxy cyclohexanecarboxilic acid Y. I r QKI COOH N N 15 r H H Me 176 To a stirred solution of methyl 4-[(1-tert-butoxycarbonyl-2-pyrrolidinyl)methoxy]cyclohexane carboxylate (900 mg, 2.6 mmol) in CH 2 Cl 2 (5.0 ml) was added TFA (5.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 Cl 2 . The extract was washed with brine, 20 dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product (200 mg, 0.83 mmol), 4-[N'-(2-bromophenyl)uredio]-3-methoxyphenylacetic acid (314 mg, 0.83 mmol), HOBt (135 mg, 1.0 mmol), and triethylamine (344 ml, 2.48 mmol) in THF (10.0 ml) and MeCN (10.0 ml) was added EDC-HCI (238 mg, 1.24 mmol) at 0 oC. The reaction mixture was stirred at room 25 temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (1:6, v/v) as eluent to give methyl [4-[1-[4-[N' (2-bromophenyl)ureido]-3-methoxylphenylacetyl]-(2S)-pyrrolidinyl]methoxy]cyclohexane 30 carboxylate (450 mg, 90%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.30-2.10 (min, 12H), 2.35-2.40 (min, 1H), 3.25-3.70 (min, 5H), 3.84 (s, 3H), 4.10-4.25 (min, 1W, 6.81-7.06 (min, 4H), 7.25-7.32 (inm, 2H), 7.50-7.52 (min, 1H), 7.90-7.92 (min, 1H), 8.13-8.15 (min, 1H). 345 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxylphenylacetyl]-(2S) pyrrolidinyl]methoxycyclohexanecarboxylate (450 mg, 0.74 mmol) in THF (10.0 ml) and MeOH (5.0 ml) was added iN NaOH (1.2 ml, 1.2 mmnol). The mixture was stirred at 70 oC for 24 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HCL. 5 The resulting solid was collected, washed with water, and dried in vacuo to give 176 (340 mg, 77%) as a white crystalline solid. MW 588.49 mp 108-111 oC; IR (KBr) 3328, 2938, 1702, 1594, 1529, 1434 cm-'; 'H-NMR (DMSO-d 6 ) 8 1.00-2.00 (m, 12H), 2.15-2.25 (m, 1H11), 3.40-3.75 (m, 5H), 3.48 (s, 2H), 3.85 (s, 3H), 4.04-4.15 (m, 1H), 6.70-6.72 (m, 1H1), 6.87 (s, 1H), 6.94-6.98 (m, 1H), 7.29-7.33 (m, 1H1), 7.59 (d, J= 8.1 Hz, IH), 7.93-7.95 (m, 2H), 8.73-8.74 (m, 1H1), 8.91-8.92 10 (m, 1H); MS (FAB) m/z 589 (M +1); Anal. calcd for C 28
H
34
N
3
O
6 Br-0.2HO 2 0: C, 56.80; H, 5.86; N, 7.10; Br, 13.49. Found: C, 56.66; H, 5.83; N, 6.97; Br, 13.66. Example 169 4-[[ 1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S)-pyrrolidinyl]methoxy]cyclohexane carboxylic acid N NCO 15 Me H H 177 To a stirred solution of methyl 4-[(1-tert-butoxycarbonyl-2-pyrrolidinyl)methoxy]cyclohexane carboxylate (450 mg, 1.3 mmol) in CH 2 Cl 2 (5.0 ml) was added TFA (5.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2
CI
2 . The extract was washed with brine, 20 dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product, 4-[N'-(2 methylphenyl)uredio]phenylacetic acid (375 mg, 1.3 mmol), HOBt (178 mg, 1.3 mmnol), and triethylamine (550 ml, 3.9 mmnol) in THF (6.0 ml) and MeCN (6.0 ml) was added EDC-HCI (380 mg, 1.9 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and 25 concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over NaSO 4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n hexane-EtOAc (1:6, v/v) as eluent to give methyl 4-[[1-[4-[N'-(2-methylphenyl)ureido] phenylacetyl]-(2S)-pyrrolidinyl]methoxy]cyclohexanecarboxylate (520 mg, 78%) as a colorless oil. 30 1 H-NMR (CDC13) 5 1.30-2.40 (m, 13H), 2.21 (s, 3H), 3.30-3.80 (m, 7H), 3.65 (s, 3H), 4.10-4.30 (m, 1H1), 6.90-7.20 (m, 8H), 7.40-7.70 (m, 2H). 346 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[[1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S) pyrrolidinyl]methoxy]cyclohexanecarboxylate (520 mg, 1.0 mmol) in THF (10.0 ml) and MeOH (5.0 ml) was added IN NaOH (1.5 ml, 1.5 mmol). The mixture was stirred at 70 oC for 24 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HCl. 5 The resulting solid was collected, washed with water, and dried in vacuo to give 177 (450 mg, 91%) as a white crystalline solid. MW 493.60 mp 107-111 oC; IR (KBr) 3353, 2938, 1704, 1540, 1454, 1240 cm-'; 'H-NMR (DMSO-d 6 ) 8 1.20-2.00 (m, 12H), 2.23 (s, 3H), 2.22-2.24 (m, 1H), 3.20-3.80 (m, 7H), 4.00-4.18 (m, 1H), 6.90-6.94 (m, 1H), 7.10-7.16 (m, 5H), 7.36-7.38 (m, 211), 7.82-7.87 (m, 2H11), 8.89 (s, 1H), 12.0 (br s, 1H); MS (FAB) m/z 494 (M+1); Anal. calcd for 10 C 2 8
H
35
N
3 Os0.2H 2 0: C, 67.64; H, 7.18; N, 8.45. Found: C, 67.66; H, 7.19; N, 8.24. Example 170 cis-4-[ -[4-[N'-(2-chlorophenyl)ureido]-3-methoxylphenylacetyl]-(2S)-octahydroindolylmethoxy] cyclohexanecarboxylic acid a-aCOOH i::NYN 0 -~-Co C HH Me 178 15 To a stirred solution of [1-tert-butoxycarbonyl-(2S)-octahydroindolyl]carboxylic acid (1.0 g, 3.7 mmol) in THF (10.0 ml) was added BH 3 -THF(1.0 M in THF, 8.0 ml) at 0 0 C. After stirred at room temperature for 1.0 h, the reaction mixture was heated under reflux for 1.5 hr. After cooled, the mixture was concentrated in vacuo. Water was added thereto at 0oC, and extracted with EtOAc. The extract was washed with water, then dried over Na 2
SO
4 , and concentrated in vacuo to give [1 20 tert-butoxycarboyl-(2S)-octahydroindolyl]methanol (947 mg, quant) as a a colorless oil. To a stirred solution of [1-tert-butoxycarboyl-(2S)-octahydroindolyl]methanol (947 mg, 3.7 mmol), methyl 4-hydroxybenzoate (565 mg, 3.7 mmol), and Ph 3 P (1.2 g, 4.5 mmol) in THF (10 ml) was added DIAD (984 mg, 4.5 mmol) at 0 'C. The reaction mixture was stirred at room temperature for 18 hr. The mixture was concentrated in vacuo. The residue was purified by 25 column chromatography on silica gel with n-hexane-EtOAc (9/1, v/v) as eluent to give methyl 4 [1-tert-butoxycarbonyl-(2S)-octahydroindolylmethoxy]benzoate (700 mg, 50%) as a pale yellow oil. 'H-NMR (CDC 3 ) 8 1.10-2.25 (m, 11H), 1.45 (s, 9H), 3.88 (s, 3H11), 3.70-4.20 (m, 3H), 4.36 (br s, 1H), 6.94 (d, J= 8.8 H z, 2H), 7.96 (d, J= 8.5 H z, 2H11). A mixture of methyl 4-[1-tert-butoxycarbonyl-(2S)-octahydroindolylmethoxy]benzoate (700 mg, 347 WO 01/00206 PCT/US00/18079 1.8 mmol) and 5% Rh on alumina (400 mg) in EtOH (10.0 ml) and AcOH (1.0 ml) was hydrogenated at room temperature at 5 atm for 48 hr. The catalyst was filtered off, and the filtrate was concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc (7:1, v/v) as eluent to give methyl cis-4-[1-tert-butoxycarbonyl-(2S)-octahydro 5 indolylmethoxy]cyclohexanecarboxylate (600 mg, 85%) as a pale yellow oil. 'H-NMR (CDC13) 8 1.10-2.35 (min, 20H), 1.44 (s, 9H), 3.45-3.90 (min, 5H), 3.80 (s, 3H). To a stirred solution of methyl cis-4-[1-tert-butoxycarbonyl-(2S)-octahydroindolylmethoxy] cyclohexanecarboxylate (600 mg, 1.5 rmmol) in CH 2 Cl 2 (6.0 ml) was added TFA (6.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in 10 vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2
CI
2 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product (221 mg, 0.75 mmol), 4-[N'-(2-chlorophenyl)uredio]-3-methoxyphenylacetic acid (250 mg, 0.75 mmol), HOBt (101 mg, 0.75 mmol), and triethylamine (312 ml, 2.3 mmol) in THF (10.0 ml) and 15 MeCN (10.0 ml) was added EDC-HCI (216mg, 1.1 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc(1:3, v/v) as eluent to give methyl cis-4-[1-[4 20 [N'-(2-chlorophenyl)ureido]-3-methoxylphenylacetyl]-(2S)-octahydroindolylmethoxy]cyclohexane carboxylate (430 mg, 94%) as a colorless oil. 'H-NMR (CDC1 3 ) 8 1.10-2.40 (min, 20H), 3.45 (br s, 1H), 3.62 (s, 2H), 3.66 (s, 3H), 3.73 (s, 3H), 3.60-3.85 (min, 2H), 4.09-4.14 (min, 2H), 6.75-6.98 (inm, 3H), 7.22-2.46 (min, 4H), 7.92 (d,J= 8.0 Hz, 1H), 8.18 (d,J = 8.3 Hz, 1H). To a stirred solution of methyl cis-4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxylphenylacetyl] 25 (2S)-octahydroindolylmethoxy]cyclohexanecarboxylate (430 mg, 0.7 mmol) in THF (10.0 ml) and MeOH (5.0 ml) was added IN NaOH (1.4 ml, 1.4 mmol). The mixture was stirred at 70 oC for 24 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HC1. The resulting solid was collected, washed with water, and dried in vacuo to givel78 (360 mg, 86%) as a white crystalline solid. MW 598.13 mp 120-121 0 C; IR (KBr) 3338, 2933, 2859, 30 1614, 1533, 1438 cm-'; 'H-NMR (CDCl 3 ) 8 1.05-2.40 (min, 20H), 3.38-3.50 (min, 2H11), 3.63 and 3.65 (each s, total 2H11), 3.71 and 3.75 (each s, total 3H), 3.70-3.80 (min, 1H), 3.93-3.97 (min, 1H), 4.15 (br s, 1H), 6.75-6.77 (min, 2H), 6.93-6.97 (min, 1H), 7.20-7.32 (min, 3H), 7.60-7.63 (min, 1H), 7.85 (d, J= 348 WO 01/00206 PCT/US00/18079 8.3 Hz, 1H), 8.16 (d, J = 8.3 Hz, 1H); MS (FAB) nm/z 598 (M+1); Anal. calcd for C 32
H
40
N
3 0 6
CI
0.5H120: C, 63.30; H, 6.81; N, 6.92; Cl, 5.84. Found: C, 63.68; H, 6.81; N, 6.81; Cl, 5.98. Example 171 cis-4-[ 1-[4-[N'-(2-bromophenyl)ureido]-3-methoxylphenylacetyl]-(2S)-octahydroindolylmethoxy] 5 cyclohexanecarboxylic acid N N N COOH 179 r H H M Me 179 To a stirred solution of methyl cis-4-[1-tert-butoxycarbonyl-(2S)-octahydroindolylmethoxy] cyclohexanecarboxylate (600 mg, 1.5 mmol) in CH2C1 2 (6.0 ml) was added TFA (6.0 ml) at 0 oC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in 10 vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2
C
2 . The extract was washed with brine ,dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product (221 mg, 0.75 mmol), 4-[N'-(2-bromophenyl)uredio]-3-methoxyphenylacetic acid (284 mg, 0.75 mmol), HOBt (101 mg, 0.75 mmol), and triethylamine (312 ml, 2.3 mmol) in THF (10.0 ml) and 15 MeCN (10.0 ml) was added EDC.HCI (216mg, 1.1 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2 M citric acid, and sat. NaHCO 3 , then dried over Na 2 SO4, and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc(1:3 ,v/v) as eluent to give methyl cis-4-[1-[4 20 [N'-(2-bromophenyl)ureido]-3-methoxylphenylacetyll-(2S)-octahydroindolylmethoxy] cyclohexane carboxylate (480 mg, 96%) as a colorless oil. 'H-NMR (CDC13) 5 1.10-2.40 (min, 20H), 3.45 (br s, 1H1), 3.61 (s, 2H), 3.66 (s, 3H), 3.76 (s, 3H), 3.60-3.80 (min, 2H), 4.11-4.14 (min, 2H), 6.76-6.92 (inm, 3H), 7.25-7.32 (min, 3H), 7.49 (d, J= 7.1 Hz, 111), 7.90 (d, J= 8.0 Hz, 1), 8.13 (d, J = 8.0 Hz, 1H). To a stirred solution of methyl cis-4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxylphenylacetyl] 25 (2S)-octahydroindolylmethoxy]cyclohexanecarboxylate (480 mg, 0.73 mmol) in THF (10.0 ml) and MeOH (5.0 ml) was added IN NaOH (1.5 ml, 1.5 mmol). The mixture was stirred at 70 oC for 24 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HCL. The resulting solid was collected, washed with water, and dried in vacuo to give 179 (400 mg, 85%) as a white crystalline solid. MW 642.58 mp 115-120 'C; IR (KBr) 3332, 2933, 2859, 30 1704, 1592, 1529, 1434 cm-'; 'H-NMR (CDC1 3 ) 8 1.10-2.40 (min, 20H), 3.40-3.50 (mn, 2H11), 3.61 and 349 WO 01/00206 PCT/US00/18079 3.63 (each s, total 2H11), 3.75 and 3.78 (each s, total 3H11), 3.70-3.80 (min, 1H), 3.90-3.93 (mn, 1), 4.15 (br s, min), 6.76-6.92 (min, 3H), 7.26-7.30 (min, 1H1), 7.43-7.52 (min, 3H), 7.84-7.86 (min, 1H), 8.10 8.12 (min, 1H); MS (FAB) nm/z 643 (M++l); Anal. calcd for C 3 2
H
40 N30,Br-0.4HO 2 0: C, 59.15; H, 6.33; N, 6.49; Br, 12.30. Found: C, 59.26; H, 6.33; N, 6.36; Br, 12.37. 5 Example 172 4-[[ 1 -[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrffolidinyllcarbonylamino] cyclohexanecarboxylic acid N H 'N N COOH Me H H- Me 180S To a stirred solution of benzyl 4-aminocyclohexanecarboxylate (900 mg, 3.9 mmol), boc-proline 10 (830 mg, 3.9 mmol), HOBt (521 mg, 3.9 mmol), and triethylamine (1.6 ml, 11.6 mmol) in CH 2 C1 2 (30.0 ml) was added EDC-HCl (1.1 g, 5.8 mmol) at 0 'C. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The residue was purified by column 15 chromatography on silica gel with n-hexane-EtOAc (6:1, v/v) as eluent to give benzylcis-4-[(1 tert-butoxycarbonyl-2-pyrrolidinyl)carbonylamino]cyclohexanecarboxylate (600 mg, 36%) as an amorphous solid. 'H-NMR (CDC13) 8 1.44 (s, 9H), 1.50-1.90 (mn, 12H), 2.20-2.26 (min, 111), 3.25 3.50 (min, 2H), 3.80-3.90 (min, 1H), 4.10-4.25 (min, 1H), 5.12 (s, 21H), 7.35-7.36 (min, 5H). To a stirred solution of benzylcis-4-[(1-tert-butoxycarbonyl-2-pyrrolidinyl)carbonylamino] 20 cyclohexanecarboxylate (600 mg, 1.4 mmol) in CH 2 C1 2 (6.0 ml) was added TFA (3.0 ml) at 0 'C. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 C1 2 . The extract was washed with brine, dried over Na 2
SO
4 , and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product 25 (300 mg, 0.7 mmol), 3-methoxy-4-[N'-(2-methylphenyl)uredio]phenylacetic acid (220 mg, 0.7 nunmmol), HOBt (94 mg, 0.7 mmol), and triethylamine (291 ml, 2.1 mmol) in THF (10.0 ml) and MeCN (10.0 ml) was added EDC-HCl (201 mg, 1.1 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. 30 NaHCO 3 , then dried over Na 2
SO
4 , and concentrated in vacuo. The resulting solid was collected and washed with EtOAc to give benzyl 4-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-2-pyrrolidinyl]carbonylamino]cyclohexanecarboxylate (380 ming, 87%) as a white 350 WO 01/00206 PCT/US00/18079 crystalline solid. 'H-NMR (CDCI 3 ) 8 1.40-2.15 (min, 12H), 2.28 (min, 3H), 2.30-2.50 (min, 2H), 3.40 3.55 (min, 2H), 3.61 (s, 2W, 3.71 (s. 3H), 3.82 (min, 1iH), 4.53 (d,J = 6.3 Hz, 1H), 5.10 (s, 2H), 6.42 (s, 111), 6.77-6.79 (min, 2W, 7.04-7.34 (min, 9H), 7.50-7.52 (min, 1iH), 8.05-8.07 (min, 1iH). To a stirred solution of benzyl 4-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2 5 pyrrolidinyl]carbonylamino]cyclohexanecarboxylate (380 mg, 0.6 mmol) in THF (10.0 ml) and EtOH (5.0 ml) was added IN NaOH (0.9 ml, 0.9 mmol). The mixture was stirred at 50 'C for 18 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HCI. The resulting solid was collected, washed with water, and dried in vacuo to give 180 (230 mg, 71%) as a white crystalline solid. MW 636.62 mp 136-142 'C; IR (KBr) 3345, 2940, 1650, 10 1625, 1535, 1454 cm-'; 'H-NMR (DMSO-d 6 ) 6 1.40-2.00 (min, 12H), 2.24 (s, 3H), 2.30-2.40 (inm, 1iH), 3.45-3.80 (min, SH), 3.86-3.87 (min, 3H), 4.30-4.43 (mn, 1iH), 6.65-7.30 (mn, SH), 7.70-7.80 (inm, 1W, 7.98-8.09 (min, 1iH), 8.46-8.57 (min, 1iH); MS (FAB) m/z 537 (M+1); Anal. calcd for
C
2 9H 36
N
4 0 6 "0.5 HO20: C, 63.84; H, 6.83; N, 10.27. Found: C, 64.18; H, 6.91; N, 9.85. Example 173 15 4-[[ 1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxylphenylacetyl]-2-pyrrffolidinyl]carbonylamino] cyclohexanecarboxylic acid N -H N COOH H H Me 181 To a stirred solution of benzylcis-4-[(1-tert-butoxycarbonyl-2-pyrrolidinyl)carbonylamino] cyclohexanecarboxylate (600 mg, 1.4 mmol) in CH 2
C
2 (6.0 ml) was added TFA (3.0 ml) at 0 oC. 20 The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO 3 was added to the residue, and extracted with CH 2 C1 2 . The extract was washed with brine, dried over Na 2 SO4, and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product (300 mg, 0.7 mmol), 4-[N'-(2-chlorophenyl)uredio]-3-methoxyphenylacetic acid (237 mg, 0.7 25 mmol), HOBt (94 mg, 0.7 mmol), and triethylamine (291 ml, 2.1 mmol) in THF (10.0 ml) and MeCN (10.0 ml) was added EDC-HCI (201 mg, 1.1 mmol) at 0 oC. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuo. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 , 2-M citric acid, and sat. NaHCO 3 , then dried over Na 2 SO4 , and concentrated in vacuo. The resulting solid was collected 30 and washed with EtOAc to give benzyl 4-[[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyl phenylacetyl]-2-pyrrolidinyl]carbonylamino]cyclohexanecarboxylate (310 inmg, 68%) as a white 351 WO 01/00206 PCT/US00/18079 crystalline solid. 'H-NMR (CDCl 3 ) 8 1.40-2.15 (min, 12H), 2.30-2.60 (mn, 2H), 3.42-3.55 (mn, 2H), 3.64 (s, 2H), 3.84 (s,3H),4.55 (d, J=6.1Hz, 1H),5.12 (s,2H), 6.81-7.35(m,12H),7.96-7.98 (mn, 1H), 8.17-8.19 (min, 111). To a stirred solution of methyl benzyl 4-[[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxylphenyl 5 acetyl]-2-pyrrolidinyl]carbonylamino]cyclohexanecarboxylate (310 mg, 0.86 mmol) in THF (10.0 ml) and MeOH (5.0 ml) was added IN NaOH (0.7 ml, 0.7 mmol). The mixture was stirred at 50 oC for 18 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HC1. The resulting solid was collected, washed with water, and dried in vacuo to give 181 (260 mg, 98%) as a white crystalline solid. MW 557.03 mp 135-140 'C; IR (KBr) 3328, 2938, 10 1594, 1533,1438, 1203 cm'; 'H-NMR (DMSO-d) 8 1.40-2.20 (min, 12H), 2.30-2.40 (m, 1H11), 3.40 3.80 (min, 5H), 3.65-3.85 (min, 3H), 4.30-4.43 (mn, 1H), 6.66-7.31 (mn, 5H), 7.42-8.10 (mn, 2H), 8.89 8.94 (min, 2H); MS (FAB) m/z 557 (M++1); Anal. calcd for C 28
H
33
N
4 0 6 CI-0.3H 2 0: C, 59.79; H, 6.02; N, 9.96; Cl, 6.30. Found: C, 59.86; H, 6.10; N, 9.60; Cl, 6.34. Example 174 15 4-[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(3S)-pyrrolidinyloxy]benzoic acid 0--/COOH ~ N6 MeH H Me 0182 To a cooled (0 oC), stirred solution of methyl 4-hydroxybenzoate (1.84 g, 12.1 immol), (R)-1 benzyl-3-pyrrolidinol (2.00 ml, 12.1 mmol), and Ph 3 P (3.81 g, 14.5 mmol) in THF (25 ml) was added DIAD (2.86 ml, 14.5 mmol) and the reaction mixture was heated under reflux for 10 hr. 20 After cooled to room temperature, the mixture was evaporated. The residue was purified by column chromatography on silica-gel with n-hexane-EtOAc (3:1, v/v) as eluent to give methyl 4 [1-benzyl-(3S)-pyrrolidinyloxy]benzoate (3.66 g, 97%) as a pale yellow oil. 'H-NMR (CDCI 3 ) 8 1.95-2.02 (min, 1 H), 2.28-2.37 (min, 1 H), 2.57-2.63 (min, 1 H), 2.72-2.81 (mn, 2 H), 2.96-3.01 (mn, 1 H), 3.63-3.71 (min, 2 H), 3.87 (s, 3 H), 4.84-4.89 (min, 1 H), 6.84 (d, J = 8.8 Hz, 2 H), 7.23-7.34 (inm, 25 5 H), 7.95 (d, J = 8.8 Hz, 2 H). A solution of methyl 4-[1-benzyl-(3S)-pyrrolidinyloxy]benzoate (3.66 g, 11.8 mmol) in MeOH (25 ml) was hydrogenated over Pd(OH) 2 /C (0.73 g, 20 wt%) overnight. The reaction mixture was filtered to remove the catalyst and the solution was evaporated to give methyl 4-[(3S) pyrrolidinyloxy]benzoate (2.60 g, q.y.) as a pale yellow oil. 'H-NMR (CDCl 3 ) 8 1.94-2.01 (mn, 2 352 WO 01/00206 PCT/US00/18079 H), 2.09-2.18 (m, 1 H), 2.91-2.97 (m, 1 H), 3.04-3.09 (m, 1 H), 3.16-3.23 (m, 2 H), 3.88 (s, 3 H), 4.88-4.91 (m, 1 H), 6.86 (m, 2 H), 7.96-7.98 (m, 2 H); MS (ESI) m/z 222 (M'+1). A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (449 mg, 1.43 mmol), methyl 4-[(3S)-pyrrolidinyloxy]benzoate (316 mg, 1.43 mmol), EDCHCI (330 mg, 1.72 mmol), 5 HOBt (193 mg, 1.43 mmol), and Et 3 N (240 ml, 1.72 mmol) in THF (5 ml) was stirred at room temperature overnight. The reaction mixture was diluted with HO 2 0, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHC13-MeOH (100:1 to 50:1, v/v) as eluent to give methyl 4-[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]lphenylacetyl]-(3S)-pyrrolidinyloxy] 10 benzoate (735 mg, 99%) as a pale yellow oil. 'H-NMR (CDC13) 8 2.03-2.31 (series of s and m, total 5 H), 3.57-3.78 (series of m, total 9 H), 3.88 (s, 3 H), 4.95-4.99 (m, 1 H), 6.73-7.00 (m, 5 H), 7.06-7.10 (m, 1 H), 7.18-7.22 (m, 2 H), 7.42-7.46 (m, 1 H), 7.57-7.62 (m, 1 H), 7.95-8.08 (m, 3 H); MS (ESI) m/z 518 (M'+1). To a stirred solution of methyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(3S) 15 pyrrolidinyloxy]benzoate (627 mg, 1.21 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 3 hr. After cooled to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was collected under a reduced pressure. The crude solid was recrystallized from MeOH-CHCl 3 -IPE to give 182 (235 mg, 39%) as a white crystalline powder. MW 503.55 mp 131-135oC; 'H-NMR (DMSO-d 6 ) 8 2.06-2.27 (series 20 of m, total 5 H), 3.57-3.64 (m, 4 H), 3.71-3.88 (series of s and m, total 5 H), 5.11 and 5.20 (each m, total 1 H), 6.73-6.77 (m, 1 H), 6.88-6.95 (m, 2 H), 7.02-7.05 (m, 2 H), 7.11-7.17 (m, 2 H), 7.79-7.81 (m, 1 H), 7.88-7.90 (m, 2 H), 7.98-8.03 (m, 1 H), 8.45-8.47 (m, 1 H), 8.55-8.57 (m,1 H), 12.66 (br s, 1 H); MS (ESI) m/z 504 (M++1); Anal. Calcd for C 28
H
29
N
3 0 6 3/4H 2 0: C, 65.04; H, 5.95; N, 8.13. Found: C, 65.11; H, 5.99; N, 7.66. 25 Example 175 4-[ 1-[4-[N'-(2-chlorophenyl)ureido]l-3-methoxyphenylacetyl]-(3S)-pyrrolidinyloxy]benzoic acid 0--/COOH ~ NN Me H M 183 A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (410 mg, 1.22 mmol), methyl 4-[(3S)-pyrrolidinyloxy]benzoate (270 mg, 1.22 mmol), EDCHCI (280 mg, 1.46 mmol), 30 HOBt (200 mg, 1.48 mmol), and Et 3 N (205 ml, 1.47 mmol) in THF (8 ml) was stirred at room 353 WO 01/00206 PCT/US00/18079 temperature overnight. The reaction mixture was diluted with HO20, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCI 3 -MeOH (100:1 to 60:1, v/v) as eluent to give methyl 4-[ 1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(3S)-pyrrolidinyloxy] 5 benzoate (652 mg, 99%) as a white solid. mp 200-203 oC; 'H-NMR (CDCl 3 ) 8 2.06-2.32 (min, 2 H), 3.60-3.82 (series of m, total 9 H), 3.88 (s, 3 H), 4.97-5.01 (mn, 1 H), 6.76-6.86 (min, 4 H), 6.95-6.99 (mn, 1 H), 7.23-7.47 (min, 4 H), 7.91-7.99 (min, 3 H), 8.19-8.21 (min, 1 H); MS (ESI) m/z 537 (M). To a stirred solution of methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(3S) pyrrolidinyloxy]benzoate (650 mg, 1.21 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and 10 the reaction mixture was heated under reflux for 5 hr. After cooled to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was collected under a reduced pressure. The crude solid was recrystallized from MeOH-CHCl 3 -IPE to give 183 (443 mg, 70%) as a pale yellow crystalline powder. MW 523.97 mp 190-193oC; 'H-NMR (DMSO-d 6 ) 8 2.06-2.27 (min, 2 H), 3.56-3.62 (min, 4 H), 3.71-3.88 (series of s and min, total 5 H), 5.11 and 5.20 (each m, total 15 1 H), 6.74-6.78 (min, 1 H), 6.89-6.91 (min, 1 H), 7.00-7.05 (m, 3 H), 7.26-7.30 (min, 1 H), 7.43-7.45 (min, 1 H), 7.88-7.98 (min, 3 H), 8.08-8.10 (min, 1 H), 8.89-8.95 (min, 2 H), 12.67 (br s, 1 H); MS (ESI) m/z 524 (M+1); Anal. Calcd for C 27
H
26
CIN
3 0 6 1/4H 2 0: C, 61.36; H, 5.05; N, 7.95; Cl, 6.71. Found: C, 61.69; H, 5.45; N, 7.29; Cl, 6.91. Example 176 20 4-[ 1-[4-[N '-(2-bromophenyl)ureido]-3-methoxyphcnylacetyl]-(3S)-pyrrolidinyloxy]benzoic acid 0- /COOH N N_ r H H Me 184 A mixture of 4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (540 mg, 1.42 mmol), methyl 4-[(3S)-pyrrolidinyloxy]benzoate (315 mg, 1.42 mmol), EDCHCI (328 mg, 1.71 mmol), HOBt (230 mg, 1.70 mmol), and Et 3 N (240 ml, 1.72 mmol) in THF (8 ml) was stirred at 25 room temperature overnight. The reaction mixture was diluted with H20, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1 to 50:1, v/v) as eluent to give methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-(3S)-pyrrolidinyloxy] benzoate (620 mg, 74%) as a white foam. 'H-NMR (CDCI 3 ) 8 2.06-2.33 (mn, 2 H), 3.60-3.82 30 (series of m, total 9 H), 3.89 (s, 3 H), 4.97-5.01 (mn, 1 H), 6.77-7.00 (min, 5 H), 7.27-7.40 (min, 3 H), 7.49-7.51 (min, 1 H), 7.91-7.99 (min, 3 H), 8.13-8.17 (min, 1 H); MS (ESI) m/z 583 (M+1). 354 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[ 1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl acetyl]-(3S)-pyrrolidinyloxy]benzoate (620 mg, 1.06 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 2.5 hr. After cooled to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was collected 5 under a reduced pressure. The crude solid was recrystallized from MeOH-CHCl 3 -IPE to give 184 (421 mg, 70%) as a white crystalline powder. MW 568.42 mp 173-175 0 C; 'H-NMR (DMSO-d 6 ) 8 2.06-2.28 (m, 2 H), 3.56-3.65 (m, 4 H), 3.71-3.88 (series of s and m, total 5 H), 5.12 and 5.20 (each m, total 1 H), 6.74-6.78 (m, 1 H), 6.89-7.05 (m, 4 H), 7.31-7.34 (m, 1 H), 7.59-7.61 (m, 1 H), 7.88-7.98 (m, 4 H), 8.73-8.74 (m, 1 H), 8.91-8.93 (m, 1 H), 12.67 (br s, 1 H); MS (ESI) m/z 10 569 (M+1); Anal. Calcd for C 27
H
26 BrN 3 06: C, 57.05; H, 4.61; N, 7.39; Br, 14.06. Found: C, 57.57; H, 5.12; N, 6.81; Br, 13.96. Example 177 4-[ 1 -[3-methoxy-4-[N '-(2-methylphenyl)ureido]phenylacetyl]-(3R)-pyrrolidinyloxy]benzoic acid O-C-COOH MeH H Me 185 15 To a cooled (0OC), stirred solution of methyl 4-hydroxybenzoate (1.78 g, 11.7 mmol), 1-benzyl (3S)-pyrrolidinol (2.07 g, 11.7 mmol), and Ph 3 P (3.68 g, 14.0 mmol) in THF (25 ml) was added DIAD (2.76 ml, 14.0 mmol) and the reaction mixture was heated under reflux for 10 hr. After cooled to room temperature, the mixture was evaporated. The residue was purified by column chromatography on silica-gel with n-hexane-EtOAc (3:1, v/v) as eluent to give methyl 4-[ 1 20 benzyl-(3R)-pyrrolidinyloxy]benzoate (3.56 g, 98%/) as a pale yellow oil. 'H-NMR (CDCl 3 ) 8 1.95-2.03 (m, 1 H), 2.28-2.37 (m, 1 H), 2.57-2.63 (m, 1 H), 2.73-2.81 (m, 2 H), 2.97-3.01 (m, 1 H), 3.63-3.72 (m, 2 H), 3.87 (s, 3 H), 4.85-4.90 (m, 1 H), 6.83-6.85 (m, 2 H), 7.27-7.34 (m, 5 H), 7.94-7.97 (m, 2 H); MS (ESI) m/z 312 (M +1). A solution of methyl 4-[1-benzyl-(3R)-pyrrolidinyloxy]benzoate (3.56 g, 11.4 mmol) in MeOH (25 25 ml) was hydrogenated over Pd(OH) 2 /C (0.72 g, 20 wt%) overnight. The reaction mixture was filtered to remove the catalyst and the solution was evaporated to give methyl 4-[(3R) pyrrolidinyloxy]benzoate (2.53 g, q.y.) as a pale yellow oil. 'H-NMR (CDCl 3 ) 8 1.94-2.18 (m, 3 H), 2.91-2.97 (m, 1 H), 3.04-3.09 (m, 1 H), 3.16-3.22 (m, 2 H), 3.88 (s, 3 H), 4.88-4.91 (m, 1 H), 6.86-6.89 (m, 2 H), 7.97-7.99 (m, 2 H); MS (ESI) m/z 263 [M++1+41, (+MeCN)]. 355 WO 01/00206 PCT/US00/18079 A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (460 mg, 1.46 mmol), methyl 4-[(3R)-pyrrolidinyloxy]benzoate (324 mg, 1.46 mmol), EDCHCI (337 mg, 1.76 mmol), HOBt (237 mg, 1.75 mmol), and Et 3 N (245 ml, 1.76 mmol) in THF (10 ml) was stirred at room temperature overnight. The reaction mixture was diluted with H 2 0, and extracted with EtOAc. 5 The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1 to 50:1, v/v) as eluent to give methyl 4-[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(3R)-pyrrolidinyloxy] benzoate (583 mg, 77%) as a colorless oil. 'H-NMR (CDC13) 8 2.06-2.23 (m, 2 H), 2.30 (s, 3 H), 3.58-3.89 (series of s and m, total 12 H), 4.95-4.99 (m, 1 H), 6.75-7.00 (m, 4 H), 7.13-7.30 (m, 5 10 H), 7.52-7.57 (m, 1 H), 7.96-8.03 (m, 3 H); MS (ESI) m/z 518 (M'+1). To a stirred solution of methyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(3R) pyrrolidinyloxy]benzoate (583 mg, 1.13 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 3 hr. After cooled to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was collected under a reduced 15 pressure. The crude solid was recrystallized from MeOH-CHC 3 -EtO to give 185 (297 mg, 52%) as a white crystalline powder. MW 503.55 mp 158-162oC; 'H-NMR (DMSO-d 6 ) 8 2.08-2.31 (series of s and m, total 5 H), 3.54-3.89 (series of m, total 9 H), 5.11 and 5.20 (each m, total 1 H), 6.72-7.17 (series of m, total 6 H), 7.78-7.80 (m, 1 H), 7.87-7.90 (m, 2 H), 7.98-8.02 (m, 2 H), 8.46-8.47 (m, 1 H), 8.55-8.57 (m,1 H), 12.66 (br s, 1 H); MS (ESI) m/z 504 (M+1); Anal. Calcd 20 for C 28
H
2 9
N
3 0 6 "1/4H 2 0: C, 66.19; H, 5.85; N, 8.27. Found: C, 66.12; H, 5.77; N, 8.21. Example 178 4-[ 1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(3R)-pyrrolidinyloxy]benzoic acid O-COOH NN 0D MeH M 186 A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (498 mg, 1.49 25 mmol), methyl 4-[(3R)-pyrrolidinyloxy]benzoate (329 mg, 1.49 mmol), EDCHCI (342 mg, 1.78 mmol), HOBt (241 mg, 1.78 mmol), and Et 3 N (250 ml, 1.79 mmol) in THF (10 ml) was stirred at room temperature overnight. The reaction mixture was diluted with HO20, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1 to 50:1, v/v) as eluent 30 to give methyl 4-[ 1-[4-[N '-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(3R)-pyrrolidinyloxy 356 WO 01/00206 PCT/US00/18079 ]benzoate (561 mg, 70%) as a white form. 'H-NMR (CDCI 3 ) 8 2.05-2.34 (min, 2 H), 3.59-4.07 (series of s and m, total 12 H), 4.97-5.02 (min, 1 H), 6.75-6.86 (min, 4 H), 6.94-7.00 (min, 1 H), 7.22 7.33 (min, 2H), 7.59-7.66 (min, 2H), 7.92-7.99 (min, 3H), 8.19-8.22 (min, 1H); MS (ESI) m/z 538 (M+1). To a stirred solution of methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl 5 acetyl]-(3R)-pyrrolidinyloxy]benzoate (561 mg, 1.04 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 5 hr. After cooled to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was collected under a reduced pressure. The crude solid was recrystallized from MeOH-CHC 3 -Et 2 O to give 186 (361 mg, 66%) as a white crystalline powder. MW 523.97 mp 193-194oC; 'H-NMR (DMSO-d,) 8 10 2.06-2.28 (min, 2 H), 3.58-3.62 (min, 4 H), 3.71-3.76 (min, 1 H), 3.83-3.89 (series of s and m, total 4 H), 5.12 and 5.20 (each m, total 1 H), 6.74-6.78 (min, 1 H), 6.90-6.91 (min, 1 H), 7.01-7.05 (mn, 3 H), 7.27-7.30 (min, 1 H), 7.43-7.45 (min, 1 H), 7.88-7.98 (min, 3 H), 8.08-8.10 (min, 1 H), 8.89-8.96 (min, 2 H), 12.67 (br s, 1 H); MS (ESI) m/z 524 (M++1); Anal. Calcd for C 27
H
26 C1N 3 0 6 1/4H 2 0: C, 61.36; H, 5.05; N, 7.95; Cl, 6.71. Found: C, 61.49; H, 5.11; N, 7.72; Cl, 7.08. 15 Example 179 4-[ 1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-(3R)-pyrrolidinyloxy]benzoic acid ,,-COOH N N rH H Me 187 A mixture of 4-[N'-(2-bromophenyl)ureidol-3-methoxyphenylacetic acid (460 rmg, 1.21 mmol), methyl 4-[(3R)-pyrrolidinyloxy]benzoate (269 rmg, 1.21 mmol), EDCHC1 (280 rmg, 1.46 mmol), 20 HOBt (197 rmg, 1.46 mmol), and Et 3 N (205 ml, 1.47 rmmol) in THF (10 ml) was stirred at room temperature overnight. The reaction mixture was diluted with H20, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1 to 60:1, v/v) as eluent to give methyl 4-[ 1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-(3R)-pyrrolidinyloxy] 25 benzoate (555 mg, 78%) as a white foam. 'H-NMR (CDC13) 8 2.05-2.33 (mn, 2 H), 3.60-4.07 (series of s and min, total 12 H), 4.96-5.01 (min, 1 H), 6.76-6.93 (mn, 5 H), 7.28-7.30 (mn, 1 H), 7.48 7.58 (min, 3 H), 7.92-7.99 (min, 3 H), 8.12-8.16 (min, 1 H); MS (ESI) m/z 582 (M). To a stirred solution of methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-(3R) pyrrolidinyloxy]benzoate (555 mg, 0.95 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and 357 WO 01/00206 PCT/US00/18079 the reaction mixture was heated under reflux for 5 hr. After cooled to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was collected under a reduced pressure. The crude solid was recrystallized from MeOH-CHCl 3 -Et 2 O to give 187 (330 mg, 61%) as a white crystalline powder. MW 568.42 mp 175-177oC; 'H-NMR (DMSO-d 6 ) 8 1.99-2.27 (m, 2 5 H), 3.56-3.63 (m, 4 H), 3.71-3.76 (m, 1 H), 3.83-3.89 (series of s and m, 4 H), 5.12 and 5.20 (each m, total 1 H), 6.74-6.78 (m, 1 H), 6.89-7.05 (m, 4 H), 7.30-7.35 (m, 1 H), 7.59-7.61 (m, 1 H), 7.88-7.98 (m, 4 H), 8.74-8.76 (m, 1 H), 8.92-8.95 (m, 1 H), 12.68 (br s, 1 H); MS (ESI) m/z 569 (M +1); Anal Calcd for C 27
H
26 BrN 3 06: C, 57.05; H, 4.61; N, 7.39; Br, 14.06. Found: C, 56.91; H, 4.66; N, 7.20; Br, 14.59. 10 Example 180 N N CO 2 H O H H 3 HN N 188 1 gram of Tentagel PHB resin (loading 0.29 mmol/gin) was taken up in DMF 25 mL and Fmoc-(4-carboxymethyl)-piperidine (318 mg, 0.87 mmol) was added. The resin was shaken for 5 min then DIC (220 mg, 0.27 mL, 1.74 mmol) and DMAP (106 mg, 0.87 mmol) were added and 15 the resin was shaken for 24 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 Cl 2 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was taken up in 20% piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x),
CH
3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a positive 20 bromophenylblue test. The resin was taken up in 25 mL of DMF and Fmoc-L- morpholine-2 carboxylic acid (307 mg, 0.87 mmol) was added. The resin was shaken for 5 min then PyBroP (406 mg, 0.87 mmol) and DIEA (123 mg, 0.15 mL, 0.87 mmol) were added and the resin was shaken for 24 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 C1 2 (3x) then 25 dried under vacuum. The resin gave a negative bromophenylblue test. The resin was taken up in 20% piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x),
CH
3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a positive 358 WO 01/00206 PCT/US00/18079 bromophenylblue test. The resin was taken up in 25 mL of DMF and 4-o-tolylureidophenylacetic acid (247 mg, 0.87 mmol) was added and the resin was shaken for 5 min. PyBroP (406 mg, 0.87 mmol) and DIEA (123 mg, 0.15 mL, 0.87 mmol) was added and the resin was shaken for 24 hr. The resin was drained and washed with DMF (3x), CHzOH (3x) and CH 2 Cl 2 (3x) then 5 dried under vacuum. The resin gave a negative bromophenylblue test. The resin was then taken up in 90% TFA in CH 2 C1 2 and shaken for 4 hr. The resin was drained and the eluate collected. The resin was taken up in fresh CH 2 Cl 2 and shaken for 30 min. The resin was drained and the eluate collected and combined with the first fraction. The solvent was removed under vacuum and the residue was recrystallized from ethyl acetate-hexane, yielding 85 mg 188. 10 Example 181 CO 2 H H H 3 HN N O 189 1 gram of Tentagel PHB resin (loading 0.29 mmol/gm) was taken up in DMF 25 mL and Fmoc-(4-carboxymethy)-piperidine (318 mg, 0.87 mmol) was added. The resin was shaken for 5 min then DIC (220 mg, 0.27 mL, 1.74 mmol) and DMAP (106 mg, 0.87 mmol) were added and 15 the resin was shaken for 24 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was taken up in 20% piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x),
CH
3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a positive 20 bromophenylblue test. The resin was taken up in 25 mL of DMF and Fmoc-L- 4-phenylproline (307 mg, 0.87 mmol) was added. The resin was shaken for 5 min then PyBroP (406 mg, 0.87 mmol) and DIEA (123 mg, 0.15 mL, 0.87 mmol) were added and the resin was shaken for 24 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was taken up in 25 20% piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x),
CH
3 OH (3x) and CH 2 Cl 2 (3x) then dried under vacuum. The resin gave a positive 359 WO 01/00206 PCT/US00/18079 bromophenylblue test. The resin was taken up in 25mL of DMF and 4-o-tolylureidophenylacetic acid (247 mg, 0.87 mmol) was added and the resin was shaken for 5 min. PyBroP (406 mg, 0.87 mmol) and DIEA (123 mg, 0.15 mL, 0.87 mmol) was added and the resin shaken for 24 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 Cl 2 (3x) then 5 dried under vacuum. The resin gave a negative bromophenylblue test. The resin was then taken up in 90% TFA in CH 2 C1 2 and shaken for 4 hr. The resin was drained and the eluate collected. The resin was taken up in fresh CH 2 C1 2 and shaken for 30 min. The resin was drained and the eluate collected and combined with the first fraction. The solvent was removed under vacuum and the residue was recrystallized from ethyl acetate-hexane, yielding 82 mg 189. 10 Example 182 rKN'CO 2 H N NJ) 0 H H 3 HNyN 0 190 1 gram of Tentagel PHB resin (loading 0.29 mmol/gm) was taken up in DMF 25 mL and Fmoc-4-carboxymethyl-piperazine (318 mg, 0.87 mmol) was added. The resin was shaken for 5 min DIC (220 mg, 0.27 mL, 1.74 mmol) and DMAP (106 mg, 0.87 mmol) were added and the 15 resin was shaken for 24 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was taken up in 20% piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x), CH30H (3x) and CH 2
CI
2 (3x) then dried under vacuum. The resin gave a positive 20 bromophenylblue test. The resin was taken up in 25 mL of DMF and Fmoc -L-proline (294 mg, 0.87 mmol) was added. The resin was shaken for 5 min. then PyBroP (406 mg, 0.87 mmol) and DIEA (123 mg, 0.15 mL, 0.87 mmol) were added and the resin was shaken for 24 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was taken up in 25 20% piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x),
CH
3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a positive 360 WO 01/00206 PCT/US00/18079 bromophenylblue test. The resin was taken up in 25 mL of DMF and 4-o-tolylureidophenyl acetic acid (247 mg, 0.87 mmol) was added and the resin was shaken for 5 min. PyBroP (406 mg, 0.87 mmol) and DIEA (123 mg, 0.15 mL, 0.87 mmol) was added and the resin was shaken for 24 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 C1 2 (3x) then 5 dried under vacuum. The resin gave a negative bromophenylblue test. The resin was then taken up in 90% TFA in CH 2 C1 2 and shaken for 4 hr. The resin was drained and the eluate collected. The resin was taken up in fresh CH 2 C1 2 and shaken for 30 min. The resin was drained and the eluate collected and combined with the first fraction. The solvent was removed under vacuum and the residue was recrystallized from ethyl acetate-hexane, yielding 78 mg 190. 10 Example 183 CO 2 H H H 3 HNyN S N191 1 gram of Tentagel PHB resin (loading 0.29 mmol/gm) was taken up in DMF 25 mL and Fmoc-isonipecotic acid (306 mg, 0.87 mmol) was added. The resin was shaken for5 min then DIC (220 mg, 0.27 mL, 1.74 mmol) and DMAP (106 mg, 0.87 mmol) were added and the resin was 15 shaken for 24 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was taken up in 20% piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x),
CH
3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a positive 20 bromophenylblue test. The resin was taken up in 25 mL of DMF and Fmoc-L- proline (294 mg, 0.87 mmol) was added. The resin was shaken for 5 min then PyBroP (406 mg, 0.87 mmol) and DIEA (123 mg, 0.15 mL, 0.87 mmol) were added and the resin was shaken for 24 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 C 2 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was taken up in 25 20% piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x),
CH
3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a positive 361 WO 01/00206 PCT/US00/18079 bromophenylblue test. The resin was taken up in 25 mL of DMF and 4-o-tolylureidophenyl acetic acid (247 mg, 0.87 mmol) was added and the resin was shaken for 5 min. PyBroP (406 mg, 0.87 mmol) and DIEA (123 mg, 0.15 mL, 0.87 mmol) was added and the resin was shaken for 24 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 C1 2 (3x) then 5 dried under vacuum. The resin gave a negative bromophenylblue test. The resin was then taken up in 90% TFA in CH 2 C1 2 and shaken for 4 hr. The resin was drained and the eluate collected. The resin was taken up in fresh CH 2 Cl 2 and shaken for 30 min. The resin was drained and the eluate collected and combined with the first fraction. The solvent was removed under vacuum and the residue was recrystallized from ethyl acetate-hexane, yielding 73 mg 191. 10 Example 184 1 NrgCO 2 H C ,H H 3 H N 192 1 gram of Tentagel PHB resin (loading 0.29 mmol/gin) was taken up in DMF 25 mL and Fmoc-(4-carboxymethy)-piperidine (318 mg, 0.87 mmol) was added. The resin was shaken for 5 min then DIC (220 mg, 0.27 mL, 1.74 mmol) and DMAP (106 mg, 0.87 mmol) were added and 15 the resin was shaken for 24 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2
CI
2 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was taken up in 20% piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x),
CH
3 OH (3x) and CH 2 Cl 2 (3x) then dried under vacuum. The resin gave a positive 20 bromophenylblue test. The resin was taken up in 25 mL of DMF and Fmoc-L- proline (294 mg, 0.87 mmol) was added. The resin was shaken for 5 min. then PyBroP (406 mg, 0.87 mmol) and DIEA (123 mg, 0.15 mL, 0.87 mmol) were added and the resin was shaken for 24 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was taken up in 25 20% piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x), CH30H (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a positive 362 WO 01/00206 PCT/US00/18079 bromophenylblue test. The resin was taken up in 25 mL of DMF and Fmoc-3-amino-2-oxo-1 pyrrolidineacetate (331 mg, 0.87 mmol) was added and the resin was shaken for 5 min. PyBroP (406 mg, 0.87 mmol) and DIEA (123 mg, 0.15 mL, 0.87 mmol) was added and the resin was shaken for 24 hr. 5 The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2
CI
2 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was taken up in 20% piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x),
CH
3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a positive bromophenylblue test. The resin was taken up in CH 2 Cl 2 and 0-tolyl isocyanate (193 mg, 10 1.45 mmol, 0.18 mL) was added. The resin was shaken for 24 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 Cl 2 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was then taken up in 90% TFA in CH 2
CI
2 and shaken for 4 hr. The resin was drained and the eluate collected. The resin was taken up in fresh CH 2 Cl 2 and shaken for 30 min. The resin was drained and the 15 eluate collected and combined with the first fraction. The solvent was removed under vacuum and the residue was recrystallized from ethyl acetate-hexane, yielding 69 mg 192. Example 185 NH OH H H 193 In a 250 mL round-bottomed flask was placed o-tolylisothiocyanate (10.0 g, 67.1 mmol) 20 in 150 mL of CH 2 C1 2 . This solution was cooled to minus 78 0 C and ammonia gas (excess) was bubbled through for 10 min. A precipitate immediately formed and was found to be the desired product o-tolylthiourea. The reaction mixture was filtered and the solid collected by washing thoroughly with cooled CH 2
CI
2 . The white solid was dried under vacuum to provide 10.12 g (92% yield) of the desired o-tolylthiourea. 25 The o-tolylthiourea (10.12 g, 61 mmol) was then methylated by addition of methyl iodide (9.1 g, 62 mmol) in anhydrous methanol (100 mL). The reaction was stirred at room temp for 6 hr and then concentrated in vacuo. The residue was poured into aqueous ammonium chloride and extracted 3x with EtOAc. The combined organics were dried and concentrated in vacuo to give 8.7 g (84% yield) of 2-methyl-2-thio-o-tolylpseudourea. The pseudourea (8.7 g, 51 mmol) was 30 dissolved in methanol (100 mL) and piperidine (8.7 g, 102 mmol) at room temp. The mixture was 363 WO 01/00206 PCT/US00/18079 stirred overnight and then concentrated in vacuo to afford 9.2 g of the product ester as a pale yellow solid. This solid was saponified with LiOH to give 9.0 g of the desired final carboxylic acid 193. Example 186 N OH 5 H H 194 Methyl-4-aminophenylacetate (4.0 g, 25 mmol) was dissolved in CH 2
CI
2 (100 mL) and to this solution was added o-tolylisothiocyante (3.7 g, 25 mmol). The reaction mixture was heated to reflux for 4 hr and then cooled to room temp. The solution was poured in to lN HCI and then extracted 3x with EtOAc, dried over MgSO 4 , and concentrated in vacuo to afford 5.2 g (67% yield) 10 of thiourea methyl ester. The ester was saponified using LiOH to give 5.0 g of the desired 4-(o tolylthioureido)phenylacetic acid 194. Example 187 H N OH N N COCH Me H H Me 195 A solution of ethyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetyl]-(2S, 3R, 15 4R)-3,4-isopropylidenedioxy-2-pyrrolidinylcarbonyl]-1-piperazinylacetate (1.27 g, 1.99 mmol) in sat. HCI (gas)- MeOH (20 mL) was stirred at room temp. for 2 hr, and MeOH was evaporated off. The residue was taken up with sat. NaHCO 3 solution, and extracted with CHCl 3 - MeOH (4 : 1, v/v). The extracts were washed with brine, dried over MgSO 4 , and concentrated to dryness. Chromatography of the residue with CHCI 3 - MeOH (5 : 1, v/v) as eluent gave ethyl 4-[1-[3 20 methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyll-(2S, 3R, 4R)-3,4-dihydroxy-2-pyrrolidinyl carbonyl]-1-piperazinylacetate (990 mg, 83 %) as a yellow amorphous solid. IR (KBr) 3338, 2937, 2830, 1743, 1625, 1600, 1532, 1454 cmr'; 'H-NMR (CDC 3 ) d 1.25 (t,J = 7.1 Hz, 3H), 2.20 (s, 3H), 2.46 - 2.56 (m, 4H), 3.15 (s, 2H), 3.40 - 3.72 (m, 9H), 3.62 (s, 3H), 4.01 - 4.08 (m, 2H), 4.16 (q, J= 7.1 Hz, 2H), 4.22 (m, 1H), 4.72 (d, J- 2.9 Hz, 111), 6.68 (d, J= 7.6 Hz, 111), 6.72 (s, 25 1H), 7.06 (t, J= 7.6 Hz, 1H), 7.15 -7.18 (m, 3H), 7.52 -7.56 (m, 2H), 7.90 (d, J= 8.3 Hz, 1H1); MS (FAB) m/z 598 (M'+1). To a solution of ethyl 4-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(2S, 3R, 4R)-3,4-dihydroxy-2-pyrrolidinylcarbonyl]-1-piperazinylacetate (870 mg, 1.46 mmol) in THF (15 mL) was added 0.25N NaOH (7.00 mL, 1.75 mmol). After being stirred at room temp. for 3.5 364 WO 01/00206 PCT/US00/18079 hr, the reaction mixture was concentrated. The residue was diluted with water and neutralized with IN HC1 at 0 oC. The mixture was concentrated and purified by ion-exchanged resin (HP-20, Mitsubishi Chemical) to give 195 (645 mg, 78 %) as a colorless amorphous solid. IR (KBr) 3330, 2937, 1627, 1535, 1454 cm'; 'H NMR (DMSO-d 6 ) d 2.25 (s, 3H), 2.38 (m, 1H), 2.42 - 2.58 (m, 5 2H), 2.64 (m, 1H), 3.01 (s, 2H), 3.13 - 3.71 (m, 8H), 3.88 (s, 3H), 3.89 (m, 1H), 4.05 (m, 1H), 4. 58 (d, J= 3.2 Hz, 1H), 6.76 (dd, J= 8.3, 1.5 Hz, 1H), 6.91 - 6.95 (m, 2H), 7.10 - 7.16 (m, 2H), 7.79 (d, J= 8.3 Hz, 1H), 8.00 (d, J= 8.3 Hz, 1H), 8.49 (s, 1H), 8.57 (s, 1H); MS (FAB) m/z 570 (M+1); Anal. Called for C 28
H
35
N
5 0 8 -2.75H 2 0: C, 54.32; H, 6.59; N, 11.31. Found: C, 54.07; H, 6.11; N, 11.00. 10 Example 188 N N'tN 4 H 0 OH 196 To a suspension of 2-amino-4-thiazoleacetic acid (4 g, 25 mmol) in 1:1 CH 2 Cl:acetone (100 mL) was added o-tolylisocyanate (3.5 g, 26 mmol). The mixture was heated to reflux for 8 hr at which time a yellow precipitate had formed. the precipitate was filtered and the solid washed 15 generously with 1:1 CH 2 Cl 2 :acetone. The solid was recrystallized with hot methanol and dried under vaccum to yield 4.8 g (66% yield) of the desired 2-(o-tolylureido)-4-thiazoleacetic acid 196. Example 189 BocHN Br 197 197 In a round bottom flask, 3-bromobenzyl amine (3.00g, 16.13mmole) was dissolved in 20 dioxane-water (1:1) and solid Na 2
CO
3 was added till the pH was 8-9. Boc 2 0 (3.87g, 17.74mmole) was added and the reaction was stirred for 12 hr at room temp. The reaction mixture was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were then washed with water, brine then dried over anhydrous MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was purified by flash 25 chromatography. (4:1 hexane-ethyl acetate) Yield 4.39g 197. Boc, N Br
CH
3 198 The Boc-protected benzyl amine (2.00g, 6.99mmole) was dissolved in dry THF under argon. The reaction was cooled to minus 780 C. Lithium bis(trimethylsilyl)amide (13.98 mL, 13.98mmole) was added over 10 min. The reaction was stirred for one hr at minus 780 C then 30 iodomethane (1.98g, 13.98mmole, 0.87 mL) was added rapidly. The reaction was allowed to slowly warm to room temp and stir overnight. The reaction was poured into IN HCI and the 365 WO 01/00206 PCT/US00/18079 aqueous layer was extracted 3x with ethyl acetate. The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (7:1 hexane ethyl acetate) Yield 1.68g 198. Boc'N CO 2
CH
3
CH
3 199 5 In a pressure tube was placed the Boc-protected 3-bromobenzyl methylamine (1.68g, 5.60mmole). The tube was then charged with DMF, sodium acetate (0.5 1g 6.16mmole), P(o tolyl) 3 (0.5 Ig, 6.16nummole), and Pd(OAc) 2 (0.25g, 1.12mmole) The tube was flushed with argon for 10 min then methyl acrylate (0.53g, 0.53mmole, 0.55mL) was added. The tube was sealed and heated to 1350 C for 24 hr. The reaction was cooled to 0oC and the tube was slowly opened. The 10 solution was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (6:1 hexane-ethyl acetate) Yield 1.60g 199. Boc'N.N i CO 2
CH
3 CH3 C 3
CH
3 - CH 3 200 15 In a pressure tube was placed the Boc-protected 3-bromobenzyl methylamine (1.00g, 3.33mmole). The tube was then charged with DMF, sodium acetate (0.30g, 3.36mmole), P(o tolyl) 3 (0.20, 0.66mmole), and Pd(OAc) 2 (0.15g, 0.66mmole). The tube was flushed with argon for 10 min then methyl methacrylate (.37g, 3.66mmole, 0.39mL) was added. The tube was sealed and heated to 1350 C for 24 hr. The reaction was cooled to 00 C and the tube was slowly opened. The 20 solution was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (6:1 hexane-ethyl acetate) Yield 1.01g 200. BOC'N CO 2
CH
3
CH
3 201 25 The a, p-unsaturated ester (1.60g, 5.49mmole) was placed in a Paar vessel and dissolved in ethyl acetate. Pd/C (0.3g) was added and the vessel was pressured to 50 psi with H 2 . The vessel was agitated for 12 hr. The Paar vessel was flushed with argon and the catalyst was removed by filtration through celite. The solvent was removed under reduced pressure. Yield 1.60g 201. Boc'N CO 2
CH
3
CH
3
CH
3 202 30 The a-methyl-, P-unsaturated ester (1.0 lg, 3.16mmole) was placed in a Paar vessel and 366 WO 01/00206 PCT/US00/18079 dissolved in ethyl acetate. Pd/C was added and the vessel was pressured to 50 psi with H 2 . The vessel was agitated for 12 hr. The Paar vessel was flushed with argon and the catalyst was removed by filtration through celite. The solvent was removed under reduced pressure. Yield 996.41mg 202. 3 H H N
CO
2
CH
3 5 203 The Boc ester (304mg, 1.04mmole) was taken up in CH 2 Cl 2 and excess trifluoroacetic acid was added. The reaction was then stirred for 2 hr. The solvent was removed and the residue was taken up in ethyl acetate and washed with sat. NaHCO 3 solution. The organic layer was washed with water, brine then dried over Na 2
SO
4 . The solution was filtered and the solvent was 10 removed under reduced pressure. The residue was taken up in CH 2 Cl 2 -DMF and HOBt (154.30mg, 1.14mmole), 4-[N'-(o-tolylurea)-phenylacetic acid (324.11mg, 1.14mmole) and EDCI (218.53mg, 1.14mmole) were added. The reaction was stirred for 24 hr. The solution was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the 15 solvent was removed under reduced pressure. The product was isolated by flash chromatography. (ethyl acetate) Yield 380.32mg 203. H H N N
CO
2 H
CH
3 204 The ester (380.32mg, 0.80mmole) was taken up in ethanol-water (4:1) and NaOH was added. The reaction was then heated to 500 C for 2 hr. The TLC (ethyl acetate) showed no 20 starting material present. The reaction was cooled to room temp. The solution was poured into IN HCI and the aqueous layer was extracted 3x ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The residue was recrystallizied from ethyl acetate-hexane. Yield 319.40mg 204. 25 Example 190 H H y No 00 N
CO
2
CH
3
CH
3 0 CH 3 205 367 WO 01/00206 PCT/US00/18079 The Boc ester (209.60, 0.65mmole) was taken up in CH 2 Cl 2 and excess trifluoroacetic acid was added. The reaction was then stirred for 2 hr. The solvent was removed and the residue was taken up in ethyl acetate and washed with sat. NaHICO 3 solution. The organic layer was washed with water, brine then dried over Na 2
SO
4 . The solution was filtered and the solvent was 5 removed under reduced pressure. The residue was taken up in CH 2 Cl 2 -DMF and HOBt (97.45mg, 0.72mmole), 4-[N'-(o-tolylurea)-phenylacetic acid (204.70mg, 0.72mmole) and EDCI (138.03mg, 0.72mmole) were added. The reaction was stirred for 24 hr. The solution was poured into IN HC1 and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was 10 removed under reduced pressure. The product was isolated by flash chromatography. (ethyl acetate) Yield 237.8mg 205. 3 H H SN C0 2 H
CH
3
CH
3 6 The ester (237.8mg, 0.49mmole) was taken up in ethanol-water (4:1) and NaOH added. The reaction was heated to 50 C for 2 hr. The TLC (ethyl acetate) showed no starting material 15 present. The reaction was cooled to room temp. The solution was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The residue was recrystalized from ethyl acetate-hexane. Yield 207.8mg 206. Example 191 0 2
N
NH 20 207 Thel1,2,3,4- tetrahydroisoquinoline (12.20g, 91.60mmole) was taken up in H 2 SO4(40mL) and cooled to minus 100 C. Concentrated HNO 3 (9.0mL) was slowly added to the solution while maintaining the internal temp at minus 100 C. On completion of the addition the reaction was allowed to stand and slowly warm to room temp over 12 hr. The reaction mixture was slowly 25 added to ice and the aqueous solution was basified with NH 4 OH. The aqueous layer was extracted 4 times with CHC 3 . The combined organic layers were washed with water then dried over Na 2
SO
4 . The solution was filtered and the solvent was removed under reduced pressure. The resulting brown oil was taken up in ethanol and concentrated HCI was added. The resulting white solid was collected by filtration and dried under vacuum. Yield 8.0g 207. 0 2 N N CO 2
CH
3 30 208 368 WO 01/00206 PCT/US00/18079 The 6- nitro-1,2,3,4-tetrahydroisoquinoline (1.00g, 5.61mmole) was taken up in ethanol. Methyl bromoacetate (0.86g, 5.61mmole, 0.53 mL) and triethylamine (1.17g, 11.59mmole, 1.62 mL) were then added and the mixture was heated to reflux for 5 hr. The solution was cooled to room temp and the solution was concentrated under vacuum. The solution was added to water and 5 the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were dried over Na 2
SO
4 , filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography (3:1 hexane-ethyl acetate). Yield 702mg 208.
H
2 N
N"CO
2
CH
3 209 209 The above ester (702mg, 2.81mmole)was placed in a Paar vessel and dissolved in 10 ethanol. Pd/C (100mg) was added and the vessel was pressured to 50 psi with H 2 . The vessel was agitated for 24 hr. The Paar vessel was flushed with argon and the catalyst was removed by filtration through celite. The solvent was removed under reduced pressure. 1H-NMR showed only desired product. Yield 587mg 209. H 0 N N -:-- N CO2CH3 NH H
CH
3
OCH
3 210 15 The aniline (587.0mg, 2.66mmole) was taken up in dry CH 2
CI
2 and pyridine under argon. The reaction was cooled to 0OC. A CH 2 C1 2 solution of 3-methoxy-4-(N'phenylureido) phenylacetyl chloride (837.70mg, 2.66mmole) was added over 5 min. The reaction was then allowed to warm to room temp and stir overnight. The reaction mixture was then poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were then washed 20 with sat. NaHCO 3 , water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography (ethyl acetate). Yield 618.36mg 210. H NH 0 N : N -CO 2 H
CH
3
OCH
3 211 The methyl ester (618.36mg, 1.20mmole) above was taken up in THF-H 2 0 and LiOH 25 (558.07mg, 13.30mmole) was added. The reaction mixture was stirred at room temp for 24 hr. The reaction was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were then washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was purified by recrystallization. (hexane-ethyl acetate). Yield 600mg 211. 369 WO 01/00206 PCT/US00/18079 Example 192 0 2 N 10"' OH ON "OH 212 The 3-nitro-phenyl propionic acid (1.00g, 5.12mmole) was taken up in dry THF under argon. The reaction was cooled to 00 C and BH 3 -THF (1.0M, 15.37mmole, 15.37 mL) was added 5 over 10 min. The reaction was stirred at 0o C for 1 hr then slowly quenched with water. The solution was slowly warmed to room temp then poured into IN HCI. The aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were then washed with sat. NaHCO 3 , water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography (1:1 hexane-ethyl acetate) 10 Yield 909.0mg 212. 0 2 N CHO 213 The 3-nitro-phenyl propanol (909.0mg, 5.02mmole) was taken up in dry CH 2
C
2 . In a second round bottom flask (COC1) 2 (700.65mg, 5.52mmole, 0.48 mL) was added to dry CH 2 C1 2 under argon. The (COCI) 2
-CH
2
CI
2 solution was then cooled to minus 600 C and DMSO 15 (862.56mg, 11.4mmole, 0.78 mL) was slowly added. The reaction was stirred at minus 600 C for 5 min then the alcohol solution was added via a cannula over 5 min. The reaction mixture was stirred at minus 600 C for 1 hr then Et3N (2.54g, 25.10mmole, 3.50 mL) was added and the reaction was allowed to slowly warm to room temp. The reaction was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with 20 sat. NaHCO 3 , water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. HI-NMR showed no starting material present. The aldehyde 213 was used as is without further purification. 0 2 N . . CO 2
CH
2
CH
3
C-
3 214 In a round bottom flask NaHl (132.48mg, 5.52mmole) was slurried in dry THF under 25 argon. Triethyl 2-phosphonopropionate (1.31 g, 5.52mmole, 1.18 mL) dissolve in dry THF was added slowly via a syringe. The reaction mixture was stirred for 30 min at room temp. The above aldehyde, dissolved in dry THF under argon, was added to the phosphonate solution via syringe over 10 min. The reaction mixture was stirred for 12 hr. The reaction was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were 30 washed with sat. NaHCO 3 , water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (1:1 ethyl acetate-hexane) Yield 992.0mg 214. 370 WO 01/00206 PCT/US00/18079
H
2 N TC0 2
CH
2
CH
3 / CH3
CH
3 215 The above a,P-unsaturated ester (992.0mg, 3.77mmole) was placed in a Paar vessel and dissolved in ethanol. The vessel was flushed with argon and Pd/C (200.0mg) was added. The argon atmosphere was replaced with H 2 at 50 psi. The Paar vessel was then shaken for 12 hr. The 5 hydrogen was flushed from the vessel with argon and the catalyst was removed by filtration through celite. The solvent was removed under reduced pressure. 1H-NMR showed only the desired product. Yield 851.2mg 215. H N N CO 2
CH
2
CH
3 N N.LN 0 H 3 ,:? H H
CH
3
OCH
3 216 The above aniline (850.0mg, 3.61mmole) was taken up in dry CH 2
CI
2 and pyridine under 10 argon. The reaction was cooled to 0o C. A CH 2
C
2 solution of 3-methoxy-4-(N'phenylureido) phenylacetyl chloride (1.14g, 3.61mmole) was added over 5 min. The reaction was then allowed to warm to room temp and stir overnight. The reaction mixture was then poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were then washed with sat. NaHCO 3 , water, brine then dried over MgSO 4 . The solution was filtered and the 15 solvent was removed under reduced pressure. The product was isolated by flash chromatography (ethyl acetate) Yield 576.0mg 216. H N NN 0 CO 2 H N .. JN.(: 0 Cr (H 3 P H H 2.
CH
3
OCH
3 217 The above ethyl ester (576.0mg, mmole) was taken up in ethanol-water and NaOH was added. The reaction mixture was heated to 500 C for 2 hr. The reaction was cooled to room temp 20 and then poured into iN HCL. The aqueous layer was extracted 3x times with ethyl acetate. The combined organic layers were then washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was purified by Sep-Pak column. Yield 534mg 217. Example 193 0 0
H
3 C ' N 25 "218 One gram of Wang resin (tentagel S-PHB 0.3mmole loading) was suspended in a solution 371 WO 01/00206 PCT/US00/18079 of 3-(Fmoc-amino)phenylpropionic acid (361.3 1mg, 0.90mmole), DMAP (109.95mg, 0.90mmole), HOBt 243.63mg, 0.90mmole), and DIC (227.16mg, 1.80mmole, 0.28 mL) in a mixture of DMF and CH 2 C1 2 . The mixture was shaken for 20 hr and drained. The resin 218 was washed with DMF, MeOH, CH 2 C1 2 and dried under reduced pressure. H H CH 3 0 0 d TN N HO N OC Y-
CH
3 013 5 CH 3 219 To the above resin (500mg, 0.15mmole) was added a solution of piperidine-DMF (50% v/v, 4 mL) and the mixture was shaken for 4 hr. The resin was washed with DMF, MeOH,
CH
2
C
2 . To the resin was added TMOF and isobutrylaldehyde (108.17mg, 1.50mmole, 0.14 mL). The mixture was shaken for 4 hr. The resin was drained and fresh TMOF and isobutrylaldehyde 10 was added. The mixture was then shaken for 12 hr. The resin was drained and taken up in MeOH-1% AcOH and NaCNBH 3 (150.0mg, 2.39mmole) was added. The resin was shaken for 6 hr. The resin was drained and washed with MeOH, MeOH-Et 3 N, MeOH, DMF, CH 2 C1 2 . The resin was taken up in DMF and 3-methoxy-4-(N'-phenylureido)phenylacetic acid (141.45mg,0.45mmole), PyBrop (209.78mg, 0.45mmole),and DIEA (58.16mg, 0.45mmole, 0.08 15 mL) were added. The resin was then shaken for 24 hr then drained. The resin was washed with DMF, MeOH, CH 2 C1 2 . To the resin was added a solution of TFA in CH 2 C1 2 (30% v/v 3 mL) and the mixture was shaken for 5 hr. The mixture was filtered and the filtrate was concentrated in vacuo. The residue was purified by Sep-Pak column. After removal of the solvent, Et 2 0 was added to the residue and the solid was collected to afford 15mg 219 as a crystalline solid 20 Example 194 OH <OH 0 N C0 2 H /N ),N\ OP
CH
3 H H OCH 3 220 Tentagel PHB resin (1.0 g, loading 0.29mmole/gm) was taken up in 25 mL of DMF and 6-bromohexanoic acid (169mg, 0.87mmol) was added. The resin was shaken for 5 min then DIC (220mg, 0.27 mL, 1.74mmoles) and DMAP (35mg, 0.29mmole) were added and the resin was 25 shaken for 14 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 Cl 2 (3x) then dried under vacuum. 372 WO 01/00206 PCT/US00/18079 To this resin was added 2,2-dimethyl-1,3-dioxolane-4-methanamine (227mg, 1.74 mmol)and lithium iodide (232mg, 1.74 mmol) in 15 mL of DMF. The resin was shaken for 14 hr at room temp. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a positive bromophenylblue test. 5 The resin was taken up in 25 mL of DMF and 4-o-tolylureido-3-methoxyphenylacetic acid (247mg, 0.87mmole) was added and the resin was shaken for 5 min. PyBrOP (406mg, 0.87mmole) and DIEA (123mg, 0.15 mL, 0.87mmole) was added and the resin was shaken for 14 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 C0 2 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. 10 The resin was then taken up in 90% TFA in CH 2 C1 2 and shaken for 4 hr. The resin was drained and the elutant collected. The resin was taken up in fresh CH 2 C1 2 and shaken for 30 min. The resin was drained and the elutant collected and combined with the first fraction. The solvent was removed under vacuum and the residue was recrystallized from ethyl acetate-hexane, yield 56mg 220. 15 Example 195 HN - CO2H 0
H
3 CO H CH 3 HN 221 1 gram of Tentagel PHB resin (loading 0.29mmole/gm) was taken up in DMF 25 mL and Fmoc- 7-aminoheptanoic acid (319mg, 0.87mmole) was added. The resin was shaken for 5 min then DIC (220mg, 0.27 mL, 1.74 mmol) and DMAP (106mg, 0.87mmole) were added and the 20 resin was shaken for 24 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and
CH
2 C1 2 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was taken up in 20% piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a positive bromophenylblue test. 25 The resin was taken up in 25 mL of DMF and 4-o-tolylureido-3-methoxyphenylacetic acid (247mg, 0.87mmole) was added and the resin was shaken for 5 min. PyBrOP (406mg, 0.87mmole) and DIEA (123mg, 0.15 mL, 0.87mmole) was added and the resin was shaken for 14 373 WO 01/00206 PCT/US00/18079 hr. The resin was drained and washed with DMF (3x), CH 3 OH (3x) and CH 2 C1 2 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was then taken up in 90% TFA in CH 2 C1 2 and shaken for 4 hr. The resin was drained and the elutant collected. The resin was taken up in fresh CH 2
CI
2 and shaken for 30 rmin. 5 The resin was drained and the elutant collected and combined with the first fraction. The solvent was removed under vacuum and the residue was recrystallized from ethyl acetate-hexane, yield 66mg 221. Example 196 N N H H3
H
3 H H
CH
3 o OH 222 10 To a solution of oxalyl chloride (3.8 g, 30 mmol) in CH 2 C1 2 (100 mL) was added DMSO (2.4 g, 31 mmol) dropwise over 30 min at minus 780 C. To this solution was added N-Boc prolinol (5.0g, 25mmol) dropwise over 15 min. The reaction was stirred at minus 780 C for 3 hr and then quenched by the cold addition of IN HC1, extracted 3x with EtOAc, dried, and concentrated in vacuo to afford the crude prolinal which was chromatographed (25% 15 EtOAc/hexanes) to yield 3.8 g of the desired product. A solution of Methyl (triphenylphosphoranylidene)butanoate (6.9 g, 19 mmol) in THF (100 mL) was generated. LiHMDS (10 mL of a 2.0M soln, 20 mmol) was added at minus 78o C and then stirred for 1 hr. The above prolinal (3.8 g, 19 mmol) was then added in one portion and the mixture was allowed to warm to room temp over 4 hr. The reaction was quenched by the 20 addition of IN HCI, extracted 3x with EtOAc, dried, and concentrated in vacuo to afford the crude alkene which was chromatographed (25% EtOAc/hexanes) to yield 2.9 g of he desired product. Hydrogenation of the alkene was performed by placing the alkene (2.9 g, 10 mmol) in ethanol (20 mL) and adding a catalytic amount of 10% Pd/C followed by Parr hydrogenation at 40 psi for 4 hr, the resulting alkane was used without purification. The Boc group was removed by 25 the addition of 1:1 TFA/CH 2
C
2 at room temp. The reaction was stirred for 2 hr and the solvent was removed in vacuo. The crude amine 1.9 g was used without further purification. A solution of the above free amine (1.9 g, 10 mmol) in CH 2 C1 2 (100 mL) was generated. 374 WO 01/00206 PCT/US00/18079 To this solution was added EDCI (2.95 g, 10 mmol), DMAP (1.2 g, 10 mmol), and 4-o tolylureido-3-methoxyphenylacetic acid (3.15g, 10 mmol) at room temp. The reaction mixture was stirred for 4 hr and then quenched by the addition of 1 N HCI, extracted 3x with EtOAc, dried, and concentrated in vacuo. The crude amide was chromatographed (5% MeOH/ CH 2 Cl 2 ) to 5 yield 1.95 g of the desired product. The ester (1.95 g, 4.2 mmol) was taken up in 1:1 THF-H 2 0 and LiOH was added at room temp. The reaction mixture was then stirred for 3 hr. The solution was poured into IN HCI and the aqueous layer was extracted 3x with EtOAc. The combine organic layers were washed with water, brine and dried over anhydrous magnesium sulfate. The solution was filtered and the 10 solvent removed under reduced pressure. The solid was then triturated with cold ether to give 1.65 g of the desired carboxylic acid 222. Example 197 C H 3 H3C, N--- OH OH 0 0
H
3 CO H CH HNN 0 223 To a solution of methyl 8-aminooctanoate (2.0 g, 12 mmol) in 1:1 dioxane:water (100 15 mL) was added Boc anhydride (2.8 g, 13 mmol) and K 2 C0 3 (10 g). This solution was allowed to stir at room temp for 14 hr. The reaction was then poured onto 1 N HC1, extracted 3x with EtOAc, dried, and concentrated in vacuo. The crude carbamnate was chromatographed (50% EtOAc/hexanes) to yield 2.7 g of the desired product. The Boc-protected amine was methylated by placing it in THF (75 mL), followed by the 20 addition of LiHMDS (25 mL of a 2.0M soln., 50 mmol) at minus 78o C, this solution was then stirred for 30 min and methyl iodide (7.2 g, 50 mmol) was added in one portion the reaction mixture was allowed to warm to room temp overnight. The reaction was quenched by the addition of 1 N HC1, extracted 3x with EtOAc, dried, and concentrated in vacuo. The crude methylated carbamate was chromatographed (50% EtOAc/hexanes) to yield 1.9 g of the desired dimethyl 25 product. The Boc group was removed by the addition of 1:1 TFA/CH 2 C1 2 at room temp. The reaction was stirred for 2 hr and the solvent was removed in vacuo. The crude amine 900 mg was 375 WO 01/00206 PCT/US00/18079 used without further purification. A solution of the above free amine (900 mg, 4.5 mmol) in CH 2
C
2 (100 mL) was generated. To this solution was added EDCI (1.33 g, 4.5 mmol), DMAP (567 mg, 4.5 mmol), and 4-o-tolylureido-3-methoxyphenylacetic acid (1.45 g, 4.6 mmol) at room temp. The reaction 5 mixture was stirred for 4 hr and then quenched by the addition of 1 N HCI, extracted 3x with EtOAc, dried, and concentrated in vacuo. The crude amide was chromatographed (5% MeOH/
CH
2 Cl 2 ) to yield 1.2 g of the desired product. The ester (1.2 g, 2.4 mmol) was taken up in 1:1 THF-H 2 0 and LiOH was added at room temp. The reaction mixture was then stirred for 3 hr. The solution was poured into IN HCI and 10 the aqueous layer was extracted 3x with EtOAc. The combine organic layers were washed with water, brine and dried over anhydrous magnesium sulfate. The solution was filtered and the solvent removed under reduced pressure. The solid was then triturated with cold ether to give 1.01 g of the desired carboxylic acid 223. Example 198 OH N N 15 CH 3 H 224 To a suspension of 4-aminophenylacetic acid (10 g, 66 mmol) in 1:1 CH 2 C1 2 :acetone (100 mL) was added o-tolyisocyanate (8.8 g, 66 mmol). The mixture was heated to reflux for 4 hr at which time a white precipitate had formed. The precipitate was filtered and the solid washed generously with 1:1 CH 2 C1 2 :acetone. The solid was recrystallized with hot methanol and dried 20 under vacuum to yield 14.1 g (75% yield) of the desired 4-(o-tolylureido)phenylacetic acid 224. Example 199
H
3 0 OH 225 To a suspension of 2-amino-4-thiazoleacetic acid (4 g, 25 mmol) in 1:1 CH 2 Cl:acetone (100 mL) was added o-tolylisocyanate (3.5 g, 26 mmol). The mixture was heated to reflux for 8 hr 25 at which time a yellow precipitate had formed. the precipitate was filtered and the solid washed generously with 1:1 CH 2 Cl 2 :acetone. The solid was recrystallized with hot methanol and dried under vacuum to yield 4.8 g (66% yield) of the desired 2-(o-tolylureido)-4-thiazoleacetic acid 225. 376 WO 01/00206 PCT/US00/18079 Example 200 NC \ Br 226 3-Bromo-4-hydroxybenzonitrile (5.00g, 25.25mmol) was taken up in DMF. Benzyl bromide (4.75g, 27.78mmol, 3.30mL) and Cs 2
CO
3 (16.45g, 50.50mmol) were added and the 5 reaction was heated to 500 C for 2 hr. The solution was cooled to room temperature and poured into 1NHCI. The aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (hexane to 8:1 hexane-ethyl acetate) Yield 8.90g 226. 0
H
3 C H Br
H
3 C \ 10 H 227 3-Bromo-4-benzyloxybenzonitrile (1.50g, 5.21mmol) was taken up in dry THF under argon and the solution was cooled to 0o C. BH 3 -THF (10.41mL, 10.41mmol) was added via syringe over 5 min. The reaction mixture was then warmed to room temp then heated to reflux for 12 hr. The solution was cooled to 00 C and methanol was slowly added. When no more gas 15 evolution was observed the solution was warmed to room temp and excess IN NaOH solution was added. Boc 2 0 (1.25g, 5.73mmol) was added and the reaction mixture was stirred at room temp for 12 hr. The solution was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was 20 isolated by flash chromatography. (7:1 ethyl acetate-hexane) Yield 1.80g 227 H3cH 3 N B r
H
3 C CH 3 228 The Boc-protected benzyl amine (l.80g, 4.59mmol) was dissolved in dry THF under argon. The reaction was cooled to minus 780 C. Lithium bis(trimethylsilyl)amide (13.77mL, 13.77mmol) was added over 10 min. The reaction was stirred for 1 hr at minus 780 C, then 25 iodomethane (1.95mL, 13.77mmol, 0.86mL) was added rapidly. The reaction was allowed to slowly warm to room temp and stir overnight. The reaction was poured into IN HCI and the 377 WO 01/00206 PCT/US00/18079 aqueous layer was extracted 3x with ethyl acetate. The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (7:1 hexane ethyl acetate) Yield 1.70g 228. HCO HH3 0
H
3 C CH 3 229 5 In a pressure tube was placed the 4-(N-methyl-Boc-aminomethyl)-2-bromobenzyloxy phenol (1.70g, 4.18mmol). The tube was then charged with DMF, sodium acetate (0.38g, 4.60mmol), dppp (0.35g, 0.84mmol), and Pd(OAc) 2 (0.19g, 0.84mmol) The tube was flushed with argon for 10 min and then methyl acrylate (0.40g, 4.60mmol, 0.41mL) was added. The tube was sealed and heated to 1350C for 24 hr. The reaction was cooled to 00 C and the tube was slowly 10 opened. The solution was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (6:1 hexane-ethyl acetate) Yield 1.12g 229
H
3 C,. CO 2 CH3 H3C H YH ( HC 230 15 The unsaturated ester (307.40mg, 0.75 mmol) was taken up in CH 2 Cl 2 and excess TFA was added. The reaction was stirred for 4 hr at room temp. The solvent was removed under reduced pressure and the residue was dried under high vacuum. The solvent was removed and the residue was taken up in ethyl acetate and washed with sat. NaHCO 3 solution. The organic layer was washed with water, brine then dried over Na 2
SO
4 . The solution was filtered and the solvent 20 was removed under reduced pressure. The residue was taken up in CH 2 C1 2 -DMF and HOBt ( 110.99 mg, 0.82 mmol), 3-methoxy-4-(N'-phenylureido) phenylacetic acid (258.31mg, 0.82 mmol) and EDCI (157.20mg, 0.82 mmol) were added. The reaction was stirred for 24 hr. The solution was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was 25 filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (ethyl acetate) Yield 296.30mg 230 378 WO 01/00206 PCT/US00/18079 H3C,
CQCH
3 H
H
3 C H CH3 231 The unsaturated ester (296.30mg, 0.49 mmol) was taken up in EtOAc and Pd/C (75mg) was added under argon. The argon atmosphere was replaced with hydrogen at 1 atmosphere and stirred for 24 hr. The hydrogen atmosphere was removed and replaced with argon. The catalyst 5 was removed by filtration through celite and the celite pad was washed with ethyl acetate 3x. The solvent was removed under reduced pressure. H'-NMR showed only the desired product. No further purification was needed. Yield 233.00mg 231
H
3 C," CO 2 H H3C H CH3 232 The ester (233.00mg, 0.45mmol) was taken up in THF-H 2 0 (4:1) and LiOH (94.41mg, 10 2.25mmol) was added. The reaction mixture was stirred at room temp for 24 hr. The solution was poured into 1N HCl and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was washed with ether-hexane (1:1) and dried under high vacuum. Yield 211.58mg 232 15 Example 201 H3 C ,N HaN 0 00
H
3 CO H CH3 HN N 233 The carboxylic acid (65.00mg, 0.13 mmol) was taken up in benzene and para toluenesulfonic acid (10.00mg, 0.06 mmol) was added. A Dean-Stark trap was added and the solution was heated to reflux for 24 hr. The reaction was cooled to room temp and poured into sat. 20 NaHCO 3 . The organic layer was seperated and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine and dried over MgSO 4 . The 379 WO 01/00206 PCT/US00/18079 solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography (4:1 hexane-ethyl acetate to ethyl acetate) Yield 29.00mg 233 Example 202 BrV CO 2 Et 234 5 5-Bromonicotinic acid (5.15g, 25.49 mmol ) was taken up in EtOH and H 2 SO4 (ImL) was added and the solution heated to reflux for 24 hr. The solution was cooled to rt and concentrated. The solution was then added to sat. NaHCO 3 and the aqueous layer were extracted 3x with Et 2 0. The combined organic layers were dried over Na 2
SO
4 , filtered and the solvent was removed under reduced pressure. The product was sufficiently pure for the next step. Yield 5.42g 234 Bryy H 10 235 The ethyl 5-Bromonicotinate (5.40g, 23.47 mmol) was taken up in 95% EtOH and NaBH 4 (8.31 g, 225.69 mmol) was added slowly at room temp. After addition the solution was stirred for 24 hr at room temp. Water was slowly added to the solution, then the mixture was stirred for 4 hr. The EtOH was removed under reduced pressure and the aqueous layer was extracted 3x with 15 CH 2
C
2 . The combined organic layers were dried over Na 2
SO
4 , filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (2:1 ethyl acetate-hexane) Yield 2.12g 235 Br - Cl HCI 236 The benzyl alcohol (2.12g, 11.28 mmol) was taken up Et 2 O and HCl) was bubbled 20 through the solution for 10 min. The solution was stirred at room temp for 1 hr and then the solid was collected by filtration. The solid was washed with Et 2 O and the then dried. The HCI salt was added to SOCl 2 and the mixture was heated to reflux for 1.5 hr. The solution was cooled to room temp and Et 2 O was added to precipitate the product. The solid was collected by filtration, washed with Et 2 O and dried under vacuum. Yield 2.42g 236 Br NCH3 25 HCI 237 The benzyl chloride (2.42g, 9.96 mmol) was added over 1 hr to CH 3
NH
2 (75.9mL, 2.5M in EtOH) at room temp. The reaction was stirred at room temp for 48 hr. The solution was concentrated and added to sat. NaHCO 3 . The aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were dried over Na 2
SO
4 , filtered and the solvent was removed under 30 reduced pressure. Yield 1.19g 237 380 WO 01/00206 PCT/US00/18079
H
3 C CH 3
H
3 C H3C -'O B 238 The 3-bromo-5-(N-methyl -aminomethyl)-pyridine(1.19g, 5.01 mmol) was taken up in DMF and triethylamine (0.90g, 1.24mL, 8.89 mmol) was added. Boc20 (1.55g, 7.10 mmol) was added and the reaction mixture was stirred at room temp for 48 hr. The solution was poured into 5 I1N HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography (2% methanol-CH 2 C1 2 ) Yield 1.6g 238 o O q 239 10 The sodium salt of a-methyl acrylic acid (5.00g, 46.27 mmol) was dissolved in DMF and benzyl bromide (8.70g, 50.89 mmol) was added at room temp. Potassium carbonate (7.03g, 50.89 mmol) was then added and the solution was heated to 500 C for 24 hr. The solution was poured into IN HC1 and the aqueous layer was extracted 3x with diethyl ether. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the 15 solvent was removed carefully under reduced pressure. The product was isolated by flash chromatography (2% ether-pentane) Yield 6.93g 239 H3C CH 3 H3C \ 0
-~CH
3 240 In a pressure tube was placed the 3-bromo-5-(N-methyl-Boc-aminomethyl)-pyridine (700.00mg, 2.33 mmol). The tube was then charged with DMF, triethylamine (260.05mg, 20 2.57mmol, 0.36mL), dppp (193.85mg, 0.47 mmol), and Pd(OAc) 2 (105.52mg, 0.47 mmol) The tube was flushed with argon for 10 min then benzyl methacrylate (452.86mg, 2.57 mmol) was added. The tube was sealed and heated to 1350 C for 24 hr. The reaction was cooled to 00 C and the tube was slowly opened. The solution was poured into lN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then 25 dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. 381 WO 01/00206 PCT/US00/18079 The product was isolated by flash chromatography. (6:1 hexane-ethyl acetate) Yield 785.23mg 240 H3CI H
H
3 C NH H3C N O 241 The unsaturated ester (392.61mg, 0.99 mmol ) was taken up in CH 2 C1 2 and excess TFA was added. The reaction was stirred for 4 hr at room temp. The solvent was removed under 5 reduced pressure and the residue was dried under high vacuum. The solvent was removed and the residue was taken up in ethyl acetate and washed with sat. NaHCO 3 solution. The organic layer was washed with water, brine then dried over Na 2
SO
4 . The solution was filtered and the solvent was removed under reduced pressure. The residue was taken up in CH 2
CI
2 -DMF and HOBt ( 147.53mg, 1.09 mmol), 3-methoxy-4-(N'-phenylureido)phenylacetic acid ( 342.64mg, 1.09 mmol) 10 and EDCI ( 208.96mg, 1.09 mmol) were added. The reaction was stirred for 24 hr. The solution was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (ethyl acetate) 363.79mg 241 H3 HN
H
3 C NH H3C-N 0 15 H 3 1 O 242 The unsaturated ester (363.00mg, 0.61 mmol) was taken up in CH 3 OH and Pd/C (100.00mg) was added under argon. The argon atmosphere was replaced with hydrogen at 1 atmosphere and stirred for 24 hr. The hydrogen atmosphere was removed and replaced with argon. The catalyst was removed by filtration through celite and the celite pad was washed with 20 ethyl acetate 3x. The solvent was removed under reduced pressure. H'-NMR showed only the desired product. The solid was washed with ether and then dried under high vacuum. Yield 254.79mg 242 382 WO 01/00206 PCT/US00/18079 Example 203 Ph CH 3 \ cH 3 NcPh 243 3-Cyanobenzaldehyde (9.41g, 71.76 mmol) was taken up in ethanol and cooled to 00 C. The NaBH 4 (2.71g, 71.76 mmol) was added in small portions. The solutions was stirred for 30 5 min at 00 C then allowed to warm to room temp and stirred for 1 hr. The reaction was slowly poured into INHCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The residue was taken up in DMF and imidazole (2.08g, 30.50 mmol) was added. TBDPSCI (4.61g, 16.78 mmol, 4.36mL) was then 10 added and the reaction was stirred at room temp for 12 hr. The solution was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (7:1 hexane-ethyl acetate to 4:1 hexane-ethyl acetate) Yield 16.23g 243
H
3 0 Ph\ ?IcH 3 O~l N O" \ LH3
H
3 -1 H Ph 15 H 3 C OH 3 244 The silyl protected 3-cyanobenzyl alcohol (8.50g, 34.36 mmol) was taken up in ethyl acetate and Boc 2 0 (8.25g, 37.79 mmol) was added. Pd/C (1.0g) was added and the Parr vessel was pressurized with hydrogen at 50 psi. The vessel was shaken for 24 hr then the hydrogen was flushed with argon and the catalyst was removed by filtration through a celite pad. The celite was 20 washed 3x with ethyl acetate. The solvent was removed under reduced pressure and the product was isolated by flash chromatography (10:1 hexane-ethyl acetate) Yield 11.10 g 244 P H 3 -. - "CH 3
H
3
H
3 Ph
H
3 0 CH 3 245 The O-silyl-N-Boc-protected benzyl alcohol (5.00g, 14.22 mmol) was dissolved in dry THF under argon. The reaction was cooled to minus 780 C. Lithium bis(trimethylsilyl)amide 25 (42.67mL, 42.67 mmol) was added over 10 min. The reaction was stirred for 1 hr at minus 78o C then iodomethane (6.06g, 42.67 mmol, 2.66mL) was added rapidly. The reaction was allowed to slowly warm to room temp and stir overnight. The reaction was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (2% ethyl 383 WO 01/00206 PCT/US00/18079 acetate-hexane) Yield 4.7g 245 H 3 H3 OH
H
3 H3 246 The O-silyl-Boc-N-methyl protected benzyl alcohol (4.7g, 9.60 mmol) was taken up in THF and TBAF (14.39mL, 1.0OM in THF) at room temp. The solution was stirred for 4 hr. TLC 5 showed no starting material present. The reaction was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (4:1 hexane-ethyl acetate to 1:1 hexane-ethyl acetate) Yield 2.39g 246 H3CXO N 'H Br 3 C COH 3 247 10 The N-methyl Boc protected benzyl alcohol (1.00g, 3.98 mmol) was taken up in dry
CH
2 C1 2 under argon. Triphenylphosphine (1.46g, 5.57 mmol) was added and the solution was cooled to 00 C. Carbon tetrabromide (1.85g, 5.57 mmol) dissolved in dry CH 2 C1 2 was added over 10 min. The solution was stirred for 1 h at 00 C then the solvent was removed under reduced pressure. The residue was taken up in Et 2 0 and the resulting solid was removed by filtration and 15 the filtrate was collected and the solvent was removed under reduced pressure. The product was isolated by flash chromatography (2/oether-pentane) Yield 1.15g 247 H3CX CH3 H3- 3C H 3
H
3
HCH
3 N 3 248 LHMDS (3.23mL, 3.23 mmol) was added to dry DME under argon at minus 780 C. Methyl butyrate (300mg, 2.94 mmol, 0.33mL) dissolved in dry DME was added to the LHMDS 20 over 15 min and the solution was stirred for lhr at minus 780 C. 3-N-methyl-N-Boc protected benzyl bromide (1.02g, 3.23 mmol) dissolved in dry DME was added to the enolate solution over 15 min then the solution was allowed to slowly warm to minus 200 C and stirred for 4 hr. The reaction was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The solution was filtered and the solvent was removed under reduced pressure. The product was 25 isolated by flash chromatography. (3/oethyl acetate-hexane) Yield 414mg 248. 384 WO 01/00206 PCT/US00/18079 N
OCH
3
H
3 H
H
3 H3 H3CO H H3 HNN 249 The Boc ester (121.60mg, 0.36 mmol) was taken up in CH 2 Cl 2 and excess trifluoroacetic acid was added. The reaction was then stirred for 2 hr. The solvent was removed and the residue was taken up in ethyl acetate and washed with sat. NaHCO 3 solution. The organic layer was 5 washed with water, brine then dried over Na 2
SO
4 . The solution was filtered and the solvent was removed under reduced pressure. The residue was taken up in CH 2 C 2 -DMF and HOBt (54.10mg, 0.40 mmol) 3-methoxy-4-(N'-phenylureido)phenylacetic acid (125.74mg, 0.40 mmol) and EDCI (77.0mg, 0.40 mmol) were added. The reaction was stirred for 24 hr. The solution was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic 10 layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (ethyl acetate) Yield 165.20mg 249 O O 1 OH H3C H H 3 H "N 250 The ester (165.20, 0.31 mmol) was taken up in ethanol-water (4:1) and NaOH was added. 15 The reaction was then heated to 500 C for 2 hr. The TLC (ethyl acetate) showed no starting material present. The reaction was cooled to room temp. The solution was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The residue was recrystallized from ethyl acetate-hexane. Yield 20 120.00mg 250 Example 204 O 0 H3
H
3
CH
3 251 Butyrolactone (250mg, 2.90 mmol, 223.20mL) was added to LHMDS (2.90mL, 1.0OM in hexane) in THF at minus 780 C under argon over 10min. The solution was stirred at minus 78oC 25 for lhr. 3-N-methyl-N-Boc protected benzyl bromide (991.24mg, 2.90 mmol) dissolved in dry 385 WO 01/00206 PCT/US00/18079 DME was added to the enolate solution over 15 min then the solution was allowed to slowly warm to room temp and stirred for 12 hr. The reaction was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (4:1 hexane-ethyl acetate to 5 1:1 ethyl acetate-hexane) Yield 501.18mg 251 0 N
H
3 C H H 3 HN N 252 The Boc ester (250.00mg, 0.78 mmol) was taken up in CH 2 C1 2 and excess trifluoroacetic acid was added. The reaction was then stirred for 2 hr. The solvent was removed and the residue was taken up in ethyl acetate and washed with sat. NaHCO 3 solution. The organic layer was 10 washed with water, brine then dried over Na 2
SO
4 . The solution was filtered and the solvent was removed under reduced pressure. The residue was taken up in CH 2 Cl 2 -DMF and HOBt (116.40mg, 0.86 mmol) 3-methoxy-4-(N'-phenylureido) phenylacetic acid (270.33mg, 0.86 mmol) and EDCI (165.06mg, 0.86 mmol) were added. The reaction was stirred for 24 hr. The solution was poured into IN HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined 15 organic layers were washed with water, brine then dried over MgSO 4 . The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (ethyl acetate) Yield 119.00mg 252 Example 205 3-methoxy-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetyl]-N-methylaminoethoxy] 20 benzoic acid (H3 I COOH P NNN%
OCH
3
CH
3 H H OCH 3 253 To a stirred and cooled (00 C) solution of N-methyl ethanolamine (3.10 g, 41.27 mmol), Et 3 N (11.80 mL, 84.66 mmol) in DMF-H20 (3:1, v/v, 40 mL) was added dropwise 30% toluene solution of benzyl chloroformate (25.40 g, 49.13 mmol) for over 15 min. The resulting mixture 25 was stirred for 1 day at room temp. The mixture was extracted with EtOAc. The extract was washed with sat. NaHCO 3 , brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with n-hexane:EtOAc (3:1, v/v) then CHC 3 as eluent to give 4.67 g (54%) N-methyl-N-(benzyloxy carbonyl)ethanolamine as a colorless oil. 'H-NMR (CDC1 3 ) d 386 WO 01/00206 PCT/US00/18079 1.82 (bs, 1 H), 3.00 (s, 3 H), 3.46 (bs, 2 H), 3.77 (bs, 2 H), 5,13 (s, 2 H), 7.29-7.36 (min, 5 H). To a stirred solution of ethyl 4-hydoxy-3-methoxybenzoate (2.01 g, 10.25 mmol), N methyl-N-(benzyloxycarbonyl)ethanolamine (2.11 g, 10.08 mmol), PPh 3 (3.26 g, 12.43 mmol) in THF was added DIAD (2.65 mL, 13.46 mmol) and the reaction mixture was heated under reflux 5 overnight. The mixture was evaporated, and the residue was subjected to short column chromatography on silica-gel with n-hexane/EtOAc (5:1, v/v) as eluent to give ethyl 3-methoxy-4 [2-methyl-2-(benzyloxycarbonyl) aminoethoxy]benzoate as a crude product. To a solution of the crude product (5.20 g, 13.42 mmol) in EtOH (50 mL) was added AcOH (5 mL) and the solution was hydrogenated over 5% Pd/C for 4 hr. The mixture was filtered 10 to remove the catalyst and the filtrate was evaporated. The residue was diluted with CHC1 3 and washed with sat. NaHCO 3 , brine, dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica-gel with CHCl 3 :MeOH (10:1, v/v) as eluent to give 510 mg (2 steps 20%) ethyl 3-methoxy-4-(2-methylamino ethoxy) benzoate as a yellow oil. 'H-NMR (CDC13) d 1.39 (t, 3 H, J=7.3 Hz), 1.82 (bs, 1 H), 2.52 (s, 3 H), 3.04 (t, 2 H, J=5.3 Hz), 3.91 (s, 3 H), 4.18 (t, 15 2 H, J=5.3 Hz), 4.36 (q, 2 H, J=7.3 Hz), 6.90 (d, 1 H, J=8.3 Hz), 7.55 (d, 1 H, J=2.0 Hz), 7.65 (dd, 1 H, J=2.0, 8.3 Hz). To a stirred solution of ethyl 3-methoxy-4-(2-methylaminoethoxy) benzoate (510 mg, 2.01 mmol) in DMF (13 mL) was added pentafluorophenyl ester of 3-methoxy-4-[N'-(2 methylphenyl) ureido] phenylacetic acid (900 mg, 1.87 mmol) and Et3N (0.420 mL, 3.01 mmol), 20 and the resulting mixture was stirred for 2 days. The mixture was diluted with EtOAc, washed with 1 N HC1, sat. NaHCO 3 , brine, and dried over Na 2
SO
4 . After being evaporated, the residue was purified by column chromatography on silica-gel with CHCl 3 :MeOH (50:1, v/v) to give 880 mg (85%) ethyl 3-methoxy-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-N methylaminoethoxy] benzoate as a colorless amorphous solid. 'H-NMR (CDCl 3 ) d 1.37-1.41 (m, 3 25 H), 2.28 (s, 3 H), 3.03 and 3.18 (s, 3 H), 3.56 (s, 2 H), 3.65 (s, 2 H), 3.75-3.87 (mn, 6 H), 4.06-4.24 (2 H, min), 4.33-4.39 (min, 2 H), 6.68-8.08 (series of m, 12 H). To a solution of ethyl 3-methoxy-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-N-methlaminoethoxy]benzoate (880 mg, 1.601 mmol) in THF (15 mL) was added 0.25 N NaOH (15 mL). Then the reaction mixture was heated under reflux overnight. The 30 mixture was poured into 1 N HCI (100 mL), and the solid was collected. The crude solid was 387 WO 01/00206 PCT/US00/18079 recrystallized from MeOH-CHC 3 to give 253 as a white powder. IR (KBr) 1700 cm-'; 'H-NMR (DMSO-d 6 ) d 2.25 (s,3 H), 2.50 (s, 2 H), 2.91 and 3.12 (s, 3 H) 3.53-3.76 (min, 2 H), 3.80 (s, 3 H), 3.84 (s, 3 H), 4.16-4.21 (mn, 2 H), 6.72-8.56 (series of m, 12 H), 12.68 (bs, 1 H); MS (FAB) m/z 522 (M++1);Anal. Calcd. for C 28
H
3 1
N
3 0 7
H
2 0: C, 62.33; H, 6.16; N, 6.63. Found: C, 62.17; H, 5 6.05; N, 7.57. Example 206 4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]methylamino]ethoxy]isophthalic acid
CH
3 'iCOOH 0N N O COOH
CH
3 H H OCH 3 254 10 To a stirred solution of N-methyl-N-benzyloxycarbonylethanolanine (1.05 g, 5.02 mmol), dimethyl 4-hydroxy isophlithalate (1.05 g, 5.00 mmol), Ph 3 P (1.59 g, 6.06 mmol) in THF (20 mL) was added DIAD (1.28 mL, 6.50 mmol) at room temp. The resulting mixture was then heated under reflux overnight. After cooling to room temp, the mixture was evaporated. The residue was dissolved in EtOH and added 5% Pd/C(200mg). The stirred resulting mixture was hydrogenated 15 for 2 hr at latmn. The mixture was filtered to remove the catalyst, and the filtrate was evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (30:1, v/v) as eluent to give 480 mg (36% for 2 steps) dimethyl 4-(2-methylaminoethoxy) isophthalate as an oil. 1 H-NMR (CDCl 3 ) d 1.68 (s, 1 H), 2.53 (s, 3 H), 3.01-3.04 (min, 2 H), 3.89 (s, 3 H), 3.90 (s, 3 H), 4.21-4.23 (min, 2 H), 7.00 (d, 1 H, J=8.8 Hz), 8.14 (dd, 1 H, J=2.4, 8.8 Hz), 8.50 (d, 1 H, J=2.4 20 Hz); MS (FAB), m/z 268 (M*+1). To a stirred solution of dimethyl 4-(2-methylaminoethoxy)isophthalate (410 mg, 1.53 mmol) in DMF (13 mL) was added pentafluorophenyl ester of 3-methoxy-4-[N'-(2 methylphenyl)ureido] phenylaceic acid (700 mg, 1.46 mmol) and Et 3 N (340 ml, 2.44 mmol), and the resulting mixture was stirred overnight. The mixture was diluted with EtOAc, washed with 1 25 N HC1, sat. NaHCO 3 , and brine. The solution was dried over Na 2
SO
4 and evaporated to give 780 mg (95%) dimethyl 4- [2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]methylamino ethoxy]isophthalate as a crystalline powder. 'H-NMR (CDC13) d 2.29 (s, 3 H), 3.24 (s, 3 H), 3.59 (s, 3 H), 3.67-3.68 (min, 2 H), 3.84 (s, 3 H), 3.91 (s, 3 H), 3.81-3.86 (min, 2 H), 4.25-4.28 (mn, 2 H), 6.51-8.48 (series of min, 12 H); MS (FAB) m/z 564 (M+I). 388 WO 01/00206 PCT/US00/18079 To a solution of dimethyl 4- [2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] methylaminoethoxy]isophthalate (780 mg, 1.384 mmol) in THF (30 mL) was added 0.25 N NaOH (30 mL). The resulting mixture was then heated under reflux overnight. The mixture was poured into ice-1 N HC (200 mL) and the solid was collected. The crude solid was recrystallized from 5 MeOH-CHC1 3 to give 420 mg (57%) 4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl acetyl]methylamino]ethoxy] isophthalic acid 254 as a white crystalline powder. mp 139-141 oC IR (KBr) 1700 cm'; 'H-NMR (DMSO-d6) d 2.94 (s, 3 H), 3.18 (s, 3 H) 3.62-3.86 (m, total 8 H), 4.24 4.28 (m, 2 H), 6.74-8.58 (series of m, total 12 H), 12.91 (bs, 1 H); MS (FAB) m/z 536 (M +1); Anal. Calcd. for C 28
H
29
N
3 0 8 s2.5HCl: C, 53.66; H, 5.07; N, 6.70. Found: C, 53.80; H, 4.64; N, 6.70. 10 Example 207 3-methoxy-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]aminoethoxy]benzoic acid H rCOOH N N-N
OCH
3
CH
3 H H OCH 3 255 To a solution of 2-ethanolamine (5.16 g, 84.48 mmol), Et 3 N (23.50 mL, 168.60 mL) in dioxane-H 2 0 (1/1, 160 mL) was added dropwise (Boc) 2 0 (23.40 mL, 101.86 mmol) at room temp. 15 The reaction mixture was stirred for 2 days at room temp. The resulting mixture was diluted with CHCl 3 , washed with 0.5 N HCI, sat. NaHCO 3 , and brine. The separated organic layer was dried over Na 2
SO
4 and evaporated to give 11.86 g (87%) N-Boc-2-ethanolamine as an oil. 'H-NMR (CDCl 3 ) d 1.45 (s, 9 H), 3.29-3.31 (m, 2 H), 3.71-3.72 (m, 2 H). To a stirred solution of ethyl 4-hydroxy-3-methoxybenzoate (1.46 g, 7.44 mmol), N-Boc 20 ethanolamine (1.19 g, 7.38 mmol), PPh 3 (2.53 g, 9.65 mmol) in THF (30 mL) was added DIAD (1.90 mL, 9.65 mmol), and the resulting mixture was then heated under reflux overnight. The mixture was evaporated to give a crude gum. The crude product was dissolved in CH 2 C1 2 (20 mL) and TFA (20 mL). The resulting mixture was stirred for 2.5 hr at room temp. The mixture was concentrated in vacuo and the residue was made basic with sat. NaHCO 3 and extracted with 25 CHC 3 . The extract was washed with brine, dried over Na 2
SO
4 , and evaporated to give the 1.61 g (90% for 2 steps) ethyl 3-methoxy-4-(2-aminoethoxy) benzoate as a yellow oil. 'H-NMR (CDCl 3 ) d 1.39 (t, 3 H, J=7.3 Hz), 3.14-3.17 (m, 2 H), 3.92 (s, 3 H), 4.09-4.11 (m, 2 H), 4.36 (q, 2 H, J=7.3 Hz), 6.89 (d, 1 H, J=8.3 Hz), 7.56 (d, 1 H, J=2.0 Hz), 7.66 (dd, 1 H, J=2.0, 8.3 Hz). 389 WO 01/00206 PCT/US00/18079 To a stirred solution of ethyl 3-methoxy-4-(2-aminoethoxy)benzoate (250mg, 1.04 mmol) and pentafluorophenyl ester of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (500 mg, 1.04 mmol) was added Et 3 N (210 pl, 3.01 mmol), and the resulting mixture was stirred for 2 days. 0.25 N NaOH (20 mL) and THF (20 mL) was added to the mixture and the resulting mixture 5 was heated under reflux overnight. After cooling, the mixture was evaporated and the residue was acidified by the addition of 1 N HCL. The mixture was extracted with CHC13, and the extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The obtained crude solid was recrystallized from CHC1 3 to give 110 mg (20% for 2 steps) 3-methoxy-4-[2-[3-methoxy-4-[AN'-(2-methylphenyl) ureido]phenylacetyl]aminoethoxy] benzoic acid 255 as a white crystalline powder. mp 180-181 10 oC; IR (KBr) 1687 cm-'; 1 H-NMR (DMSO-de) d 2.24 (s, 3 H), 3.37 (s, 2 H), 3.38 (s, 2 H), 3.41 3.50 (min, 2 H), 3.81 (s, 3 H), 3.83 (s, 3 H), 4.06-4.08 (min, 2 H), 6.76-8.55 (series of m, total 12 H); MS (FAB) m/z 508 (M+1); Anal. Calcd for C 27
H
29
N
3 07-1/2H 2 0: C, 62.78; H, 5.85; N, 8.13. Found: C, 62.46; H, 5.69; N, 8.03. Example 208 15 3-methoxy-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]ethylaminoethoxy] benzoic acid
CH
3 COOH -N YNO
OCH
3
CH
3 H H OCH 3 256 To a cooled (0o C) solution of ethyl 3-methoxy-4-(2-aminoethoxy)benzoate (1.93 g, 8.07 mmol) and Et 3 N (2.00 mL, 14.35 mmol) was added TFAA (1.35 mL, 9.56 mmol) and the resulting 20 mixture was stirred overnight at room temp. The resulting mixture was diluted with Et 2 0 and washed successively with sat. NaHCO 3 , 1 N HC1, H 2 0, and brine. The extract was dried over Na 2
SO
4 and evaporated to give 1.22 g (45%) ethyl 3-methoxy-4-(2-N-trifluoroacetamidoethoxy) benzoate as an oil. 'H-NMR (CDCl 3 ) d 1.39 (t, 3 H, J=7.3 Hz), 3.77-3.81 (min, 2 H), 3.92 (s, 3 H), 4.18-4.20 (min, 2 H), 4.37 (q, 2 H, J=7.3 Hz), 6.92 (d, 1 H, J=8.7 Hz), 7.59 (d, 1 H, J=2.0 Hz), 7.67 25 (dd, 1 H, J=2.0, 8.7 Hz); MS (FAB) m/z 335 (M), 290 (M*-OEt). To a stirred solution of ethyl 3-methoxy-4-(2-N-trifluoroacetamidoethoxy)benzoate (1.20 g, 3.58 mmol) in DMF (15 mL) was added K 2
CO
3 (0.98 g, 7.09 mmol) and EtI (0.43 mL, 5.38 mmol) at room temp. The resulting mixture was stirred for 2 days at 600 C. The mixture was diluted with EtOAc, washed successively with 1 N HC1, brine, and dried over Na 2
SO
4 . The solvent 30 was evaporated and the residue was purified by column chromatography on silica-gel with n 390 WO 01/00206 PCT/US00/18079 hexane-EtOAc (2:1, v/v) as eluent to give 990 mg (76%) ethyl 3-methoxy-4-[2-(N-ethyl-N trifluoroacetamido)ethoxybenzoate as a yellow crystalline solid. 'H-NMR (CDCl 3 ) d 1.28-1.31 (m, 3 H), 1.37-1.40 (m, 3 H), 3.64-3.69 (m, 2 H), 3.81-3.84 (m, 2 H), 3.92 (s, 3 H), 4.27-4.30 (m, 2 H), 4.34-4.39 (m, 2 H), 6.89 (d, 1 H, J=8.3 Hz), 7.55 (d, 1 H, J=2.0 Hz), 7.66 (dd, 1 H, J=2.0, 8.3 5 Hz); MS (FAB) m/z 364 (M++1). To a stirred solution of ethyl 3-methoxy-4-[2-(N-ethyl-N-trifluoroacetamido) ethoxybenzoate (990 mg, 2.73 mmol) in THF-MeOH-HO 2 0 (2:1:1, v/v, 20 mL) was added K 2 C0 3 (560 mg, 4.05 mmol), and the resulting mixture was stirred overnight. The resulting mixture was diluted with H 2 0 and extracted with EtOAc. The extract was washed successively with sat. 10 NaHCO 3 , brine, dried over Na 2
SO
4 , and evaporated to give 800 mg (q.y.) ethyl 3-methoxy-4-(2 ethylaminoethoxy)benzoate as an oil. 'H-NMR (CDCl 3 ) d 1.15 (t, 3 H, J=7.3 Hz), 1.39 (t, 3 H,J=7.3 Hz), 1.76 (bs, 1 H), 2.74 (q, 2 H, J=7.3 Hz), 3.08 (t, 2 H, J=5.4 Hz), 3.91 (s, 3 H), 4.18 (t, 2 H, J=5.4 Hz), 4.36 (q, 2 H, J=7.3 Hz), 6.90 (d, 1 H, J=8.3 Hz), 7.55 (d, 1 H, J=2.0 Hz), 7.66 (dd, 1 H, J=2.0, 8.3 Hz); MS (FAB) m/z 268 (M +1). 15 To a stirred solution of ethyl 3-methoxy-4-(2-ethylaminoethoxy)benzoate (290 mg, 1.08 mmol) and pentafluorophenyl ester of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (502 mg, 1.05 mmol) in DMF (7 mL) was added Et 3 N (250 pl, 1.79 mmol), and the resulting mixture was stirred overnight. The mixture was diluted with EtOAc, washed with 0.5 N HC1, brine, and dried over Na 2
SO
4 . The solvent was evaporated and the residue was purified by column 20 chromatography on silica-gel with CHC13-MeOH (40:1, v/v) as an eluent to give 550 mg (93%) ethyl 3-methoxy-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]ethylaminoethoxy] benzoate as an amorphous solid. 'H-NMR (CDCl 3 ) d 1.11-1.18 (m, 3 H), 1.37-1.41 (m, 3 H), 2.30 (s, 3 H), 3.47-3.53 (m, 2 H), 3.61-3.75 (m, 7 H), 3.84 (s, 3 H), 4.03-4.27 (m, 2 H), 4.33-4.39 (m, 2 H), 6.34-8.07 (series of m, total 12 H). 25 To a solution of ethyl 3-methoxy-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl acetyl]ethylaminoethoxy]benzoate (550 mg, 0.98 mmol) in THF (15 mL) was added 0.25 N NaOH (15 mL). The resulting mixture was then heated under reflux for 2 days. The mixture was poured into 1 N HCI and the solid was collected. The crude solid was recrystallized from EtOH-CHC 3 to give 182 mg (35%) 3-methoxy-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] 30 ethylaminoethoxy]benzoic acid 256 as a white crystalline powder. mp 115-118 oC; IR (KBr) 1707 391 WO 01/00206 PCT/US00/18079 cm'; 'H-NMR (DMSO-d) d 1.02-1.12 (min, 3 H), 2.25 (s, 3 H), 2.50 (s, 2 H), 3.35-3.89 (mn, 10 H), 4.11-4.16 (min, 2H), 6.71-8.56 (series of m, total 12H), 12.65 (br s, 1H); MS (FAB) m/z 536 (M+I); Anal. Calcd for C 2 9H 3 3
N
3 07-3/4H20: C, 63.43; H,6.33; N, 7.65. Found: 63.34; H, 6.28; N, 7.28. Example 209 5 3-nitro-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]aminoethoxy]benzoic acid H NOCOOH 0 "-h'N i"_' . N )LN- Q
NO
2 H3H H OCH 3 257 To a stirred solution of 4-hydroxy-3-nitrobenzoic acid (5.18 g, 28.29 mmol) in benzene MeOH (4:1, v/v, 140 mL) was added TMSCHN 2 (14.10 mL, 28.20 mmol, 2 M solution in hexane) at room temp, and the resulting mixture was stirred overnight. The mixture was evaporated and 10 the residue was purified by column chromatography on silica-gel with CHC 3 as eluent to give 4.18 g (75%) methyl 3-nitro-4-hydroxybenzoate as a yellow crystalline solid. 'H-NMR (CDCl 3 ) d 3.95 (s, 3H), 7.22 (d, 1H, J=8.8 Hz), 8.24 (dd, 1, J=2.0, 8.8 Hz), 8.83 (d, 1H, J=2.0 Hz), 10.89 (s, 1H). To a stirred solution of methyl 3-nitro-4-hydroxybenzoate (1.98 g, 10.04 mmol), N-Boc ethanolamine (1.63 g, 10.11 mmol) and PPh 3 (3.43 g, 13.08 mmol) in THF (40 mL) was added 15 DIAD (2.57 mL, 13.05 mmol), and the reaction mixture was then heated under reflux overnight. The resulting mixture was evaporated to give a gum. The residual crude gum was dissolved in
CH
2 Cl 2 (30 mL) and TFA (30 mL), and the mixture was stirred for 1 hr at room temp. The mixture was concentrated in vacuo and made basic with sat. NaHCO 3 . The mixture was extracted with CHC1 3 , washed with brine, and dried over Na 2
SO
4 . The solvent was evaporated in vacuo to 20 give the oily residue, which was purified by column chromatography on silica -gel with CHC1 3 then CHCl 3 -MeOH (20:1, v/v) as eluent to give 930 mg (27% for 2 steps ) methyl 3-nitro-4-(2 aminoethoxy) benzoate as gum. 'H-NMR (CDCl 3 ) d 3.16-3.19 (min, 1 H), 3.53-3.57 (min, 1 H), 3.90 and 3.94 (s, 3 H), 3.95-3.98 (min, 1 H), 4.21-4.24 (min, 1 H), 6.89-6.91 and 7.11-7.13 (min, 1 H), 8.03 8.19 and 8.21 (min, 1 H), 8.52 and 8.86 (min, 1 H). 25 To a stirred solution of pentafluorophenyl ester of 3-methoxy-4-[N'-(2-methylphenyl) ureido] phenylacetic acid (1.86 g, 3.87 mmol) and methyl 3-nitro-4-(2-aminoethoxy)benzoic acid (0.93 g, 3.87 mmol) in DMF (27 mL) was added Et 3 N (0.90 mL, 6.46 mmol), and the resulting mixture was stirred overnight. The mixture was poured into 0.5 N HCI and the resulting solid was collected. The crude solid was dissolved in THF-0.25 N NaOH (1/1, 20 mL) and the resulting 392 WO 01/00206 PCT/US00/18079 mixture was heated under reflux overnight. The mixture was extracted with EtOAc, washed with brine, dried over Na 2
SO
4 , and evaporated. The crude solid was recrystallized from CHCl 3 -EtOH to give 60 mg (3% for 2 steps) 3-nitro-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] aminoethoxy]benzoic acid 257 as a yellow crystalline solid. mp 112-115 oC; 'H-NMR (DMSO-d 6 ) 5 d 2.24 (s, 3 H), 3.37-3.66 (min, 7 H), 3.84 (s, 3 H), 4.27-4.30 (min, 1 H), 6.74-8.56 (series of m, total 12 H); MS (FAB) m/z 523 (M++1); Anal. Calcd for C 26
I
26
N
4 0s-3/2H 2 0: C, 56.83; H, 5.32; N, 10.20. Found: C, 56.66; H, 4.90; N, 9.33. Example 210 3-methoxy-4-[2-[3-methoxy-4-[N'-(2-fluorophenyl)ureido]phenylacetyl]ethylaminoethoxy] benzoic 10 acid
CH
3 ~ COOH r\.N K a 0OCH 3 F H H OCH 3 258 To a stirred solution of pentafluorophenyl ester of 3-methoxy-4-[N'-(2-fluorophenyl) ureido] phenylacetic acid (135 mg, 0.28 mmol) and ethyl 3-methoxy-4-(2-ethylaminoethoxy) benzoate (78 mg, 0.29 mmol) was added Et 3 N (0. lmL, 0.72 mmol), and the resulting mixture was 15 stirred overnight. The mixture was diluted with EtOAc, washed successively with 0.5N HCI, brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica gel with CHCl 3 -MeOH (50:1, v/v) as eluent to give 160 mg(q.y.) ethyl 3-methoxy-4-[2-[3-methoxy -4-[N'-(2-fluorophenyl) ureido]phenylacetyl]ethylaminoethoxy]benzoate as an oil. 'H-NMR (CDCl 3 ) d 1.13-1.23 (min, 3 H), 1.37-1.40 (min, 3 H), 2.90-3.89 (min, 12 H), 4.09-4.28 (min, 2 H), 4.33 20 4.39 (min, 2 H), 6.70-8.21 (series of m, total 12 H). To a stirred solution of ethyl 3-methoxy-4-[2-[3-methoxy-4-[N'-(2-fluorophenyl)ureido] phenylacetyl]ethylaminoethoxy]benzoate (160 mg, 0.28 mmol) in THF (5 mL) was added 0.25 N NaOH (5 mL) and the resulting mixture was the heated under reflux overnight. The mixture was poured into 1 N HCI and the solid was collected. The crude solid was recrystallized from EtOH 25 CHCl 3 -n-hexane to give 70 ming (46%) 3-methoxy-4-[2-[3-methoxy-4-[N'-(2-fluorophenyl)ureido] phenylacetyl]ethylaminoethoxy] benzoic acid 258 as a yellow crystalline powder. mp 105-110 oC; IR (KBr) 1687 cm- 1 ; 1 H-NMR (DMSO-d 6 ) d 1.00-1.10 (min, 3 H), 2.48 (s, 2 H), 3.35-3.81 (mn, 10 H), 4.13-4.14 (min, 2 H), 6.70-9.15 (series of m, 12 H); MS (FAB) m/z 540 (M'+1); Anal. Calcd for
C
28
H
30
FN
3 0 7 -1/2H 2 0: C, 61.31; H, 5.82; N, 7.47. Found: C, 61.05; H, 5.82; N, 7.47. 393 WO 01/00206 PCT/US00/18079 Example 211 4-[4-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl]acetyl- 1 -piperazinylbenzoic acid 0 H CH 0 Nj
H
3 H H CH 3 259 A stirred mixture of tert-butyl 1-piperazinecarboxylate (1.00 g, 5.37 mmol), ethyl 4 5 fluorobenzoate (903 mg, 5.37 mmol), and K 2 C0 3 (1.11 g, 8.06 mmol) in DIF (10 mL) was heated at 120 0 C overnight. After cooling, the mixture was diluted with EtOAc (300 mL), followed by washing with brine (2 x 200 mL), drying over MgSO 4 , and evaporation. The residue was chromatographed on silica-gel with CHCl 3 -EtOAc (20:1 to 4:1, v/v) as eluent to give 257 mg (14%) ethyl 4-[4-(tert-butyloxycarbonyl)-1-piperazinyl]benzoate as a pale yellow amorphous solid. 10 IR (KBr) 1701, 1612 cm-'; 'H-NMR (CDC1 3 ) d 1.37 (3 H, t, J= 7.3 Hz), 1.49 (9 H, s), 3.30 (4 H, t, J= 5.4 Hz), 3.58 (4 H, t, J= 5.4 Hz), 4.33 (2 H, q, J= 7.3 Hz), 6.87 (2 H, d, J= 8.8 Hz), 7.94 (2 H, dt, J= 8.8, 2.4 Hz); MS (FAB)m/z 335 (M'+l);Anal. Calcd for C 8
H
26
N
2 0 4 : C, 64.54; H, 7.84; N, 8.38. Found: C, 64.39; H, 7.89; N, 8.38. To a stirred solution of ethyl 4-[4-(tert-butyloxycarbonyl)-1l-piperazinyl]benzoate (240 15 mg, 0.718 mmol) in CH 2 Cl 2 (5 mL) was added TFA (5 mL), and the resulting mixture was stirred for 3 hr. The mixture was concentrated in vacuo and the residue was made basic by the addition of sat. NaHCO 3 , followed by extraction with CHC13 (2 x 100 mL). The combined extracts were dried over Na 2
CO
3 and evaporated to give 168 mg ethyl 4-(1-piperazinyl)benzoate (100%) as a yellow oil. 'H-NMR (CDC13) d 1.37 (3 H, t, J 7.3 Hz), 3.03 (4 H, t, J= 4.9 Hz), 3.29 (4 H, t, J= 20 4.9 Hz), 4.33 (2 H, q, J = 7.3 Hz), 6.87 (2 H, dt, J = 8.8, 2.4 Hz), 7.91-7.94 (2 H, m). To a stirred solution of ethyl 4-(1-piperazinyl)benzoate (170 mg, 0.730 mmol) and 3 methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (229 mg, 0.730 mmol) in DMF (10 mL) was added EDC-HCl (210 mg, 1.10 mmol), DMAP (catalytic amount), and HOBt (catalytic amount), and the mixture was stirred overnight. The mixture was poured into H 2 0 (100 mL) and 25 the solid was collected with suction. The residue was recrystallized from CHCl 3 -n-hexane to give 290 mg (75%) ethyl 4-[4-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl]acetyl-1-piperazinyl benzoate as a colorless crystalline powder. mp 208-210 oC; IR (KBr) 1711, 1695 cm-'; 'H-NMR (CDCl 3 ) d 1.37 (3 H, t, J= 7.3 Hz), 2.29 (3 H, s), 3.14 (2 H, t, J= 4.9 Hz), 3.28 (2 H, t, J= 4.9 Hz), 3.62 (2 H, t, J= 4.9 Hz), 3.71 (3 H, s), 3.72 (2 H, s), 3.79 (2 H, t, J= 4.9Hz), 4.33 (2 H, q, J 30 = 7.3 Hz), 6.38 (1 H, s), 6.78-6.99 (4 H, m), 7.13-7.24 (4 H, m), 7.50 (1 H, d, J = 7.8 Hz), 7.92 (2 394 WO 01/00206 PCT/US00/18079 H, d, J = 8.8 Hz), 8.12 (1 H, d, J = 7.8 Hz); MS (FAB) nm/z 531 (M'+1); Anal. Calcd for
C
30
H
3 4
N
4 0 5 -0.5H 2 0: C, 66.77; H, 6.54; N, 10.38. Found: C, 66.89; H, 6.39; N, 10.45. To a stirred solution of ethyl 4-[4-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl] acetyl-1-piperazinylbenzoate (290 mg, 0.547 mmol) in MeOH-THF (2:1, v/v, 15 mL) was added 5 0.25 N NaOH (5 mL, 1.25 mmol) and the mixture was heated under reflux for 3 hr. The mixture was poured into ice-IN HCI (100 mL) and the solid was collected with suction. The residue was recrystallized from CHCl 3 -MeOH to give 190 mg (69%) 4-[4-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenyl]acetyl-1-piperazinylbenzoic acid 259 as a yellow crystalline powder. mp 240-245 OC; 'H-NMR (DMSO) d 2.24 (3 H, s), 3.17-3.50 (8 H, m), 3.72 (2 H, s), 3.86 (3 H, s), 6.77 (1 H, d, 10 J= 8.3 Hz), 6.90 (1 H, s), 6.91-6.96 (3 H, m), 7.11-7.17 (2 H, m), 7.76-7.80 (3 H, m), 8.03 (1 H, d, J= 8.3 Hz), 8.47 (1 H, s), 8.58 (1 H, s), 12.30 (1 H, s);Anal.Calcd for C 28
H
30
N
4
O-H
2 0: C, 64.60; H, 6.20; N, 10.76. Found: C, 64.64; H, 5.85; N, 10.51. Example 212 (R)-3-methoxy-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetylamino]-1-propoxy] 15 benzoic acid H COOH -N N CH 3
OCH
3
CH
3 H H OCH 3 260 To a cooled (0OC) solution of (R)-2-amino-1-propanol (3.01 g, 0.04 mmol) and Et 3 N (6.70 mL, 0.05 mmol) in DMF-H 2 0 (1:1, v/v)(40 mL) was added (Boc) 2 0 (10.0 mL, 0.04 mmol), and the resulting mixture was stirred at room temp for 2 days. The mixture was diluted with 20 EtOAc, washed with H 2 0, brine, dried over Na 2
SO
4 and evaporated to give 6.91 g (98%) (R)-2-N tert-butoxycarbonylamino-1-propanol as a colorless oil. 'H-NMR (CDCl 3 ) 51.15 (d, 3 H, J=6.8 Hz), 1.45 (s, 9 H), 3.48-3.53 (m, 1 H), 3.62-3.66 (m, 1 H), 3.76-3.77 (m, 1 H); MS (FAB) m/z 176 (M++1), 120 (Mt Bu). To a stirred solution of ethyl 4-hydroxy-3-methoxybenzoate (7.74 g, 0.04 mmol), (R)-2-N 25 tert-butoxycarbonylamino-1-propanol (6.91 g, 0.04 mmol) and Ph 3 P (13.44 g, 0.05 mmol) in THF (70 mL) was added diisopropyl azodicarboxylate(DIAD)(10.0 mL, 0.05 mmol), and the resulting mixture was heated under reflux overnight. After cooling to room temp, the solvent was evaporated. The mixture was dissolved in CH 2 C1 2 (50 mL) and TFA (30 mL) and the solution was stirred at room temp for 1 hr. After concentration in vacuo, the residue was poured into sat. 395 WO 01/00206 PCT/US00/18079 NaHCO 3 and extracted with CHC1 3 . The extract was washed with brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with 5% MeOH in
CHCI
3 as eluent to give 7.93g (2 steps 79%) ethyl (R)-3-methoxy-4-(2-amino-1-propoxy)benzoate as a yellow oil. 'H-NMR (CDCl 3 ) 8 1.90 (d, 3 H, J=6.8 Hz), 1.39 (t, 3 H, J=7.3 Hz), 1.72 (bs, 2 5 H), 3.42-3.47 (m, 1 H), 3.74-3.89 (m, 1 H), 3.91 (s, 3 H), 3.96-4.00 (m, 1 H), 4.35 (q, 2 H, J=7.3 Hz), 6.88 (d, 1 H, J=8.3 Hz), 7.55 (d, 1 H, J=2.0 Hz), 7.65 (dd, 1 H, J=2.0, 8.3 Hz); MS (FAB) m/z 254 (M
+
+1). To a stirred solution of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetate (459 mg, 0.96 mmol) and ethyl (R)-3-methoxy-4-(2-aminopropoxy)benzoate (242 10 mg, 0.96 mmol) in DMF (5 mL) was added Et 3 N (200 ml, 1.43 mmol), and the resulting mixture was stirred for 2 hr. The mixture was diluted with EtOAc, washed with 0.5 N HCI, brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (50:1, v/v) as eluent to give 360 mg (69%) ethyl (R)-3-methoxy-4-[2-[3 methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetylamino]-1-propoxy]benzoate as a colorless 15 crystalline solid. 'H-NMR (CDCl 3 ) 5 1.23-1.28 (m, 3 H), 1.38-1.41 (t, 3 H, J=7.3 Hz), 2.32 (s, 3 H), 3.50-4.13 (m, total 11 H), 4.36 (q, 2 H, J=7.3 Hz), 6.65-8.13 (series of m, total 12 H). To a stirred solution of ethyl (R)-3-methoxy-4-[2-[3-methoxy-4-[N'-(2-methylphenyl) ureido] phenylacetylamino]-1-propoxy]benzoate (360 mg, 0.66 mmol) in THF-MeOH (20 mL, 9:1, v/v) was added 0.25 N NaOH (10 mL), and the resulting mixture was heated under reflux 20 overnight. The mixture was poured into ice-1 N HCI, and precipitate was collected. The crude solid was recrystallized from CHC13-n-hexane to give 172 mg(50%) 260 as a white crystalline powder. mp 168-169 oC; IR (KBr) 1687 cm'; 'H-NMR (DMSO-d 6 ) 8 1.18 (d, 3 H, J=6.8 Hz), 2.24 (s, 3 H), 2.50-2.51 (m, 2 H), 3.80 (s, 3 H), 3.84 (s, 3 H), 3.87-4.06 (m, 2 H), 4.07-4.14 (m, 1 H), 6.76-8.57 (series of m, total 12 H), 12.66 (bs, 1 H); MS (FAB) m/z 522 (Mf+1); Anal. Calcd 25 for C 28
H
3 1
N
3 0 7 ,-3/4 H 2 0: C, 62.85; H, 6.12; N, 7.85. Found: C, 62.77; H, 5.95 N, 7.79. Example 213 4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]allylamino]ethoxy]benzoic acid H2 N NN OCH 3
CH
3 H H OCH 3 261 396 WO 01/00206 PCT/US00/18079 To a stirred mixture of ethyl 4-(2-N-trifluoroacetylaminoethoxy)-3-methoxy benzoate (3.5g, 10.4mmol)and K 2 C0 3 (2.3g, 16.4mmol) in DMF (20mL) was added allyl bromide (14.2mL, 16.5mmol), and the resulting mixture was stirred for 45 min at 650 C. After cooling, water was added to the mixture and extracted with EtOAc. The extract was washed with brine, dried over 5 Na 2
SO
4 , and evaporated in vacuo. The residue was dissolved in THF-MeOH-H 2 0 (1:1: 1,v/v/v) (30 mL) and added K 2
CO
3 (2.3 g, 16.4 mmol). The resulting mixture was stirred for 16 hr at room temp. The mixture was diluted with EtOAc, washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica-gel with CHCl 3 :MeOH (95:5 to 95:5, v/v) as eluent to give 2.9 g (100%) ethyl 4-(2-allylaminoethoxy)-3-methoxybenzoate as a pale-yellow 10 oil. 'H-NMR(CDC 3 , 400MHz) 8 1.39 (t, 3H, J=7.3Hz), 3.07 (t, 2H, J=5.3Hz), 3.34 (d, 2H, J=5.9Hz), 3.91 (s, 3H), 4.18 (t, 2H, J=5.4Hz), 4.35 (dd, 2H, J=7.3Hz, 14.1Hz), 5.12 (d, 1H, J=10.3Hz), 5.22 (dd, 1H, J=1.5Hz, 17.1Hz), 5.92 (m, 2H), 6.90 (d, 1H, J=8.3Hz), 7.55 (d, 1H, J=1.5Hz), 7.65 (dd, 1H, J=2.0Hz, 8.3Hz); MS(FAB) nm/z 278, 280(M+H)*. To a stirred mixture of ethyl 4-(2-allylaminoethoxy)-3-methoxy benzoate (578mg, 15 2. Immol), 3-methoxy-4-[N-(2-methylphenyl)ureido]phenylaceticacid (650mg,2. Immol), HOBt (420mg, 3.1 1mmol), and DMAP (catalytic amount) in DMF (4 mL) was added EDC (596mg,3.1 1mmol) at room temp. The resulting mixture was stirred for a further 18 hr at room temp. The mixture was poured into IN HCI and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated in vacuo. The residue was chromatographed on 20 silica-gel with CHCl 3 :EtOAc (95:5 to 1:1, v/v) as eluent to give Ig (84%) 3-methoxy-4-[[2-[3 methoxy-4-[NM-(2-methylphenyl)ureido] phenylacetyl]allylamino]ethoxy] benzoic acid as a pale yellow gum. 'H-NMR(CDC13, 400MHz) 8 1.39 (t, 3H, J=7.3Hz), 2.29 (s, 3H), 3.58 and 3.63 (s, total 3H1), 3.70-3.77 (m, 2H), 3.83 and 3.87 (s, 3H11), 4.05-4.13 (m, 2H), 4.25 (m, 1H), 4.36 (q, 1H, J=7.0Hz), 5.04-5.22 (m, 2H), 5.73 (m, 1H), 6.32 and 6.47 (s, 1IH), 6.69-6.85 (m, 2H11), 7.12 (m, 25 2H), 7.23 (m, 2H11), 7.50-7.65 (m, 2H), 8.05 (d, 1H, J=7.8Hz); MS (FAB) m/z 576(M+H)*. A mixture of 3-methoxy-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] allylamino]ethoxy]benzoic acid (50mg, 0.09mmol) in THF-MeOH (1:1, v/v) (2mL) and IN NaOH (0.135mL, 0.135mmol was stirred for 15 hr at room temp and 3 hr at 50oC. The mixture was poured into ice-IN HCI. Solid was collected, washed with water, and air-dried. The crude solid 30 was recrystallized from CHCl 3 -n-hexane to give 38mg (77%) 261 as a white crystalline material. mp 125-130 oC; IR(KBr), 3319, 2939, 1687, 1647, 1601, 1535, 1456, 1417, 1269, 1223, 1034, 760cm-'; 'H-NMR(DMSO-d 6 , 400MHz) 8 2.29 (s, 3H), 3.68 (s, 2H11), 3.75-3.85 (m, 8H), 4.05 (br, 397 WO 01/00206 PCT/US00/18079 1H1), 4.19 (min, 3H), 5.10-5.25 (min, 2H), 5.65-5.90 (min, 1H), 6.75 (min, 1H), 6.85 (s, 1H), 6.92 (min, 1H1), 7.02-7.20 (min, 3H), 7.48 (d, 1H, J=10.2Hz), 7.56 (min, 1H), 7.79 (d, 1H, J=6.8Hz), 8.01 (min, 1H1), 8.46 (s, 1H), 8.56 (d, 1H, J=4.4Hz), 12.7 (br, 1H); MS (FAB) m/z 548(M+H)+; Anal. calcd. for
C
30
H
3 3
N
3 07-0.5H 2 0, C, 64.74; H, 6.16; N, 7.55. Found, C, 64.72; H, 6.07; N, 7.55. 5 Example 214 3-methoxy-4-[[2-[3-methoxy-4-[N-(2-methylphenyl)ureido]phenylacetyl]-(2-morpholino) ethylamino]ethoxy]benzoic acid COOH O N\ N O OCH 3
CH
3 H H CH 3 262 To a stirred solution of 3-methoxy-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido] 10 phenylacetyl]allylamino]ethoxy]benzoic acid (950mg, 1.65mmol) in TIHIF:H 2 0 (7mL) was added N-methylmorpholine-N-oxide (579mg,4.95mmol) and osmium tetroxide (0.2M solution in water) (0.413mL, 0.08mmol). The resulting mixture was stirred for 3 hr at room temp. Sat. NaHSO 3 was added to the mixture, and the mixture was filtered through Celite. The filtrate was extracted with EtOAc. The extract was washed with brine, dried over MgSO 4 , and evaporated in vacuo. The 15 residue was dissolved in MeOH-THF-H 2 0 (1:1:1, v/v) (12mL) and added sodium periodate (318mg,1.5mmol). The resulting mixture was stirred at an ambient temp for 1 hr. The mixture was diluted with EtOAc, washed with brine, and dried over MgSO 4 . Solvent was evaporated in vacuo to afford 862mg (90%) ethyl 3-methoxy-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-N-formylmethylamino]ethoxy] benzoate as a pale-yellow gum. 'H-NMR (CDCI 3 , 20 400MHz) 8 1.39 (t, 3H, J=7.3Hz), 2.29 (s, 3H), 3.31-3.95 (min, llH), 4.10-4.42 (min, 5H11), 6.51-6.82 (mn, 3H11), 7.10-7.25 (m, 3H), 7.50 (m, 2H), 7.60 (min, 1H1), 8.10 (min, 1H), 9.50 (mn, 1H11); MS (FAB) m/z 578 (M+H) +. To a stirred mixture of ethyl 3-methoxy-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-N-formnylmethylamino]ethoxy]benzoate (265mg, 0.46mmol), morpholine (0.40mL, 25 4.59mmol), and AcOH (0.263mL, 4.6mmol) in EtOH (3mL) was added NaBH 3 CN (288mg, 4.6mmol) at room temp. The resulting mixture was stirred for 15 hr at room temp and the mixture was diluted with EtOAc and added sat. NaHCO 3 at 0' C. The resulting mixture was stirred for 0.5 hr at 00 C. The mixture was extracted with EtOAc. The extract was washed with brine, dried over MgSO 4 , and evaporated in vacuo. The residue was chromatographed on silica 398 WO 01/00206 PCT/US00/18079 gel with CHCI 3 :MeOH (95:5, v/v) as eluent to give 213mg (71%) ethyl 3-methoxy-4-[[2-[3 methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-(2-morpholino)ethylamino]ethoxy]benzoate as an oil. 'H-NMR (CDCl 3 , 400MHz) 5 1.28 (t, 3H, J=7.0Hz), 2.31(s, 3H), 2.48 (brs, 4H), 2.52 (m, 2H), 3.60-3.91 (m, 16H), 4.11 and 4.28 (m, total 2H1), 4.39 (q, 2H, J=7.0Hz), 6.70-6.85 (m, 5 4H), 7.15 (m, 2H), 7.50-7.63 (m, 3H), 8.08 (d, 1H, J=8.0Hz); MS (FAB) nm/z 649(M+H)*. A mixture of ethyl 3-methoxy-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl acetyl]-(2-morpholino)ethylamino]ethoxy]benzoate (265mg, 0.46mmol) in THF (4mL) and IN NaOH (0.984mL) was stirred at 500 C for 15 hr. The pH of the mixture was adjusted to 7.4 by the addition of IN HCI, and extracted with CHCl 3 :MeOH(9:1, v/v). The extract was washed with 10 brine, dried over MgSO 4 , and evaporated in vacuo. The residue was crystallized with Et 2 O to give 160 mg(78%) 262 as a white crystalline material. mp 125-130 oC; IR (KBr), 3346, 2956, 2937, 1705, 1622, 1599, 1537, 1456, 1417, 1299, 1114, 1032, 752cmr'; 1 H-NMR (CD 3 OD, 400MHz) 8 2.29 (s, 3H1), 2.49-2.64 (m, 6H), 3.65-3.85 (m, 16H), 4.13 (m, 1H), 4.26 (m, 1H11), 6.78-7.04 (m, 4H), 7.18 (m, 2H), 7.55-7.64 (m, 3H11), 7.99 (m, 2H); MS (FAB) m/z 621(M+H); Anal. Calcd. for 15 C 33 H40N 4 Os-2.5H 2 0, C, 59.54; H, 6.81; N, 8.42. Found, C, 59.71; H, 6.35; N, 7.98. Example 215 3-methoxy-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-[2-[4-methyl- 1 piperazinyl]ethylamino]ethoxy]benzoic acid
CH
3 0 ~ ~- OOH NON OCH 3
CH
3 H H OCH 3 263 20 To a stirred mixture of ethyl 3-methoxy-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-N-formylmethylamino]ethoxy]benzoate (242mg, 0.42mmol), N-methylpiperazine (0.465mL, 4.2mmol), and AcOH (0.240mL, 4.2mmol) in EtOH (3mL) was added NaBH 3 CN (263mg, 4.2mmol) at room temp. The resulting mixture was stirred for 15 hr at room temp. The mixture was diluted with EtOAc and added sat. NaHCO 3 at 0o C. The resulting mixture was 25 stirred for 0.5 hr at 0o C. The mixture was extracted with EtOAc. The extract was washed with brine, dried over MgSO 4 , and evaporated in vacuo. The residue was chromatographed on silica gel with CHCl 3 :MeOH (95:5, v/v) as eluent to give 195mg (70%) ethyl 3-methoxy-4-[[2-[3 methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-[2-(4-methyl-1-piperazinyl]ethylamino] 399 WO 01/00206 PCT/US00/18079 ethoxy]benzoate as an oil. 'H-NMR (CDC1 3 , 400MHz) 8 1.23 (t, 3H, J=7.0Hz), 2.25 (s, 3H), 2.29 (s, 3H), 2.50 (br m, 12H), 3.44-3.85 (m, 12H), 4.10 (br, 1H), 4.22 (br, 1H), 4.35 (m, 2H), 6.70 6.85 (m, 3H), 6.98 (s, 1H), 7.10 (m, 1H), 7.20 (m, 2H), 7.40 (m, 1H), 7.60-7.70 (m, 3H), 8.05( d, 1H, J=7.8Hz); MS (FAB) m/z 662(M+H) . 5 A mixture of ethyl 3-methoxy-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-[2-(4-methyl-l1-piperazinyl]ethylamino]ethoxy]benzoate (195mg, 0.30mmol) in THF:MeOH(4:1, v/v) (5mL) and IN NaOH (0.885mL) was stirred at 500 C for 15 hr. The pH of the mixture was adjusted to 7.4 by the addition of IN HC1, and extracted with CHCl 3 :MeOH(9:1, v/v). The extract was washed with brine, dried over MgSO 4 , and evaporated in vacuo. The 10 residue was crystallized with Et 2 0 to give 141 mg(75%) 263 as a white crystalline material. mp 155-160 oC; IR (KBr), 2937, 1537, 783cm-'; 'H-NMR (CD 3 OD, 400MHz) 8 2.29 (s, 3H), 2.49-2.80 (m, 15H), 3.60-3.85 (m, 9H), 3.92 (s, 1H), 4.12 (m, 1H), 4.25 (m, 1H), 6.78-7.20 (m, 6H), 7.61 (m, 3H), 8.00 (m, 1H); MS (FAB) m/z 632(M)+; Anal. Calcd. for C 34
H
43 N507-2.5H 2 0, C, 60.16; H, 7.13; N, 10.32. Found, C, 59.72; H, 6.86; N, 9.97. 15 Example 216 3-methoxy-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-[2-cyclopropylamino] ethylamino]ethoxy]benzoic acid HA r) O- COOH \ N N OCH 3 CH3H H CH3 264 To a stirred mixture of ethyl 3-methoxy-4-[[2-[3-methoxy-4-[N-(2-methylphenyl)ureido] 20 phenylacetyl]-N-formylmethylamino]ethoxy]benzoate (267mg, 0.46mmol), cyclopropylamine (0.32mL, 4.6mmol), and AcOH (0.264mL, 4.6nunmmol) in EtOH (3mL) was added NaBHzCN (290mg, 4.6mmol) at room temp. The resulting mixture was stirred for 15 hr at room temp. The mixture was diluted with EtOAc and added sat. NaHCO 3 at 0o C. The resulting mixture was stirred for 0.5 hr at 0oC. The mixture was extracted with EtOAc. The extract was washed with 25 brine, dried over MgSO 4 , and evaporated in vacuo. The residue was chromatographed on silica gel with CHC1 3 :MeOH (95:5, v/v) as eluent to give 156mg (55%) ethyl 3-methoxy-4-[[2-[3 methoxy-4-[N-(2-methylphenyl)ureido]phenylacetyl]-[2-cyclopropylamino]ethylamino] ethoxy]benzoate as an oil. 'H-NMR (CDCl 3 , 400MHz) 8 0.35 (m, 4H), 1.22 (br s, 3H), 2.10 (m, 1H), 2.20 (s, 3H), 2.42 (br, 2H), 2.90 (br s, 2H), 3.60-3.80 (m, 10H), 4.10 (br, 1H), 4.22 (br, 1H), 400 WO 01/00206 PCT/US00/18079 4.33 (br, 2H), 6.72 (m, 3H), 7.05-7.30 (m, 4H), 7.55 (m, 4H), 8.06 (br s, 1H); MS (FAB) m/z 619(M+H), A mixture of ethyl 3-methoxy-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-N-[2-cyclopropylamino]ethylamino]ethoxy]benzoate (195mg, 0.30mmol)) in 5 THF:MeOH(4: 1, v/v) (5mL) and IN NaOH (0.756mL) was stirred at 500 C for 15 hr. The pH of the mixture was adjusted to 7.4 by the addition of IN HCI, and extracted with CHC1 3 :MeOH(9: 1, v/v). The extract was washed with brine, dried over MgSO 4 , and evaporated in vacuo. The residue was crystallized with Et20 to give 57 mg (38%) 3-methoxy-4-[[2-[3-methoxy-4-[N-(2 methyl phenyl)ureido]phenylacetyl]-[2-cyclopropylamino] ethylamino]ethoxy]benzoic acid 264 as 10 a white crystalline material. mp 135-140 oC; IR (KBr), 3324, 2937, 1535, 1032, 754cm-'; 'H NMR (CD 3 OD, 400MHz) 5 0.50-0.73 (m, 4H), 2.29 (s, 3H), 2.53 (m, 1H), 2.98 (m, 1H), 3.21 (m, 1H), 3.58-3.88 (m, 11H), 3.91 (s, 1H), 4.09 (m, 1H), 4.25 (m, 1H), 6.76-6.92 (m, 3H), 7.01 (m, 1H), 7.18 (m, 2H), 7.60 (m, 3), 8.00 (d, J=8.3Hz, 1H); MS (FAB) m/z 591(M+H)+; Anal. Calcd. for C 32
H
38
N
4 0 7 3.0H 2 0, C, 59.62; H, 6.88; N, 8.69. Found, C, 59.25; H, 6.29; N, 8.29. 15 Example 217 4-[[2-[3-methoxy-4-[N '-(2-fluorophenyl)ureido]phenylacetyl]-N-methylamino]ethoxy]benzoic acid
CH
3 O COOH F H H OCH 3 265 3-chloro-4-[[2-[3-methoxy-4-[N '-(2-fluorophenyl)ureido]phenylacetyl]-N-methylaminoethoxy] benzoic acid
NH
3 N COOH N-ZN 0 CI 20 F H H OCH 3 266 To a stirred cold (0oC) solution of 2-(N-benzyloxycarbonyl-N-methyl)ethanolamine (3.01 g, 14.4 mmol), methyl 3-chloro-4-hydroxybenzoate (2.68 g, 14.4 mmol), Ph 3 P (5.65 g, 21.5 mmol) in THF (30 mL) was added diisopropyl azodicarboxylate (DIAD) (4.25 mL, 21.6 mmnol), and the resulting mixture was heated under reflux overnight. The solution was evaporated off and the 25 residue was purified by column chromatography on silica-gel with CHC1 3 as eluent to give 3.90g (72%) methyl 3-chloro-4-[2-(N-benzyloxycarbonyl-N-methylamino)ethoxy]benzoate as a pale yellow solid. 'H-NMR (CDC13) 8 3.15 (s, 3 H), 3.74-3.76 (m, 2 H), 3.89 (s, 3 H), 4.17-4.27 (m, 2 H), 5.14 (s, 2 H), 6.81-6.94 (m, 1 H), 7.33-7.36 (m, 5 H), 7.85-7.92 (m, 1 H), 8.05 (bs, 1 H). 401 WO 01/00206 PCT/US00/18079 A solution of methyl 3-chloro-4-[2-(N-benzyloxycarbonyl-N-methylamino)ethoxy] benzoate (3.90 g, 10.3 mmol) in EtOAc-AcOH (40 mL, 1:1, v/v) was hydrogenated over 5% Pd-C (1.95 g, 50 wt%) at 3 atm for 2 hr. The mixture was filtered and the filtrate was washed with sat. NaHCOz and the basic aqueous layer was extracted with CHCl 3 , washed with brine and evaporate 5 to give unseparable mixture of methyl 3-chloro-4-(N-methylaminoethoxy)benzoate and methyl 4 [2-(N-methylamino) ethoxy]benzoate the title compound (1.61 g) as a pale yellow oil. 'H-NMR (CDC13) 8 2.52-2.52 and 2.53-2.54 (each m, 3 H), 2.98-3.00 and 3.03-3.05 (each m, each 2 H), 3.88 and 3.99 (each s, each 3 H), 4.11-4.14 and 4.18-4.20 (each m, each 2 H), 6.91-6.96 and 7.90 8.05 (series of m, total 7 H). 10 A mixture of 3-methoxy-4-[N'-(2-fluorophenyl)ureido]phenylacetic acid (392 mg), a mixture of methyl 3-chloro-4-[2-(N-methylamino)ethoxy]benzoate and methyl 4-[2-(N methylamino)ethoxy] benzoate (305 mg), EDC(hydrochloride) (354 mg), HOBt (250 mg), and DMAP (250 mg) in DMF (8 mL) was stirred at room temp for 6 hr. The mixture was diluted with 15 EtOAc, washed with 0.5 N HCI, brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with 1% MeOH in CHCI 3 as eluent to give a mixture of methyl 3-chloro-4-[[2-[3-methoxy-4-[N'-(2-fluorophenyl) ureido]phenylacetyl]-N methylaminoethoxy]benzoate and methyl 4-[[2-[3-methoxy-4-[N'-(2-fluorophenyl)ureido] phenylacetyl]-N-methylaminoethoxy]benzoate (550 mg) as a brown amorphous solid. 20 To a stirred solution of this mixture (550 mg) of methyl 3-chloro-4-[[2-[3-methoxy-4-[N' (2-fluorophenyl)ureido]phenylacetyl]-N-methylaminoethoxy]benzoate and methyl 4-[[2-[3 methoxy-4-[N'-(2-fluorophenyl)ureido]phenylacetyl]-N-methylaminoethoxy]benzoate in THF MeOH (20 mL, 1:1, v/v) was added 0.5 N NaOH (10 mL), and the resulting mixture was heated under reflux for 6 hr. The mixture was poured into ice-1 N HCI, and the solid was collected. The 25 crude solid was purified by preparative TLC with 10% MeOH in CHC1 3 as eluent to give 265 (56 mg, as a white amorphous solid) and 266 (88 mg, as a brown amorphous solid). Example 218 4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]methylamino]ethoxy]benzoic acid
CNH
3 N COOH
CH
3 H H OCH 3 267 30 3-chloro-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]methylamino]ethoxy] 402 WO 01/00206 PCT/US00/18079 benzoic acid
CH
3 - COOH NON C O
CH
3 H H OCH 3 268 A mixture of methyl 4-[2-(N-methyl-2-amino)ethoxy]-3-chlorobenzoate (292mg, 1.2mmol), 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylaceticacid (377mg,1.2mmol), EDC 5 (345mg,1.8mmol), HOBt(243mg,1.8mmol), and DMAP(29mg, 0.24mmol) in DMF(2.7mL) was stirred for 6 hr at room temp. The mixture was poured into ice-IN HCI and extracted with EtOAc. The extract was washed with brine, dried over MgSO 4 , and evaporated in vacuo. The residue was chromatographed on silica-gel with CHC1 3 :EtOAc (95:5 to 0:100, v/v) as eluent to give unseparable mixture (489mg) of methyl 3-chloro-4-[[2-[3-methoxy-4-[N'-(2 10 methylphenyl)ureido]phenylacetyl] methylamino]ethoxy]benzoate and methyl 4-[[2-[3-methoxy-4 [N'-(2-methylphenyl)ureido]phenyl acetyl]methylamino]ethoxy]benzoate as pale-yellow oil. A mixture (480mg as mixture) of methyl 3-chloro-4-[[2-[3-methoxy-4-[N'-(2-methyl phenyl)ureido] phenylacetyl]methylamino]ethoxy]benzoate and methyl 4-[[2-[3-methoxy-4-[N'-(2 methylphenyl)ureido] phenylacetyl]methylamino]ethoxy]benzoate in THF-MeOH(4mL, 1:1, v/v) 15 was stirred at 500 C for 15 hr. The mixture was poured into ice-IN HCL. The solid was collected, washed with water, and air-dried. The crude solid was purified by preparative TLC CHCl3:MeOH (93:7, v/v) as eluent to afford 267 (180mg, 2steps 31% as a crystalline material) and 268 (280mg, 2steps 44% as a crystalline material). 267: mp 145-150 oC; 'H-NMR (DMSO-d, 400MHz) 8 2.31 (s, 3H), 3.05 and 3.19 (s, 3H), 3.35 and 3.38 (s, 3H), 3.72-3.85 (mn, 7H), 4.09 and 4.23 (min, total 20 2H11), 6.79-7.20 (min, 7H11), 7.60 (min, 1H), 7.86-8.09(m, 3H); MS (FAB) m/z 493(M+H)*; Anal. calcd. for C 27
H
29
N
3 0 6 -1.75H 2 O, C, 62.00; H, 6.26; N, 8.03. Found, C, 62.16; H, 5.88; N, 7.82. 268: mp 145-150 oC; 'H-NMR (DMSO-d,, 400MHz) 8 2.29 (s, 3H), 3.06 and 3.26 (s, 3H11), 3.31 and 3.35 (s, 3H), 3.85-3.94 (min, 4H11), 4.18 and 4.32 (min, total 2H11), 6.75-6.85 (min, 2H), 6.99-7.20 (min, 4H), 7.59 (min, 1H), 7.90-8.02 (min, 3H); MS (FAB) m/z 526(M+H)+; Anal. calcd. for C 27
H
2 8
CIN
3 0 6 -2.0H 2 0, C, 25 57.70; H, 5.74; N, 7.48. Found, C, 57.99; H, 5.53; N, 7.07. Example 219 4-[3-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-N-methylamino]-1-propyl]benzoic acid 403 WO 01/00206 PCT/US00/18079
CH
3 NN N'aCOOH
CH
3 H H OCH 3 269 To a stirred cold (minus 780 C) solution of triethyl 4-phosphonomethylbenzoate (1.22 g, 4.05 mmol) in THF (10 mL) was added NaHMDS (1.0 M in THF) (4.0 mL, 4.0 mmol), and the resulting mixture was stirred for 1 hr at the same temp. A solution of 2-(N-benzyloxycarbonyl-N 5 methylamino) acetaldehyde (700 mg, 3.38 mmol) in THBF (5 mL) was slowly added to this solution at that temp, and the mixture was allowed to warm to room temp for over 2 hr with stirring. The solution was quenched by the addition of sat. NH 4 Cl (100 mL), and extracted with EtOAc. The extract was washed with brine (200 mL), dried over MgSO 4 , and evaporated. The residue was chromatographed on silica gel with CHCl 3 -EtOAc (20:1, v/v) as eluent to give 810 mg (68%) ethyl 10 (E)-4-[3-(N-benzyloxycarbonyl-N-methylamino)-1-propenylbenzoate as a yellow oil. 'H-NMR (CDCl 3 ) 8 1.40 (t, J= 7.3 Hz, 3 H), 2.95 (min, 2 H), 4.09 (min, 2 H), 4.35-4.40 (min, 2 H), 5.17 (s, 2 H), 6.26-6.64 (series of m, 2 H), 7.36 (min, 7 H), 7.99 (d, J = 8.3 Hz, 2 H). A stirred solution of ethyl (E)-4-[3-(N-benzyloxycarbonyl-N-methylamino)-1-propenyl benzoate (810 mg, 2.29 mmol) in EtOH-AcOH (10:1, v/v, 22 mL) was hydrogenated over 5% Pd 15 C (1 g) for 3 days. The mixture was filtered and the filtrate was evaporated. The residue was made basic with sat. NaHCO 3 and extracted with CHCI 3 . The extract was dried over Na 2
CO
3 and evaporated to give 438 mg (86%) ethyl 4-(3-methylamino-1-propyl)benzoate as a yellow oil. 'H NMR (CDCl 3 ) 5 1.39 (t, J=7.3 Hz, 3 H), 1.82 (min, 2 H), 2.43 (s, 3 H), 2.61 (t, J= 7.3 Hz, 2 H), 2.72 (t, J= 7.3 Hz, 2 H), 3.33 (br s, 1 H), 4.36 (q, J= 7.3 Hz, 2 H), 7.25 (d, J= 8.3 Hz, 2 H), 7.96 20 (d, J= 8.3 Hz, 2 H). To a stirred solution of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (456 mg, 1.45 mmol) and ethyl 4-(3-methylamino-1-propyl)benzoate (220 mg, 1.45 mmol) were added EDC-HCI (417 mg, 2.16 mmol), HOBt (cat.), and DMAP (catalytic amount) in DMF (10 mL), and the resulting mixture was stirred overnight. The mixture was diluted with EtOAc (300 mL), 25 washed with brine, dried over MgSO 4 , and evaporated. The residue was chromatographed on silica-gel with CHC13-EtOH (10:1) to give 503 mg (71%) ethyl 4-[3-[3-methoxy-4-[N'-(2 methylphenyl) ureido]phenylacetyl-N-methylamino]-1-propyl] benzoate as a yellow oil. MS (FAB) m/z 518(M+H) t . To a stirred solution of ethyl 4-[3-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl 30 acetyl-N-methylamino]-1-propyl]benzoate (500 mg, 0.966 mmol) in THF (8 mL) was added 0.25 404 WO 01/00206 PCT/US00/18079 N NaOH (8 mL), and the mixture was heated under reflux overnight. The resulting solution was poured into ice-1 N HCI (100 mL) and the solid was collected with suction. The solid was dissolved in CHC1 3 (100 mL) and dried over MgSO 4 . After removal of the solvent, the residue was chromatographed on silica gel with CHCl 3 -MeOH (10:1 to 5:1, v/v) to give 131 mg (28%) 4-[3-[3 5 methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl-N-methylamino]-1-propyl]benzoic acid 269 as a pale yellow amorphous solid. 'H-NMR (DMSO-d 6 ) 8 1.68-1.80 (m, 2 H), 2.24 (s, 3 H), 2.57 (m, 2 H), 2.81 and 2.97 (s, each, total 3 H), 3.33 (m, 2 H), 3.57-3.61 (m, 2 H), 3.84 (s, 3 H), 6.71 (dd, J = 29.8, 8.3 Hz, 1 H), 6.87 (d,J= 11.2 Hz, 1 H), 6.93 (t,J= 7.3 Hz, 1 H), 7.15 (m, 2H), 7.28 (m, 2 H), 7.79 (d, J = 8.3 Hz, 1 H), 7.84-7.87 (m, 2 H), 8.02 (d, J = 8.3 Hz, 1 H), 8.49 (d, J= 10 7.3 Hz, 1 H), 8.58 (d, J = 4.9 Hz, 1 H); MS (FAB) m/z 490 (M +1); Anal. Calcd for
C
28
H,
31
N
3 0 5 -1/2H 2 0: C, 67.45; H, 6.47; N, 8.43. Found: C, 67.27; H, 6.51; N, 8.02. Example 220 4-[[2-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]-N-methylamino]ethoxy]benzoic acid
H
3 OCOOH _N N
CH
3 H H 270 15 3-chloro-4-[[2-[4-[N '-(2-methylphenyl)ureido]phenylacetyl]-N-methylamino]ethoxy]benzoic acid
NH
3 fO' COOH 9HA .'.-N..".N.x OCI
CH
3 H H 271 A solution of pentafluorophenyl 4-[N'-(2-methylphenyl)ureido]phenylacetate (562 mg, 1.29mmol), a mixture (304 mg) of methyl 3-chloro-4-[2-(N-methylamino)ethoxy]benzoate and methyl 4-[2-(N-methyl amino)ethoxy]benzoate and Et 3 N (260 mL) in DMF (8 mL) was stirred at 20 room temp for 4 hr. The mixture was diluted with EtOAc, washed with 0.5 N HC1, brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (50:1, v/v) as eluent to give a mixture (670 mg) of methyl 3-chloro-4-[[2-[4 [N'-(2-methylphenyl)ureido) phenylacetyl]-N-methylamino) ethoxy]benzoate and methyl 4-[[2-[4 [N'-(2-methylphenyl)ureido] phenylacetyl]-N-methylamnino)ethoxy] benzoate as an oil. 25 To a stirred suspension of this mixture (670 mg) in THF-MeOH (20 mL, 1:1, v/v) was added 0.5 N NaOH (10 mL) and the resulting mixture was heated under reflux for 6 hr. The solution was poured into ice- 1 N HCI and the solid was collected. The crude solid was purified by 405 WO 01/00206 PCT/US00/18079 preparative thin layer chromatography (TLC) with 10% MeOH in CHCl 3 as eluent to give 73mg 270 as an amorphous solid and 110 mg 271 as a white amorphous solid. 270 MS (FAB) m/z 462 (M +1). 271 MS (FAB) m/z 496 (M'+1). Example 221 5 (S)-4-[2-[3-methoxy-4-[N '-(2-methylphenyl)ureido]phenylacetylaminol]-1-propoxy]benzoic acid H N
"N
C O O H
PW
1 ANJC CH 3
CH
3 H H OCH 3 272 To a cooled (00 C) solution of (S)-2-amino-1-propanol (2.08 g, 27.7 mmol) and Et 3 N (4.63 mL, 33.2 mmol) in DMF-H 2 0 (40 mL, 1:1, v/v) was added (Boc) 2 0 (6.36 mL, 27.7 mmol), and the resulting solution was stirred at room temp for 2 days. H 2 0 was added to the mixture and 10 extracted with EtOAc. The extract was washed with brine and dried over Na 2
SO
4 . The solvent was evaporated to give 4.24 g (87%) (S)-2- (N-tert-butoxycarbonylamino)-1-propanol as a colorless oil. 'H-NMR (CDC13) 6 1.14 (d, 3 H, J=6.8 Hz), 1.45 (s, 9 H), 3.51-3.52 (min, 1 H), 3.63 3.66 (min, 1 H), 3.77 (min, 1 H), 4.62 (min, 1 H). To a cooled (00 C) solution of (S)-2-(N-tert-butoxycarbonylamino)-l1-propanol (1.02 g, 15 5.82 mmol), methyl 4-hydroxybenzoate (0.89 g, 5.85 mmol), and Ph 3 P (1.98 g, 7.55 mmol) in THF (20 mL) was added diisopropyl azodicarboxylate (DIAD) (1.49 mL, 7.57 mmol), and the resulting mixture was heated under reflux overnight. The solution was evaporated and the residue was dissolved in CH 2 C1 2 (20 mL) and TFA (10 mL). The mixture was stirred at room temp for 1.5 hr. The solution was concentrated in vacuo and the residue was treated with sat. NaHCO 3 . The 20 mixture was extracted with CHC1 3 , washed with brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHC 3 :MeOH (50:1, v/v) to give 480 mg (2 steps 39%) methyl (S)-4-(2-amino-1-propoxy) benzoate as a pale yellow oil. 'H-NMR (CDCl 3 ) 8 1.19 (d, 3 H, J=6.4 Hz), 3.35-3.39 (min, 1 H), 3.72-3.76 (mn, 1 H), 3.89 (s, 3 H), 3.90-3.94 (min, 1 H), 6.92 (d, 2 H, J=8.8 Hz), 7.99 (d, 2 H, J=8.8 Hz). 25 A mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetate (505 mg, 1.05 mmol), methyl (S)-4-(2-amino-1-propoxy)benzoate (220 mg, 1.05 mmol), and Et 3 N (0.220mL, 1.58 mmol) in DMF (8 mL) was stirred at room temp for 3 hr. The mixture was diluted with EtOAc, washed with 0.5 N HC1, brine, and dried over Na 2
SO
4 . After removal of the solvent, the residue was recrystallized from MeOH-CHCl 3 -n-hexane to give 290 mg (55%) methyl 406 WO 01/00206 PCT/US00/18079 (S)-4-[2-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenylacetylamino]-l1-propoxy]benzoate as a white crystalline powder. 'H-NMR (DMSO-d 6 ) 8 1.18 (d, 3 H, J=6.8 Hz), 2,24 (s, 3 H), 3.36 (s, 2H), 3.80 (s, 3H), 3.82 (s, 3H), 3.93-4.03 (min, 2H), 4.09-4.14 (min, 1H), 6.75-8.57 (series of m, total 13 H). 5 To a stirred solution of methyl (S)-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl acetylamino]-1-propoxylbenzoate (290 mg, 0.57 mmol) in THF-MeOH (20 mL, 1:1, v/v) was added 0.5 N NaOH (20 mL) and the solution was heated under reflux for 2 hr. The mixture was poured into ice-1 N HCI and extracted with CHCl 3 -MeOH (10 :1, v/v). The extract was washed with brine, dried over Na 2
SO
4 and evaporated. The residue was recrystallized from MeOH-CHCl1 3 10 n-hexane to give 158 mg (56%) 272 as a white crystalline powder. mp 198-201 oC; 'H-NMR (DMSO-d 6 ) 5 1.18 (d, 3 H, J=6.3 Hz), 2.24 (s, 3 H), 3.36 (s, 2 H), 3.82 (s, 3 H), 3.87-4.10 (min, 2 H), 4.10-4.16 (min, 1 H), 6.75-6.78 (min, 1 H), 6.92-7.02 (min, 4 H), 7.11-7.18 (min, 2 H), 7.78-7.80 (inm, 1 H), 7.86-7.89 (min, 2 H), 7.98-8.00 (min, 1 H), 8.12-8.14 (min, 1 H), 8.46 (s, 1 H), 8.55 (s, 1 H), 12.62 (bs, 1 H); MS (FAB) m/z 492 (M +1); Anal. Calcd for C 27
H
2 9
N
3 0 6 -1/2H 2 0: C, 64.79; H, 15 6.04; N, 8.21. Found: C, 64.36; H, 5.85; N, 8.21. Example 222 (S)-4-[2-[4-[N'-(2-methylphenyl)ureido]phenylacetylamino]-1-propoxy]benzoic acid H NCOOH -,N..N
CH
3
CH
3 H H 273 A mixture of pentafluorophenyl 4-[N'-(2-methylphenyl)ureido]phenylacetate (560 mg, 20 1.24 mmol), methyl (S)-4-(2-amino-1-propoxy)benzoate (260 mg, 1.24 mmol), Et 3 N (0.260mL, 1.87 mmol) in DMF (8 mL) was stirred at room temp for 3 hr. The mixture was diluted with EtOAc and the solution was washed with 0.5 N HCI, brine, and dried over Na 2
SO
4 . After removal of the solvent, the residue was purified by recrystallization from MeOH-CHCls-n-hexane to give 210 mg (36%) (S)-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetylamino]-1 25 propoxy]benzoic acid as a white crystalline powder. 'H-NMR (DMSO-d 6 ) 5 1.17 (d, 3 H, J=6.8 Hz), 2.24 (s, 3 H), 3.32 (s, 2 H), 3.81 (s, 3 H), 3.92-4.03 (mn, 2 H), 4.08-4.15 (mn, 1 H), 6.92-6.95 (min, 1 H), 7.04-7.06 (min, 2 H), 7.12-7.18 (min, 4 H), 7.35-7.39 (mn, 2 H), 7.83-7.85 (mn, 1 H), 7.89 7.92 (min, 3 H), 8.12-8.14 (min, 1 H), 8.97 (s, 1 H). To a stirred solution of methyl (S)-4-[2-[4-[N'-(2-methylphenyl)ureido]phenylacetyl 407 WO 01/00206 PCT/US00/18079 amino]-1- propoxy]benzoate (200 mg, 0.42 mmol) in THF-MeOH (10 mL, 1:1, v/v) was added 0.5 N NaOH (10 mL), and the mixture was heated under reflux for 2 hr. The mixture was poured into ice-1 N HCI, and the solid was collected. The crude solid was recrystallized from MeOH-CHCl 3 -n hexane to give 68 mg (34%) 273 as a white crystalline powder. mp 262-265 oC; 'H-NMR 5 (DMSO-d) 8 1.17 (d, 3 H, J=6.8 Hz), 2.24 (s, 3 H), 3.32 (s, 2 H), 3.91-4.02 (m, 2 H), 4.09-4.15 (m, 1 H), 6.92-6.96 (m, 1 H), 7.01-7.03 (m, 2 H), 7.12-7.20 (m, 4 H), 7.36-7.40 (m, 2 H), 7.83 7.95 (m, 4 H), 8.12-8.14 (m, 1 H), 8.99 (s, 1 H), 12.63 (bs, 1 H); MS (FAB) m/z 462 (M*+1); Anal. Calcd for C 26
H
2 7 N30 5 -1/4H 2 0: C, 67.01; H, 5.95; N, 9.02. Found: C, 67.13; H, 5.90; N, 9.02. Example 223 10 (S)-3-chloro-4-[2-[4-[N'-(2-methylphenyl)ureido]phenylacetylamino]- 1-propoxy]benzoic acid H , COOH IN N NCH3
CH
3 H H 274 274 To a cooled (00 C) solution of (S)-2-(N-tert-butoxycarbonylamino)-1-propanol (1.05 g, 5.99 mmol), methyl 3-chloro-4-hydroxybenzoate (1.12 g, 6.00 mmol), and Ph 3 P (2.36 g, 9.00 mmol) in THF (20 mL) was added diisopropyl azodicarboxylate (DIAD) (1.77 mL, 8.99 mmol), 15 and the resulting mixture was heated under reflux for 2 days. The solution was evaporated off and the residue was dissolved in CH 2 Cl 2 (20 mL) and TFA (10 mniL). The resulting mixture was stirred at room temp for 1.5 hr. The solution was concentrated in vacuo, and the residue was dissolved in CHC1 3 . The mixture was extracted with H 2 0, and the aqueous layer was made basic by the addition of sat. NaHCO 3 . This basic aqueous layer was extracted with CHC1 3 . The extract was 20 washed with brine, dried over Na 2
SO
4 and evaporated to give 660 mg (2 steps, 32%) methyl (S)-3 chloro-4-(2-amino-1-propoxy)benzoate as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.21 (d, 3 H, J=6.4 Hz), 3.41-3.48 (m, 1 H), 3.77-3.81 (m, 1 H), 3.89 (s, 3 H), 3.98-4.01 (m, 1 H), 6.91-6.94 (m, 1 H), 7.90-7.93 (m, 1 H), 8.05-8.06 (m, 1 H). A mixture of pentafluorophenyl 4-[N'-(2-methylphenyl)ureido]phenylacetate (508 mg, 25 1.13 mmol), methyl (S)-3-chloro-4-(2-amino-1-propoxy)benzoate (275 mg, 1.13 mmol) and Et 3 N (0.240 mL, 1.72 mmol) in DMF (10 mL) was stirred at room temp overnight. The mixture was diluted with EtOAc, and the solution was washed with 0.5 N HCI, sat. NaHCO 3 , brine, , dried over Na 2
SO
4 , and evaporated. The residue was recrystallized from MeOH-CHCl 3 -n-hexane to give 240 mg (42%) methyl (S)-3-chloro-4-[2-[4-[N'-(2-methylphenyl)ureido]phenylacetylamino]-1 30 propoxy]benzoate as a white crystalline powder. 'H-NMR (DMSO-d 6 ) 8 1.21 (d, 3 H, J=6.4 Hz), 2.25 (s, 3 H), 3.33 (s, 2 H), 3.82 (s, 3H), 4.04-4.14 (m, 3H), 6.90-6.94 (m, 1H), 7.11-7.16 (m, 4H), 7.29-7.38 (m, 3H), 7.83-7.94 (m, 3H), 8.13-8.17 (m, 2H), 9.34 (s, 1 H); MS(FAB) m/z 510 (M'). 408 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl (S)-3-chloro-4-[2-[4-[N'-(2-methylphenyl)ureido] phenylacetylamino]-1-propoxy]benzoate (240 mg, 0.47 mmol) in THF-MeOH (10 mL, 1:1, v/v) was added 0.5 N NaOH (10 mL), and the resulting mixture was heated under reflux overnight. The mixture was poured into ice-1 N HCI and the solid was collected. The crude solid was 5 recrystallized from MeOH-CHCl 3 -n-hexane to give 98 mg (42%) 274 as a white crystalline powder. mp 228-231 oC; 'H-NMR (DMSO-d) 8 1.20 (d, 3 H, J=6.3 Hz), 2.24 (s, 3 H), 3.34 (s, 2 H), 4.02-4.18 (m, 3 H), 6.92-6.95 (m, 1 H), 7.12-7.42 (series of m, total 7 H), 7.82-8.18 (series of m, total 5 H), 9.12 (s, 1 H); MS (FAB) m/z 496 (M), 497 (M+I); Anal. Calcd for C 26
H
26 C1N 3 0 5 1/2H 2 0: C, 61.84; H, 5.39; C1,7.02; N,8.32. Found: C,61.76; H,5.25; C1,7.09; N,8.25. 10 Example 224 (S)-3-chloro-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetylamino]-1-propoxy] benzoic acid H N COOH I~ I P '%-NN 0 CH- 3 CI
CH
3 H H OCH 3 275 A mixture ofpentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetate 15 (513 mg, 1.07 mmol), methyl (S)-3-chloro-4-(2-amino-1-propoxy)benzoate (260 mg, 1.07 mmol) and Et 3 N (220 gl, 1.58 mmol) in DMF (10 mL) was stirred at room temp overnight. The mixture was diluted with EtOAc and the solution was washed with sat. NaHCO 3 , dried over Na 2
SO
4 , and evaporated. The residue was recrystallized from MeOH-CHC13-EtOAc-n-hexane to give 400 mg (69%) methyl (S)-3-chloro-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetylamino]-l 20 propoxy]benzoate as pale brown crystalline powder. 'H-NMR (DMSO-d 6 ) 8 1.21 (d, 3 H, J=6.4 Hz), 2.24 (s, 3 H), 3.37 (s, 2 H), 3.82 (s, 3 H), 3.83 (s, 3 H), 4.04-4.12 (m, 3 H), 6.75-6.77 (m, 1 H), 6.91-6.95 (m, 2 H), 7.11-7.17 (m, 2 H), 7.29-7.31 (m, 1 H), 7.78-7.99 (m, 4 H), 8.12-8.13 (m, 1 H), 8.46 (s, 1 H), 8.55 (s, 1 H). To a stirred solution of methyl (S)-3-chloro-4-[2-[3-methoxy-4-[N'-(2-methylphenyl) 25 ureido]phenylacetylamino]-l-propoxy]benzoate (400 mg, 0.74 mmol) in THF-MeOH (20 mL, 1:1, v/v) was added 0.5 N NaOH (20 mL), and the resulting mixture was heated under reflux overnight. The mixture was poured into ice-1 N HCI and the solid was collected. The crude solid was recrystallized from MeOH-CHC 3 -Et 2 O to give 200 mg (51%) 275 as a pale brown crystalline powder. mp 198-201 oC; 'H-NMR (DMSO-d 6 ) 8 1.21 (d, 3 H, J=6.8 Hz), 2.24 (s, 3 H), 3.37 (s, 2 30 H), 3.84 (s, 3 H), 4.00-4.15 (m, 3 H), 6.76-7.28 (series of m, total 6 H), 7.77-8.14 (series of m, total 5 H), 8.46 (s, 1 H), 8.56 (s, 1 H); MS (FAB) m/z 526 (M), 528 (M++2); Anal. Calcd for 409 WO 01/00206 PCT/US00/18079
C
27
H
2 8
CIN
3 0 6 1/4H 2 0: C,61.13;H,5.42;CI,6.68;N, 7.92. Found: C, 60.97; H,5.48; CI,6.86; N, 7.89. Example 225 3-dimethylamino-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] methylamino]ethoxy]benzoic acid
?H
3 l COOH NH3 "N N" H 3 C NCH 3 5 CH 3 H H OCH 3 276 To a stirred and cooled (00 C) solution of 2-(N-Boc-N-methylamino)ethanol (3g, 17mmol), methyl 4-hydroxy-3-nitro benzoate (3.38g, 17mmol), and Ph 3 P (5.4g, 21mmol) in THF (20 mL) was added diisopropyl azodicarboxylate (DIAD) (4mL, 21 mmol), and the resulting mixture was heated under reflux for 15 hr. The solution was evaporated off. The residue was 10 chromatographed on silica-gel with CHC13:MeOH (100:0 to 4:1, v/v) as eluent to give 2.5g (39%) methyl 4-[2-(N-methyl-2-amino)ethoxy]-3-nitro benzoate as a pale yellow oil. 'H-NMR (CDC1 3 , 400MHz) 8 2.82 (s, 3H), 3.50 (t, 2H, J=4.5Hz), 3.95 (s, 3H), 4.54 (t, 2H, J=4.5Hz), 7.26 (d, 1H, J=8.8Hz), 8.25 (d, 1H, J=8.8Hz), 8.56 (s, 1H); MS (FAB) m/z 255 (M +1). A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (992mg, 15 3.1 Immol), methyl 4-(N-methyl-2-aminoethoxy)-3-nitro benzoate (800mg, 3.1 Immol) and 4 DMAP (77mg, 0.63mmol), HOBt (640mg,4.7mmol), and EDC (904mg,4.7mmol) in DMF (20 mL) was stirred at room temp overnight. The mixture was diluted with EtOAc, and the solution was washed with 0.5 N HC1, sat. NaHCO 3 , brine, , dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica-gel with CHCl 3 :EtOAc (95:5 to 0:100, v/v) as eluent to 20 give 587mg (34%) methyl 3-nitro-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] methylamino]ethoxy]benzoate as a pale yellow oil. 'H-NMR (CDC1 3 , 400MHz) 8 2.31 (s, 3H), 3.05 (s, 1H), 3.23 (s, 2H), 3.71 (s, 2H), 3.83 (s, 3H), 3.85 (m, 3H), 3.94 (s, 3H), 4.19 and 4.39 (m, total 2H), 6.80 (m, 2H), 7.05 (m, 1H), 7.22 (m, 3H), 7.62 (d, 1H, J=8.2Hz), 8.02 (d, 1H, J=8.2Hz), 8.21 (dd, 1H,J=2.1Hz, 8.8Hz, 8.55 (d, 1H,J=2.1Hz); MS (FAB)m/z551(M++1). 25 A mixture of methyl 3-nitro-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl acetyl]methylamino]ethoxy]benzoate (587mg, 1. Immol) and 5%-Pd-C (600mg) in THF-MeOH AcOH (1:1:1, v/v, 150mL) was hydrogenated at 45psi for 18hr. Insoluble catalyst was removed with suction, and the filtrate was evaporated in vacuo to afford 555mg (100%) methyl 3-amino-4 [[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]methylaminolethoxy]benzoate as a 30 pale yellow gum. 'H-NMR (CDC 3 , 400MHz) 8 2.29 (s, 3H), 3.08 (m, 1H), 3.19 (s, 2H), 3.65 (s, 410 WO 01/00206 PCT/US00/18079 1H), 3.73-3.80 (m, 3H), 3.84 (s, 3H), 3.87(s, 3H), 4.19 and 4.40 (m, total 2H), 6.70-6.82 (m, 2H), 7.02-7.29 (m, 6H), 7.60 (d, 1H, J=7.8Hz), 7.92-7.99 (m, 3H); MS (FAB) m/z 521 (M+1). To a stirred solution of methyl 3-amino-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]methylamino]ethoxy]benzoate (555mg, 1. Immol), formaldehyde (10mL), and AcOH 5 (0.58mL, 10mmol) in MeCN (10mL) was added NaBH 3 CN (0.67g, 10mmol) at room temp, and the resulting mixture was stirred for 15 hr at the same temp. Sat. NaHCO 3 was added to the mixture and extracted with CHC1 3 . The extract was washed with brine, dried over MgSO 4 , and evaporated in vacuo. The residue was chromatographed on silica-gel with toluene:acetone (7:3 to 1:1, v/v) as eluent to give 123mg (21%) methyl 3-dimethylamino-4-[[2-[3-methoxy-4-[N'-(2 10 methylphenyl) ureido]phenylacetyl] methylamino]ethoxy]benzoate as an oil. 'H-NMR (CDC1 3 , 400MHz) 6 2.30 (s, 3H), 2.70 (s, 3H), 2.75 (s, 3H), 3.05 (s, 1H), 3.18 (s, 2H), 3.61 (s, 3H), 3.70 (s, 1H), 3.80 (m, 3H), 3.86 (s, 3H), 4.07 and 4.22 (m, total 2H), 6.28 (m, 1H), 6.70-6.80 (m, 3H), 7.03 (m, 1H), 7.15-7.25 (m, 4H), 7.46-7.65 (m, 2H), 8.02 (m, 1H); MS (FAB) m/z 548 (M + 1). A stirred mixture of methyl 3-dimethylamino-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl) 15 ureido] phenylacetyl]methylamino]ethoxy]benzoate (123mg, 0.22mmol) in THF (15mL) and IN NaOH (0.885mL, 0.885mmol) was heated under reflux for 15 hr. The pH of the mixture was adjusted to 5.0 by the addition of IN HC1, and extracted with CHCI 3 -MeOH(9:1, v/v). The extract was washed with brine, dried over MgSO 4 , and evaporated in vacuo. The residue was crystallized with Et20-n:hexane to give 118 mg(100%) 276 as a white crystalline material. mp 125-130 oC; IR 20 (KBr) 3346, 2940, 1620, 1597, 1535, 1456, 1417, 1227, 1039, 754cm-'; 'H-NMR (CD 3 OD, 400MHz) 8 2.29 (s, 3H), 2.70 (s, 3H), 2.79 (s, 2H), 3.05 (s, 1H), 3.22 (s, 2H), 3.75 (s, 3H), 3.85 (m, 4H), 4.15 and 4.28 (m, 2H), 6.78-7.05 (m, 4H), 7.18 (m, 2H), 7.55-7.70 (m, 3H), 7.98 (m, 1H); MS (FAB) m/z 535(M + 1); Anal. calcd. for C 2
H
34 N40 6 -2.0H20: C, 61.04; H, 6.71; N, 9.82. Found: C, 61.15; H, 6.43; N, 8.94. 25 Example 226 3-dimethylamino-4-[[2-[3-methoxy-4-[N-(2-fluorophenyl)ureido]phenylacetyl]methylamino] ethoxy]benzoic acid
CH
3 COOH N" N H 3 C 3 OCH 3 H H OCH 3 277 A mixture of 3-methoxy-4-[N'-(2-fluorophenyl)ureido]phenylacetic acid(lg,3.1 Immol), 30 methyl 4-[2-(N-methyl-2-amino)ethoxy]-3-nitrobenzoate (800mg, 3.1 Immol) and 4-DMAP 411 WO 01/00206 PCT/US00/18079 (77mg, 0.63mmol), HOBt(640mg,4.7mmol), and EDC (904mg,4.7nummol) in DMF (20 mL) was stirred at room temp overnight. The mixture was diluted with EtOAc, and the solution was washed with 0.5 N HCI, sat. NaHCO 3 , brine, , dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica-gel with CHC1 3 -EtOAc (95:5 to 0:100, v/v) as eluent to give 5 420mg (19%) methyl 3-nitro-4-[[2-[3-fluoro-4-[N'-(2-fluorophenyl)ureido]phenylacetyl] methylamino] ethoxy]benzoate as a pale yellow oil. 'H-NMR (CDCl 3 , 400MHz) 8 3.04 (s, 1H), 3.24 (s, 2H), 3.72 (s, 1H), 3.85 (s, 3H), 3.90 (min, 3H), 3.93 (s, 3H), 4.16 and 4.39 (2m, 3H), 6.80 (min, 2H), 6.99 (min, 1H), 7.05 (min, 2H), 7.22 (d, 1H, J=8.8Hz), 7.51 (s, 2H), 8.00 (min, 1H), 8.08 (inm, 1H), 8.21 (d, 1H, J=8.6Hz), 8.51 (s, 1H); MS (FAB) m/z 555 (M' + 1). 10 A mixture of methyl 3-nitro-4-[[2-[3-methoxy-4-[AN'-(2-fluorophenyl)ureido]phenylacetyl] methylamino]ethoxy]benzoate (420mg, 0.76mmol) and 5%-Pd-C (1g) in THF-MeOH-AcOH (1:1:1, v/v, 150mL) was hydrogenated at 45psi for 18 hr. Insoluble catalyst was removed with suction, and the filtrate was evaporated in vacuo to afford 397mg (100%) methyl 3-amino-4-[[2 [3-methoxy-4-[NAP-(2-fluorophenyl)ureido]phenylacetyl]methylamino]ethoxy]benzoate as a pale 15 yellow gum. 'H-NMRv (CDCI 3 , 400MHz) 8 3.05 and 3.13 (s, total 3H), 3.66 (s, 3H), 3.70 (s, 2H), 3.65-3.90 (min, 4H), 3.86 (s, 3H), 4.10 and 4.23 (mn, 2H), 6.70-6.83 (mn, 3H), 6.98-7.15 (mn, 6H), 7.25-7.43 (min, 2H), 7.99 (min, 1H), 8.13 (min, 1H); MS (FAB) m/z 525 (M'+1). To a stirred solution of methyl 3-amino-4-[[2-[3-methoxy-4-[N'-(2-fluorophenyl)ureido] phenylacetyl]methylamino]ethoxy]benzoate (397mg, 0.76mmol), formaldehyde (O10mL), and 20 AcOH (0.43mL, 7.6mmol) in MeCN (10mniL) was added NaBH 3 CN (0.48g, 7.6mmol) at room temp, and the resulting mixture was stirred for 15 hr at the same temp. Sat. NaHCO 3 was added to the mixture and extracted with CHCl 3 . The extract was washed with brine, dried over MgSO 4 , and evaporated in vacuo. The residue was chromatographed on silica-gel with toluene:acetone (7:3 to 1:1, v/v) as eluent to give 123mg (21%) methyl 3-dimethylamino-4-[[2-[3-methoxy-4-[N-(2 25 methylphenyl)ureido]phenylacetyl] methylamino]ethoxy]benzoate as an oil. 'H-NMR (CDCl1 3 , 400MHz) 8 2.74 (s, 3H), 2.77 (s, 3H), 3.08 (s, 1H), 3.22 (s, 2H), 3.52 (s, 3H), 3.61 (s, 1H), 3.83 (min, 3H), 3.88 (s, 3H), 4.12 and 4.23 (min, total 2H), 6.68 (s, 1H), 6.78 (mn, 2H), 6.98 (mn, 2H), 7.10 (min, 1H), 7.55-7.68 (min, 4H), 7.99 (mn, 1H), 8.16 (t, 1H, J=8.3Hz); MS (FAB) m/z 553 (M +1). A stirred mixture of methyl 3-dimethylamino-4-[[2-[3-methoxy-4-[N-(2-fluorophenyl) 30 ureido] phenylacetyl]methylamino]ethoxy]benzoate (61mg, 0.11 mmol) in THF (15mL) and IN NaOH (0.22mL, 0.22mmol) was heated under reflux for 15 hr. The pH of the mixture was adjusted to 5.0 by the addition of IN HCI, and extracted with CHCl 3 :MeOH(9: 1, v/v). The extract 412 WO 01/00206 PCT/US00/18079 was washed with brine, MeOH:acetone (93:7, v/v) as eluent to give 37 mg (63%) 277 as a white crystalline material. mp 120-125 oC; 'H-NMR (CD 3 OD, 400MHz) 8 2.60 (s, 4H), 2.78 (s, 2H), 3.06 (s, 1H), 3.22 (s, 2H), 3.75 (s, 3H), 3.85-3.92 (m, 4H), 4.17 and 4.29 (m, total 2H), 6.80-7.12 (m, 6H), 7.61-7.70 (m, 2H), 8.00 (m, 1H), 8.08 (m, 1H); MS (FAB) 539 (M'+1); Anal. calcd. for 5 C 28 H3 1
FN
4 0 6 -2.75H 2 0: C, 57.18; H, 6.26; N, 9.53. Found, C, 57.20; H, 5.62; N, 9.06. Example 227 3-dimethylamino-4-[[2-[4-[N/-(2-methylphenyl)ureido]phenylacetyl]methylamino]ethoxy] benzoic acid CH11 3 COOH N~N i N"Nf O H3 C N H 3
CH
3 H H 278 10 A mixture of pentafluorophenyl [4-[N-(2-methylphenyl)ureido]phenyl]acetate (1.42g, 3.15mmol), methyl 4-[2-(N-methyl-2-amino)ethoxy]-3-nitro benzoate (800mg, 3.15mmol) and triethylamine (0.66mL,4.73mmol) in DMF (8 mL) was stirred at 500 C for 15 hr. The mixture was poured into ice-lN HC1 and extracted with CHC13. The extract was brine, dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica-gel with CHCl 3 :EtOAc (95:5 to 15 0:100, v/v) as eluent to give 1.04g (63%) methyl 3-nitro-4-[[2-[4-[N'-(2-methylphenyl)ureido] phenylacetyl]methylamino]ethoxy]benzoate as a pale yellow oil. 'H-NMR (CDCl 3 , 400MHz) 8 2.30 (s, 3H), 2.90, 3.02 and 3.05 (s, total 1H), 3.22 (s, 2H), 3.71 (s, 1H), 3.85 (3H), 3.93 (s,3H), 4.19 and 4.39 (m, 2H), 7.02 (m, 1H), 7.19 (m, 4H), 7.35 (d, 1H, J=8.3Hz), 7.40 (d, 1H, J=8.0Hz), 7.55 (s, 2H), 7.70 (m, 1H), 8.22 (d, 1H, J=6.7Hz), 8.51 (s, 1H); MS (FAB) m/z 521(M+1). 20 A mixture of methyl 3-nitro-4-[[2-[4-[N'-(2-methylphenyl)ureido]phenylacetyl] methylamino]ethoxy]benzoate (1.04g, 2mmol) and 5%-Pd-C (1.2g) in THF-MeOH-AcOH (1:1:1, v/v, 150mL) was hydrogenated at 45 psi for 18 hr. Insoluble catalyst was removed with suction, and the filtrate was evaporated in vacuo to afford methyl 3-amino-4-[[2-[4-[N'-(2-methylphenyl) ureido]phenylacetyl]methylamino]ethoxy]benzoate as a pale yellow gum. 25 To a stirred solution of 3-amino-4-[[2-[4-[N'-(2-methylphenyl)ureido]phenylacetyl] methylamino]ethoxy]benzoate, formaldehyde (5mL), and AcOH (1.14mL, 20mmol) in MeCN (5mL) was added NaBH 3 CN (1.26g, 20mmol) at room temp, and the resulting mixture was stirred for 15 hr at the same temp. Sat. NaHCO 3 was added to the mixture and extracted with CHCl 3 . The extract was washed with brine, dried over MgSO 4 , and evaporated in vacuo. The residue was 30 chromatographed on silica-gel with toluene:acetone (7:3 to 1:1, v/v) as eluent to give 85mg (2 413 WO 01/00206 PCT/US00/18079 steps, 8%) methyl 3-dimethylamino-4-[[2-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]methyl amino]ethoxy]benzoate as an oil. 'H-NMR (CDCl 3 , 400MHz) 8 2.12 (s, 3H), 2.73 (s, 3H), 2.75 (s, 3H), 3.05 (s, 1H), 3.20 (s, 2H), 3.60 (s, 1H), 3.80 (m, 3H11), 3.88 (s, 3H), 4.17 (m, 2H), 6.95-7.28 (m, 8H), 7.55-7.75 (m, 3H); MS (FAB) m/z 518 (M'+1). 5 A stirred mixture of methyl 3-dimethylamino-4-[[2-[4-[N'-(2-methylphenyl)ureido] phenylacetyl] methylamino]ethoxy]benzoate(ap315201)(85mg, 0.16mmol) in THF (15mL) and IN NaOH (0.32mL, 0.32mmol) was heated under reflux for 15 hr. The pH of the mixture was adjusted to 5.0 by the addition of IN HC1, and extracted with CHC1 3 :MeOH(9:1, v/v). The extract was washed with brine, dried over MgSO 4 , and evaporated in vacuo. The residue was crystallized 10 from Et 2 O to give 53 mg(65%) 278 as a white crystalline material. mp 110-115 oC; 'H-NMR
(CD
3 OD, 400IMHz) 8 2.29 (s, 3H), 2.75 (s, 3H), 2.76 (s, 3H), 3.02 (s, 1H), 3.20 (s, 2H1), 3.72 (s, 1H1), 3.85 (m, 3H), 4.18 and 4.28 (m, total 2H), 6.95-7.03 (m, 2H), 7.18 (m, 4H), 7.34 (d, 1H, J=8.3Hz), 7.38 (d, 1H, J=8.8Hz), 7.62 (d, 1H, J=8.3Hz), 7.66 (s, 1H1), 7.80 (m, 1H); MS (FAB) m/z 505 (M'+1). 15 Example 228 3-isopropylamiino-4-[[2-[3-methoxy-4-[N-(2-fluorophenyl)ureido]phenylacetyl]methylamino] ethoxy]benzoic acid
CNH
3 ,COOH .0g,,. H 3 C NH H H OCH 3
CH
3 279 To a stirred cold (00 C) solution of methyl 3-amino-4-[[2-[3-methoxy-4-[N'-(2-fluoro 20 phenyl)ureido]phenylacetyl]methylamino]ethoxy]benzoate (100mg, 0.19mmol) in acetone / AcOH/DMF (13 mL, 6:6:1, v/v/v) was added NaBH 3 CN (300 mg), and the resulting mixture was stirred for 60 hr at room temp. The mixture was pored into sat. NaHCO 3 and the solid was collected with suction. The precipitate was dissolved in CHCl 3 (20mL), and the solution was washed with brine, dried over MgSO 4 , and evaporated under a reduced pressure. The residue was 25 chromatographed on silica-gel plate with toluene:acetone (2:1, v/v) as eluent to give 108 mg (100%) methyl 3-isopropylamino-4-[[2-[3-methoxy-4-[N-(2-fluorophenyl)ureido]phenylacetyl] methylamino]ethoxy]benzoate as a colorless oil. 'H-NMR (CDCl 3 , 400MHz) 8 1.20 (m, 1H), 2.88 (s, 6H), 3.02 and 3.12 (s, total 3H), 3.53 (s, 2H), 3.60-3.80 (m, 7H), 3.85 (s, 3H), 4.10 and 4.20 (m, 2H), 6.65-675 (m, 2H), 6.90-7.08 (m, 2H), 7.22-7.35 (m, 2H), 8.02 (s, 2H), 8.10 (m, 2H1), 8.21 30 (br, 1H1), 8.33 (br, 1H); MS (FAB) m/z 566 (M+1). 414 WO 01/00206 PCT/US00/18079 A stirred mixture of methyl 3-isopropylamino-4-[[2-[3-methoxy-4-[N'-(2-fluorophenyl) ureido]phenylacetyl]methylamino]ethoxy]benzoate (116mg, 0.2mmol), 0.25 N NaOH (6 mL), and THF (6 mL) was heated under reflux for 8 hr. The mixture was poured into water (200 mL), acidified by 1N HCI and the solid was collected with suction. The solid was recrystallized from 5 CHCl 3 -n-hexane-diisopropylether to give 56mg (50%) 279 as a colorless crystalline powder. mp 195-200 oC; 'H-NMR (CD 3 OD, 400MHz) 8 1.15-1.20 (mn, 6H), 3.01 (s, 1H11), 3.12 (s, 2H), 3.48 3.60 (min, 1H), 3.68 (s, 3H), 3.75 (s, 2H), 3.82 (s, 1H), 3.86 (min, 3H11), 4.15-4.23 (mn, 2H), 6.80 (inm, 3H), 4.15-4.23 (min, 2H), 6.80 (min, 3H), 6.98 (min, 1H), 7.10 (min, 2H), 7.20 and 7.25 (s, total 1H11), 7.32 (min, 1H), 7.98 (min, 1H), 8.05 (min, 1H); MS (FAB) m/z 552 (M+H); Anal. calcd. for 10 C 29
H
33 FN40 6 "1.0H 2 0: C, 61.04; H, 6.18; N, 9.82. Found, C, 61.36; H, 6.25; N, 9.45. Example 229 4-[[1-[4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenylacetyl]-2-methylamino]-2-methyl-2 propoxy]benzoic acid
H
3 C H 3 C CH3 COOH N NC H H OCH3 280 280 15 To a stirred solution of 2-amino-2-methyl-1-propanol (8.4g, 93.89mmol) and triethylamine (11.4g, 0.113mol) in DMF-water (1:1, v/v, 100mL) was added di-tert-butyl dicarbonate (25g, 0.1 15mol) at 5 to 100 C. The resulting solution was stirred for 2 hr at room temp. The mixture was diluted with water(100miL) and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on 20 silica-gel with CH 2 C1 2 as eluent to give 12g (68%) 2-tert-butoxycarbonylamino-2-methyl-1 propanol as a syrup. 'H-NMR (CDCl 3 ) 8 1.25 (s, 6H), 1.43 (s, 9H), 3.59 (d, J=8.3Hz, 2H11), 4.68 (br, 11). To a stirred suspension of 2-tert-butoxycarbonylamnino-2-methyl-1-propanol (5.7g, 30.1 1mmol) and powdered NaOH(6.7g, 0.15 1mol) in Et 2 0(200mL) was added p-toluenesulfonyl 25 chloride (6.9g, 36.14mmol) at room temp. The stirred resulting mixture was heated under reflux for 8 hr. After cooling, ice-water(100mL) was added to the solution. Separated Et20 layer was washed with brine, dried over Na 2
SO
4 , and evaporated. To the residue was added n-hexane and triturated. The solid was collected to afford 8.5g (82.2%) 2-tert-butoxycarbonylamino-2-methyl-1 propyl p-toluenesulfonate as a crystalline material. 'H-NMR (CDC 3 ) 8 1.26 (s, 6H), 1.38 (s, 9H), 30 2.37 (s, 3H), 4.05 (s, 2H), 4.49 (br, 1H), 7.34 (d, J=7.8Hz, 2H), 7.78 (d, J=7.8Hz, 2H). A stirred mixture of 2-tert-butoxycarbonylamino-2-methyl- 1-propyl p-toluenesulfonate 415 WO 01/00206 PCT/US00/18079 (8.2g, 23.88mmol) and powdered NaOH (6.7g, 0.15 mol) in Et20 (200mL) was heated under reflux for 10 hr. After cooling, the mixture was filtered. And the filtrate was washed with water, brine, dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica-gel with n hexane:EtOAc (6:1, v/v) as eluent to give 2.7g (66.2%) 1-tert-butoxycarbonyl-2-methylpropylene 5 imine as an oil. 'H-NMR (CDCl 3 ) 8 1.29 (s, 6H), 1.47 (s, 9H), 2.05(s, 2H). To a stirred solution of 1-tert-butoxycarbony-2-methylpropyleneimine (1.03g, 6.0 1mmol) and methyl 4-hydroxybenzoate (800mg, 6.26mmol) in CH 2 C1 2 (10mL) was added boron trifluoride diethyl ether (0.127mL, Immol) at ambient temp. The resulting solution was stirred for a further 3 hr at the same temp. The mixture was washed with water, brine, dried over Na 2
SO
4 , and 10 evaporated. The residue was chromatographed on silica-gel with n-hexane:EtOAc (6:1, v/v) as eluent to give 550mg (33%) methyl 4-(1-tert-butoxycarbonylamino-2-methyl-2-propoxy)benzoate as a gum. 'H-NMR (CDCl 3 ) 8 1.33 (s, 6H), 1.47 (s, 9H), 3.36 (d, J=6.3Hz, 2H), 3.90 (s, 3H), 5.05 (br, 1H), 6.99 (d, J=8.8Hz, 2H), 7.97 (d, J=8.8Hz, 2H). A mixture of methyl 4-(1-tert-butoxycarbonylamino-2-methyl-2-propoxy)benzoate 15 (460mg, 1.42mmol) and anisole (0.155mL, 1.42mmol) in CH 2 C12 (15mL) and TFA (3mL) was stirred for 3 hr at room temp. The mixture was evaporated off. The residue was dissolved in
CH
2 Cl 2 (30mL) and made basic by the addition of 0.5N NaOH. The CH 2 Cl 2 layer was separated, dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica-gel with CH 2 C1 2 as eluent to give 370mg (100%) methyl 4-(1-amino-2-methyl-2-propoxy)benzoate as a gum. 'H 20 NMR (CDCl 3 ) 8 1.34 (s, 6H), 2.87(s,2H),3.90 (s,3H),7.10 (d, J=8.8Hz,2H), 7.97 (d, J=8.8Hz, 2H). To a stirred solution of methyl 4-(1-amino-2-methyl-2-propoxy)benzoate (370mg, 1.66mmol) and triethylamine (0.35mL, 2.49mmol) in CH 2 C1 2 (15mL) was added trifluoroacetic anhydride (0.316mL, 2.24mmol) at 0o C. After stirred for 1 hr at the same temp, water was added to the solution. CH 2 C1 2 layer was separated, washed with water, dried over Na 2
SO
4 , and 25 evaporated. The residue was chromatographed on silica-gel(20mL) with CH 2 C1 2 as eluent to give 530mg (100%) methyl 4-(1-trifluoroacetamido-2-methyl-2-propoxy)benzoate as a gum. This compound was used to the subsequent reaction without further purification. To a stirred mixture of methyl 4-(1-trifluoroacetamido-2-methyl-2-propoxy)benzoate (530mg, 1.66mmol) and K 2 CO3(345mg, 2.49mmol) in DMF (10mL) was added Mel (0.14mL, 30 2.37mmol) at room temp. The resulting mixture was stirred for 18 hr at room temp. The mixture was poured into water, and extracted with EtOAc. The extract was washed with washed with 416 WO 01/00206 PCT/US00/18079 brine, dried over Na 2
SO
4 , and evaporated. The residual gum was used to the subsequent reaction without further purification. The above crude residue was dissolved in MeOH(O10mL). To the stirred solution was added water (5mL) and Na 2
CO
3 (352mg, 3.32mmol), and the resulting mixture was stirred for 5 hr 5 at room temp. The mixture was poured into water and extracted with CHCl 3 . The extract was washed with water, dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica-gel with CHCI 3 as eluent to give 390mg (100%) methyl 4-(1-methylamino-2-methyl-2 propoxy)benzoate as a gum. 'H-NMR (CDCI 3 ) 8 1.37 (s, 6H), 2.51 (s, 3H), 3.89 (s, 3H), 6.92 (d, J=8.8Hz, 2H), 7.97 (d, J=8.8Hz, 2H). 10 To a stirred mixture of methyl 4-(1-methylamino-2-methyl-2-propoxy)benzoate (200mg, 0.84mmol), 4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenylacetic acid (268mg, 0.84mmol), 4 DMAP(125mg, 1.0mmol) in DMF(5mL) was added EDC(220mg, 1.14mmol) at ambient temp. The resulting mixture was stirred for a further 10 hr at ambient temp. The mixture was poured into water, and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and 15 evaporated. The residual gum was triturated with CH 2
CI
2 and Et 2 0 to give 200mg (44.1%) methyl 4-[[ 1-[4-[N-(2-fluorophenyl)ureido]-3-methoxyphenylacetyl]methylamino]-2-methyl-2 propoxy]benzoate as a crystalline material. 'H-NMR (CDCl 3 ) 5 1.36 and 1.31 (each s, 6H11), 3.24 3.88 (series of s, 13 H), 6.65-8.20 (seris of m, 14H). A mixture of methyl 4-[[1-[4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenylacetyl] 20 methylamino]-2-methyl-2-propoxy]benzoate(180mg, 0.335mmol) in THF(3mL) and 0.25N NaOH(4mL) was stirred for 5 hr at room temp. The mixture was poured into ice-IN HCl(5mL). The solid was collected, washed with water, and air-dried. The crude solid was recrystallized from EtOH-CHCl 3 -n-hexane to give 70 mg (40%)280 as fine needles. mp 200-207 oC; 'H-NMR (DMSO-d) 8 1.26 and 1.33 (each s, 6H), 3.70-3.81 (series of s, 7H), 3.83 (s, 3H), 6.75-8.20 (series 25 of m, 10H11), 8.71 (s, 1H), 9.17 (br s, 1H), 12.72 (s, 1H11). Example 230 (S)-3-chloro-4-[2-[3-methoxy-4-[N'-(2-fluorophenyl)ureido]phenylacetylamino]-1 -propoxy] benzoic acid H I COOH 0 -0Y" _P1"O N N N
H
3 CI H H OCH 3 281 30 A mixture of 3-methoxy-4-[N'-(2-fluorophenyl)ureido]phenylacetic acid (327 mg, 1.03 417 WO 01/00206 PCT/US00/18079 mmol), methyl (S)-3-chloro-4-(2-amino-1-propoxy)benzoate (250 mg, 1.03 mmol), EDC(hydrochloride) (295 mg, 1.54 mmol), HOBt (208 mg, 1.54 mmol), and DMAP (25 mg, 0.20 mmol) in DMF (8 mL) was stirred at room temp overnight. The mixture was diluted with EtOAc, washed with 0.5 N HC1, brine, dried over Na 2
SO
4 and evaporated. The residue was recrystallized 5 from CHC13-EtOAc to give 308 mg (55%) as a white crystalline powder. 'H-NMR (CDCl 3 ) 8 1.29 (d, 3 H, J=6.8 Hz), 3.51 (s, 2 H), 3.84 (s, 3 H), 3.88 (s, 3 H), 4.01-4.08 (m, 2 H), 4.38-4.39 (m, 1 H), 6.25 (d, 1 H, .J=8.3 Hz), 6.78-6.83 (m, 2 H), 6.90-6.95 (m, 2 H), 7.02-7.11 (m, 2 H), 7.89 (dd, 1 H, J=2.0, 8.8 Hz), 8.02 (d, 1 H, J.=2.4 Hz), 8.20 (d, 1 H, J=8.3 Hz), 8.27-8.31 (m, 1 H), 8.53 (s, 1 H), 8.84 (s, 1 H). 10 To a stirred solution of methyl (S)-3-chloro-4-[2-[3-methoxy-4-[N'-(2-fluorophenyl) ureido]phenylacetylamino]-1-propoxy]benzoate (308 mg, 0.57 mmol) in THF-MeOH (10 mL, 1:1, v/v) was added 0.5 N NaOH (10 mL), and the reaction mixture was heated under reflux for 1 hr. The mixture was poured into ice-1 N HC1 and the solid was collected. The crude solid was purified by recrystallization from MeOH-CHCl 3 -n-hexane to give 196 mg (65%) 281 as a white 15 crystalline powder. mp 188-191 oC; 'H-NMR (DMSO-d 6 ) 8 1.21 (d, 3 H, J=6.4 Hz), 3.38 (s, 2 H), 3.83 (s, 3 H), 4.02-4.18 (m, 3 H), 6.78-7.29 (series of m, total 6 H), 7.85-8.00 (m, 3 H), 8.12-8.19 (m, 2 H), 8.70 (s, 1 H), 9.17 (s, 1 H), 12.98 (bs, 1 H); MS (FAB) m/z 530 (M), 531 (M+1), 532 (M+2); Anal. Calcd for C 26
H
25
CIFN
3 0 6 -1/4H 2 0: C, 58.43; H, 4.81; Cl, 6.63; F, 3.55; N, 7.86. Found: C, 58.45; H, 4.83; Cl, 6.68; F, 3.38; N, 7.79. 20 Example 231 4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]lmethylamino]ethylamino]benzoic acid
CH
3 ,yCOOH LN~ ( H
CH
3 H H OCH 3 282 A stirred solution of N-benzyloxycarbonyl-N-methylaminoacetaldehyde (1.77g, 1. 1mmol) 25 in toluene (3mL) was added methyl-4-aminobenzoate (1.29g, 8.5mmol), and the mixture was stirred for 1 hr at room temp. The reaction mixture was evaporated under a reduced pressure and the residue was dissolved into MeCN(15 mL). To the solution was added AcOH (2.44mL, 43mmol) and NaBH 3 CN (2.67g, 43mmol), and the resulting mixture was stirred for 15 hr at room temp. The reaction was quenched by the addition of sat. NaHCO 3 . The mixture was extracted 30 with EtOAc, washed with brine, dried over MgSO 4 , and evaporated. The residue was chromatographed on silica-gel with n-hexane: EtOAc (7:3, v/v) as eluent to give 1.26g (43%) 418 WO 01/00206 PCT/US00/18079 methyl 4-[2-(N-benzyloxy carbonyl-N-methylamino)-ethylamino] benzoate as a colorless oil. 'H NMR (CDCl 3 , 400MHz) 5 2.96 (s, 3H), 3.32 (br m, 2H), 3.50-3.60 (min, 2H), 3.82 (s, 3H), 5.15 (each d, 2H, J=14.6Hz), 6.35 and 6.58 (min, total 2H), 7.36 (s, 5H), 7.76 and 7.84 (min, total 2H1); MS (FAB) m/z 342 (M + 1). 5 To a stirred solution of methyl 4-[2-(N-benzyloxycarbonyl-N-methylamino)ethylamino] benzoate (1.26g, 3.68mmol) in MeOH(20 mL) was added 5 wt. Pd -C (700mg), and the mixture was hydrogenated (3 atm) for 4 hr at room temp. The mixture was filtered, and the filtrate was evaporated under a reduced pressure to give 600mg (78%) methyl 4-[2-(N-methylamino) ethylamino]benzoate as a colorless oil. 'H-NMR (CDCl 3 , 400MHz) 8 2.46 (s, 3H), 2.88 (t, 2H, 10 J=5.5Hz), 3.27 (br s, 2H11), 3.85 (s, 3H), 6.57 (d, 2H, J=8.8Hz), 7.85 (d, 2H, J=8.8Hz); MS (FAB) m/z 209 (M + 1). To a stirred solution of methyl 4-[2-(N-methylamino)ethylamino]benzoate (590mg, 2.83mmol)3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (891mg,2.83mmol) in DMF(14mL) was added EDC (815mg,4.25mmol), HOBt (574mg,4.25mmol),and 4-DMAP 15 (519mg, 4.25mmol), and the resulting mixture was stirred overnight at room temp. The mixture was poured into IN HCI and the solid was collected with suction. The crude solid was purified by chromatography on silica-gel (middle pressure) with CHCl 3 -EtOAc (10:0 to 7:3, v/v) as eluent to give 1.25g (87%) methyl 4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] methylamino]ethylamino]benzoate as a light yellow oil. 'H-NMR (CDC13, 400MHz) 8 3.18 (s, 20 2H), 3.05 and 3.32 (s, total 2H), 3.72-3.85 (min, 9H), 4.12 and 4.23 (min, total 2H), 6.78 (min, 3H), 7.05 (t, 1H, J=7.5Hz), 7.20 (min, 2H), 7.43-7.50 (min, 3H), 7.62 (d, 1H, J=8.8Hz), 8.05 (d, 1H, J=8.8Hz); MS (FAB) m/z 505 (M+1). A stirred mixture of methyl 4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] methylamino]ethylamino]benzoate (300mg, 0.6mmol) in 0.25 N NaOH (6 mL) and THF (6 mL) 25 was heated under reflux for 8 hr. The mixture was poured into water, and acidified with IN HCI. The solid was collected with suction. The crude solid was recrystallized from n-hexane diisopropylether to give 202mg (69%) 282 as a pale yellow crystalline powder. mp 115-120 oC; IR(KBr) 3346, 2935, 1603, 1531, 1454, 1417, 1257, 1174, 1036, 754cm-'; 'H-NMR (CD 3 OD, 400MHz) 5 2.28 (s, 3H), 2.98 (s, 1H), 3.10 (s, 2H), 3.35-3.42 (min, 2H), 3.53-3.65 (min, 3H), 3.70 (s, 30 1H), 3.80 and 3.82 (s, 3H), 6.60-6.85 (min, 4H), 7.02 (min, 1H), 7.18 (min, 2H), 7.58 (mn, 1H), 7.82 (inm, 2H), 7.96 (min, 1H); MS (FAB) m/z 491 (M+1). 419 WO 01/00206 PCT/US00/18079 Example 232 (S)-3-chloro-4-[2-[N-methyl-N-[3-methoxy-4-[N '-(2-fluorophenyl)ureido]phenylacetyl] amino]-1-propoxy]benzoic acid
CH
3 , COOH NI~N C6H 3 CI H H OCH 3 283 5 To a cooled (00 C) solution of methyl (S)-3-chloro-4-(2-amino-1-propoxy)benzoate (1.74 g, 7.14 mmol) in CH 2 C1 2 (20 mL) was added Et3N (1.19 mL, 8.54 mmol) and trifluoroacetic anhydride (TFAA) (1.11 mL, 7.86 mmol), and the reaction mixture was stirred at room temp overnight. The mixture was diluted with CHCl 3 , washed with 0.5 N HCI, brine, dried over Na 2
SO
4 and evaporated. The residue was recrystallized from CHCl 3 -n-hexane to give 1.59 g (66%) (S)-3 10 chloro-4-(2-trifluoroacetamido-1-propoxy)benzoate as a white crystalline material. mp 122-125 oC; 'H-NMR (CDCl 3 ) 8 1.48 (d, 3 H, J=6.8 Hz), 3.91 (s, 3 H), 4.10-4.19 (m, 2 H), 4.47-4.52 (m, 1 H), 6.68 (bs, 1 H), 6.92-6.94 (m, 1 H), 7.93-7.95 (m, 1 H), 8.07-8.08 (m, 1 H); MS (FAB) m/z 340 (M+1); Anal. Calcd for C 3
H,
3 C1F 3
NO
4 : C, 45.96; H, 3.86; Cl, 10.44; F, 16.78; N, 4.12. Found: C, 45.88; H, 3.97; Cl, 10.24, F, 16.72; N, 4.18. 15 To a stirred solution of methyl (S)-3-chloro-4-(2-trifluoroacetamido-1l-propoxy)benzoate (800 mg, 2.36 mmol) in DMF (5 mL) was added K 2
CO
3 (651 mmol, 4.71 mmol) and Mel (0.22 mL, 3.53 mmol), and the reaction mixture was stirred at 600 C overnight. The mixture was diluted with EtOAc, washed with 0.5 N HC1, brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with 5% EtOAc in CHCl 3 as eluent 20 to give 850 mg (100%) methyl (S)-3-chloro-4-[2-(N-methyl-N-trifluoroacetamido)-l1-propoxy] benzoate as a colorless oil. 'H-NMR (CDC1 3 ) 8 1.43-1.46 (m, 3 H), 3.03 and 3.21 (s, 3 H), 3.90 (s, 3 H), 4.04-4.22 (m, 2 H), 4.81-4.87 (m, 1 H), 6.91 (d, 1 H, J=8.3 Hz), 7.93 (dd, 1 H, J=2.0, 8.3 H), 8.06 (d, 1 H, J=2.0 Hz). To a stirred solution of methyl (S)-3-chloro-4-[2-(N-methyl-N-trifluoroacetamido)-1 25 propoxy] benzoate (880 mg, 2.49 mmol) in MeOH-H 2 0 (10 mL, 1:1, v/v) was added K 2
CO
3 (516 mg, 3.73 mmol), and the resulting mixture was stirred at room temp overnight. The mixture was diluted with EtOAc, washed with H 2 0, brine, dried over Na 2
SO
4 and evaporated to give 460 mg (72%) (S)-3-chloro-4-(2-methylamino-1-propoxy)benzoate as a colorless oil. 'H-NMR (CDCI 3 ) 8 1.20(d, 3 H, J=6.4 Hz), 2.51 (s, 3 H), 3.07-3.12 (m, 1 H), 3.89 (s, 3 H), 3.92-4.04 (m, 2 H), 6.93 30 6.95 (m, 1 H), 7.90-7.93 (m, 1 H), 8.05-8.06 (m, 1 H). 420 WO 01/00206 PCT/US00/18079 A mixture of 3-methoxy-4-[N'-(2-fluorophenyl)ureido]phenylacetic acid (296 mg, 0.93 mmol), methyl (S)-3-chloro-4-(2-methylamino-1-propoxy)benzoate (240 mg, 0.93 mmol), EDC(hydrochloride) (268 mg, 1.40 mmol), HOBt (189 mg, 1.40 mmol), and DMAP (23 mg, 0.19 mmol) in DMF (8 mL) was stirred at room temp for 1.5 hr. The mixture was diluted with EtOAc, 5 washed with 0.5 N HCI, brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with 5% MeOH in CHC13 as eluent to give 520 mg (100%) methyl (S)-3-chloro-4-[2-[N-methyl-N-[3-methoxy-4-[N'-(2-fluorophenyl)ureido]phenylacetyl] amino]-1-propoxy]benzoate as a red-brown amorphous solid. To a stirred solution of this product (520 mg, 0.93 mmol) in THF (10 mL) was added 0.5 10 N NaOH (10 mL), and the resulting mixture was heated under reflux for 3 hr. The mixture was poured into ice-1 N HCI and the solid was collected. The crude solid was recrystallized from CHCl 3 -Et20 to give 170 mg (2 steps, 37%) 283 as a white crystalline powder. mp 142-147 oC; 'H NMR (DMSO-d 6 ) 8 1.13-1.20 (m, 3 H), 2.74 and 2.94 (s, 3 H), 3.65 (s, 2 H), 3.82 and 3.84 (s, 3 H), 4.13-4.22 (m, 2 H), 4.53 and 4.91-4.92 (m, 1 H), 6.71-7.29 (series of m, total 6 H), 7.86-8.02 15 (m, 3 H), 8.15-8.19 (m, 1 H), 8.71 (s, 1 H), 9.17 (s, 1 H), 13.00 (bs, 1 H); MS (FAB) m/z 544 (M), 545 (M+1); Anal. Calcd for C 27
H
27
CIFN
3 0-1/4H 2 0: C, 59.13; H, 5.05; Cl, 6.46; F, 3.46; N, 7.66. Found: C, 59.19; H, 4.99; Cl, 6.64; F, 3.23; N, 7.55. Example 233 (S)-3-chloro-4-[2-[N-methyl-N-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetyl] 20 amino-1-propoxy]benzoic acid
CH
3 N OOH I , P-NN% 0 CH 3 CI
CH
3 H H OCH 3 284 A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (261 mg, 0.83 mmol), methyl (S)-3-chloro-4-(2-methylamino-1-propoxy)benzoate (214 mg, 0.83 mmol), EDC(hydrochloride) (239 mg, 1.25 mmol), HOBt (168 mg, 1.24 mmol), and DMAP (20 mg, 0.16 25 mmol) in DMF (8 mL) was stirred at room temp for 2 hr. The mixture was diluted with EtOAc, washed with 0.5 N HC1, brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (50:1, v/v) as eluent to give 470 mg (100%) methyl (S)-3-chloro-4-[2-[N-methyl-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]amino]-1-propoxy]benzoate as a pale yellow amorphous solid. 30 To a stirred solution of the above product (470 mg, 0.95 mmol) in THF (10 mL) was 421 WO 01/00206 PCT/US00/18079 added 0.5 N NaOH (10 mL) and the reaction mixture was heated under reflux for 3 hr. The mixture was poured into ice-1 N HCI and the solid was collected. The crude solid was recrystallized from CHCl 3 -Et 2 O to give 168 mg (2 steps, 33%) 284 as a pale yellow crystalline powder. mp 125-130 oC; 'H-NMR (DMSO-d 6 ) 8 1.13-1.19 (min, 3 H), 2.24 (s, 3 H), 2.74 and 2.94 5 (s, 3 H), 3.65 (s, 2 H), 3.83 and 3.85 (s, 3 H), 4.14-4.22 (min, 2 H), 4.91-4.93 (min, 1 H), 6.70-6.74 (min, 1 H), 6.84 (min, 1 H), 6.92-6.95 (min, 1 H), 7.11-7.17 (min, 2 H), 7.25-7.29 (min, 1 H), 7.78-7.80 (inm, 1 H), 7.85-7.93 (mn, 2 H), 7.99-8.02 (m ,1 H), 8.46 (s, 1 H), 8.56 (s, 1 H), 12.99 (bs, 1 H); MS (FAB) m/z 540 (M), 541 (M++1); Anal. Calcd for C 28
H
30 C1N 3 0 6 -1/2H 2 0: C, 61.26; H, 5.69; N, 7.65. Found: C, 61.15; H, 5.58; N, 7.51. 10 Example 234 3-isopropylamino-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]methylamino] ethoxy]benzoic acid
CH
3 COOH NN N 3 CNH
CH
3 H H OCH 3
CH
3 285 To a cold (00 C), stirred solution of methyl 3-amino-4-[[2-[3-methoxy-4-[N-(2-methyl 15 phenyl) ureido]phenylacetyl]methylamino]ethoxy]benzoate (300mg, 0.58mmol) in acetone-AcOH DMF (13 mL, 6:6:1, v/v/v) was added NaBH 3 CN (300 mg) and the resulting mixture was stirred for 60 hr at room temp. The mixture was poured into sat. NaHCO 3 and extracted with CHC1 3 . The extract was washed with brine, dried over MgSO 4 , and evaporated to give 280 mg (86%) methyl 3-isopropylamino-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] 20 methylamino]ethoxy]benzoate as a colorless oil. 'H-NMR (CDC1 3 , 400MHz) 8 1.21 (mn, 6H), 2.30 (s, 3H), 3.05 and 3.10 (s, total 2H), 3.59 (s, 3H), 3.62-3.81 (mn, 6H), 3.89 (s, 3H), 4.10 and 4.21 (min, 2H), 6.65-6.80 (min, 4H), 7.10 (min, 1H), 7.20 (s, 3H), 7.32 (min, 2H), 7.54 (d, 1H, J=8.3Hz), 8.00 (s, 1H), 8.07 (d, 1H, J=8.3Hz); MS (FAB) m/z 563 (M+1). A stirred mixture of methyl 3-isopropylamino-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl) 25 ureido]phenylacetyl]methylamino]ethoxy]benzoate (280mg, 0.5mmol), 0.25 N NaOH (6 mL), and THF (6 mL) was heated under reflux for 8 hr. The mixture was poured into water (200 mL), acidified with IN HC1 and the solid was collected with suction. The solid was recrystallized from CHCl 3 -n-hexane-diisopropylether to give 202mg (74%) 285 as a light yellow crystalline powder. mp 130-135 oC; 'H-NMR (CD 3 OD, 400MHz) 8 1.18 and 1.22 (d, total 6H, J=6.3Hz), 2.29 and 30 2.32 (s, total 3H), 3.04 and 3.14 (s, total 2H), 3.60-3.90 (min, 9H), 4.16 and 4.25 (min, total 2H11), 6.80 (min, 3H), 7.02 (min, 1H), 7.12-7.24 (min, 3H), 7.29-7.38 (min, 1H), 7.59 (min, 1H), 8.02 (mn, 1H); MS 422 WO 01/00206 PCT/US00/18079 (FAB) m/z 548 (M+H); Anal. calcd. for C 30
H
36
N
4 0 6 : C, 65.68; H, 6.61. Found: C, 65.80; H, 6.83. Example 235 4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]methylamino]ethyl]methylamino benzoic acid
CH
3 "COOH NNN ~ CH 3 5 CH 3 H H OCH 3 286 To a stirred cold (0o C) solution of methyl 4-[[2-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenylacetyl]methylamino]ethyl]aminobenzoate (300mg, 0.6mmol) in MeCN formaldehyde-AcOH (11 mL, 8:2:1, v/v/v) was added NaBH 3 CN (187 mg, 2.40 mmol), and the resulting mixture was stirred for 18 hr at room temp. The mixture was poured into sat. NaHCO 3 10 and extracted with CHC 3 . The extract was washed with brine, dried over MgSO 4 , and evaporated to give 309 mg (100%) methyl 4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-N' methylamino]ethyl]N-methylaminobenzoate as a colorless oil. 'H-NMR (CDCl 3 , 400MHz) 8 2.30 (min, 3H), 2.98 (min, 7H), 3.46-3.72 (min, 9H), 3.82 (min, 3H), 6.32 (min, 1H), 6.55-6.75 (mn, 4H), 7.05 7.50 (min, 2H), 7.82-8.02 (min, 4H); MS (FAB) m/z 518 (M+1). 15 A stirred mixture of methyl 4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] -2-N'-methylamino]ethyl]methylaminobenzoate (309mg, 0.6mmol) in 1 N NaOH (2.4 mL), and THF (10 mL) was heated under reflux for 8 hr. The mixture was poured into water (200 mL), acidified by the addition of IN HC1. The solid was collected with suction. The crude solid was recrystallized from CHCl 3 -n-hexane-diisopropylether to give 211mg (70%) 286 as a light yellow 20 crystalline powder. mp 125-130 oC; IR(KBr) 3338, 2933, 1601, 1529, 1182, 1036, 752cm-'; 1
H
NMR (CD 3 OD, 400MHz) 8 2.28 and 2.29 (s, 3H1), 2.95-3.02 (min, 6H), 3.60 (br, 6H11), 3.80 (s, 1H), 3.86 (s, 2H11), 6.60-6.82 (min, 4H11), 7.01-7.18 (min, 3H), 7.58 (min, 1H), 7.82-7.99 (min, 3H); MS (FAB) m/z 504 (M + 1). Example 236 25 (S)-3-chloro-4-[2-[N-benzyl-N-[3-methoxy-4-[N'-(2-fluorophenyl)ureido]phenylacetyl]amino]-1 propoxy]benzoic acid NO COOH N N (;N 6H 3 C1 H H OCH 3 287 To a stirred solution of NaBH 3 CN (985 mg, 15.68 mmol) in MeOH (5 mL) was added a 423 WO 01/00206 PCT/US00/18079 solution of methyl (S)-3-chloro-4-(2-amino-1-propoxy)benzoate (382 mg, 1.57 mmol) and benzaldehyde (0.19 mL) in MeOH (5 mL), and the resulting mixture was stirred at room temp overnight. The mixture was quenched by H 2 0 and extracted with CHCI 3 . The extract was washed with brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column 5 chromatography on silica-gel with CHCl 3 :MeOH (30:1, v/v) as eluent to give 336 mg (64 %) (S)-3 chloro-4-(2-N-benzylamino-l1-propoxy)benzoate as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.22 (d, 3 H, J=6.4 Hz), 3.21-3.25 (min, 1 H), 3.84-4.03 (min, total 7 H), 6.89-6.91 (mn, 1 H), 7.23-7.37 (min, 5 H), 7.89-7.91 (min, 1 H), 8.05 (min, 1 H). A mixture of 3-methoxy-4-[N'-(2-fluorophenyl)ureido]phenylacetic acid (320 mg, 1.01 10 mmol), methyl (S)-3-chloro-4-(2-N-benzylamino-1-propoxy)benzoate (336 mg, 1.01 mmol), EDC (hydrochloride) (289 mg, 1.51 mmol), HOBt (204 mg, 1.51 mmol) and DMAP (25 mg, 0.20 mmol) in DMF (7 mL) was stirred at room temp for 2 days. The mixture was diluted with EtOAc, washed with 0.5 N HC1, brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 :MeOH (50:1, v/v) as eluent to give 607 mg 15 (95%) methyl (S)-3-chloro-4-[2-[N-benzyl-N-[3-methoxy-4-[N'-(2-fluorophenyl)ureido]phenyl acetyl]amino]-1-propoxy]benzoate as a pale yellow amorphous solid. 'H-NMR (CDCl 3 ) 6 1.20 1.27 (min, 3 H), 3.62 (s, 2 H), 3.73-4.16 (series of m, total 9 H), 4.71 (bs, 2 H), 6.67-7.38 (series of min, total 12 H), 7.77-8.17 (series of m, total 5 H). To a stirred solution of methyl (S)-3-chloro-4-[2-[N-benzyl-N-[3-methoxy-4-[N'-(2 20 fluorophenyl) ureido]phenylacetyl]amino]-l-propoxy]benzoate (607 mg, 0.96 mmol) in THF (6 mL) was added 0.5 N NaOH (6 mL), and the resulting mixture was heated under reflux overnight. The mixture was poured into ice-1 N HCI and the solid was collected. The crude solid was recrystallized to give 192 ming (32%) 287 as a white crystalline powder. mnp 125-130 oC; 'H-NMR (DMSO-ds) 8 1.07-1.23 (min, 3 H), 3.76 (s, 2 H), 3.85 (s, 3 H), 3.90-4.26 (m, 3 H), 4.56 (s, 2 H), 25 6.66-7.38 (series of m, total 10 H), 7.82-8.20 (series of min, total 5 H), 8.70-8.74 (mn, 1 H), 9.18-9.20 (mn, 1 H), 13.02 (bs, 1 H); MS (FAB)m/z 621 (M+1); Anal. Calcd for C 3 zH 3
,CFN
3 0 6 .1/4H 2 0: C, 63.46; H, 5.08; Cl, 5.68; F, 3.04; N, 6.73. Found: C, 63.67; H, 5.16; Cl, 5.75; F, 2.95; N, 6.55. Example 237 3-(N-isopropyl-N-methylamino)-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl] 30 methylamino]ethoxy]benzoic acid 424 WO 01/00206 PCT/US00/18079
H
3 N NCOOH P-NY-N-( O H3 CYIN..CH
CH
3 H H OCH 3
CH
3 288 To a stirred cold (00 C) solution of methyl 3-isopropylamino-4-[[2-[3-methoxy-4-[N'-(2 methylphenyl)ureido]phenylacetyl]methylamino]ethoxy]benzoate (324mg, 0.58mmol) in CH 3
CN
formaldehyde-AcOH (11 mL, 8:2:1, v/v/v) was added NaBH 3 CN (145 mg, 2.30 mmol), and the 5 resulting mixture was stirred for 18 hr at room temp. The mixture was poured sat. NaHCO 3 and the solid was collected with suction. The crude solid was dissolved in CHCl 3 (20mL), and the solution was washed with brine, dried over MgSO 4 , and evaporated to give 317 mg (95%) methyl 3-(N-isopropyl-N-methylamino)-4-[[2-[3-methoxy-4-[N-(2-methylphenyl)ureido]phenylacetyl] methylamino]ethoxy] benzoate as a colorless oil. 'H-NMR (CDC13) 8 1.02 (min, 6H11), 2.29 (s, 3H1), 10 2.63 (min, 3H11), 3.02-3.20 (min, 3H11), 3.49-3.80 (min, 8H11), 3.88 (s, 3H), 4.06 and 4.21 (min, total 2H11), 6.59 (min, 1H), 6.76 (min, 2H), 7.11-7.23 (min, 3H11), 7.50-7.62 (min, 3H), 8.05 (d, 1H11, J=8.3Hz); MS (FAB) m/z 576 (M++1). A stirred mixture of methyl 3-(N-isopropyl-N-methylamino)-4-[[2-[3-methoxy-4-[N'-(2 methylphenyl)ureido]phenylacetyl]methylamino]ethoxy]benzoate(317mg, 0.55mmol) in 0.25 N 15 NaOH (8.2 mL) and THF (8 mL) was heated under reflux for 8 hr. The mixture was poured into water (200 mL) , acidified by the addition of IN HCI and the solid was collected with suction. The crude solid was recrystallized from CHCl 3 -n-hexane-diisopropylether to give 288 as a light yellow crystalline powder. mp 130-135 'C; IR (KBr) 2970, 1537, 1038, 754 cm~'; 'H-NMR (CD 3 OD) 8 1.12(m, 6H), 2.29 (s, 3H11), 2.76 (min, 1H1), 2.89 (s, 2H11), 3.02 (s, 1H), 3.21 (s, 2H11), 3.72 (s, 2H), 3.80 20 (s, 3H), 3.84 (min, 3H), 4.13 and 4.40 (min, total 2H), 6.76 (d, 1H, J=7.8Hz), 6.86 (s, 111), 7.00 (inm, 2H11), 7.18 (min, 3H11), 7.57 (d, 1H, J=7.8Hz), 7.95 (min, 2H); MS (FAB) m/z 562 (M'+1); Anal. Calcd for C 3
,
1
H
3
,N
4 0 6 -2.011H 2 0: C, 62.19; H, 7.07; N, 9.36. Found: C, 62.54; H, 6.85; N, 8.90. Example 238 3-(1 -piperidinyl)-4-[[2-[3-methoxy-4-[N-(2-fluorophenyl)ureidolphenylacetyllmethylamino] 25 ethoxy]benzoic acid
CH
3 COOH 0 N rN H OCH 3 289 To a stirred solution of methyl 3-amino-4-[[2-[3-methoxy-4-[N-(2-fluorophenyl)ureido] phenylacetyl]methylamino]ethoxy]benzoate (300mg, 0.57mmol) in THF (2 mL) was added a solution of glutaraldehyde (50% aqueous solution) (0.13mL, 0.69mmol) in MeOH (1.4 mL), THF 425 WO 01/00206 PCT/US00/18079 (0.993mL), and 3N H 2
SO
4 ( 0.952 mL) at 00 C. To the solution was added NaBH 3 CN (150 mg), DMF (5 mL), and MeOH (2 mL) at room temp, and the resulting mixture was stirred for 15 hr. The mixture was poured into sat. NaHCO 3 and extracted with CHCl 3 . The extract was washed with brine, dried over MgSO 4 , and evaporated under a reduced pressure. The residue was 5 chromatographed on silica gel with toluene:acetone (7:3, v/v) as eluent to give 145 mg (43%) methyl 3-(1-piperidinyl)-4-[[2-[3-methoxy-4-[N'-(2-fluorophenyl)ureido]phenylacetyl]lmethyl amino]ethoxy]benzoate as a colorless oil. 'H-NMR (CDCl 3 , 400MHz) 8 1.58-1.72 (min, 6H), 2.80 2.92 (min, 4H), 3.26 (s, 2H), 3.58 (s, 2H), 3.69 (s, 2H), 3.80-3.89 (min, 7H), 4.22 (min, 2H), 6.72 (s, 1H), 6.78 (min, 2H), 6.95-7.18 (min, 2H), 7.32 (s, 1H1), 7.60 (s, 1H), 7.63 (d, 1H, J=7.8Hz), 7.98 (d, 10 1H, J=7.8Hz), 8.18 (min, 1H); MS (FAB) m/z 593 (M+1). A stirred mixture of methyl 3-(1-piperidinyl)-4-[[2-[3-methoxy-4-[N'-(2-fluorophenyl) ureido]phenylacetyl]methylamino]ethoxy]benzoate (290mg, 0.49mmol), IN NaOH (1.95 mL) in MeOH(5mL) and THF (10 mL) was heated under reflux for 4 hr. The mixture was poured into water (200 mL), acidified with IN HCI (pH=4.0), and extracted with CHCI 3 -MeOH(9:1, v/v). The 15 combined extract was dried over MgSO 4 , and evaporated. The residue was crystallized from diisopropylether-hexane to give 201mg (71%)289 as a light yellow crystalline powder. mp 125 130 0 C; IR (KBr) 3338, 2935, 1599, 1537, 1041, 752 cm-'; 'H-NMR (CD 3 OD) 8 1.52-1.73 (mn, 6H), 2.88 (s, 3H), 2.98 (br, 11), 3.09 (s, 1H), 3.23 (s, 2H), 3.66 (s, 2H), 3.75 (s, 1H), 3.82 (s, 4H), 4.13 and 4.29 (min, total 2H), 6.78 (min, 2H), 6.99 (min, 2H), 7.10 (min, 2H), 7.60-7.70 (min, 2H), 7..96-8.07 20 (min, 2H); MS (FAB) m/z 578 (M+1); Anal. Calcd for C 3 lH 3 5
FN
4 0 6 -0.5H 2 0: C, 63.36; H, 6.17; N, 9.53. Found: C, 63.22; H, 6.15; N, 9.16. Example 239 3-amino-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]methylamino]ethoxy] benzoic acid
CH
3 -COOH .- N-N NH 2 25 CH 3 H H CH 3 290 To a stirred solution of methyl 3-amino-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]methylamino]ethoxy]benzoate (300 mg, 0.58 mmnol) in MeOH-THF (2:1, v/v, 12 mL) was added 0.25 N NaOH (9 mL), and the resulting mixture was heated under reflux for 16 hr. The mixture was poured into water (100 mL) and acidified by the addition of IN HCL. The solid was 30 collected with suction. The residue was recrystallized from diisopropylether to give 243 mg (83%) 290 as a yellow crystalline powder. mp 125-130 'C; IR (KBr) 3346, 2935, 1533, 1211, 1034, 756, 426 WO 01/00206 PCT/US00/18079 637cm'; 'H-NMR (DMSO-d 6 ) 5 2.29 (s, 3H), 3.05 (s, 1H), 3.72-3.85 (m, 7H), 4.12 and 4.25 (m, total 2H), 6.76-6.89 (m, 3H), 7.00 (m, 1H), 7.18 (m, 2H), 7.39 (m, 2H), 7.60 (d, 1H, J=7.8Hz), 7.96 (m, 1H1); Anal.Calcd for C 27
H
30
N
4 0 6 -0.5H 2 0: C, 62.90; H, 6.06; N, 10.87. Found: C, 63.03; H, 6.14; N, 10.56. 5 Example 240 3-(1-piperidinyl)-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]methylamino] ethoxy]benzoic acid
CH
3 COOH NON N o I
CH
3 H H OCH 3 291 To a cooled and stirred solution of methyl 3-aino-4-[[2-[3-methoxy-4-[AP-(2 10 methylphenyl) ureido]phenylacetyl]methylamino]ethoxy]benzoate (573mg, 1. Immol) in THF MeOH (2:1, v/v, 6mL) was added glutaraldehyde (0.423mL, 2.2mmol), 3N H 2
SO
4 (1.83mL, 5.5mmol), and NaBH 3 CN (276mg, 4.4mmol). The resulting mixture was stirred for 18 hr at room temp. The mixture was poured into sat. NaHCO 3 (100mL) and the solid was collected with suction. The crude solid was purified by column chromatography on silica-gel with toluene-acetone (10:0 15 to 4:1, v/v) to give 240mg (37%) methyl 3-(1-piperidinyl)-4-[[2-[3-methoxy-4-[N'-(2-methyl phenyl)ureido]phenylacetyl]methylamino]ethoxy] benzoate as a gum. 'H-NMR (CD 3 OD) 8 1.52 1.72 (m, 6H), 2.30 (s, 3H), 2.36 (s, 2H), 2.89 (br s, 3H), 2.98 (m, 1H1), 3.05 (s, 1H), 3.20 (s, 2H), 3.68 (s, 2H), 3.69 (m, 2H), 3.79 (m, 2H), 3.88 (s, 2H), 4.08 and 4.21 (m, 2H), 6.35 and 6.42 (s, total 1H), 6.70 (s, 1H), 6.70 (s, 1H), 6.79 (m, 2H), 7.09-7.24 (m, 4H), 7.50-7.65 (m, 3H), 8.05 (d, 20 1H, J=8.3Hz); MS (FAB) m/z 588 (M+1). A stirred mixture of methyl 3-(1-piperidinyl)-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenylacetyl]methylamino]ethoxy]benzoate (240 mg, 0.41 mmol), 1 N NaOH (1.63 mL) in MeOH (6.5 mL), HO20 (5 mL), and THF (6.5 mL) was heated under reflux for 11 hr. The mixture was poured into water (200 mL), acidified by the addition of IN HCI until pH=4.0, and the solid 25 was collected with suction. The aqueous layer was extracted with CHC1 3 -MeOH (9:1, v/v, 30mL x 3). The precipitate was dissolved in the combined organic extract. The organic layer was dried over MgSO 4 and evaporated. The residue was crystallized from diisopropylether to give 151 mg (65 %) 291 as a light yellow crystalline powder. mp 120-125 0 C; IR (KBr) 3354, 2935, 1535, 1252, 1217, 1034, 754, 638 cm-'; 'H-NMR (CD 3 OD) 8 1.55-1.72 (m, 6H11), 2.30 (s, 3H), 2.89 (br, 30 2H), 2.98 (br, 1H), 3.09 (s, 1H), 3.22 (s, 2H), 3.69 (s, 3H), 3.74 (s, 1H), 3.83 (m, 4H), 4.12 and 4.28 (m, total 2H1), 6.75 (m, 2H), 6.88-7.02 (m, 1H), 7.17 (m, 2H), 7.58-7.73 (m, 4H1), 7.99 (d, 1H, 427 WO 01/00206 PCT/US00/18079 J=8.3Hz); MS (FAB) m/z 574 (M++1); Anal. Calcd for C 32
H
38
N
4 0 6 -0.5H 2 0: C, 65.85; H, 6.73; N, 9.60. Found: C, 65.94; H, 6.88; N, 9.03. Example 241 3-amino-4-[[2-[3-methoxy-4-[N'-(2-fluorophenyl)ureido]phenylacetyl]methylamino]ethoxy] 5 benzoic acid
H
3 COOH ' _N
NH
2 H H OCH 3 292 A stirred mixture of methyl 3-amino-4-[[2-[3-methoxy-4-[N'-(2-fluorophenyl)ureido] phenylacetyl]methylamino]ethoxy]benzoate (250 mg, 0.48 mmol) in IN NaOH (1.91 mL), MeOH (8 mL), H20 (6 mL), and THF (8 mL) was heated under reflux for 10 hr. The mixture was poured 10 into water (200 mL), acidified by the addition of IN HCI (pH=4.8), and the solid was collected with suction. The crude solid was recrystallized from diisopropylether to give 209 mg (86 %) 292 as a pale yellow crystalline powder. mp 125-130 0 C; IR (KBr) 3325, 2935, 1537, 1209, 1032, 752, 449 cmu'; 'H-NMR (CD 3 OD) 5 3.05 (s, 1H11), 3.31 (s, 2H11), 3.77 (min, 3H11), 3.80 (s, 1H), 3.84 (mn, 3H), 4.14-4.26 (min, 2H11), 6.80-6.87 (min, 3H), 7.01 (min, 1H), 7.10 (min, 2H11), 7.39 (min, 2H11), 7.80 (min, 1H), 15 8.07 (min, 1H11); MS (FAB) m/z 510 (M+I); Anal. Calcd for C 2 ,6H 27
FN
4 0 6 "0.5H 2 0: C, 60.11; H, 5.43; F, 3.66; N, 10.78. Found: C, 60.29; H, 5.40; F, 3.60; N, 10.59. Example 242 (S)-3-amino-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetylamino]-1-propoxy] benzoic acid H ~, COOH N- q NN 0 CH 3
NH
2 20 CH 3 H H OCH 3 293 To a stirred solution of (S)-2-(N-tert-butoxycarbonylamino)-1-propanol (0.90 g, 5.13 mmol), methyl 4-hydroxy-3-nitrobenzoate (1.01 g, 5.12 mmol), and Ph 3 P (1.75 g, 6.67 mmol) in THF (15 mL) was added diisopropyl azodicarboxylate (DIAD) (1.31 mL, 6.65 mmol) at room temp, and the resulting mixture was heated under reflux overnight. The solution was evaporated 25 and the residue was dissolved in CH 2 C1 2 (20 mL) and TFA (10 mL). The mixture was stirred at room temp for 3 hr. The mixture was concentrated in vacuo. The residue was dissolved in CHC1 3 , washed with sat. NaHCO 3 , brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with 5% MeOH in CHC1 3 as eluent to give 313 mg (24%) methyl(S)-3-nitro-4-(2-amino-l-propoxy)benzoate as a pale yellow oil. 'H-NMR 30 (CDCl 3 ) 8 1.22 (d, 3 H, J=6.8 Hz), 3.43-3.48 (min, 1 H), 3.69-3.87 (min, 2 H), 3.89 (s, 3 H), 6.93-6.95 (min, 1 H), 7.99-8.02 (min, 1 H), 8.18-8.21 (min, 1 H). 428 WO 01/00206 PCT/US00/18079 A mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetate (591 mg, 1.23 mmol), methyl (S)-3-nitro-4-(2-amino-1-propoxy)benzoate (313 mg, 1.23 mmol), and Et 3 N (257 mL, 1.84 mmol) in DMF (5 mL) was stirred at room temp for 2 days. The mixture was diluted with EtOAc, washed with 0.5 N HC1, brine, dried over Na 2
SO
4 , and evaporated. The 5 residue was purified by column chromatography on silica-gel with 5% MeOH in CHCl 3 as eluent to give 310 mg (46%) methyl (S)-3-nitro-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl acetylamino]-1-propoxy]benzoate as a gum. 'H-NMR (CDCl 3 ) 8 1.28 (d, 3 H, J=6.8 Hz), 2.31 (s, 3 H), 3.49 (s, 2 H), 3.71 (s, 3 H), 3.92 (s, 3 H), 4.14-4.16 (min, 2 H), 4.38-4.41 (mn, 1 H), 5.88-5.90 (min, 1 H), 6.49 (s, 1 H), 6.70-6.71 (min, 1 H), 6.77-6.79 (min, 1 H), 7.05-7.30 (min, 4 H), 7.55-7.57 (inm, 10 1 H), 8.02-8.06 (min, 2 H), 8.16-8.19 (min, 1 H), 8.51-8.52 (min, 1 H); MS (FAB) m/z 551 (M+1). A stirred solution of methyl (S)-3-nitro-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetylamino]-1-propoxy]benzoate (310 mg, 0.56 mmol) in MeOH-THF (10 mL, 1:1, v/v) was hydrogenated over 5% Pd-C (50 mg, 16 wt%) overnight. The mixture was filtered to remove the catalyst and the filtrate was evaporated to give methyl (S)-3-amino-4-[2-[3-methoxy-4-[N'-(2 15 methylphenyl) ureido]phenylacetylamino]-1-propoxy]benzoate (240 mg) as a gum. To a stirred solution of the above crude product (220 mg) in THF-MeOH (10 mL, 1:1, v/v) was added 0.5 N NaOH (10 mL), and the resulting mixture was heated under reflux for 2 hr. The mixture was poured into ice-H 2 0, and the aqueous layer was made acidic (pH 4.8) by the addition of 1 N HC1. The solid was collected and the crude solid was recrystallized from MeOH 20 CHCl 3 -n-hexane to give 116 mg (2 steps, 44%) 293 as a pale yellow crystalline powder. mp 200 204 oC; 'H-NMR (DMSO-d) 8 1.21 (d, 3 H, J=6.8 Hz), 2.24 (s, 3 H), 3.37 (s, 2 H), 3.80 (s, 3 H), 3.83-3.99 (min, 2 H), 4.10-4.19 (min, 1 H), 4.99 (bs, 2 H), 6.75-7.23 (series of m, total 8 H), 7.79 (d, 1 H, J=7.8 Hz), 7.98 (d, 1 H, J=7.8 Hz), 8.17 (d, 1 H, J=8.3 Hz), 8.46 (s, 1 H), 8.55 (s, 1 H); MS (FAB) m/z 507 (M'+1);Anal. Called for C 27
H
3 0
N
4 0 6 -1/2H 2 0: C, 62.90; H, 6.06; N, 10.87. Found: 25 C, 62.85; H, 6.10; N, 10.51. Example 243 (S)-4-[2-[3-methoxy-4-[N'-(2-bromophenyl)ureido]phenylacetylamino]- 1 -propoxy]benzoic acid H rCOOH NN N OCH 3 Br H H OCH 3 294 To a stirred and cooled (0o C) solution of (S)-2-(N-tert-butoxycarbonylamino)-1-propanol 30 (6.74 g, 0.04 mol), benzyl 4-hydroxybenzoate (8.78 g, 0.04 mol), and Ph 3 P (15.13 g, 0.06 mol) in 429 WO 01/00206 PCT/US00/18079 THF (100 mL) was added diisopropyl azodicarboxylate (DIAD) (11.4 mL, 0.06 mol), and the resulting mixture was heated under reflux overnight. The solution was evaporated and the residue was dissolved in CH 2 C0 2 (30 mL) and TFA (30 mL). The resulting solution was stirred at room temp for 30 min., and the solution was evaporated in vacuo. The residue was dissolved in CHC1 3 , 5 washed with sat. NaHCO 3 , dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 to CHCl 3 :MeOH (9:1, v/v) as eluent to give 6.63 g (2 steps, 60%) benzyl (S)-4-(2-amino-1-propoxy) benzoate as a yellow oil. 'H-NMR (CDCl 3 ) 8 1.18 (d, J= 6.3 Hz, 3 H), 3.35-3.39 (min, 1 H), 3.71-3.75 (min, 1 H), 3.90-3.93 (mn, 1 H), 5.34 (s, 2 H), 6.90-6.93 (min, 2 H), 7.32-7.46 (min, 5 H), 8.01-8.04 (min, 2 H). 10 A mixture of 3-methoxy-4-[N'-(2-bromophenyl)ureido]phenylacetic acid (480 mg, 1.27 mmol), benzyl (S)-4-(2-amino-1-propoxy)benzoate (361 mg, 1.27 mmol), EDC(hydrochloride) (364 mg, 1.90 mmol), HOBt (256 mg, 1.89 mmol), and 4-DMAP (31 mg, 0.25 mmol) in DMF (8 mL) was stirred at room temp. overnight. The mixture was diluted with EtOAc, washed with 0.5 N HCI, brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column 15 chromatography on silica-gel with CHCl 3 to 5% MeOH in CHCl 3 as eluent to give 476 mg (58%) benzyl (S)-4-[2-[3-methoxy-4-[N'-(2-bromophenyl)ureido]phenylacetylamino]-1-propoxy]benzoate as a brown solid. 'H-NMR (CDCl 3 ) 8 1.25-1.28 (min, 3 H), 3.51 (s, 2 H), 3.74 (s, 3 H), 3.95-3.97 (mn, 2 H), 4.36-4.39 (min, 1 H), 5.33 (s, 2 H), 6.75-6.94 (min, 5 H), 7.26-7.70 (mn, 8 H), 7.99-8.26 (inm, 5H). 20 To a stirred solution of benzyl (S)-4-[2-[3-methoxy-4-[N'-(2-bromophenyl)ureido] phenylacetylamino]-1-propoxy]benzoate (476 mg, 1.39 mmol) in THF (10 mL) was added 0.5 N NaOH (10 mL), and the resulting mixture was heated under reflux for 2 hr. The mixture was poured into ice-1 N HC1, and the solid was collected. The crude solid was recrystallized from MeOH-CHCl 3 -n-hexane to give 240 ming (59%) 294 as a white crystalline powder. mp 202-205 oC; 25 'H-NMR (DMSO-d 6 ) 8 1.18 (d, J= 6.8 Hz, 3 H), 3.37 (s, 2 H), 3.82 (s, 3 H), 3.92-4.00 (mn, 2 H), 4.03-4.15 (min, 1 H), 6.77-6.79 (min, 1 H), 6.93-7.03 (min, 4 H), 7.30-7.34 (min, 1 H), 7.59-7.61 (mn, 1 H), 7.87-7.97 (min, 4 H), 8.13-8.15 (min, 1 H), 8.73 (s, 1 H), 8.91 (s, 1 H), 12.63 (bs, 1 H); MS (FAB) m/z 557 (M+1); Anal. Calcd for C 2 6
H
26 BrN 3 06: C, 56.12; H, 4.71; Br, 14.36; N, 7.55. Found: C, 56.11; H, 4.74; Br, 14.56; N, 7.49. 30 Example 244 (S)-4-[[2-[3-methoxy-4-[N '-(2-methylphenyl)ureido]phenylacetylj-(2-aminobenzyl)amino]-1 propoxy] benzoic acid 430 WO 01/00206 PCT/US00/18079 H2N N l'1" " C
O
O
H P-N-O C CH 3 NON
CH
3 H H OCH 3 295 To a stirred cooled (00 C) solution of benzyl (S)-4-(2-amino-1 -propoxy)benzoate (1.50 g, 5.26 mmol) and 2-nitrobenzaldehyde (0.87 g, 5.76 mmol) in MeOH-AcOH (16 mL, 15:1, v/v) was added NaBH 3 CN (1.65 g, 26.3 mmol), and the resulting mixture was stirred at room temp 5 overnight. The mixture was quenched by sat. NaHCO 3 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with CHC1 3 to 5% MeOH in CHC1 3 as eluent to give 931 mg (42%) benzyl (S)-4-[2-(2-nitrobenzylamino) -1-propoxy]benzoate as a yellow oil. 1H-NMR (CDCl 3 ) 8 1.21 (d,J= 6.4 Hz, 3 H), 3.13-3.18 (min, 1 H), 3.88-3.97 (min, 2 H), 4.06-4.20 (mn, 2 H), 5.34 (s, 2 10 H), 6.89-6.94 (min, 2 H), 7.29-7.65 (min, 8 H), 7.94-8.03 (mn, 3 H); MS (FAB) m/z 421 (M +1). A mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetate (460 mg, 0.96 mmol), benzyl (S)-4-[2-(2-nitrobenzylamino)-1-propoxy]benzoate (403 mg, 0.96 mmol), and Et 3 N (200 mL, 1.43 mmol) in DMF (8 mL) was stirred at room temp overnight. The mixture was diluted with EtOAc, washed with 0.5 N HC1, brine, dried over Na 2
SO
4 , and 15 evaporated. The residue was purified by column chromatography on silica-gel with 1% MeOH in CHC1 3 as eluent to give 504 mg (73%) benzyl (S)-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-(2-nitrobenzyl)amino]-1-propoxy]benzoate as a brown amorphous solid. MS (FAB) m/z 717 (M++1). A stirred solution of benzyl (S)-4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl 20 acetyl]-(2-nitrobenzyl)amino]-1-propoxy]benzoate (504 mg, 0.70 mmol) in MeOH-THF (11 mL, 10:1, v/v) was hydrogenated over 5% Pd-C (100 mg, 20 wt%) at 3 atm overnight. The mixture was filtered to remove the catalyst and the filtrate was evaporated. The residue was purified by preparative TLC with 5% MeOH in CHCI 3 as eluent to give 115 mg (27%) 295 as a white powder. MS (FAB) m/z 597 (M'+I). 25 Example 245 (S)-4-[[2-[3-methoxy-4-[N'-(2-bromophenyl)ureido]phenylacetyl]-(2-nitrobenzyl)amino]- 1 propoxy]benzoic acid 431 WO 01/00206 PCT/US00/18079 02. O OOH N N CH 3 rH H OCH 3 296 To a stirred and cooled (00 C) solution of benzyl (S)-4-(2-amino-1-propoxy)benzoate (1.50 g, 5.26 mmol) and 2-nitrobenzaldehyde (0.87 g, 5.76 mmol) in MeOH-AcOH (16 mL, 15:1, v/v) was added NaBH 3 CN (1.65 g, 26.3 mmol), and the resulting mixture was stirred at room temp 5 overnight. The mixture was quenched by sat. NaHCO 3 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHC1 3 to 5% MeOH in CHC1 3 as eluent to give 931 mg (42%) benzyl (S)-4-[2-(2-nitrobenzylamino) -1-propoxy]benzoate as a yellow oil. 'H-NMR (CDCl 3 ) 8 1.21 (d, 3 H, J=6.4 Hz), 3.13-3.18 (min, 1 H), 3.88-3.97 (min, 2 H), 4.06-4.20 (min, 2 H), 5.34 (s, 2 H), 10 6.89-6.94 (min, 2 H), 7.29-7.65 (min, 8 H), 7.94-8.03 (min, 3 H); MS (FAB) m/z 421 (M +1). A mixture of 3-methoxy-4-[N'-(2-bromophenyl)ureido]phenylacetic acid (476 mg, 1.26 mmol), benzyl (S)-4-[2-(2-nitrobenzylamino)-1-propoxy]benzoate (528 mg, 1.26 nummol), EDC(hydrochloride) (361 mg, 1.88 mmol), HOBt (255 mg, 1.89 mmol), and DMAP (30 mg, 0.25 mmol) in DMF (10 mL) was stirred at room temp. overnight and at 600 C for 1 day. The mixture 15 was diluted with EtOAc, washed with 0.5 N HC1, brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHCI 3 to 2% MeOH in CHCI 3 as eluent to give benzyl (S)-4-[[2-[3-methoxy-4-[N'-(2-bromophenyl)ureido]phenylacetyl]-(2 nitrobenzyl)amino]-1-propoxy]benzoic acid as an oil, which is used to the subsequent reaction without further purification. 20 To a stirred solution of the above crude in THF-MeOH (10 mL, 1:1, v/v) was added 0.5 N NaOH (10 mL), and the resulting mixture was heated under reflux for 3 hr. The mixture was poured into ice-H 2 0, and the basic aqueous layer was made acidic (pH 4.3) with 1 N HC1. The solid was collected, and the crude solid was purified by preparative TLC with 5% MeOH in CHCI 3 as eluent to give 162 mg (2 steps, 19%) 296 as a white amorphous solid. MS (FAB) m/z 692 25 (M +1); Anal. Calcd for C 33
H
31 BrN 4 Os 8 7/4H 2 0: C, 54.82; H, 4.81; N, 7.75. Found: C, 54.80; H, 4.61; N, 7.24. Example 246 4-[2-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]lphenylacetamido]lethoxy]benzoic acid 432 WO 01/00206 PCT/US00/18079 H ,aCOOH NNy Me H H OMe 297 To a cooled (0 0 C) solution of 2-(N-tert-butoxycarbonylamino)ethanol (3.20 g, 19.9 mmol), methyl 4-hydroxybenzoate (3.02 g, 19.9 mmol) and Ph3P (6.25 g, 23.8 mmol) in THF (50 ml) was added dropwise DIAD (4.69 ml, 23.8 mmol) over for 5 min. The reaction mixture was heated 5 under reflux for 3 hr. The mixture was evaporated and the residue was dissolved in CH 2 C1 2 (30 ml) and TFA (30 ml). The reaction mixture was stirred at room temperature overnight. The mixture was concentrated in vacuo and the residue was dissolved in CHC1 3 and H 2 0. The solution was made basic by sat. NaHCO 3 and extracted with CHC13. The extract was washed with brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica 10 gel with CHCl 3 -MeOH (30:1, v/v) as eluent to give methyl 4-(2-aminoethoxy)benzoate (1.03 g, 34% for 2 steps) as a colorless oil. 'H-NMR (CDCl 3 ) 8 3.10-3.13 (mn, 2 H), 3.89 (s, 3 H), 4.03 4.06 (min, 2 H), 6.92-6.94 (min, 2 H), 7.98-8.00 (min, 2 H). A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (557 mg, 1.77 mmol), methyl 4-(2-aminoethoxy)benzoate (346 mg, 1.77 mmol), EDCHCI (408 mg, 2.13 mmol), 15 HOBt (287 mg, 2.12 mmol) and DMAP (52 mg, 0.43 mmol) in DMF (10 ml) was stirred at room temperature for 2 days. The mixture was diluted with EtOAc and H 2 0. The resulting precipitate was collected to give methyl 4-[2-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetamido] ethoxy]benzoate (598 mg, 69%) as a white crystalline powder. 'H-NMR (DMSO-d 6 ) 8 2.23 (s, 3 H), 3.36 (s, 2 H), 3.42-3.45 (min, 2 H), 3.79 (s, 3 H), 3.81 (s, 3 H), 4.02-4.09 (mn, 2 H), 6.73-6.75 20 (min, 1 H), 6.90-6.94 (min, 2 H), 7.02-7.04 (mn, 2 H), 7.09-7.15 (min, 2 H), 7.77-7.79 (mn, 1 H), 7.88 7.90 (min, 2 H), 7.95-7.98 (min, 1 H), 8.23-8.24 (min, 1 H), 8.44 (s, 1 H), 8.53 (s, 1 H); MS (FAB), m/z 492 (M +1); Anal. Calcd for C 27
H
2 9
N
3 0 6 1/4H 2 0: C, 65.38; H, 5.99; N, 8.47. Found: C, 65.26; H, 5.99; N, 8.49. To a stirred solution of methyl 4-[2-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido] 25 phenylacetamido]ethoxy]benzoate (280 mg, 0.57 mmol) in THF-MeOH (10 ml, 1:1, v/v) was added 0.5 N NaOH (10 ml) and the reaction mixture was heated under reflux for 5 hr. The mixture was cooled to room temperature and poured into ice-1 N HC1. The resulting precipitate was collected and recrystallized from Et20-CHCl 3 -MeOH to give 297 (135 mg, 50%) as a white crystalline powder. MW 477.51 'H-NMR (DMSO-d 6 ) 5 2.24 (s, 3 H), 3.38 (s, 2 H), 3.43-3.47 (inm, 30 2 H), 3.82 (s, 3 H), 4.06-4.09 (min, 2 H), 6.76-6.78 (min, 1 H), 6.92-7.17 (mn, 6 H), 7.79 (d, J = 8.1 Hz, 1 H), 7.88-7.89 (min, 2 H), 7.98 (d, J= 8.1 Hz, 1 H), 8.24-8.27 (min, 1 H), 8.45 (s, 1 H), 8.55 (s, 433 WO 01/00206 PCT/US00/18079 1 H); MS (FAB) m/z 478 (MW+1); Anal. Calcd for C 2
H
27
N
3 0 6 1/4H 2 0: C, 64.79; H, 5.75; N, 8.72. Found: C, 64.67; H, 5.63; N, 8.60. Example 247 (S)-4-[2-N-[4-[N '-(2-bromophenyl)ureido]-3-methoxyphenylacetamido]-1 -propoxy]benzoic acid H 5~COOH 0 N N 5r H H 6Me 298 To a cooled (0 0 C), stirred solution of (S)-2-(N-tert-butoxycarbonylamnino)-1-propanol (6.74 g, 0.04 mol), benzyl 4-hydroxybenzoate (8.78 g, 0.04 mol), and Ph 3 P (15.13 g, 0.06 mol) in THF (100 ml) was added DIAD (11.4 ml, 0.06 mol), and the reaction mixture was heated under reflux overnight. The solution was evaporated and the residue was dissolved in CH 2 C0 2 (30 ml) 10 and TFA (30 ml). The solution was stirred at room temperature for 30 min. and the solution was concentrated in vacuo. The residue was dissolved in CHC1 3 , washed with sat. NaHCO 3 , dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHC1 3 to CHCl 3 -MeOH (9:1, v/v) as eluent to give benzyl (S)-4-(2-amino-1-propoxy)benzoate (6.63 g, 2 steps, 60%) as a yellow oil. 'H-NMR (CDC 3 ) 8 1.18 (d,J = 6.3 Hz, 3 H), 3.35-3.39 (inm, 15 1 H), 3.71-3.75 (min, 1 H), 3.90-3.93 (min, 1 H), 5.34 (s, 2 H), 6.90-6.93 (min, 2H), 7.32-7.46 (mn, 5H), 8.01-8.04 (min, 2H). A mixture of 4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (480 mg, 1.27 mmol), benzyl (S)-4-(2-amino-1-propoxy)benzoate (361 mg, 1.27 mmol), EDCHCI (364 mg, 1.90 mmol), HOBt (256 mg, 1.89 mmol) and DMAP (31 mg, 0.25 mmol) in DMF (8 ml) was stirred at 20 room temperature overnight. The mixture was diluted with EtOAc, washed with 0.5 N HC1, brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica gel with CHCl 3 to 5% MeOH in CHCl 3 as eluent to give benzyl (S)-4-[2-[4-[N'-(2-bromophenyl) ureido]-3-methoxyphenylacetamido] -1-propoxy]benzoate (476 mg, 58%) as a brown solid. 'H-NMR (CDCl 3 ) 5 1.25-1.28 (min, 3 H), 3.51 (s, 2 H), 3.74 (s, 3 H), 3.95-3.97 (mn, 2 H), 4.36-4.39 25 (min, 1 H), 5.33 (s, 2 H), 6.75-6.94 (min, 5 H), 7.26-7.70 (min, 8 H), 7.99-8.26 (mn, 5 H). The a stirred solution of benzyl (S)-4-[2-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl acetamido]-l-propoxy]benzoate (476 mg, 1.39 mmol) in THF (10 ml) was added 0.5 N NaOH (10 ml), and the reaction mixture was heated under reflux for 2 hr. The mixture was poured into ice-1 N HCI, and the resulting precipitate was collected. The crude solid was purified by 30 recrystallization from MeOH-CHCl 3 -n-hexane to give 298 (240 mg, 59%) as a white crystalline powder. MW 556.41 'H-NMR (DMSO-d6) 8 1.18 (d, J= 6.8 Hz, 3 H), 3.37 (s, 2 H), 3.82 (s, 3 434 WO 01/00206 PCT/US00/18079 H), 3.92-4.00 (min, 2 H), 4.03-4.15 (min, 1 H), 6.77-6.79 (min, 1 H), 6.93-7.03 (min, 4 H), 7.30-7.34 (inm, 1 H), 7.59-7.61 (min, 1 H), 7.87-7.97 (min, 4 H), 8.13-8.15 (min, 1 H), 8.73 (s, 1 H), 8.91 (s, 1 H), 12.63 (bs, 1 H); MS (FAB) m/z 557 (M+ 1); Anal. Calcd for C 26
H
26 BrN 3 06: C, 56.12; H, 4.71; Br, 14.36; N, 7.55. Found: C, 56.11; H, 4.74; Br, 14.56; N, 7.49. 5 Example 248 (S)-4-[2-N-[[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl]-N-methylacetamido]-1-propoxy] benzoic acid N O COOH Me H H OMe 299 To a cooled (0OC) solution of (S)-2-[(N-tert-butoxycarbonyl)amino]-1-propanol (3.08 g, 10 17.6 mmol), methyl 4-hydroxybenzoate (2.67 g, 17.6 mmol) and Ph 3 P (5.53 g, 21.1 mmol) in THF (35 ml) was added dropwise DIAD (4.15 ml, 21.1 mmol). The reaction mixture was heated under reflux overnight. The mixture was evaporated and the residue was purified by column chromatography on silica-gel with n-hexane-EtOAc (5:1, v/v) as eluent to give methyl (S)-4-[2 [(N-tert-butoxycarbonyl)amino]-1-propoxylbenzoate (1.24 g, 23%/) as a white solid. 'H-NMR 15 (CDC1 3 ) 8 1.30 (d, J = 6.8 Hz, 3 H), 1.45 (s, 9 H), 3.89 (s, 3 H), 3.98-3.99 (min, 2 H), 4.07 (min, 1 H), 4.76 (min, 1 H), 6.91-6.93 (min, 2 H), 7.98-8.00 (min, 2 H); MS (FAB) m/z 310 (M'+1). To a stirred solution of methyl (S)-4-[2-[(N-tert-butoxycarbonyl)amino]-1-propoxy] benzoate (1.24 g, 4.01 mmol) in CH 2 C1 2 (10 ml) was added TFA (10 ml) and the reaction mixture was stirred at room temperature overnight. The mixture was concentrated in vacuo, and made 20 basic by sat. NaHCO 3 . The mixture was extracted with CHCl 3 , washed with brine, dried over
K
2 C0 3 and evaporated to give methyl (S)-4-(2-amino-l-propoxy) benzoate (790 mg, 94%) as a yellow oil. 'H-NMR (CDC13) 8 1.19 (d, J = 6.7 Hz, 3 H), 3.34-3.42 (min, 1 H), 3.70-3.77 (min, 1 H), 3.86-3.94 (series of s and m, total 4 H), 6.92 (d, J = 9.0 Hz, 2 H), 7.98 (d, J = 9.0 Hz, 2 H). To a cooled (0oC) solution of methyl (S)-4-(2-amino-1-propoxy)benzoate (790 mg, 3.78 25 mmol) and Et 3 N (630 ml, 4.52 mmol) in THF (10 ml) was added TFAA (640 ml, 4.53 mmol) and the reaction mixture was stirred at room temperature for 4 days. The mixture was diluted with 0.5 N HCI and extracted with CHCI 3 . The extract was washed with brine, dried over K 2 CO3, and evaporated. The residue was purified by recrystallization from n-hexane-CHCl 3 to give methyl (S) 4-[2-(trifluoroacetamido)-1-propoxy]benzoate (790 mg, 69%) as a white crystalline material. 'H 30 NMR (CDCl 3 ) 8 1.42 (d, J = 6.8 Hz, 3 H), 3.89 (s, 3 H), 4.01-4.13 (min, 2 H), 4.44-4.50 (min, 1 H), 6.57-6.61 (min, 1 H), 6.92 (d, J = 9.0 Hz, 2 H), 8.00 (d, J = 9.0 Hz, 2 H); MS (FAB) m/z 306 435 WO 01/00206 PCT/US00/18079 (M*+1); Anal. Calcd for C 13
H,
4
F
3
NO
4 : C, 51.15; H, 4.62; F, 18.67; N, 4.59. Found: C, 51.14; H, 4.60; F, 18.50; N, 4.54. To a stirred solution of methyl (S)-4-[2-(trifluoroacetamniido)-1-propoxy]benzoate (695 mg, 2.28 mmol) and K 2
CO
3 (630 mg, 4.56 mmol) in DMF (10 ml) was added Mel (210 ml, 3.37 5 mmol) and the reaction mixture was stirred at room temperature for 2 days. The mixture was diluted with H20 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was purified by column chromatography on silica-gel with EtOAc-CHCI 3 (1:19, v/v) as eluent to give methyl (S)-4-[2-[(N-methyl)trifluoroacetamido]-1 propoxy]benzoate (720 mg, 99%) as a white solid. mp 73-75oC; 'H-NMR (CDCl 3 )8 1.36-1.39 (inm, 10 3 H), 2.96 and 3.10 (each s, total 3 H), 3.89 (s, 3 H), 3.99-4.14 (min, 2 H), 4.84-4.92 (mn, 1 H), 6.88 6.92 (min, 2 H), 7.98-8.00 (m, 2 H); MS (FAB) m/z 320 (M++1); Anal. Calcd for C 4
H
16
,F
3
NO
4 : C, 52.67; H, 5.05; F, 17.85; N, 4.39. Found: C, 52.76; H, 5.09; F, 17.53; N, 4.32. To a stirred solution of methyl (S)-4-[2-[(N-methyl)trifluoroacetamido]-1-propoxy] benzoate (710 mg, 2.22 mmol) in MeOH-H 2 0 (10 ml, 1:1, v/v) was added K 2 CO3 (460 mg, 3.33 15 mmol) and the reaction mixture was stirred at room temperature overnight. The mixture was diluted with H 2 0 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated to give methyl (S)-4-[2-(N-methylamino)-1-propoxy]benzoate (430 mg, 87%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.17 (d, J = 6.6 Hz, 3 H), 2.48 (s, 3 H), 2.98-3.06 (inm, 1 H), 3.86-3.96 (series of s and m, total 5 H), 6.92 (d, J = 9.0 Hz, 2 H), 7.98 (d, J = 9.0 Hz, 2 H). 20 A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (605 mg, 1.93 mmol), methyl (S)-4-[2-(N-methylamino)-1-propoxy]benzoate (430 mg, 1.93 mmol), EDCHC1 (444 mg, 2.32 mmol), HOBt (313 mg, 2.32 mmol), Et 3 N (320 ml, 2.30 mmol) in THF (13 ml) was stirred at room temperature overnight. The mixture was diluted with H20 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was 25 purified by column chromatography on silica-gel with CHCl 3 -MeOH (100:1 to 75:1, v/v) as eluent to give methyl (S)-4-[2-N-[[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl]-N-methylacetamido] -1-propoxy]benzoate (953 mg, 95%) as a white foam. 'H-NMR (CDCl 3 ) 8 1.12-1.13 and 1.25-1.27 (each min, total 3 H), 2.27 (s, 3 H), 2.84 and 2.92 (each s, total 3 H), 3.63 (s, 3 H), 3.67 (s, 2 H), 3.71-4.05 (series of s and m, total 5 H), 4.39-4.44 and 4.96-5.01 (each min, total 1 H), 6.66-6.85 (m, 30 5 H), 7.09-7.27 (min, 4 H), 7.53-7.55 (min, 1 H), 7.92-7.98 (min, 2 H), 8.04-8.08 (min, 1 H); MS (FAB) m/z 520 (M++1); Anal. Calcd for C 2 9H 33
N
3 0 6 11/4H 2 0: C, 61.20; H, 6.82; N, 7.38. Found: C, 61.14; H, 5.86; N, 7.16. 436 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl (S)-4-[2-N-[[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenyl]-N-methylacetamido]-1 -propoxy]benzoate (663 mg, 1.28 mmol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction mixture was heated under reflux for 5 hr. After cooled to room temperature, the mixture was poured into ice-1 N HCI and the resulting precipitate was 5 collected under a reduced pressure. The crude solid was dissolved in CHCl 3 and evaporated. The residue was washed with Et 2 O to give 299 (465 mg, 72%) as a white amorphous solid. MW 505.56 'H-NMR (DMSO-d 6 ) 8 1.11-1.15 (min, 3 H), 2.25 (s, 3 H), 2.73 and 2.88 (each s, total 3 H), 3.60-3.76 (min, 2 H), 3.83 (s, 3 H), 4.03-4.12 (min, 2 H), 4.41-4.50 and 4.48-4.94 (each m, total 1 H), 6.71-6.76 (min, 1 H), 6.84-6.86 (min, 1 H), 6.91-7.01 (min, 3 H), 7.11-7.12 (min, 2 H), 7.78-7.80 (min, 1 10 H), 7.86-7.90 (min, 2 H), 8.01-8.03 (min, 1 H), 8.45 (s, 1 H), 8.54 (s, 1H); MS (FAB) m/z 520 (M++1); Anal. Calcd for C2 8
H
31
N
3 0 6 3/2H 2 0: C, 63.15; H, 6.43; N, 7.89. Found: C, 63.09; H, 5.99; N, 7.64. Example 249 4-[2-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetamido]-2-methyl-1-propoxy]benzoic acid H COOH NNN 15 Me H H OMe 300 A stirred mixture of methyl 4-hydroxybenzoate (3 g, 19.72 mmol), K 2
CO
3 (6.8 g, 49.3 mmol), 3-chloropivalic acid (2.9 g, 21.69 mmol) and catalytic amount of KI (200 mg) in DMF (70 ml) was heated at 100 oC for 14days under a current of nitrogen. The mixture was poured into ice water, and extracted with EtOAc. The extracts were washed with brine, dried over Na 2
SO
4 , and 20 evaporated. The residue was chromatographed on silica gel (50ml) with CHC1 3 -EtOH (10:1, v/v) as eluent to give the title compound (1 g, 23%) as an amorphous solid. 'H-NMR (CDCl 3 ) 8 1.37 (s, 6 H), 3.89 (s, 3 H), 4.03 (s, 2 H), 6.92 (d, J =9 Hz, 2 H), 7.98 (d, J = 9Hz, 2 H). To a stirred mixture of 2,2-dimethyl-3-(4-methoxycarbonyl)phenoxypropionic acid (720 mg, 2.85 mmol) and triethylamine (0.46 ml, 3.28 mmol) in tert-BuOH (10 ml) and benzene (10 25 ml) was added a solution of diphenyl phosphoryl azide (870 mg, 3.14 mmol) in benzene (3 ml) at room temperature. The resulting mixture was heated at reflux for 20 hr. After cooling, ice and IN HCI (5ml) was added to the mixture and extracted with toluene. The extracts were washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica gel (50ml) with toluene-EtOAc (10:1, v/v) as eluent to give methyl 4-[1-(2-amino-2-methyl)prpoxy]benzoate 30 as a gum (520 mg), which was used to the subsequent reaction without further purification. 'H NMR (CDCl 3 ) 8 1.41 (s, 9 H), 3.89 (s, 3 H), 4.04 (s, 2 H), 4.69 (br s, 1H), 6.94 (dd, J=2 and 7 Hz, 2 H), 7.98 (dd, J =2 and 7 Hz). 437 WO 01/00206 PCT/US00/18079 A solution of methyl 4-[1-(2-methyl-2-tert-butoxycarbonylamino-)prpoxy]benzoate (520 mg) and anisole (0.175 ml, 1.61 mmol) in CH 2 C1 2 (5 ml) and TFA (3 ml) was stirred at room temperature for 18 hr. The mixture was evaporated off. The residue was dissolved in CH 2 Cl 2 , and the mixture was made basic by the addition of sat. NaHCO 3 . Separated CH 2 Cl 2 layer was dried 5 over Na 2
SO
4 and Na 2
CO
3 , and evaporated. The residue was chromatographed on silica gel with CHCl 3 -EtOH (10:1, v/v) as eluent to give methyl 4-[1-(2-amino-2-methyl)prpoxy]benzoate (250 mg, 39% in two steps) as a gum. 1H-NMR (CDCl 3 ) 8 3.75 (s, 2 H), 3.89 (s, 3 H), 6.93 (d, J=8.8 Hz, 2 H), 7.98 (d, J=8.8 Hz, 2 H). To a stirred mixture of methyl 4-[1-(2-amniino-2-methyl)prpoxy]benzoate (250 mg, 1.12 10 mmol), 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (352 mg, 1.12 mmol), 4 DMAP (165 mg, 1.34 mmol) in DMF (10 ml) was added EDC-HCI (290 mg, 1.51 mmol) at room temperature. The resulting mixture was stirred at room temperature for 18 hr. The mixture was pored into ice-water. The solid was collected, washed with water and air-dried. The crude solid was purified by silica gel column chromatography with CHCI 3 -EtOH (4:1, v/v) as eluent to give 15 methyl 4-[2-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetamido]-2-methyl-1 propoxy]benzoate (580mg, q.y.) as a crystalline material. IR (KBr) 3350, 3286, 1712, 1687, 1637, 1606cm-'; 1 H-NMR (CDCl 3 ) 8 1.39 (s, 6 H), 2.33 (s, 3 H), 3.41 (s, 2 H), 3.63 (s, 3 H), 3.88 (s, 3H11), 4.05 (s, 2 H), 5.44 (br s, 1 H), 6.33 (br s, 1 H), 6.79 (d, J= 8.3 Hz, 2 H), 7.12 (s, 1 H), 7.18 (t, J= 7.5 Hz, 1 H), 7.53 (d, J= 7.8 Hz, 1 H), 7.94 (d, J= 7.8 Hz, 2 H), 8.14 (d, J= 8.3 Hz, 1 H); MS 20 (FAB) m/z 520 (M' +1); Anal. Calcd for C 29
H
3 3
N
3 0 6 : C, 67.04; H, 6.40; N, 8.09. Found: C, 66.86; H, 6.36; N, 8.22. To a stirred solution of methyl 4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl acetamido]-2-methyl-2-propoxy]benzoate (510 mg, 0.98 mmol) was added 0.25N NaOH (8 ml, 2 mmol) at room temperature. The resulting mixture was stirred at an ambient temperature for 18 25 hr. The mixture was pored into ice-IN HCI (5 ml). the solid was collected, washed with water and air-dried. The crude solid was recrystallized from CHCl 3 -EtOH-Et20 to give 300 (480 mg, 97%) as fine needles. MW 505.56 IR (KBr) n 3346, 3294, 1687, 1637, 1604 cm'; 'H-NMR (DMSO-d 6 ) 8 1.35 (s, 6 H), 2.24 (s, 3 H), 3.33 (s, 2 H), 3.80 (s, 3 H), 4.15 (s, 2 H), 6.75 (d, J = 8.3 Hz, 1 H), 6.88 (s, 1 H), 6.95-6.99 (min, 3 H1), 7.11-7.17 (min, 3 H), 7.80 (d,J= 8.3 Hz, 1 H), 7.82 30 (s, 1 H), 7.87 (d, J = 8.8 Hz, 2 H), 7.98 (d, J= 7.8 Hz, 1 H), 8.45 (s, 1 H), 8.54 (s, 1 H), 12.62 (br s, 1 H); MS (FAB) m/z 506 (M +1); Anal. Calcd for C 28
H
3 1
N
3 0 6 : C, 66.52; H, 6.18; N, 8.31. Found: C, 66.22; H, 6.28; N, 8.11. 438 WO 01/00206 PCT/US00/18079 Example 250 3-amino-4-[2-N-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetamido]-2-methyl-1 propoxy]benzoic acid H C C O O H I HH 6Me 301 5 To a stirred solution of 4-amino-2-nitrophenol (10 g, 64.88 mmol) in AcOH (70 ml) and DMSO (20 ml) was added c. H 2
SO
4 at 0-5 oC. To a stirred this solution was added dropwise a solution of NaNO 2 (5.4 g, 77.9 mmol) in water (5 ml) below 20 oC for over 10 min. The resulting mixture was further stirred for 0.5 hr at 5 oC. This mixture was poured into a stirred solution of KI (30 g, 0.182 mol) and catalytic amount of Cu powder (200 mg) in ice-water (200 ml) for over 10 10 min. The resulting mixture was for a further 1 hr at an ambient temperature. The mixture was extracted with CH 2
CI
2 . The extracts were washed successively with sat. Na 2
S
2 0 3 and brine. The organic layer was dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica gel (50ml) with CHClI-EtOAc (3:1, v/v) as eluent to give 4-iodo-2-nitrophenol (2.5 g, 15%) as a yellow crystalline material. 'H-NMR (CDCl 3 ) 8 6.94 (d, J= 8.8 Hz, 1 H), 7.82 (dd, J= 2 and 15 8.8Hz, 1 H), 8.42 (d, J= 2 Hz, 1H), 10.49 (s, 1 H). To a stirred solution of 4-iodo-2-nitrophenol (2 g, 7.75 mmol), hydroxypivalic acid methyl ester (1.05 g, 7.92 mmol) and PPh 3 (2.3 g, 8.68 mmol) in THF (10 ml) was added dropwise a solution of DIAD (1.77 g, 8.30 mmol) in THF (2 ml) under ice-water bath cooling. The resulting mixture was then heated under reflux for 18hr. After cooling, the mixture was 20 evaporated off. The residue was chromatographed on silica gel (100 ml) with with toluene-EtOAc (4:1, v/v) as eluent to give methyl 3-(4-iodo-2-nitro)phenoxy-2,2-dimethylpropionate (2.9 g, q.y.) as a crystalline material. 'H-NMR (CDCl 3 ) 8 1.34 (s, 6 H), 3.71 (s, 3 H), 4.08 (s, 2 H), 6.86 (d, J = 8.8 Hz, 1 H), 7.78 (dd, J= 2 and 8.8Hz, 1 H), 8.12 (d, J= 2 Hz, 1H). A mixture of methyl 3-(4-iodo-2-nitro)phenoxy-2,2-dimethylpropionate (2.8 g, 7.38 25 mmol) in THF (15 ml) and 0.25 N NaOH (60 ml, 15 mol) at an ambient temperature for 18 hr. The mixture was poured into ice-iN HCI (20 ml). The solid was collected, washed with water and air-dried. The crude solid was recrystallized from CHC13-EtOH-IPE to afford 3-(4-iodo-2 nitro)phenoxy-2,2-dimethylpropionic acid (2.0 g, 74%) as a crystalline material. Mp 165-182oC; IR (KBr) n 1716, 1525, 1344 cm~'; 'H-NMR (CDCl 3 ) 8 1.38 (s, 6 H), 4.10 (s, 2 H), 6.86 (d, J= 8.8 30 Hz, 1 H), 7.79 (dd, J= 2.2 and 8.8Hz, 1 H), 8.12 (d, J= 2 Hz, 1H); MS (FAB) m/z 366 (M +1); Anal. Calcd for C 29
H
33
N
3 0 6 : C, 36.18; H,3.18; N, 3.84. Found: C, 36.85; H, 3.35; N, 3.79. 439 WO 01/00206 PCT/US00/18079 To a stirred mixture of 3-(4-iodo-2-nitro)phenoxy-2,2-dimethylpropionic acid (1.93 g, 5.29 mmol) and triethylamine (590 mg, 5.81 mmol) in tert-BuOH (15 ml) and toluene (15 ml) was added a solution of diphenyl phosphoryl azide (1.53 g, 5.55 mmol) in toluene (3 ml) at room temperature. The resulting mixture was then heated at reflux for 20 hr. After cooling, ice and IN 5 HCI (5ml) was added to the mixture and extracted with toluene. The extracts were washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica gel (50ml) with toluene-EtOAc (10:1, v/v) as eluent to give 3-nitro-4-(2-tert-butoxycarbonylamino-2-methyl 1-propoxy)iodobenzene (1.91 g, 83%) as a gum. 'H-NMR (CDCl 3 ) 8 1.38 (s, 9 H), 1.39 (s, 6 H), 4.19 (s, 2 H), 4.67 (br s, 1 H), 6.88 (d, J= 8.8 Hz, 1 H), 7.77 (dd, J= 2.0 and 8.8Hz, 1 H), 8.12 (d, 10 J= 2.0 Hz, 1H). A mixture of 3-nitro-4-(2-tert-butoxycarbonylamino-2-methyl-1-propoxy)iodobenzene (1.9 g, 4.36 mmol), Pd(OAc) 2 and 1,3-bis(diphenylphosphino)propane (dppp) (90 mg, 0.22 mmol) in triethylamine-MeOH-DMSO (1:2:5, v/v, 48 ml) was stirred under a current of CO (gas) at 70 oC for 6 hr. After cooling, the mixture was poured into water and extracted with EtOAc. The 15 extracts were washed with brine, dried over Na 2
SO
4 , and evaporated. The residue was chromatographed on silica gel (50ml) with toluene-EtOAc (6:1, v/v) as eluent to give methyl 4-(2 tert-butoxycarbonyl amino-2-methyl-1-propoxy)-3-nitrobenzoate (820 mg, 51%) as a gum. 'H NMR (CDCl 3 ) 8 1.38 (s, 9 H), 1.42 (s, 6 H), 3.93 (s, 3 H), 4.29 (s, 2 H), 4.67 (br s, 1 H), 7.15 (d, J = 8.8 Hz, 1 H), 8.18 (dd, J= 1.7 and 8.8Hz, 1 H), 8.52 (d, J= 1.7 Hz, 1H). 20 A stirred mixture of methyl 4-(2-tert-butoxycarbonylamino-2-methyl-1l-propoxy)-3 nitrobenzoate (350 mg, 0.95 mmol) and 5% Pd-C (70 mg) in EtOH (30 ml) was hydrogenated in an atmospheric hydrogen pressure at room temperature for 20 hr. Insoluble Pd-catalyst was removed with suction and washed with EtOH. The filtrate was evaporated off to afford methyl 4 (2-tert-butoxycarbonyl amino-2-methyl)-1-propoxy-3-aminobenzoate as a gum, which was used to 25 the subsequent reaction without further purification. 'H-NMR (CDCl 3 ) 8 1.41 (s, 9 H), 1.43 (s, 6 H), 3.86 (s, 3 H), 4.07 (s, 2 H), 4.67 (br s, 1 H), 6.80 (d, J= 8.5 Hz, 1 H), 7.39 (d, J = 2.2 Hz, 1H), 7.44 (dd, J = 2.2 and 8.5Hz, 1 H). To a stirred mixture of the above methyl 4-(2-tert-butoxycarbonylamino-2-methyl)-1 propoxy-3-aminobenzoate and triethylamine (0.20 ml, 1.43 mmol) in CH 2 C1 2 (10 ml) was added a 30 solution of trifluoroacetic anhydride (0.182 ml, 1.28 mmol) in CH 2 C1 2 (3 ml) at 0-5 oC. The resulting mixture was stirred at room temperature for 1 hr. Ice-sat. NaHCO 3 was added to the mixture, and extracted with CH 2 C1 2 . The extracts were washed with brine, dried over Na 2
SO
4 , and 440 WO 01/00206 PCT/US00/18079 evaporated. The residue was dissolved in CH 2 Cl 2 (5 ml) and added anisole (0.105 ml, 0.95 mmol) and TFA (2 ml). The resulting mixture was stirred at room temperature for 18 hr. The mixture was evaporated in vacuo, and the residue was diluted with CH 2 C1 2 and made basic by the addition of sat. NaHCO 3 . The separated CH 2 Cl 2 layer was dried over Na 2
SO
4 , and evaporated. The residue 5 was chromatographed on silica gel (50ml) with CHCl 3 -EtOH (99:1, v/v) as eluent to give methyl 4 (2-amino-2-methyl-1-propoxy)-3-trfluoroacetamidobenzoate (631 mg, 63% in 3-steps) as a gum. 'H-NMR (CDCl 3 ) 8 1.29 (s, 9 H, tert-Bu), 3.86 (s, 2 H, CH 2 ), 3.91 (s, 3 H, ), 6.99 (d, J= 8.5 Hz, 1 H), 7.91 (dd, J= 2.0 and 8.5 Hz, 1H), 8.83 (d, J= 8.5Hz, 1 H). To a stirred mixture of methyl 4-(2-amino-2-methyl)-1-propoxy-3-trfluoroacetamido 10 benzoate (200 rmg, 0.598 mmol), 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (210 mg, 0.598 mmol), 4-DMAP (90 mg, 0.72 mmol) in DMF (7 ml) was added EDC-HCI (160 mg, 0.81 mmol) at room temperature. The resulting mixture was stirred at room temperature for 20 hr. The mixture was pored into ice-water. The solid was collected, washed with water and air-dried. The crude solid was purified by silica gel column chromatography with CHCl 3 - EtOH (98:2, v/v) 15 as eluent to give methyl 4-[2-N-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetamido]-2 methyl-l-propoxy]-3-trfluoroacetamidobenzoate (580mg, q.y.) as a crystalline material. 'H-NMR (CDCl 3 ) 5 1.42 (s, 6 H), 3.42 (s, 2 H), 3.69 (s, 3 H), 3.89 (s, 3 H), 4.23 (s, 2 H), 5.33 (br s, 1 H), 6.66 (s, 1 H), 6.71 (min, 1 H), 6.92 (d, J= 8.5 Hz, 1 H), 7.02 (min, 1 H), 7.09 (min, 2 H), 7.29 (mn, 1 H), 7.37 (d,J= 8.0 Hz, 1H), 7.87 (dd, J= 2 and 8.5Hz, 1 H), 7.97 (d,J= 8.0 Hz, 1 H), 8.19 (d,J= 20 8.2 Hz, 1 H), 8.59 (br s, 1 H), 8.74 (d, J = 2.0 Hz, 1 H). To a stirred solution of methyl 4-[2-N-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl acetamido]-2-methyl-l-propoxy]-3-trfluoroacetamidobenzoate (320 mg, 0.492 mmol) in THF (2 ml) was added 0.25N NaOH (6 ml, 1.5 mmol) at an ambient temperature. And the resulting mixture was stirred for 20 hr. The mixture was poured into ice-lN HCI (2 ml). The solid was 25 collected, washed with water and air-dried. The crude solid was recrystallized from CHCl 3 - EtOH Et 2 0 to give 301 (240mg, 89%) as fine needles. MW 541.00 IR (KBr) n 3338, 3296, 1691, 1641 cm'; IH-NMR (CDCl 3 ) 8 'H-NMR (DMSO d 6 ) 5 1.37 (s, 6 H), 3.35 (s, 2 H), 3.77 (s, 3 H), 4.05 (s, 2 H), 4.96 (br s, 1 H), 6.75 (br d, J= 8.3 Hz, 1 H), 6.78 (d, J= 8.3 Hz, 1 H), 6.87 (d, J= 1.7 Hz, 1 H), 7.01 (m, 1H), 7.15 (dd, J= 2 and 8.5 Hz, 1H), 7.24 (d,J= 2 Hz, 1 H), 7.27 (dt, J = 2.0 and 30 8.5 Hz, 1 H), 7.43 (dd, J= 2 and 8.0 Hz, 1 H), 7.78 (br s, 1 H), 7.90 (d, J= 8.0 Hz, 1 H), 8.08 (dd, J= 2 and 8.3 Hz, 1 H), 8.85 (s, 1 H), 8.89 (s, 1 H), 12.23 (br s, 1 H); MS (FAB) m/z 541 (M +1); Anal. Calcd for C 27
H,,
29 C1N 4 0, 6 : C, 58.00; H, 5.59; N, 10.02. Found: C, 57.97; H, 5.39; N, 10.01. 441 WO 01/00206 PCT/US00/18079 Example 251 2-acetylamino-4-[2-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl] -N-methylacetamido] ethylaminobenzoateic acid ,COOH N N NHAc j Q H NN% Me H H Me 302 5 To a stirred solution of 2-acetylamino-4-nitrobenzoic acid (1.28 g, 5.71 mmol) in benzene-MeOH (4:1, v/v, 25 mL), was added trimethylsilyldiazomethane (2.0 M solution in n hexane, 4.28 ml, 8.56 mmol) at 0 oC. The stirring was continued for 18 hours at rt. The reaction was poured into hexane, and the resulting precipitate was collected by filtration to give methyl 2 acetylamino-4-nitrobenzoate (1.32 g, 97%) as a white solid; mp no data; 'H-NMR (400 MHz, 10 CDCl 3 ) 6 2.30 (s, 3 H), 4.00 (s, 3 H), 7.88 (min, 1 H), 8.18 (dd, J= 2.0 Hz, 8.8 Hz, 1 H), 9.60 (t, J= 2.2 Hz, 1 H), 11.10 (s, 1 H); MS (ESI) m/z 238 (M). To a solution of methyl 2-acetylamino-4-nitrobenzoate (1.31 g, 5.50 mmol) in MeOH (30 mL) was added 5% Pd on carbon (195 mg), and the stirring under H 2 gas (3 atm) was continued for 18 hours at rt. The catalyst was filtered off and the mixture was evaporated. The resulting 15 crude solid was recrystallized with CHCl 3 -MeOH-hexane to give methyl 2-acetylamino-4 aminobenzoate (1.03 g, 90%) as a white solid. 'H-NMR (400 MHz, CDCl 3 ) 6 2.23 (s, 3 H), 3.82 (s, 3 H), 4.20 (s, 2 H), 6.30 (dd, J = 2.5 Hz, 8.8 Hz, 1 H), 7.80 (d, J = 8.8 Hz, 1 H), 8.06 (s, 1 H), 11.26 (s, 1 H); MS (FAB), m/z 208 (M). To a cooled solution of methyl 2-acetylamino-4-aminobenzoate (300 mg, 1.44 mmol) and 20 N-tert-butoxycarbonyl-N-methylglycinal (499 mg, 2.88 mmol) in 1,2-dichloroethane (30 ml), was added NaBH(OAc) 3 (964 mg, 4.32 mmol) and the stirring was continued for 64 h at 0 oC. The mixture was poured into sat. NaHCO 3 and was extracted with CHC13 (50 ml x 3), washed with brine, and dried over MgSO 4 . After removal of the solvent in vacuo, the residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC 25 5404-FC, linear gradient of hexane-EtOAc from 9:1 to 2:1) to give methyl 2-acetylamino-4-[2-(N tert-butoxycarbonyl-N-methylamino)ethylamino]benzoate (451 mg, 86%) as a colorless oil. 1
H
NMR (CDCl 3 , 400 MHz) 8 1.48 (s, 9 H), 2.22 (s, 3 H), 2.90 (s, 3 H), 3.32 (min, 2 H), 3.50 (min, 2 H), 3.80 (s, 3 H), 6.20 (dd, J= 2.2 Hz, 8.8 Hz, 1 H), 7.80 (min, 1 H), 7.95 (min, 1 H), 11.30 (brs, 1 H); MS (FAB) m/z 366 (M++I). 30 To a stirred solution of methyl methyl 2-acetylamino-4-[2-(N-tert-butoxycarbonyl-N 442 WO 01/00206 PCT/US00/18079 methylamino)ethylamino]benzoate (450 mg, 1.23 mmol) in dichloromethane (5 mL), was added TFA (5 mL) and the stirring was continued for 18 h at rt. After removal of the solvent in vacuo, the residue was dissolved in CHC 3 (200 mL), washed with brine, sat. NaHCO 3 , and dried over MgSO 4 . The solvent was removed to give methyl 2-acetylamino-4-[2-(N-methylamino) 5 ethylamino] benzoate (298 mg, 88%) as a colorless oil. 'H-NMR (CDCl 3 , 400 MHz) 8 2.21 (s, 3 H), 2.46 (s, 3 H), 2.88 (min, 2 H), 3.31 (mn, 2 H), 3.83 (s, 3 H), 4.85 (br, 1 H), 6.24 (dd, J = 2.5 Hz, 8.8 Hz, 1 H), 7.80 (d, J = 8.8 Hz, 1 H), 7.99 (d, J = 2.5 Hz, 1 H); MS (FAB), m/z 266 (M+1). A mixture of methyl 2-acetylamino-4-[2-(N-methylamino)ethylamino]benzoate (145 mg, 0.55 mmol), 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (172 mg, 0.55 mmol), 10 EDC-HCl (158 mg, 0.83 mmol), HOBt (141 mg, 1.05 mmnol), and DMAP (13 mg, 0.11 mmol) in DMF (10 ml) was stirred for 18 hours. The mixture was diluted with EtOAc (300 ml), washed with brine, and dried over MgSO 4 . After removal of the solvent, residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC-5404-FC, linear gradient of CHCI 3 -MeOH from 100:0 to 70:30) to give methyl 2-acetylamnino-4-[2-N-[3-methoxy-4-[N'-(2 15 methylphenyl)ureido]phenyl]-N-methylacetamido]ethylaminobenzoate (309 mg, 100%) as an amorphous foam. 'H-NMR (CDCl 3 )5 2.22 (s, 3 H), 2.30 (s, 3 H), 3.02 (s, 3 H), 3.35 (mn, 2 H), 3.58 (s, 3 H), 3.50-3.74 (min, 4 H), 3.85 (s, 3 H), 6.20 (min, 1 H), 6.58 (s, 1 H), 6.65-6.75 (min, 3 H), 7.13 (min, 2 H), 7.40-7.50 (min, 2 H), 7.75 (min, 2 H), 7.90 (min, 1 H), 8.00 (min, 1 H), 11.32 (s, 1 H); MS (FAB) m/z 562 (M +1). 20 To a solution of methyl 2-acetylamino-4-[2-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenyl]-N-methylacetamido]ethylaminobenzoate (309 mg, 0.55 mmol) in THF-MeOH (1: 1, v/ v, 9 ml), was added 0.25 N NaOH (4.4 ml, 1.1 mmol) at rt, and heated to reflux. The stirring was continued for 18 hours at reflux. The reaction mixture was poured into water, and acidified to pH 5.0 with 1.0 N HC1. The resulting precipitate was recrystallized with hexane-diethylether to give 25 302 (175 mg, 58%) as a pale red powder. MW 547.60 'H-NMR (CD 3 OD) 8 2.16 (d, J = 4.8 Hz, 3 H), 2.28 (d, J = 4.2 Hz, 3 H), 2.98 and 3.10 (2 s, total 3 H), 3.35 (min, 2 H), 3.68 (min, 4 H), 3.81 and 3.84 (2 s, total 3 H), 6.30 (min, 1 H), 6.60-6.82 (mn, 2 H), 7.00 (mn, 1 H), 7.15 (mn, 2 H), 7.53 (mn, 1 H), 7.77-7.96 (min, 3 H); MS (FAB) m/z 548 (M+1); Anal. Calcd for C 29
H
33 NsO 5 6 0.5 H 2 0: C, 62.58; H, 6.16; N, 12.58. Found: C, 62.55; H, 6.31; N, 12.15. 30 Example 252 2-acetylamino-4-[2-N-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl]-N-methylacetamido] ethylaminobenzoic acid 443 WO 01/00206 PCT/US00/18079 COOH " N NHAc Br H H Me 303 A mixture of methyl 2-acetylamino-4-(2-N-methylamino-1-ethylamino)benzoate (145 mg, 0.55 mmol), 3-methoxy-4-[N'-(2-bromophenyl)ureido]phenylacetic acid (209 mg, 0.55 mmol), EDC-HCl (158 mg, 0.83 mmol), HOBt (141 mg, 1.05 mmol), and DMAP (13 mg, 0.11 mmol) in 5 DMF (10 ml) was stirred for 18 hours. The mixture was diluted with EtOAc (300 ml), washed with brine, and dried over MgSO 4 . After removal of the solvent, residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC-5404-FC, linear gradient of CHC1 3 -MeOH from 100:0 to 70:30) to give methyl 2-acetylamino-4-[2-N-[4-[N'-(2-bromo phenyl)ureido]-3-methoxyphenyl]-N-methylacetamido]ethylaminobenzoate (294 mg, 85%) as an 10 amorphous foam. 'H-NMR (CDCl 3 ) 8 2.21 (s, 3 H), 3.05 (s, 3 H), 3.40 (m, 2 H), 3.65-3.70 (m, 4 H), 3.78 (s, 3 H), 3.86 (s, 3 H), 6.21 (m, 1 H), 6.79 (m, 2 H), 6.93 (m, 1 H), 7.10 (d, J = 10.3 Hz, 1 H), 7.30 (m, 1 h), 7.42 (m, 1 H), 7.61 (m, 1 H), 7.78-7.85 (m, 2 H), 7.93 (m, 2 H), 8.13 (m, 1 H); MS (FAB), m/z 627 (MW). To a solution of methyl 2-acetylamino-4-[3-methoxy-4-[N'-(2-bromophenyl)ureido] 15 phenylacetamido]-2-N-methylamino-1-ethylaminobenzoate (294 mg, 0.47 mmol) in THF-MeOH (1: 1, v/ v, 8 ml), was added 0.25 N NaOH (3.8 ml, 0.94 mmol) at rt, and heated to reflux. The stirring was continued for 18 hours at reflux. The reaction mixture was poured into water, and acidified to pH 5.0 with 1.0 N HC1. The resulting precipitate was recrystallized with hexane diethylether to give 303 as a white powder (210 mg, 73%). MW 612.47 mp 155-160 0 C; 'H 20 NMR (CD 3 OD) 8 22.18 (d, J= 5.5 Hz, 3 H), 3.00 and 3.12 (2 s, total 3 H), 3.39 (m, 1 H), 3.60 (m, 4 H), 3.70 (s, 1 H), 3.85 and 3.86 (2 s, total 3 H), 6.31 (m, 1 H), 6.68 (m, 1 H), 6.78 (m, 1 H), 6.78 and 6.85 (2 m, total 1 H), 6.97 (m, 1 H), 7.30 (m, 1 H), 7.56 (m, 1 H), 7.80-7.95 (m, 4 H); MS (ESI) m/z 613 (MW); Anal. Calcd for C 2 8
H
31 BrN 5 0 6 0.75 H 2 0: C, 53.64; H, 5.22; N, 11.17. Found: C, 53.89; H, 5.23; N, 10.69. 25 Example 253 4-[2-N-[[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl]-N-phenylacetamido]ethoxy]benzoic acid COOH N N4) " "O-- 0 I H H Me 304 To a solution of methyl 4-[2-(methanesulfonyloxy)ethoxy]benzoate (2.74 g, 10 mmol) in MeCN (50 ml), was added aniline (9.1 ml, 100 mmol) at rt. The reaction was stirred for 64 hours 444 WO 01/00206 PCT/US00/18079 at reflux. The mixture was poured into 1H120 (200 mL), extracted with EtOAc (100 mL x 2), dried over MgSO 4 . After removal of the solvent in vacuo, the unreacted aniline was removed in vacuo by co-evaporation with toluene (10 mL x 3) at 80 'C. The residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC-5404-FC, f50 mm x 150 mm, 5 CHC1 3 ) to give methyl 4-[2-(N-phenylamino)ethoxy]benzoate (2.23 g, 82%) as a colorless oil. 'H NMR (CDC 3 ) 8 3.56 (t,J = 5.1 Hz, 2 H), 3.90 (s, 3 H), 4.21 (t,J= 5.1 Hz, 2 H), 6.68 (dd, J= 1.0 Hz, 8.6 Hz, 2 H), 6.75 (t, J= 7.3 Hz, 1 H), 7.20 (AB type d, J= 7.3 Hz, 2 H), 8.00 (d, J= 9.1 Hz, 2 H); MS (ESI) m/z 272 (M*+1). A mixture of methyl 4-[2-(N-phenylamino)ethoxy]benzoate (136 mg, 0.5 mmol), 3 10 methoxy-4-[N'-(2-chlorophenyl)ureido]phenylacetic acid (167 mg, 0.5 mmol) and PyBOP (781 mg, 0.75 nummol), i-PrNEt 2 (261 ml, 0.96 mmol) in DMF (10 ml) was stirred for 18 hours. The mixture was diluted with EtOAc (100 mL) , washed with 1 N HC1, brine and dried over MgSO 4 . The residue was co-evaporated with toluene (10 ml x 3) to remove DMF. The residue was chromatographed on TLC (MERCK, silicagel 60, 2 mm, 2 plates, CHCl 3 -MeOH, 20:1) to give 15 methyl 4-[2-N-[[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl]-N-phenylacetamidoJ]ethoxy] benzoate (129 mg, 44%) as a white amorphous foam. 'H-NMR (CDCl 3 ) 8 3.42 (s, 1 H), 3.69 (d, J = 8.3 Hz, 1 H), 3.74 (s, 3 H), 3.87 (s, 3 H), 4.10 (m, 2 H), 4.23 (mn, 2 H), 6.48-7.44 (min, 13 H), 7.93 (d, J= 9.3 Hz, 2 H), 8.18 (dd, J= 1.5 Hz, 8.3 Hz, 1 H); MS (ESI) m/z 588 (M). To a solution of methyl 4-[2-N-[[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl]-N 20 phenyl acetamido]ethoxy]benzoate (124 mg, 0.19 mmol) in THF-MeOH (6: 1, v/ v, 3 ml), was added 0.5 N NaOH (2 ml, 1 mmol) at rt, and heated to reflux in a sealed bottle. The stirring was continued for 15 hours at reflux. The reaction mixture was poured into water, acidified with 1.0 N HC1, extracted with CHCl 3 -MeOH (2:1, 20 mL x 3), and dried over MgSO 4 . After removal of the solvent, the residue was crystallized with CHCl 3 -hexane-diethylether to give 304 (77 mg, 64%) 25 as a white powder. MW 574.02 'H-NMR (CD 3 OD) 8 3.45 (s, 2 H), 3.79 (s, 3 H), 4.12 (mn, 2 H), 4.22 (mn, 2 H), 6.48 (dd, J= 2.0 Hz, 8.3 Hz, 1 H), 6.61 (d, J= 2.0 Hz, 1 H), 6.87 (d, J= 8.8 Hz, 2 H), 7.00 (min, 1 H), 7.22 (min, 3 H), 7.36 (min, 1 H), 7.43 (min, 3 H), 7.90 (d, J= 8.3 Hz, 1 H), 7.95 (d, J = 8.8 Hz, 2 H), 8.02 (dd, J= 1.5 Hz, 8.3 Hz, 1 H); MS (ESI) m/z 574 (MW); Anal. Calcd for
C
31
H
28 C1N 3 0 6 -0.5 H20: C, 63.86; H, 5.01; N, 7.21. Found: C, 63.67; H, 4.91; N, 6.99. 30 Example 254 (S)-4-[2-N-[[3-methoxy-4-[N '-(2-methylphenyl)ureido]phenyl]-N-(2-aminobenzyl)acetamido]- 1 propoxy]benzoic acid 445 WO 01/00206 PCT/US00/18079
H
2 N -, COOH NN MeH H Me 305 To a cooled (0OC) solution of benzyl (S)-4-(2-amino-1-propoxy)benzoate (1.50 g, 5.26 mmol) and 2-nitrobenzaldehyde (0.87 g, 5.76 mmol) in MeOH-AcOH (16 ml, 15:1, v/v) was added NaBH 3 CN (1.65 g, 26.3 mmol) and the reaction mixture was stirred at room temperature 5 overnight. The mixture was quenched by sat. NaHCO 3 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica gel with CHC13 to 5% MeOH in CHC1 3 as eluent to give benzyl (S)-4-[2 (2-nitrobenzylamino) -1-propoxy]benzoate (931 mg, 42%) as a yellow oil. 'H-NMR (CDCl 3 ) 8 1.21 (d,J= 6.4 Hz, 3 H), 3.13-3.18 (min, 1 H), 3.88-3.97 (min, 2 H), 4.06-4.20 (min, 2 H), 5.34 (s, 2 10 H), 6.89-6.94 (min, 2 H), 7.29-7.65 (min, 8 H), 7.94-8.03 (min, 3 H); MS (FAB) m/z 421 (M'+1). A mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetate (460 mg, 0.96 mmol), benzyl (S)-4-[2-(2-nitrobenzylamino)-1-propoxy]benzoate (403 mg, 0.96 mmol) and Et 3 N (200 ml, 1.43 mmol) in DMF (8 ml) was stirred at room temperature overnight. The mixture was diluted with EtOAc, washed with 0.5 N HC1, brine, dried over Na 2
SO
4 , and 15 evaporated. The residue was purified by column chromatography on silica-gel with 1% MeOH in CHCl 3 as eluent to give benzyl (S)-4-[2-N-[[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenyl]-N (2-nitrobenzyl) acetamido]-1-propoxy]benzoate (504 mg, 73%) as a brown amorphous solid. MS (FAB), m/z 717 (M'+1). A stirred solution of benzyl (S)-4-[2-N-[[3-methoxy-4-[N'-(2-methylphenyl)ureido] 20 phenyl]-N-(2-nitrobenzyl)acetamido]-1-propoxylbenzoate (504 mg, 0.70 mmol) in MeOH-THF (11 ml, 10:1, v/v) was hydrogenated over 5% Pd-C (100 mg, 20 wt%) at 3 atm overnight. The mixture was filtered to remove the catalyst and the filtrate was evaporated. The residue was purified by preparative thin layer chromatography with 5% MeOH in CHC1 3 as eluent to give 305 (115 mg, 27%) as a white powder. MW 596.67 MS (FAB), m/z 597 (M +l). 25 Example 255 (S)-4-[2-N-[[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl]-N-(2-nitrobenzyl) acetamido]- 1 propoxy]benzoic acid 446 WO 01/00206 PCT/US00/18079 0 2
N
COOH NN r H H Me 306 To a cooled (0) solution of benzyl (S)-4-(2-amino-1-propoxy)benzoate (1.50 g, 5.26 mmol) and 2-nitrobenzaldehyde (0.87 g, 5.76 mmol) in MeOH-AcOH (16 ml, 15:1, v/v) was added NaBH 3 CN (1.65 g, 26.3 mmol) and the reaction mixture was stirred at room temperature 5 overnight. The mixture was quenched by sat. NaHCO 3 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica gel with CHC1 3 to 5% MeOH in CHCl 3 as eluent to give benzyl (S)-4-[2 (2-nitrobenzylamino) -1-propoxy]benzoate (931 mg, 42%) as a yellow oil. H-NMR (CDCl 3 ) 8 1.21 (d,J= 6.4 Hz, 3 H), 3.13-3.18 (min, 1 H), 3.88-3.97 (m, 2 H), 4.06-4.20 (min, 2 H), 5.34 (s, 2 10 H), 6.89-6.94 (min, 2 H), 7.29-7.65 (min, 8 H), 7.94-8.03 (min, 3 H); FAB-MAS, m/z 421 (M'+1). A mixture of 4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (476 mg, 1.26 mmol), benzyl (S)-4-[2-(2-nitrobenzylamino)-1-propoxy]benzoate (528 mg, 1.26 mmol), EDCHCI (361 mg, 1.88 mmol), HOBt (255 mg, 1.89 mmol) and DMAP (30 mg, 0.25 mmol) in DMF (10 ml) was stirred at room temperature overnight. And the reaction could not be completed, so the 15 reaction mixture was stirred at 60oC for 1 day. The mixture was diluted with EtOAc, washed with 0.5 N HC1, brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHCl 3 to 2% MeOH in CHCl 3 as eluent to give the title compound as a crude oil. To a stirred solution of the crude product in THF-MeOH (10 ml, 1:1, v/v) was added 0.5 N NaOH (10 ml) and the reaction mixture was heated under reflux for 3 hr. 20 The mixture was poured into ice-H 2 0 and the basic aqueous layer was acidified (pH 4.3) with 1 N HC1. The resulting precipitate was collected and the crude solid was purified by preparative thin layer chromatography with 5% MeOH in CHC1 3 l as eluent to give 306 (162 mg, 2 steps, 19%) as a white amorphous solid. MW 691.53 MS (FAB), m/z 692 (M +1); Anal. Calcd for CzzH 3 zBrN 4 Os7/4H 2 0: C, 54.82; H, 4.81; N, 7.75. Found: C, 54.80; H, 4.61; N, 7.24. 25 Example 256 4-[2-N-cyclopropyl-N-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl]acetamido] ethoxybenzoic acid Y 0j COOH H H Me 307 A mixture of methyl 4-(2-cyclopropylaminoethoxy)benzoate (290 mg, 1.23 mmol), 4-[N'-(2 447 WO 01/00206 PCT/US00/18079 chlorophenyl)ureido]-3-methoxyphenylacetic acid (412 mg, 1.23 mmol), EDC.HCI (354 mg, 1.85 mmol), HOBt (cat.), and DMAP (cat.) in DMF (10 ml) was stirred overnight. The mixture was partitioned between EtOAc (300 ml) and H 2 0 (100 ml). The organic phase was separated, washed with brine (2 x 100 ml), dried over MgSO 4 , and evaporated. The residue was chromatographed on 5 silica gel with CHCl 3 -MeOH (20:1) as eluent to give methyl 4-[2-N-cyclopropyl-N-[4-[N'-(2 chlorophenyl)ureido]-3-methoxyphenyl]acetamido]ethoxybenzoate (506 mg, 75%) as a yellow viscous oil. 'H-NMR (CDCl 3 ) 8 0.90-0.97 (m, 4 H), 2.75 (m, 1 H), 3.61 (s, 3 H), 3.79 (t, J= 5.4 Hz, 2 H), 3.87 (s, 3 H), 3.88 (s, 2 H), 4.16 (t, J = 5.4 Hz, 2 H), 6.76-6.80 (m, 4 H), 6.95 (dt, J= 7.8, 1.5 Hz, 1 H), 7.21-7.31 (m, 2 H), 7.53 (s, 1 H), 7.56 (s, 1 H), 7.93 (d, J= 8.3 Hz, 3 H), 8.19 10 (dd, J= 8.3, 1.5 Hz, 1 H). To a stirred solution of methyl 4-[2-N-cyclopropyl-N-[4-[N'-(2-chlorophenyl)ureido]-3 methoxyphenyl]acetamido]ethoxybenzoate (506 mg, 0.917 mmol) in THF (7 ml) was added 0.25 N NaOH (7.3 ml, 1.83 mmol). After stirring overnight, the mixture was poured into 1 N HCI (50 ml) and extracted with CHCl 3 -MeOH (4:1, 2 x 200 ml). The combined extracts were dried over 15 MgSO 4 and evaporated. The residue was chromatographed on silica gel with CHC13-MeOH (20:1 to 10:1) as eluent to give 307 (403 mg, 82%) as a colorless amorphous solid. MW 537.99 'H NMR (DMSO) 6 0.86-0.91 (m, 4 H), 2.75 (m, 1 H), 3.69 (t, J= 5.5 Hz, 2 H), 3.81 (s, 3 H), 3.84 (s, 2 H), 4.16 (t, J= 5.5 Hz, 2 H), 6.76 (d, J= 8.3 Hz, 1 H), 6.88 (s, 1 H), 6.97-7.04 (m, 3 H), 7.28 (t, J= 7.8 Hz, 1H), 7.44 (d,J= 7.8 Hz, 1 H), 7.88 (d, J = 8.8 Hz, 2 H), 7.96 (d, J = 8.1Hz, 1 H), 20 8.10 (d, J =8.3 Hz, 1 H), 8.89 (s, 1 H), 8.93 (s, 1 H), 12.65 (s, br s); MS (FAB), m/z 538 (M'+1); Anal. Calcd for C 28
H
2 8 C1N 3 0 6 : C, 62.51; H, 5.25; N, 7.81. Found: C, 61.85; H, 5.42; N, 7.41. Example 257 4-[2-N-cyclohexyl-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl] acetamido]ethoxybenzoic acid COOH N N 25 Me H H Me 308 To a solution of methyl 4-[(2-methanesulfonyloxy)-1-ethoxy]benzoate (2.74 g, 10 mmol) in MeCN (50 ml), was added cyclohexylamine (5.72 ml, 50 mmol) at rt. The reaction was stirred for 18 hours at reflux. The mixture was poured into H 2 0 (200 mL), extracted with EtOAc (100 mL x 2), dried over MgSO 4 . After removal of the solvent, residue was chromatographed on silica 30 gel (middle pressure chromatography system: YAMAZEN YFLC-5404-FC, f50 mm x 150 mm, linear gradient of CHCl 3 -EtOAc from 10:0 to 1:1) to give methyl 4-(2-N-cyclohexylamino)ethoxy 448 WO 01/00206 PCT/US00/18079 benzoate (2.43 g, 88%) as a colorless oil. 'H-NMR (CDCl 3 ) 8 1.10 (m, 2 H), 1.25 (m, 2 H), 1.60 (br, 2 H), 1.73 (m, 2 H), 1.90 (br, 2 H), 2.49 (m, 1 H), 3.02 (t,J = 5.2 Hz, 2 H), 3.88 (s, 3 H), 4.12 (t, J = 5.2 Hz, 2 H), 6.90 (d, J = 6.90 Hz, 2 H), 7.99 (d, J= 7.99 Hz, 2 H); MS (ESI) m/z 278 (M++1). 5 A mixture of methyl 4-(2-N-cyclohexylamino)ethoxybenzoate (139 mg, 0.5 mmol), 3 methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (157 mg, 0.5 mmol), EDC-HCI (144 mg, 0.75 mmol), HOBt (128 mg, 0.95 mmol), and DMAP (12 mg, 0.1 mmol) in DMF (2.5 ml) was stirred for 18 hours. The mixture was diluted with EtOAc (200 ml), washed with IN HCI and brine, and dried over MgSO 4 . After removal of the solvent, residue was chromatographed on silica 10 gel (middle pressure chromatography system: YAMAZEN YFLC-5404-FC, linear gradient CHCl 3 EtOAc 10:0 to 1:4) to give methyl 4-[2-N-cyclohexyl-N-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenyl] acetamido] ethoxybenzoate (247 mg, 86%) as an amorphous foam. 'H-NMR
(CDCI
3 ) 8 1.08-1.80 (m, 10 H), 2.30 (s, 3 H), 3.60-3.79 (m, 8 H), 3.88 (s, 3 H), 4.16 (m, 2 H), 6.30 (s, 1 H), 6.70-6.83 (m, 2 H), 6.88 (d, 2 H, J= 9.0 Hz), 7.12 (m, 2 H), 7.23 (m, 1 H), 7.60 (d, 1 H, J 15 = 8.3 Hz), 7.92 (d, 2 H, J= 9.0 Hz), 8.10 (d, 1 H, J = 8.0 Hz); MS (ESI) m/z 574 (M'+1). To a solution of methyl 4-[2-N-cyclohexyl-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenyl] acetamido]ethoxybenzoate (247 mg, 0.43 mmol) in THF-MeOH (6: 1, v/ v, 7 ml), was added 0.5 N NaOH (3.4 ml, 0.84 mmol) at rt, and heated to reflux in a sealed bottle. The stirring was continued for 18 hours at reflux. The reaction mixture was poured into water, acidified with 20 1.0 N HCI, extracted with CHC1 3 -MeOH (2:1, 20 mL x 3), and dried over MgSO 4 . After removal of the solvent, the residue was crystallized with CHC13-hexane-diethylether to give 308 (196 mg, 81%) as a white powder. MW 559.62 'H-NMR (CD 3 OD) 8 0.90-1.82 (m, 10 H), 2.29 (s, 3 H), 3.62 (m, 2 H), 3.78 (s, 3 H), 3.80 (m, 3 H), 4.12 (m, 2 H), 6.82 (m, 2 H), 6.96 (m, 3 H), 7.16 (m, 2 H), 7.58 (d, J = 7.7 Hz, 1 H), 7.92 (m, 3 H); MS (ESI) m/z 560 (M'+1); Anal. Calcd for 25 C 32
H
37
N
3 0 6 -0.5 H20: C, 67.59; H, 6.74; N, 7.39. Found: C, 67.83; H, 6.80; N, 7.13. Example 258 4-[2-N-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl]-N-propargylacetamido] ethoxybenzoic acid
-
COOH i HH 6Me 309 30 To a solution of methyl 4-[(2-methanesulfonyloxy)-1-ethoxy]benzoate (2.74 g, 10 mmol) 449 WO 01/00206 PCT/US00/18079 in MeCN (50 ml), was added propargylamine (3.43 ml, 50 mmol) at rt. The reaction was stirred for 18 hours at reflux. The mixture was poured into H 2 0 (200 mL), extracted with EtOAc (100 mL x 2), dried over MgSO 4 . After removal of the solvent in vacuo, the residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC 5 5404-FC, f50 mm x 150 mm, linear gradient of CHCl 3 -EtOAc from 10:0 to 9:1) to give methyl 4 (2-N-propargylamino) ethoxybenzoate (2.33 g, 100%) as a colorless oil. 1 H-NMR (CDCl 3 ) 8 2.28 (d,J= 2.4 Hz, 1 H), 3.11 (t,J= 5.1 Hz, 2H), 3.52 (d,J= 2.4 Hz, 2 H), 3.88 (s, 3 H), 4.15 (t,J= 5.1 Hz, 2 H), 6.90 (d, J= 8.8 Hz, 2 H), 7.98 (d, J= 8.8 Hz, 2 H); MS (ESI) mnz 234 (M+I). A mixture of methyl 4-(2-N-propargylamino)ethoxybenzoate (117 mg, 0.5 mmol), 3 10 methoxy-4-[N'-(2-chlorophenyl)ureido]phenylacetic acid (167 mg, 0.5 mmol), EDC-HCI (144 mg, 0.75 mmol), HOBt (128 mg, 0.96 mmol), and DMAP (12 mg, 0.1 mmol) in DMF (10 ml) was stirred for 18 hours. The mixture was diluted with EtOAc (100 mL) , washed with 1 N HC1, brine and dried over MgSO 4 . The residue was co-evaporated with toluene (10 ml x 3) to remove DMF. The residue was chromatographed on TLC (MERCK, silicagel 60, 2 mm, 2 plates, CHCI 3 -MeOH, 15 20:1) to give methyl 4-[2-N-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl]-N-propargyl acetamido] ethoxybenzoate (244 mg, 89%) as a white amorphous foam. 'H-NMR (CDCI 3 ) 8 2.20 and 2.32 (2 m, total 1 H), 3.72 (s, 2 H), 3.83 (mn, 5 H), 3.88 (s, 3 H), 4.09-4.35 (min, 4 H), 6.77-6.86 (mn, 4 H), 6.99 (min, 1 H), 7.11 (min, 2 H), 7.24 (min, 1 H), 7.34 (d, J= 7.9 Hz, 1 H), 7.96 (mn, 3 H), 8.18 (dd, J= 1.5 Hz, 8.3 Hz, 1 H); MS (ESI) m/z 550 (M). 20 To a solution of methyl 4-[2-N-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl]-N propargyl acetamido]ethoxybenzoate (240 mg, 0.44 mmol) in THF-MeOH-H 2 0 (2:2:1, v/v, 10 ml), was added NaOH (500 mg, 12.5 mmol) at rt. The stirring was continued for 2 hours at rt. The reaction mixture was poured into water, acidified with 1.0 N HC1, extracted with CHCI 3 -MeOH (2:1, 20 mL x 3), and dried over MgSO 4 . The residue was chromatographed on TLC (Whatman, 25 1 mm, 3 plates, CHC13-MeOH, 92:8) to give 309 (202 mg, 86%) as a white solid. MW 535.98 1
H
NMR (CD 3 OD) 8 2.60 and 2.81 (2d, J = 2.5 Hz, total 1 H), 3.79-3.94 (min, 4 H), 3.85 (s, 3 H), 4.15 (min, 1 H), 4.24 (min, 1 H), 4.32 (min, 2 H), 6.80 (d, J= 8.3 Hz, 1 H), 6.85 (d, J= 4.3 Hz, 1 H), 6.94 (mn, 2 H), 7.02 (min, 1 H), 7.25 (min, 1 H), 7.38 (min, 1 H), 7.87-8.02 (min, 4 H); MS (ESI) m/z 536 (M+1); Anal. Calcd for C 28
H
26
CIN
3 0 6 -2.25 H20: C,58.33; H,5.33; N,7.29. Found: C,58.23; 30 H,4.77; N,6.91. Examples 259 and 260 4-[2-N-allyl-N-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl]acetamido]ethoxybenzoic acid 450 WO 01/00206 PCT/US00/18079 [ ]N o -COOH N N4)" O I H H Me 310 A mixture of methyl 4-(2-N-allylamino)ethoxybenzoate (118 mg, 0.5 mmol), 3-methoxy 4-[N'-(2-chlorophenyl)ureido]phenylacetic acid (167 mg, 0.5 mmol), EDC-HCI (144 g, 0.75 mmol), HOBt (128 mg, 0.95 mmol), and DMAP (12 mg, 0.1 mmol) in DMF (2.5 ml) was stirred 5 for 18 hours. The mixture was diluted with EtOAc (300 ml), washed with IN HCI and brine, and dried over MgSO 4 . After removal of the solvent, residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC-5404-FC, linear gradient CHCl 3 EtOAc 100:0 to 85: 15) to give methyl 4-[2-N-allyl-N-[4-[N'-(2-chlorophenyl)ureido]-3 methoxyphenyl]acetamido] ethoxybenzoate (253 mg, 92%) as an amorphous foam. 'H-NMR 10 (CDCl 3 ) 8 3.65-3.85 (m, 4 H), 3.73 (s, 3 H), 3.88 (s, 3 H), 5.08 (m, 2 H), 4.22 (m, 2 H), 5.10-5.24 (m, 2 H), 5.76 (m, 1 H), 6.77 (m, 2 H), 6.85 (m, 2 H), 6.99 (m, 1 H), 7.06 (m, 2 H), 7.26 (m, 1 H), 7.34 (d, 1 H, J = 8.1 Hz), 7.94 (d, 2 H, J = 8.8 Hz), 7.98 (mn, 1 H), 8.18 (d, 1 H, J = 6.9 Hz); MS (FAB) m/z 552 (M). To a solution of methyl 4-[2-N-allyl-N-[4-[AN'-(2-chlorophenyl)ureido]-3-methoxyphenyl] 15 acetamido]ethoxybenzoate (250 mg, 0.45 mmol) in THF-MeOH (1: 1, v/ v, 8 ml), was added 0.25 N NaOH (3.6 ml, 0.91 mmol) at rt, and heated to reflux. The stirring was continued for 18 hours at reflux. The reaction mixture was poured into water, and acidified with 1.0 N HC1. The resulting precipitate was collected by filtration. The precipitate was recrystallized with hexane diethylether to give 310 as a white powder (195 mg, 80%). MW 537.99 'H-NMR (CD 3 OD)6 3.61 20 (s, 1 H), 3.76 (s, 3 H), 3.82 (m, 1 H), 3.85 (s, 1 H), 3.88 (m, 1 H), 4.11-4.25 (m, 4 H), 5.12-5.25 (m, 2 H), 5.81 (m, 1 H), 6.78 (d, 1 H, J= 8.3 Hz), 6.82 (m, 1 H), 6.92 (m, 2 H), 7.01 (m, 1 H), 7.26 (m, 1 H), 7.37 (m, 1 H), 7.96 (m, 3 H), 8.02 (m, 1 H); MS (FAB) m/z 537 (M); Anal. Called for C 2 8H 28
CIN
3 0 6 1/4 H20: C, 61.99; H, 5.30; N, 7.75. Found: C, 62.00; H, 5.56; N, 7.76. Example 261 25 4-[2-N-allyl-N-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyljacetamido]ethoxybenzoic acid N O.. COOH N N N O31 r H H M e 311 A mixture of methyl 4-(2-N-allylamino)ethoxybenzoate (118 mg, 0.5 mmol), 4-[N'-(2 bromophenyl)ureido]-3-methoxyphenylacetic acid (190 mg, 0.5 mmol), EDC-HCl (144 mg, 0.75 rnmol), HOBt (128 mg, 0.95 mmol), and DMAP (12 mg, 0.1 mmol) in DMF (2.5 ml) was stirred 451 WO 01/00206 PCT/US00/18079 for 18 hours. The mixture was diluted with EtOAc (300 mnl), washed with IN HCI and brine, and dried over MgSO 4 . After removal of the solvent, residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC-5404-FC, linear gradient CHCl 3 EtOAc 100:0 to 70:30) to give methyl 4-[2-N-allyl-N-[4-[N'-(2-bromophenyl)ureido]-3 5 methoxyphenyl] acetamido]ethoxybenzoate (251 mg, 84%) as an amorphous foam. 'H-NMR (CDCl 3 )6 3.65-3.85 (m, 4 H), 3.73 (s, 3 H), 3.88 (s, 3 H), 4.08 (m, 2 H), 4.22 (m, 2 H), 5.10-5.25 (m, 2 H), 5.78 (m, 1 H), 6.79 (m, 1 H), 6.85 (m, 3 H), 6.93 (m, 1 H), 7.02 (m, 2 H), 7.30 (m, 1 H), 7.51 (m, 1 H), 7.94 (d, 2 H, J = 8.8 Hz), 7.97 (m, 1 H), 8.14 (m, 1 H); MS (FAB) m/z 596 (M). To a solution of methyl 4-[2-N-allyl-N-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl] 10 acetamido]ethoxybenzoate (251 mg, 0.42 mmol) in THF-MeOH (1: 1, v/ v, 8 ml), was added 0.25 N NaOH (3.4 ml, 0.84 mmol) at rt, and heated to reflux. The stirring was continued for 18 hours at reflux. The reaction mixture was poured into water, and acidified with 1.0 N HCl. The resulting precipitate was collected by filtration. The precipitate was recrystallized with Hexane diethylether to give 311 (192 mg, 78%) as a white powder. MW 582.44 'H-NMR (CD 3 OD) 8 3.61 15 (s, 3 H), 3.77 (s, 3 H), 3.80 (m, 1 H), 3.85 (s, 1 H), 3.88 (s, 1 H), 4.12-4.25 (m, 4 H), 5.12-5.23 (m, 2 H), 5.81 (m, 1 H), 6.76 (m, 1 H), 6.82 (m, 1 H), 6.93 (m, 3 H), 7.29 (m, 1 H), 7.56 (m, 1 H), 7.94 (m, 4 H); MS (FAB), m/z 582 (M); Anal. Calcd for C28H 2 sBrN30 6 : C, 57.74; H, 4.85; N, 7.21. Found: C, 57.40; H, 5.07; N, 7.04. For HCI salt of 311 : Anal. Calcd for C 28
H
27 BrN30 6 0.25 H 2 0: C, 55.23; H, 4.55; N, 6.90. Found: C, 54.98; H, 4.71; N, 6.53. 20 Example 262 4-[2-N-allyl-N-[3-methyl-4-[N'-(2-methylphenyl)ureido]phenyl]acetamido]ethoxybenzoic acid 'I . O COOH Me"H H 312 A mixture of methyl 4-(2-N-allylamino-1-ethyl)ethoxybenzoate (87 mg, 0.37 mmol), 3 methyl-4-[N'-(2-methylphenyl)ureidolphenylacetic acid (100 mg, 0.37 mmol), EDC-HCI (105 mg, 25 0.56 mmol), HOBt (95 mg, 0.70 mmol), and DMAP (9 mg, 0.07 mmol) in DMF (7.4 ml) was stirred for 18 hours. The mixture was diluted with EtOAc (100 ml), washed with IN HCI and brine, and dried over MgSO 4 . The residue was co-evaporated with toluene (10 ml x 3) to remove DMF. The residue was chromatographed on TLC (Whatman, PLK-5F, 2 plates, CHC13-MeOH, 97:3) to give methyl 4-[2-N-allyl-N-[3-methyl-4-[N'-(2-methylphenyl)ureido]phenyl]acetamido 30 ethoxybenzoate (190 mg, 100%) as a white amorphous foam. 'H-NMR (CDC1 3 )8 2.05 and 2.09 (2s, total 3 H), 2.20 and 2.21 (s, total 3 H), 3.62 (s, 2 H), 3.76 (m, 2 H), 3.90 (s, 3 H), 3.88 (m, 1 452 WO 01/00206 PCT/US00/18079 H), 4.08 (m, 2 H), 4.11 (m, 1 H), 6.28 (m, 2 H), 5.78 (m, 1 H), 6.88 (d, J = 8.8 Hz, 2 H), 7.05 (m, 1 H), 7.12 (m, 1 H), 7.21 (m, 1 H), 7.58 (m, 2 H), 7.95 (d, J = 8.8 Hz, 2 H), 8.02 (m, 1 H); MS (FAB), m/z 516 (M*+1). To a solution of methyl 4-[2-N-allyl-N-[3-methyl-4-[N'-(2-methylphenyl)ureido]phenyl] 5 acetamido]ethoxybenzoate (217 mg, 0.43 mmol) in THF-MeOH (6: 1, v/ v, 7 ml), was added 0.5 N NaOH (1.9 ml, 0.86 mmol) at rt, and heated to reflux. The stirring was continued for 2 hours at reflux in a sealed bottle. The reaction mixture was poured into water, acidified with 1.0 N HCI, extracted with CHCl 3 -MeOH (2:1, 20 mL x 3), and dried over MgSO 4 . After removal of the solvent, the residue was crystallized with CHCl 3 -hexane-diethylether to give 312 (84 mg, 38%) as 10 a white powder. MW 501.57 1 H-NMR (CD 3 OD)8 2.18 and 2.24 (s, total 3 H), 2.30 (d, J= 4.9 Hz, 3 H), 3.70 (s, 1 H), 3.78 (m, 2 H), 3.88 (s, 1 H), 4.12 (m, 4 H), 5.20 (m, 2 H), 5.81 (m, 1 H), 6.92 7.20 (m, 7 H), 7.58 (m, 2 H), 7.96 (m, 2 H); MS (ESI) m/z 502 (M*); Anal. Calcd for C 2 9 31 N 3 0 5 : C, 69.44; H, 6.23; N, 8.38. Found: C, 68.99; H, 6.39; N, 8.03. Example 263 15 4-[2-N-allyl-N-[3-chloro-4-[N'-(2-methylphenyl)ureidolphenyl]acetamido]ethoxybenzoic acid COOH MeH H 1 313 To a stirred solution of methyl 4-(2-N-allylaminoethoxy)benzoate (141 mg, 0.60 mmol) and 3-chloro-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (191 mg, 0.60 mmol) in DMF (5 mL) were added EDC-HCl (172.5 mg, 0.90 mmol), HOBt (154 mg, 1.14 immol), and DMAP (15 20 mg, 0.12 mmol), and the stirring was continued overnight at rt. The mixture was diluted with EtOAc (50 mL) and washed with 1M HCI (x 3), IM NaOH (x 1), and brine. The mixture was dried over anhydrous MgSO 4 and concentrated under a reduced pressure to give methyl 4-[2-N allyl-N-[3-chloro-4-[N'-(2-methylphenyl)ureido] phenyl]acetamido]ethoxybenzoate (350 mg, 109%) as a white powder. 'H-NMR (CDCI 3 ) 8 2.35 (s, 3 H), 3.60 (s, 1 H), 3.75 (m, 2 H), 3.90 (s, 25 3 H), 4.10 (m, 4 H), 4.21 (m, 1 H), 5.20 (m, 2 H), 5.80 (m, 1 H), 6.50 (s, 1 H) , 6.85 (m, 2 H), 7.08 (m, 2 H), 7.20 (m, 4 H), 7.50 (d,J= 8.1 Hz, 1H), 7.95 (d,J= 8.1 Hz, 2 H), 8.12 (d,J= 8.1 Hz, 1 H); MS (ESI) m/z: 536 (M+H). To a stirred solution of methyl 4-[2-N-allyl-N-[3-chloro-4-[N'-(2-methylphenyl)ureido] phenyl]acetamido]ethoxybenzoate (321 mg, 0.6 mmol) in THF-MeOH-H 2 0 (2:2:1, v/v, 30 ml), 30 was added NaOH (500 mg, 12.5 mmol) at rt. The stirring was continued for 18 hours at rt. The 453 WO 01/00206 PCT/US00/18079 reaction mixture was poured into water, washed with diethyl ether, acidified with 1M HCI, extracted with CHCI 3 -MeOH (2:1, 20 mL x 3), dried over anhydrous MgSO 4 , and concentrated under a reduced pressure. The residue was solidified with CHClJn-hexane to give 313 (283 mg, 83%) as a white solid. IR (KBr): 3345, 1581, 1529, 1243, 1167 cm-'; 'H-NMR (CD 3 OD)8 2.30 (s, 5 3 H), 3.71 (s, 1 H), 3.78 (m, 1 H), 3.82 (m, 1 H), 3.89 (s, 1 H), 4.10 (m, 1 H), 4.19 (m, 2 H), 4.21 (t, J= 5.4 Hz, 1 H), 5.20 (m, 2 H), 5.82 (m, 1 H), 6.95 (m, 2 H), 7.03 (m, 1 H), 7.18 (m, 3 H), 7.28 (s, 1 H), 7.60 (d, J= 8.1 Hz, 1 H), 7.99 (m, 3 H); MS (ESI) m/z 522 (MW+l);Anal. Calcd for
C
28
H
2 1C1N 3 0 5 -1.75 H 2 0: C, 60.76; H, 5.74; N, 7.59. Found: C, 60.43; H, 5.34; N, 7.17. Example 264 10 methyl 4-[2-N-[2-(4-morpholinyl)ethyl]-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylI] acetamido]ethoxybenzoate O/n COOMe N N Me H H OMe 314 To a stirred solution of methyl 4-[2-N-(2,3-dihydroxy-l1-propyl)-[3-methoxy-4-[N'-(2 methylphenyl) ureido]phenylacetamido]ethoxy]benzoate (1.83 g, 3.24 mmol) in THF-MeOH-H 2 0 15 (1: 1: 1, v / v / v, 15 mL) was added sodium periodate (2.08 g, 9.71 mmol), and stirred for 18 hours at rt. A saturated Na 2
S
2 0 3 (50 ml) was added to the reaction mixture and the mixture was stirred for 1 hour. The mixture was extracted with EtOAc (100 ml x 3), washed with brine, dried over MgSO 4 . The solvent was removed to give the title compound (1.73 g, 100%) as an amorphous foam. 'H-NMR (CDCl 3 ) 8 2.32 (t, 3 H, J= 2.8 Hz), 3.33-4.30 (m, 8 H), 3.72 (s, 3 H), 20 3.86 (s, 3 H), 6.20 (m, 1 H), 6.70 (m, 1 H), 6.80 (m, 4 H), 7.06 (m, 1 H), 7.18 (m, 1 H), 7.26 (m, 1 H), 7.49 (d, 1 H, J = 7.4 Hz), 7.96 (m, 2 H), 8.10 (m, 1 H), 9.57 and 9.63 (2 s, total 1 H); MS (FAB), m/z 534 (MW+ 1). To a stirred solution of methyl 4-[2-N-formnylmethyl-N-[3-methoxy-4-[N'-(2-methyl phenyl)ureido]phenylacetamido]ethoxy]benzoate (400 mg, 0.75 mmol) in EtOH (7.5 ml), were 25 added morpholine (654 ml, 7.5 mmol) and acetic acid (429 ml, 7.5 mmol) at rt. The reaction was stirred for 5 min. at rt, then cooled to 0 oC. To the cooled solutionl, was added NaBH 3 CN (471 mg, 7.5 mmol) and the stirring was continued for 1 h at rt. The mixture was poured into sat. NaHCO 3 and was extracted with EtOAc (50 ml x 3), washed with brine, and dried over MgSO 4 * After removal of the solvent in vacuo, the residue was chromatographed on silica gel (middle 30 pressure chromatography system: YAMAZEN YFLC-5404-FC, linear gradient toluene-acetone 100:0 to 1: 1) to give 314 (346 mg, 76%) as a white amorphous foam. 'H-NMR (CDCl 3 , 400 454 WO 01/00206 PCT/US00/18079 MHz) 8 2.31 (s, 3H), 2.46 (min, 4 H), 3.52-3.79 (mn, 12 H), 3.70 (d, 3 H, J= 2.7 Hz), 4.05 and 4.22 (min, total 2 H), 6.24 and 6.29 (s, total 1 H), 6.73 (min, 2 H), 6.85 (min, 2 H), 7.07 (s, 1 H), 7.17 (mn, 1 H), 7.25 (min, 2 H), 7.50 (t, 1 H, J = 7.3 Hz), 7.96 (min, 2 H), 8.08 (min, 1 H); MS (FAB), m/z 605 (M+ 1); Anal. Calcd for C 33
H
40
N
4 0 7 1/2 H 2 0: C, 64.58; H, 6.73; N, 9.13. Found: C, 64.95; H, 5 6.88; N, 8.82. HCI salt of 314: Anal. Calcd for C 33
H
4 C1N 4 0 7 -2.5 H 2 0: C, 57.76; H, 6.76; N, 8.16; Cl, 5.17; Found: C, 58.29; H, 6.81; N, 7.42; Cl, 5.05. Example 265 4-[2-N-[2-(4-morpholinyl)ethyl]-N-[3-methoxy-4-[N '-(2-methylphenyl)ureido]phenyl]acetamido] ethoxybenzoic acid OCOOH NN O 10 Me H H OMe 315 To a solution of methyl 4-[2-N-[2-(4-morpholinyl)ethyl]-N-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenyl]acetamniido]ethoxybenzoate (146 mg, 0.24 nunmmol) in THF-MeOH (1: 1, v/ v, 6 ml), was added 0.25 N NaOH (1.9 ml, 0.48 mmol) at rt, and heated to reflux. The stirring was continued for 18 hours at reflux. The reaction mixture was poured into water, and acidified with 15 1.0 N HCL. The mixture was extracted with CHCl 3 -MeOH (3: 1, v/ v, 30 ml x 5). The combined organic solvent was dried over MgSO 4 . After removal of solvent, the residue was crystallized with diethylether to give 315 (102 mg, 71%) as a white powder. 'H-NMR (CD 3 OD) 8 2.28 (d, J= 3.0 Hz, 3 H), 2.46 (min, 1 H), 2.40 (min, 1 H), 2.56 (min, 1 H), 2.63 (mn, 1 H), 3.62-3.80 (mn, 12 H), 3.85 (s, 3 H), 4.12 (min, 1 H), 4.26 (min, 1 H), 6.82 (min, 2 H), 6.96 (min, 2 H), 7.01 (mn, 1 H), 7.17 (min, 2 H), 20 7.58 (d, J= 7.8 Hz, 1 H), 7.93 (min, 3 H); MS (FAB) m/z 591 (M); Anal. Calcd for C 32
H
3 8
N
4 071.0
H
2 0: C, 63.14; H, 6.62; N, 9.20. Found: C, 63.48; H, 6.66; N, 8.79. Example 266 4-[2-N-cyclopropyl-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl]acetamido] ethoxybenzoic acid Y COOH N-N 25 Me H H Me 316 To a stirred solution of methyl 4-(2-hydroxyethyloxy)benzoate (5.00 g, 25.5 mmol), DMSO (18.1 ml, 255 mmnol), Et 3 N (17.7 ml, 127.5 mmol) in CH 2 C1 2 (200 ml) was added SO3-Py (12.2 g, 76.5 mmol). After stirring for 5 h, the mixture was concentrated in vacuo and the residue was diluted with H 2 0 (100 ml). The mixture was extracted with EtOAc (2 x 200 ml). The 30 combined extracts were washed with brine (100 ml), dried over (MgSO 4 ), and evaporated. The 455 WO 01/00206 PCT/US00/18079 residue was chromatographed on silica gel with hexane-EtOAc (4:1) to give 4:1 mixture of methyl 4-formyl methyloxybenzoate and methyl 4-hydroxybenzoate (2.00 g) as a white solid. 1 H-NMR (CDCl 3 )8 3.90 (s, 3 H), 4.64 (d, J= 1.0 Hz, 2 H), 6.92 (d, J= 9.0 Hz, 2 H), 8.02 (d, J= 9.0 Hz, 2 H), 9.86 (d, J = 1.0 Hz, 1 H). 5 To a stirred solution of 4:1 mixture of methyl 4-formylmethyloxybenzoate and methyl 4 hydroxybenzoate (1.00 g) and cyclopropylamine (425 ml, 6.18 mmol) in MeOH-AcOH (10:1, 11 ml) was added NaBH 3 CN (681 mg, 10.3 mmol). After stirring overnight, the mixture was quenched by addition of sat. NaHCO 3 (50 ml) and extracted with CHCl 3 (2 x 200 ml). The combined extracts were dried over MgSO 4 and evaporated. The residue was chromatographed on 10 silica gel with CHCl 3 -MeOH (20:1) to give methyl 4-(2-cyclopropyl aminoethoxy)benzoate (595 mg, 49%) as a colorless oil. 'H-NMR (CDCl 3 )8 0.37-0.49 (m, 4 H), 1.91 (m, 1 H), 2.18-2.23 (m, 1 H), 3.11 (t, J= 5.2 Hz, 2 H), 3.88 (s, 3 H), 4.12 (t, J= 5.2 Hz, 2 H), 6.92 (d, J= 8.8 Hz, 2 H), 7.98 (d, J= 8.8 Hz, 2 H). A mixture of methyl 4-(2-cyclopropylaminoethoxy)benzoate (290 mg, 1.23 mmol), 3 15 methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (387 mg, 1.23 mmol), EDC-HCI (354 mg, 1.85 mmol), HOBt (cat.), and DMAP (cat.) in DMF (10 ml) was stirred overnight. The mixture was partitioned between EtOAc (300 ml) and H 2 0 (100 ml). The organic phase was separated, washed with brine (2 x 100 ml), dried over MgSO 4 , and evaporated. The residue was chromatographed on silica gel with CHCl 3 -MeOH (50:1) to give the title compound (426 mg, 20 65%) as a yellow viscous oil. 'H-NMR (CDC1 3 )5 0.90-0.97 (m, 4 H), 2.28 (s, 3 H), 3.60 (s, 3 H), 3.77 (t, J= 5.4 Hz, 2 H), 3.85 (s, 2 H), 3.87 (s, 3 H), 4.15 (t, J= 5.4 Hz, 2 H), 6.60-6.81 (m, 5 H), 7.09-7.23 (m, 4 H), 7.57 (d, J= 8.3 Hz, 1 H), 7.92-7.95 (m, 2 H), 8.04 (d, J= 8.3 Hz, 1 H). To a stirred solution of methyl 4-[2-[N-cyclopropyl-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenyl]acetamido]ethoxy]benzoate (426 mg, 0.801 mmol) in THF (7 ml) was added 0.25 N 25 NaOH (6.4 ml, 1.60 mmol). After stirring overnight, the mixture was poured into 1 N HCI (50 ml) and extracted with CHCl 3 -MeOH (4:1, 2 x 200 ml). The combined extracts were dried over MgSO 4 and evaporated. The residue was chromatographed on silica gel with CHCl 3 -MeOH (20:1 to 10:1) as eluent to give 316 (333 mg, 80%) as a colorless amorphous solid. 1 H-NMR (DMSO) 8 0.86-0.91 (m, 4 H), 2.25 (s, 3 H), 2.74 (m, 1 H), 3.69 (t, J= 5.5 Hz, 2 H), 3.81 (s, 3 H), 3.83 (s, 2 30 H), 4.15 (t,J= 5.5 Hz, 2 H), 6.75 (d,J= 8.3 Hz, 1 H), 6.87 (s, 1 H), 6.92-6.99 (m, 3 H), 7.11-7.17 (m, 2 H), 7.81 (d, J= 8.1 Hz, 1 H), 7.88 (d, J= 8.5 Hz, 2 H), 8.01 (d, J=8.1 Hz, 1 H), 8.47 (s, 1 H), 8.55 (s, 1 H), 12.96 (s, br s); MS (FAB), m/z 518 (M++1); Anal. Calcd for C29H3 1
N
3 06: C, 456 WO 01/00206 PCT/US00/18079 67.30; H, 6.04; N, 8.12. Found: C, 66.71; H, 6.26; N, 7.82. Example 267 4-[2-N-[2-(N', N'-dimethylamino)-1-ethyl)-N-[-4-[N"'-(2-chlorophenyl)ureido]-3-methoxphenyl] acetamido]ethoxybenzoic acid N- OH NN O 5 I H H Me 317 To a solution of methyl 4-[2-N-[2-(N', N'-dimethylamino)-1-ethyl)-N-[-4-[N"-(2 chlorophenyl) ureido]-3-methoxphenyl]jacetamido]ethoxybenzoate (100 mg, 0.17 mmol) in THF MeOH (1: 1, v/v, 3 ml), was added 0.25 N NaOH (2.0 ml, 0.5 mmol) at rt, and heated to reflux. The stirring was continued for 3 hours at reflux. The reaction mixture was poured into water, and 10 acidified to pH 5.0 with 1.0 N HC1. After removal of the organic solvent in vacuo, the resulting mixture was chromatographed with HP-20 (H 2 0-MeOH 100:0 to 0:100) to give 317 (63 mg, 64%) as a white powder. 'H-NMR (CD 3 OD) 8 2.41 (s, 2 H), 2.65 (s, 3 H), 2.69 (s, 3 H), 3.02 (min, 2 H), 3.62-3.85 (min, 4 H), 3.84 (s, 3 H), 4.05 and 4.22 (min, total 2 H), 6.82-6.88 (min, 4 H), 7.02 (min, 1 H), 7.25 (min, 1 H), 7.38 (min, 1 H), 7.92 (min, 2 H), 8.01 (min, 2H); MS (FAB), m/z 569 (M); Anal. Calcd 15 for C 2
,H
33 C1N 4 0 6 3.0 H 2 0: C, 55.90; H, 6.31; N, 8.99. Found: C, 56.40; H, 6.50; N, 8.08. Example 268 isopropyl 4-[2-N-[2-(4-morpholinyl)ethyl]-N-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenyl] acetamido]ethoxybenzoate ON COOiPr Me H H Me 318 20 To a stirred solution of 4-[2-N-[2-(4-morpholinyl)ethyl]-N-[3-methoxy-4-[N'-(2 methylphenyl) ureido]phenyl]acetamido]ethoxybenzoic acid (250 mg, 0.42 mmol) in DMF (2 mnL), were added isopropyl iodide (264 ml, 2.53 mmol) and K 2 C0 3 (88 mg, 0.64 mmol) at rt. The reaction was stirred for 2 hours at 50 oC. The mixture was poured into brine and was extracted with CHC13 (50 ml x 3), washed with brine, and dried over MgSO 4 . After removal of the solvent 25 in vacuo, the residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC-5404-FC, linear gradient toluene-acetone 100:0 to 40: 60) to give 318 (261 mg, 97%/) as a white amorphous foam. 'H-NMR (CDC13, 400 MHz) 8 1.35 (s, 3 H), 1.36 (s, 3 H), 2.31 (s, 3 H), 2.45 (min, 4 H), 2.50 (min, 2 H), 3.55-3.78 (m, 10 H), 3.70 (s, 3 H), 4.05 and 4.20 (t, J= 5.2 Hz, total 2 H), 5.22 (min, 1 H), 6.24 and 6.33 (2 s, total 1 H), 6.70-6.83 (min, 4 H), 7.08 (s, 1 H), 30 7.15 (min, 1 H), 7.22 (min, 1 H), 7.40 (t, J= 9.0Hz, 1 H), 7.93 (d,J= 8.8Hz, 1 H), 7.97 (d, J= 8.8 457 WO 01/00206 PCT/US00/18079 Hz, 1 H), 8.07 (t, J= 7.8 Hz, 1 H); MS (FAB), m/z 633 (M+ 1);Anal..Calcd for C 3 H44N 4 07-0.75 H20: C, 65.05; H, 7.10; N, 8.67. Found: C, 65.19; H, 7.09; N, 8.50. Example 269 4-[2-N-[2-(3,3-difluoro-1-pyrrolidinyl)ethyl]-N-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl] 5 acetamido]ethoxybenzoic acid sodium salt F tF NCOONa N Nn r H H 6Me 319 To a stirred solution of methyl 4-[2-N-formylmethyl-N-[3-methoxy-4-[N'-(2 bromophenyl)ureido] phenyl]acetamido]ethoxybenzoate (300 mg, 0.5 mmol) in 1,2-dichloroethane (3.6 ml), was added 3,3-difluoropyrrolidine AcOH salt (420 mg, 2.5 mmol) at rt. The reaction 10 was stirred for 5 min. at rt, then cooled to 0 'C. To the cooled solution, was added NaBH(OAc) 3 (530 mg, 2.5 mmol), and the stirring was continued for 4 h at rt. The mixture was poured into sat. NaHCO 3 , was extracted with CHCl 3 (50 mL x 3), washed with brine, and dried over MgSO 4 . After removal of the solvent in vacuo, the residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC-5404-FC, linear gradient CHCl 3 -MeOH 10:0 15 to 97:3) to give methyl 4-[2-N-[2-(3,3-difluoro-1-pyrrolidinyl)ethyl]-N-[4-[N'-(2-bromophenyl) ureido]-3-methoxyphenyl] acetamido]ethoxybenzoate (345 mg, 100%) as a white amorphous foam. 'H-NMR (CDC13, 400 MHz) 8 2.20-2.80 (m, 6 H), 3.48 (m, 2 H), 3.70-3.80 (m, 9 H), 3.89 (s, 3 H), 4.09 (m, 1 H), 4.21 (m, 1 H), 6.79-6.85 (m, 4 H), 6.93 (m, 1 H), 7.08 (m, 2 H), 7.30 (m, 1 H), 7.50 (m, 1 H), 7.96 (m, 3 H), 8.13 (dd, J= 1.5 Hz, 8.3 Hz, 1 H); MS (ESI) m/z 689 (MW). 20 To a solution of methyl 4-[2-N-[2-(3,3-difluoro-l1-pyrrolidinyl)ethyl]-N-[4-[N'-(2 bromophenyl) ureido]-3-methoxyphenyl]acetamido]ethoxybenzoate (345 mg, 0.5 mmol) in THF MeOH (1: 1, v/ v, 8 ml), was added 0.25 N NaOH (4 ml, 1.0 mmol) at rt, and heated to reflux. The stirring was continued for 6 hours at reflux. The solvent was removed, and the residue was chromatographed on HP-20 (H 2 0-MeOH, 0:100 to 100: 0) to give 319 (306 mg, 91%) as a pale 25 red powder. 'H-NMR (CD 3 OD, 400 MHz) 8 2.20-2.90 (m, 6 H), 3.60 (m, 2 H), 3.70-3.92 (m, 9 H), 4.10 (m, 1 H), 4.22 (m, 1 H), 6.84 (m, 4 H), 6.96 (m, 1 H), 7.30 (m, 1H), 7.55 (m, 1H), 7.93 (m, 3H), 7.97 (m, 1 H); MS (ESI) m/z 676 (M'+1); Anal. Calcd for C 31
H
3 2 BrF 2
N
4 0 6 2.5 H 2 0: C, 52.85; H, 5.29; N, 7.95. Found: C, 52.67; H, 5.20; N, 8.11. Example 270 30 4-[2-N-(N'-methoxy-N'-methylamino)ethyl-N-[3-methoxy-4-[N" '-(2-methylphenyl)ureido] 458 WO 01/00206 PCT/US00/18079 phenyl]acetamido]ethoxybenzoic acid sodium salt -o -0O" Na+ NNN MeN Me 320 To a stirred solution of methyl 4-[2-N-formylmethyl-[3-methoxy-4-[N'-(2-methylphenyl) ureido] phenylacetamido]ethoxylbenzoate (350 mg, 0.66 mmol) in EtOH (13 ml), was added N 5 methoxy-N-methylamine hydrochloride (637 mg, 6.6 mmol) at rt. The reaction was sonicated for 5 min. at rt, then cooled to 0 °C. To the cooled solution, was added NaBH 3 CN (105 mg, 1.65 mmol) and the stirring was continued for 18 h at rt. The mixture was poured into sat. NaHCO 3 and was extracted with CHC13 (50 ml x 3), washed with brine, and dried over MgSO 4 . After removal of the solvent in vacuo, the residue was chromatographed on silica gel (middle pressure 10 chromatography system: YAMAZEN YFLC-5404-FC, linear gradient toluene-acetone 9: 1 to 2: 3) to give the title compound (344 mg, 91%) as a white amorphous foam. 'H-NMR (CDCl 3 , 400 MHz) 8 2.29 (s, 3 H), 2.52 and 2.58 (s, total 3 H), 2.79 (m, 2 H), 3.49-3.76 (m, 9 H), 3.88 (s, 3 H), 4.05 and 4.20 (m, total 2 H), 6.69-6.86 (mn, 5 H), 7.10 (m, 1 H), 7.20 (m, 2 h), 7.29 (m, 1 H), 7.44 (m, 1 H), 7.94 and 7.99 (d, J = 8.6 Hz, total 2 H), 8.06 (m, 1 H); MS (FAB), nm/z 579 (M+ 1); 15 Anal. Calcd for C 31
H
38
N
4 0 7 2.5H 2 0: C, 59.70; H, 6.95; N, 8.98. Found: C, 59.58; H,6.65; N, 8.90. To a solution of methyl 4-[2-N-(N'-methoxy-N'-methylamino)ethyl-N-[3-methoxy-4-[N" (2-methylphenyl)ureido]phenyl]acetamido]ethoxybenzoate (138 mg, 0.24 mmol) in THF-MeOH (1: 1, v/ v, 4 ml), was added 0.25 N NaOH (1.9 ml, 0.48 mmol) at rt, and heated to reflux. The stirring was continued for 18 hours at reflux. The solvent was removed, and the residue was 20 chromatographed on HP-20 (H 2 0O-MeOH, 100: 0 to 0: 100) to give 320 (140 mg, 100%) as a white powder. 'H-NMR (CD 3 OD) 8 2.29 (s, 3 H), 2.54 and 2.56 (2 s, total 3 H), 2.82 (m, 2 H), 3.48 (m, 2 H), 3.65-5.80 (m, 9 H), 4.09 and 4.21 (2 m, total 2 H), 6.80 (m, 4 H), 7.00 (t, J= 7.5 Hz, 1 H), 7.18 (m, 2 H), 7.57 (d, J= 7.8 Hz, 1 H), 7.88 (m, 2 H), 7.99 (m, 1 H); MS (FAB), m/z 565 (M+1); Anal. Calcd for C 30
H
35
N
4 0 7 Na-1.0 H 2 0: C, 59.59; H, 6.17; N, 9.27. Found: C, 59.10; H, 6.28; N, 25 8.86. Example 271 4-[2-N-(N'-methoxy-N'-methylamino)ethyl-N-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl acetamido]ethoxy]benzoic acid 459 WO 01/00206 PCT/US00/18079 -0 N OH N N O 32 r H H 6Me 321 To a stirred solution of methyl 4-[2-N-formylmethyl-N-[3-methoxy-4-[N'-(2 bromophenyl)ureido] phenyl]acetamido]ethoxybenzoate (209 mg, 0.35 mmol) in EtOH (7 ml), was added N-methoxy-N-methylamine HCI salt (341 mg, 3.5 mmol) at rt. The reaction was sonicated 5 for 5 min. at rt, then cooled to 0 C. To the cooled solution, was added NaBH(OAc) 3 (370 mg, 1.75 mmol) and the stirring was continued for 18 h at rt. The mixture was poured into sat. NaHCO 3 , extracted with CHCl 3 (50 ml x 3), and dried over MgSO 4 . After removal of the solvent, the residue was chromatographed on TLC (Whatman, PLK-5F, 2 plates, CHC13-MeOH, 98:2) to give methyl 4-[2-N-(N'-methoxy-N'-methylamino)ethyl-N-[4-[N'-(2-bromophenyl)ureido]-3 10 methoxyphenyl acetamido]ethoxy]benzoate (89 mg, 40%) as a white amorphous foam. 1 H-NMR
(CDC
3 , 400 MHz) 6 2.52 and 2.60 (2 s, 3 H), 2.80 (m, 2 H), 3.48 and 3.50 (2 s, total 3 H), 3.65 (m, 2 H), 3.72 (s, 3 H), 3.77 (in, 4 H), 3.90 (s, 3 H), 4.08 (m, 1 H), 4.22 (m, 1 H), 6.82 (m, 5 H), 7.12 (s, 2 H), 7.30 (m, 1 H), 7.52 (d,J= 8.1 Hz, 1 H), 7.94 (mi, 3 H), 8.15 (d,J= 8.3 Hz, 1 H); MS (ESI) m/z 643 (M). 15 To a solution of methyl 4-[2-N-(N'-methoxy-N'-methylamino)ethyl-N-[4-[N'-(2 bromophenyl)ureido]-3-methoxyphenylacetamido]ethoxy]benzoate (89 mg, 0.14 mmol) in THF MeOH (5: 1, v/ v, 6 ml), was added 0.5 N NaOH (1.4 ml, 0.7 mmol) at rt, and heated to reflux in a glass sealed bottle. The stirring was continued for 3 hours at reflux. The reaction was poured into water, and was acidified with 1 N HCI to pH 5, extracted with CHCl 3 -MeOH (2:1, v/ v, 30 mL 20 x 3), dried over MgSO 4 . The solvent was removed in vacuo to give 321 (53 mg, 60%) as a white powder. 'H-NMR (CD30D, 400 MHz)8 2.62 and 2.64 (2 s, total 3 H), 2.80 (m, 2 H), 3.50 (d, J= 7.3 Hz, 3 H), 3.63-3.88 (m, 6 H), 4.12 (m, 1 H), 4.25 (m, 1 H), 6.82-7.00 (m, 5 H), 7.30 (m, 1 H), 7.58 (m, 1H), 7.95 (m, 4 H); MS (FAB), m/z 629 (M+); Anal. Calcd for C 29
,H
33 BrN 4 O0 7 0.25 H 2 0: C, 54.94; H, 5.33; N, 8.84. Found: C, 55.39; H, 5.53; N, 8.23. 25 Example 272 4-[2-N-(N', N'-diallyl)ethyl-N-[3-methoxy-4-[N"-(2-methylphenyl)ureido]phenyl]acetamido] ethoxybenzoic acid sodium salt O Na+ - 0 NK; N 32 Me H H 4Me 322 460 WO 01/00206 PCT/US00/18079 To a stirred solution of methyl 4-[2-N-formylmethyl-N-[3-methoxy-4-[N'-(2 methylphenyl)ureido] phenyl]acetamido]ethoxybenzoate (400 mg, 0.75 mmol) in EtOH (15 ml), were added diallylamine (926 ml, 7.5 mmol) and acetic acid (429 ml, 7.5 mmol) at rt. The reaction was stirred for 5 min. at rt, then cooled to 0 *C. To the cooled solution, was added 5 NaBH 3 CN (118 mg, 1.9 mmol) and the stirring was continued for 1 h at rt. The mixture was poured into sat. NaHCO 3 and was extracted with EtOAc (50 ml x 3), washed with brine, and dried over MgSO 4 . After removal of the solvent in vacuo, the residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC-5404-FC, linear gradient toluene-acetone 9: 1 to 1: 1) to give methyl 4-[2-N-(N', N'-diallyl)ethyl-N-[3-methoxy-4-[N"-(2 10 methylphenyl)ureido] phenyl]acetamido] ethoxybenzoate (385 mg, 84%) as a white amorphous foam. 'H-NMR (CDCl 3 , 400 MHz) 8 2.30 (s, 3 H), 2.60 (m, 2 H), 3.09 (m, 4 H), 3.49 (m, 2 H), 3.60 (s, 2 H), 3.70 (m, 5 H), 3.89 (s, 3 H), 4.02 and 4.19 (2 m, total 2 H), 5.01-5.20 (m, 4 H), 4.79 (m, 2 H), 6.30 and 6.32 (2 s, total 1 H), 6.70-6.84 (m, 5 H), 7.08 (m, 1 H), 7.14 (m, 1 H), 7.25 (m, 1 H), 7.60 (m, 1 H), 7.93 and 7.98 (2 d, J= 8.8 Hz, 2.H), 8.03 and 8.06 (2 d, J = 8.3 Hz, 1 H); MS 15 (FAB), m/z 615 (M+ 1). To a stirred solution of methyl 4-[2-N-(N', N'-diallyl)ethyl-N-[3-methoxy-4-[N"'-(2 methylphenyl) ureido]phenyl]acetamido]ethoxybenzoate (385 mg, 0.63 mmol) in MeOH (3 ml), was added IN HCI (756 ml, 0.76 mmol) at rt. The reaction was stirred for 5 min. at rt, then evaporated to give methyl 4-[2-N-(N', N'-diallyl)ethyl-N-[3-methoxy-4-[N"-(2-methylphenyl) 20 ureido]phenyl] acetamido]ethoxybenzoate HCI salt (385 mg, 99%) as an amorphous foam. Anal. Calcd for C 3
,I
43 C1N 4 0-0.5 H 2 0: C, 63.67; H, 6.72; N, 8.49. Found: C, 63.67; H, 6.69; N, 8.43. To a solution of 4-[2-N-(N', N'-diallyl)ethyl-N-[3-methoxy-4-[N '-(2-methylphenyl) ureido]phenyl]acetamido]ethoxybenzoate (175 mg, 0.29 mmol) in THF-MeOH (1: 1, v/ v, 20 ml), was added 0.25 N, NaOH (2.5 ml, 0.63 mmol) at rt, and heated to reflux. The stirring was 25 continued for 1 hours at reflux. The solvent was removed, and the residue was chromatographed on HP-20 (H 2 0-MeOH, 100: 0 to 0: 100) to give 322 (160 mg, 94%) as a white powder. 'H-NMR
(CD
3 OD) 8 2.29 (s, 3 H), 2.60 (t, J= 6.9 Hz, 1 H), 2.67 (t, J= 7.0 Hz, 1 H), 3.10 (d, J= 6.6 Hz, 2 H), 3.14 (d, J = 6.6 Hz, 2 H), 3.59 (m, 2 H), 3.69-3.80 (m, 4 H), 3.80 (s, 3 H), 4.06 (t,J= 5.2 Hz, 4.21 (t, J= 5.1 Hz, 1 H), 5.15 (m, 4 H), 5.80 (m, 2 H), 6.79 (m, 2 H), 6.84 (d, J= 8.8 Hz, 2 H), 30 7.00 (t, J= 7.5 Hz, 1 H), 7.14 (m, 2 h), 7.48 (m, 1 H), 7.91 (dd, J= 6.1 Hz, 8.8Hz, 2 H), 8.00 (m, 1 H); MS (FAB), m/z 601 (M); Anal. Calcd for C 34
H
39 N40 6 Na -0.5 HO 2 0: C, 64.65; H, 6.38; N, 8.87. Found: C, 64.53; H, 6.58; N, 8.78. 461 WO 01/00206 PCT/US00/18079 Example 273 4-[2-N-(N', N'-diallyl)ethyl-N-[4-[N '-(2-chlorophenyl)ureido]-3-methoxyphenylacetamido]ethoxy] benzoic acid sodium salt N N O C O O N a I H H (Me 323 5 To a stirred solution of methyl 4-[2-N-formylmethyl-N-[4-[N'-(2-chlorophenyl)ureido]-3 methoxy phenylacetamido]ethoxy]benzoate (100 mg, 0.18 mmol) in EtOH (3.6 ml), was added diallylamine (223 ml, 1.81 mmol) at rt. The reaction was stirred for 5 min. at rt, then cooled to 0 oC. To the cooled solution, were added AcOH (104 ml, 1.81 mmol) and NaBH 3 CN (28 mg, 0.45 mmol), and the stirring was continued for 18 h at rt. The mixture was poured into sat. NaHCO 3 , 10 was extracted with CHC1 3 (30 mL x 3), washed with brine, and dried over MgSO 4 . After removal of the solvent in vacuo, the residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC-5404-FC, linear gradient CHCl 3 -MeOH 10:0 to 20: 1) to give methyl 4-[2-N-(N', N'-diallyl)ethyl-N-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenyl acetamido]ethoxy] benzoate (96 mg, 83%) as a white amorphous foam. 'H-NMR (CDCl 3 , 400 15 MHz) 8 2.60 (m, 2 H), 3.09 (d, J = 6.4 Hz, 1 H), 3.11 (d, J= 6.4 Hz, 3 H), 3.52 (m, 2 H), 3.61 (s, 2 H), 3.70-3.80 (m, 5 H), 3.86 (s, 3 H), 4.05 and 4.20 (2 m, total 2 H), 5.06-5.21 (m, 4 H), 5.80 (m, 2 H), 6.71-6.85 (m, 4 H), 6.98 (m, 1 H), 7.22 (m, 1 H), 7.32 (m, 3 H), 7.93 (d, J = 7.8 Hz, 2 H), 7.98 (m, 1 H), 8.18 (dd, J= 1.5 Hz, 8.2 Hz, 1 H); MS (ESI) m/z 635 (M). To a solution of methyl 4-[2-N-(N', N'-diallyl)ethyl-N-[4-[N'-(2-chlorophenyl)ureido]-3 20 methoxy phenylacetamido]ethoxy]benzoate (96 mg, 0.15 mmol) in THF-MeOH (1: 1, v/ v, 8 ml), was added 0.25 N NaOH (3.91 ml, 0.98 mmol) at rt, and heated to 50 *C. The stirring was continued for 6 hours at reflux. The solvent was removed, and the residue was chromatographed on HP-20 (H 2 0-MeOH, 0:100 to 100: 0) to give 323 (88 mg, 94%) as a white powder. 'H-NMR
(CD
3 OD, 400 MHz) 8 2.61 (m, 2 H), 3.10 (s, 2 H), 3.12 (s, 2 H), 3.59 (m, 2 H), 3.70 (s, 2 H), 3.78 25 (m, 2 H), 3.82 (s, 3 H), 4.10 (m, 1 H), 4.21 (m, 1 H), 5.18 (m, 4 H), 5.81 (m, 2 H), 6.82 (m, 4 H), 7.01 (t, J = 7.8 Hz, 1 H), 7.25 (m, 1 H), 7.39 (d, J = 7.8 Hz, 1 H), 7.90 (m, 2 H), 8.02 (m, 2 H); MS (ESI) m/z 621 (M); Anal. Calcd for C 33
H
3 6 CIN40 6 Na-1.25 H 2 0: C, 59.55; H, 5.83; N, 8.42. Found: C, 59.90; H, 5.74; N, 7.96. Example 274 30 4-[2-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl]-N-methylacetoamido]ethyl-1 piperazinylacetic acid 462 WO 01/00206 PCT/US00/18079 N N NCOOH Me H H CMe 324 To a stirred suspension of 1-(2-hydroxyethyl)piperazine (5.21g, 40.0mmol) and K 2 C0 3 (8.76g, 63.4mmol) in CH 3 CN (100ml) was added ethyl bromoacetate (5.60ml, 50.5mmol) at 0 0 C. The reaction mixture was heated under reflux for 5h, diluted with EtOAc, and washed with water 5 and brine. The extract dried over Na 2
SO
4 , concentrated to dryness and afforded ethyl 4-(2 hydroxyethyl)-1-piperazinylacetate (9.65g, 100%) as a yellow oil. 'H-NMR (CDCl 3 ) 8 1.23 (t, 3H, J=7.3Hz), 2.51-2.61(m, 11H), 3.22(s, 2H), 3.61(t, 2H, J=5.4Hz), 4.19(q, 2H, J=7.3Hz). To a solution of 2,4-dinitrobenzenesulfonyl chloride (1.0g, 3.75mmol) and pyridine (0.34ml, 4.20mmol) in THF (19ml) was added dropwise methylamine (2.0M THF solution, 2.3ml, 10 4.60mmol) at 0 0 C. The reaction mixture was stirred for lhr, quenched by the addition of IN HCI solution, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 solution and brine, dried over Na 2
SO
4 , and concentrated to dryness. The residue was recrystallized from EtOAc-Et20 to give methyl 2,4-dinitrobenzenesulfonamide (546mg, 56%) as a colorless solid. 'H-NMR (DMSO) 8 2.60 (d, 3H, J=4.9Hz), 8.22 (d, 1H, J=8.8Hz), 8.31 (q, 1H, J=4.9Hz), 8.66 (dd, 1H, 15 J=8.8, 2.0Hz), 8.90 (d, 1H, J=2.0Hz). To a solution of ethyl 4-(2-hydroxyethyl)-l1-piperazinylacetate (452mg, 2.09mmol), methyl 2,4-dinitrobenzenesulfon-amide (546mg, 2.09mmol) and PPh 3 (658mg, 2.5 lmmol) in THF was added DIAD (0.50ml, 251mmol) at 0'C. After stirring 17h at room temperature, the reaction mixture was concentrated to dryness. Chromatography of the residue with EtOAc-MeOH (10:1) to 20 afford ethyl 4-[2-[N-(2,4-dinitrobenzensulfonyl)-N-methylamino]ethyl]-1l-piperazinylacetate (864mg, 90%) as a reddish oil. 'H-NMR (CDCl 3 ) 5 1.27 (t, 3H, J=6.8Hz), 2.35-2.63 (m, 10H), 2.98 (s, 3H), 3.20 (s, 2H), 3.41 (t, 2H, J=6.8Hz), 4.17 (q, 2H, J=6.8Hz), 8.33 (d, 1H, J=8.3Hz), 8.46 (d, 1H, J=2.0Hz), 8.50 (dd, 1H, J=8.3, 2.0Hz). A solution of ethyl 4-[2-[N-(2,4-dinitrobenzensulfonyl)-N-methylamino]ethyl]- 1 25 piperazinylacetate (864mg, 1.88 mmol) ,mercaptoacetic acid (0.17ml, 2.44mmol) and Et 3 N (0.53ml, 3.76mmol) in CH 2 C1 2 (25ml) was stirred at rt for 3hr. The reaction mixture ethyl 4-(2 methylaminoethyl)-1-piperazinylacetate (388mg, 90%) as reddish oil. 'H-NMR (CDC13) 8 1.27 (t, 3H, J=6.8Hz), 2.50 (s, 3H), 2.53-2.60 (m, 8H), 2.75 (t, 2H, J=5.9Hz), 3.20 (s, 2H), 4.18 (q, 2H, J=6.8Hz). 463 WO 01/00206 PCT/US00/18079 To a solution of ethyl 4-(2-methylaminoethyl)-1-piperazinylacetate (388mg, 1.69mmol), Et 3 N (0.32ml, 2.25mmol) and DMAP (46mg, 0.38mmol) in DMF (15ml) was stirred for 15min at room temperature, then (532mg, 1.69mmol), HOBt (103mg, 0.76mmol) and EDC-HCI (486mg, 2.53mmol) was added to the reaction mixture which was stirred for 15h at rom temperature The 5 reaction mixture was diluted with EtOAc, which was washed with brine, dried over Na 2
SO
4 , and concentrated to dryness. Chromatography of the residue with CHClI-MeOH (10:1, v/v) to afford ethyl 4-[2-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl]-N-methylacetoamido]ethyl- 1 piperazinylacetate (889mg, mixture of DMF) as a reddish oil. 'H-NMR (CDCl 3 ) 8 1.25-1.29 (m, 3H), 2.29 (s, 3H11), 2.42-2.63 (m, 10H), 3.20, 3.18 (each s, total 3H11), 3.55, 3.40 (each t, total 2H, 10 J=6.8Hz), 3.65, 3.69 (each s, total 2H), 3.72 (s, 3H), 4.15-4.21 (m, 2H), 6.50 (m, 1H), 6.77-6.81 (m, 8H11), 7.11-7.24 (m, 3H), 7.53 (d, 1H, J=8.3Hz), 8.02 (s, 1H), 8.06 (d, 1H, J=7.8Hz). To a stirred solution of ethyl 44-[2-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl] N-methylacetoamido]ethyl-1-piperazinylacetate (889mg, 1.69mmol) in THF-EtOH (5:1, v/v, 18 ml) was added 4N NaOH (0.84ml, 3.38mmol). The reaction mixture was stirred at rt for 4h, 15 adjusted to pH 7.5 with IN HCI and extracted with CHClI-MeOH (4:1, v/v). The combined extracts were dried over MgSO 4 and concentrated to afforded 324 (218mg, 26% 2steps) as a brown amorphous foam. IR(KBr) n 3299, 3004, 1700, 1627, 1598, 1536cur'; 'H-NMR (DMSO) 5 2.25 (s, 3H), 2.36-2.62 (m, 10H), 2.84, 2.99 (each s, total 3H11), 3.13, 3.14 (each s, total 2H11), 3.38-3.45 (m, 2H), 3.61, 3.65 (each s, total 2H), 3.86 (s, 3H), 6.74 (t, 1H, J=7.8H11z), 6.87 (s, 1H11), 6.93 (t, 1H, 20 J=7.8Hz), 7.11-7.17 (m, 2H), 7.79 (d, 1H, J=7.8Hz), 8.01 (d, 1H, J=7.8H1z), 8.47 (s, 1H), 8.57 (s, 1H1); MS (FAB) m/z 498 (M +1); Anal. Calcd for C 2 6I 3 5
N
5 0,2HCl-H 2 0: C, 53.06; H, 6.67; N, 11.89. Found: C, 53.04; H, 6.15; N, 11.09. Example 275 1 -[2-[N-methyl-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]lphenyl]acetamido] ethyl] -4 25 piperidinylacetic acid N NCOOH Me H H 6Me 325 To a stirred solution of 2-(N-benzyloxycarbonyl-N-methylamino)acetaldehyde (2.07 g, 10.0 mmol) and ethyl 4-piperidinylideneacetate (1.69 g, 10.0 mmol) in MeOH-AcOH (10:1, v/v, 22 ml) was added NaBH 3 CN (1.32 g, 20 mmol) and the stirring was continued overnight. The 30 mixture was quenched by addtion of sat. NaHCO 3 (200 ml) and extracted with CHCl 3 (3 x 150 ml). The combined extracts were dried over MgSO 4 and evaporated. The residue was crhomatographed on silica-gel with CHCl 3 -EtOH (40:1, v/v) to give ethyl 1-[2-(N-benzyloxycarbonyl-N 464 WO 01/00206 PCT/US00/18079 methylamino) ethyl]-4-piperidinylideneacetate (1.71 g, 47%) as a colorless oil. 'H-NMR (CDCI 3 ) 8 1.25 (t, J= 7.3 Hz, 3 H), 2.16 (m, 2 H), 2.57 (m, 4 H), 2.95 (m, 7 H), 3.44 (m,2 H), 4.13 (q, J= 7.3 Hz, 2 H), 5.12 (s, 2 H), 5.49-5.53 (m, 1 H), 7.35 (m, 5 H). A solution of ethyl 1-[2-(N-benzyloxycarbonyl-N-methylamino)ethyl]-4-piperidinylidene 5 acetate (1.70 g, 4.72 mmol) in EtOH-AcOH (20:1, v/v, 21 ml) was hydrogenated over 5% Pd/C (2 g) for 3 days with stirring. The mixture was filtered and the filtrate was concentrated in vacuo. The residue was made basic with sat. NaHCO 3 and extracted with CHCl 3 (300 ml). The extract was dried over Na 2
CO
3 and evaporated to give ethyl 1-(2-methylaminoethyl)-4-piperidinylacetate (813 mg, 75%) as a yellow oil. 'H-NMR (CDC 3 ) 8 1.25 (t, J= 7.3 Hz, 3 H), 1.68-1.81 (m, 5 H), 10 1.97 (t, J= 11.2 Hz, 2 H), 2.22 (d, J= 7.3 Hz, 2 H), 2.43-2.47 (m, 5 H), 2.66 (t, J= 6.4 Hz, 2 H), 2.85-2.90 (m, 2 H), 4.13 (q, J= 7.3 Hz, 2 H). To a stirred solution of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (550 mg, 1.75 mmol) and ethyl 1-(2-methylaminoethyl)-4-piperidinylacetate (400 mg, 1.75 mmol) in DMF (10 ml) were added EDC-HCI (503 mg, 2.63 mmol), HOBt (cat.), and DMAP (cat.) and the 15 stirring was continued overnight. The mixture was diluted with EtOAc (300 ml), washed with brine (200 ml), dried over MgSO 4 , and evaporated. The residue was crhomatographed on silica gel with CHCl 3 -EtOH (10:1, v/v) to give ethyl 1-[2-[N-methyl-N-[3-methoxy-4-[N'-(2 methylphenyl) ureido]phenyl]acetamido]ethyl]-4-piperidinylacetate (697 mg, 76%) as a yellow gum. 'H-NMR (CDCl 3 ) 8 1.19-2.06 (series of m, 12 H), 2.21 (t, J = 7.8 Hz, 2 H), 2.28 (s, 3 H), 20 2.41 (t, J= 7.3 Hz, 1 H), 2.46 (t, J= 7.3 Hz, 1 H), 2.80-2.89 (m, 2 H), 2.95 and 3.01 (s, each, total 3 H), 3.40 and 3.50 (t, J= 6.8 Hz, each, total 2 H), 3.64-3.75 (m, 5 H), 4.09-4.16 (m, 2 H), 6.59 (s, 1 H), 6.77-6.79 (m, 2 H), 7.12 (t,J= 7.3 Hz, 1 H), 7.21-7.27 (m, 3 H), 7.54 (d,J= 8.3 Hz, 1 H), 8.06 (dd, J= 8.3, 2.4 Hz, 1 H). To a stirred solution of ethyl 1-[2-[N-methyl-N-[3-methoxy-4-[N'-(2-methylphenyl) 25 ureido]phenyl]acetamido]ethyl]-4-piperidinylacetate (690 mg, 1.32 mmol) in THF (11 ml) was added 0.25 N aq. NaOH (11 ml, 2.75 mmol) and the stirring was continued overnight. The mixture was diluted with H 2 0 (50 ml), neutralized with 1 N HC1, and extracted with CHCl 3 -MeOH (2:1, v/v, 3 x 100 ml). The combined extracts were dried over MgSO 4 and evaporated. The residue was dissolved in MeOH (50 ml) and activated carbon (2 g) was added to this solution. The 30 suspension was refluxed for 30 min with stirring and filtered through Celite. The filtrate was evaporated and the residue was triturated by taking up CHCl 3 and adding hexane until a precipitate fomed. This precipitate was collected and dried in vacuo to give 325 (75 mg, 11%) as a white 465 WO 01/00206 PCT/US00/18079 amorphous solid. 'H-NMR (DMSO) 8 1.19-2.99 (series of m, total 17 H), 3.32-3.43 (m, 4 H), 3.62-3.65 (m, 2H), 3.86 (s, 3 H), 6.73 (t, J= 8.3 Hz, 1 H), 6.87 (s, 1 H), 6.93 (t, J= 7.8 Hz, 1 H), 7.11-7.17 (m, 2 H), 7.79 (d,J= 8.3 Hz, 1 H), 8.01 (d,J= 8.3 Hz, 1 H), 8.47 (s, 1 H), 8.57 (s, 1 H); MS-FAB m/z 497 (M'+1); Anal. Calcd for C 27
H
36
N
4 0 5 s-HCl: C, 60.84; H, 7.00; N, 10.51. Found: 5 C, 60.97; H, 7.14; N, 10.17. Example 276 1-[2-[N-methyl-N-[4-[N'-(2-methylphenyl)ureido]phenyl]acetamido]ethyl]-4-piperidinylacetic acid N N No COOH Me H H 326 To a stirred solution of ethyl 1-(2-methylaminoethyl)-4-piperidinylacetate (400 mg, 1.75 10 mmol) and Et 3 N (366 ul, 2.63 mmol) in DMF (10 ml) was added pentafluorophenyl 4-(N'-(2 methylphenyl)ureido)phenylacetate (788 mg, 1.75 mmol) and the stirring was continued overnight. The mixture was diluted with EtOAc (300 ml), washed with brine (200 ml), dried over MgSO 4 , and evaporated. The residue was crhomatographed on silica-gel with CHCl 3 -EtOH (10:1, v/v) to give ethyl 1-[2-[N-methyl-N-[4-[N'-(2-methylphenyl)ureido]phenyl]acetamido]ethyll-4-piperidiny 15 acetate (630 mg, 73%) as a colorless oil. To a stirred solution of ethyl 1-[2-[N-methyl-N-[4-[N'-(2-methylphenyl)ureido]phenyl] acetamido]ethyl]-4-piperidinylacetate (630 mg, 1.27 mmol) in THF (10 ml) was added 0.25 N aq. NaOH (10 ml) and the stirring was continued overnight. The reaction mixture was diluted with
H
2 0 (100 ml), neutralized with 1 N HC1, and extracted with CHCl 3 -MeOH (2:1, v/v, 3 x 100 ml). 20 The combined extracts were dried over MgSO 4 and evaporated. The residue was triturated by taking up CHCl 3 and adding hexane until precipitate formed. This precipitate was collected and dried in vacuo to give 326 (20 mg, 3%) as a white amorphous solid. 'H-NMR (DMSO)8 1.69 (m, S H), 2.15 (m, 4 H), 2.24 (s, 2 H), 2.50 (m, 2 H), 2.83 and 2.99 (s, each, total 3 H), 3.32-3.49 (m, 4 H), 3.63 (d, J= 6.8 Hz, 2 H), 6.91-6.95 (m, 1 H), 7.13 (m, 4 H), 7.39 (d, J= 8.3 Hz, 2 H), 7.83 (d, 25 J= 7.3 Hz, 1 H), 7.97 (m, 1 H), 9.12 (m, 1 H); MS (FAB): m/z 467 (M +1). Example 277 4-[2-N-[4-[N'-(2-methylphenyl)ureido]phenyl]-N-methylacetamido]ethyl-1-piperazinylacetic acid NN0,_,COOH N N< Me H H 327 To a solution of ethyl 4-(2-methylaminoethyl)-1-piperazinylacetate (700mg, 3.05mmol), 30 Et 3 N (0.64ml, 4.58mmol) and DMAP (75mg, 0.61mmol) in THF (15ml) was stirred for 30min at 466 WO 01/00206 PCT/US00/18079 rt, then 4-[N'-(2-methylphenyl)ureido]phenylacetic acid (917mg, 3.05mmol), HOBt (82mg, 0.61mmol) and EDC-HCI (879mg, 4.58mmol) was added to the reaction mixture which was stirred for 12h at rt. The reaction mixture was diluted with EtOAc, which was washed with brine, dried over Na 2
SO
4 , and concentrated to dryness. Chromatography of the residue with CHCl 3 -MeOH 5 (10:1, v/v) afforded ethyl 4-[2-N-[4-[N'-(2-methylphenyl)ureido]phenyl]-N-methylacetamido]ethyl 1-piperazinylacetate (996mg, 66%) as a yellow amorphous foam. 'H-NMR (CDCl 3 ) 8 1.25-1.29 (m, 3H), 2.20 (s, 3H), 2.47-2.58 (m, 10H), 2.97, 3.05 (each s, total 3H), 3.17, 3.20 (each s, total 2H), 3.45, 3.52 (each d, total 2H, J=6.8Hz), 3.64, 3.68 (each s, 2H), 4.15-4.21 (m, 2H), 7.01-7.19 (m, 8H), 7.48 (m, 1H), 7.64 (m, 1H); MS (FAB) m/z 496 (M+1). 10 To a stirred solution of ethyl 4-[2-N-[4-[N'-(2-methylphenyl)ureido]phenyl]-N-methyl acetamido]ethyl-1 -piperazinylacetate (996mg, 2.0 1 mmol) in THF-EtOH (5:1, 12 ml) was added 4N NaOH (1.0ml, 4.00mmol). The reaction mixture was stirred at rt for 4h, adjusted to pH 7.5 with IN HCI and extracted with CHCl 3 -MeOH (4:1, v/v). The combined extracts were dried over MgSO 4 and concentrated to afforded 327 (73mg, 8%) as a yellow amorphous foam. IR(KBr) n 15 3338, 2925, 2850, 2821, 1704, 1627, 1540cm'; 'H-NMR (DMSO) 8 2.24 (s, 3H), 2.33-2.61 (m, 10H), 2.82, 2.95 (each s, total 3H), 3.00, 3.02 (each s, total 2H), 3.39 (t, 2H, J=6.8Hz), 3.60, 3.62 (each s, total 2H), 6.92 (t, 1H, J=7.8Hz), 7.09-7.16 (m, 4H), 7.41-7.44 (m, 2H), 7.76, 7.77 (each d, 2H, J=7.8Hz), 8.46, 8.53 (each s, 1H), 9.54, 9.59 (each s, 1H); MS (FAB) m/z 468 (M+1); Anal. Calcd for C 25
H
3 3 NsOs-2HCl: C, 55.56; H, 6.53; N, 12.96. Found: C, 54.99; H, 6.45; N, 11.58. 20 Example 278 4-[2-N-[4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenyl]-N-methylacetamido]ethyl- 1 piperazinylacetic acid N N< (N, COOH H H OMe 328 To a solution of ethyl 4-(2-methylaminoethyl)-1-piperazinylacetate (695mg, 3.03mmol), 25 Et 3 N (0.64ml, 4.58mmol) and DMAP (75mg, 0.61mmol) in DMF (15ml) was stirred for 15min at rt, then 4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenylacetic acid (965mg, 3.03mmol), HOBt (82mg, 0.61mmol) and EDC-HCI (872mg, 4.54mmol) was added to the reaction mixture which was stirred for 12h at rt. The reaction mixture was diluted with EtOAc, which was washed with brine, dried over Na 2
SO
4 , and concentrated to dryness. Chromatography of the residue with CHCI 3 30 MeOH (10:1, v/v) afforded ethyl 4-[2-N-[4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenyl]-N methyl acetamido]ethyl-1-piperazinylacetate (1.21g, mixture of DMF) as a black oil. 'H-NMR (CDCl 3 ) 8 1.24-1.29 (m, 3H), 2.45-2.59 (m, 10H), 2.98, 3.05 (each s, total 3H), 3.17, 3.20 (each s, 467 WO 01/00206 PCT/US00/18079 total 2H), 3.44, 3.52 (each t, total 2H, J=6.8Hz), 3.66 (s,2H), 4.15-4.21 (m, 2H), 6.77-6.78 (m, 2H), 6.79-7.11 (m, 3H), 7.68-7.95 (m, 2H11), 7.64 (broad s, 1H), 8.20 (t, 1H, J=7.8Hz); MS (FAB) m/z 530 (M'+I). To a stirred solution of ethyl 4-[2-N-[4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenyl]-N 5 methyl acetamido]ethyl-1-piperazinylacetate (1.21g, mixture of DMF) in THF-EtOH (5:1, v/v, 12 ml) was added 4N NaOH (1.0ml, 4.00mmol). The reaction mixture was stirred at rt for 4h, adjusted to pH 7.5 with IN HCI and extracted with CHCl 3 -MeOH (4:1, v/v). The combined extracts were dried over MgSO 4 and concentrated to afforded 328 (78mg, 5% 2steps) as a brown amorphous foam. IR(KBr) n 3299, 2940, 2830, 1704, 1627, 1598, 1536cm'; 'H-NMR (DMSO) 5 10 2.36-2.61 (m, 10H), 2.83, 2.98 (each s, total 3H11), 3.11 (s, 2H), 3.37-3.43 (m, 2H), 3.62, 3.65 (each s, total 2H), 3.85 (s, 3H), 6.75 (m, 1H), 6.87 (s, 1H), 6.98 (s, 1H1), 7.12 (t, 1H, J=7.8Hz), 7.20, 7.23 (each d, 2H, J=7.8Hz), 8.01 (d, 1H, J=7.8Hz), 8.17 (t, 1H, J=7.8Hz), 8.72 (s, 1H), 9.19 (s, 1H); MS (FAB) m/z 502 (M+1); Anal. Calcd for C 2 1 32
FN
5 0 5 s-2HCl-0.5H 2 0: C, 51.46; H, 6.05; N, 12.00. Found: C, 51.08; H, 5.69; N, 11.27. 15 Example 279 3-fluoro-1 -[2-N-methyl-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl]acetamido]ethyl-4 piperidinylacetic acid IF N N N COOH Me H H OMe 329 To a stirred solution of 1-tert-butoxycarbonyl-4-piperidone (14.9 g, 74.8 mmol) in DMF 20 (35 mL) was added TMSC1 (11.4 mL, 89.7 mmol) and then Et 3 N (25.0 mL, 179 mmol) dropwise at room temperature, and the reaction mixture was heated at 80 oC for 18 hr. Hexane was added to the reaction mixture, and the resulting mixture was washed with sat. NaHCO 3 and brine, dried over Na 2
SO
4 , and concentrated to dryness. Chromatography of the residue with hexane - EtOAc (5: 1, v/v) as eluent gave 1-tert-butoxycarbonyl-1,2,3,6-tetrahydro-4-(trimethylsilyloxy)pyridine 25 (20.4 g, 99 %) as a yellow oil. 'H-NMR (CDC13) 5 0.19 (s, 9H11), 1.46 (s, 9H11), 2.05 -2.15 (m, 2H), 3.48 - 3.56 (m, 2H), 3.83 - 3.91 (m, 2H), 4.79 (broad s, 1H1). To a solution of 1-tert-butoxycarbonyl-1,2,3,6-tetrahydro-4-(trimethylsilyloxy)pyridine (20. 4 g, 75.0 mmol) in CH 3 CN (500 mL) was added SelectfluorTM (29.2 g, 82.5 mmol) at room temperature, and the reaction mixture was stirred for 2 hr. EtOAc was added to the reaction 30 mixture, and the mixture was washed with brine, dried over Na 2
SO
4 , and concentrated to dryness. Chromatography of the residue with CHC1 3 - MeOH (6 : 1, v/v) as eluent gave 1-tert 468 WO 01/00206 PCT/US00/18079 butoxycarbonyl-3-fluoro-4-piperidone (14.5 g, 89 %) as a colorless oil. 'H-NMR (CDCl 3 ) 5 1.50 (s, 9H), 2.44 (t, J = 6.9 Hz, 1H), 2.48 - 2.63 (min, 2H), 3.26 (ddd, J= 13.5, 10.5, 3.9 Hz, 1H), 3.72 (t, J= 6.9 Hz, 1H), 4.16 (min, 1H), 4.42 (min, 1H11), 4.83 (dt, 49.2, 6.9 Hz, 1H). To a solution of triethyl phosphonoacetate (3.72 g, 16.6 mmol) in THF (70 mL) was 5 added lithium bis(trimethylsilyl)amide (1.0M THF solution, 15.5 mL, 15.5 mmol) at -78 oC. After being stirred at the same temperature for 1 hr, 1-tert-butoxycarbonyl-3-fluoro-4-piperidone (3.02 g, 13.9 mmol) was added to the reaction mixture. The mixture was stirred for 30 min at the same temperature, quenched by the addition of sat. NH 4 CI solution and extracted with EtOAc. The extracts were washed with brine, dried over Na 2
SO
4 , and concentrated to dryness. 10 Chromatography of the residue with hexane - EtOAc (8 : 1, v/v) as eluent gave ethyl (1 butoxycarbonyl-3-fluoropiperidin-4-yliden)acetate (3.23 g, 81%) as a colorless solid. 'H-NMR 5 1.30 (t,J= 7.1 Hz, 3H), 1.48 (s, 9H11), 2.10 (min, 111), 2.56 (min, 1H), 2.77 (mn, 1H), 3.13 -3.54 (inm, 2H), 3.70 (min, 1H), 4.17 4.18 (each q, J= 7.1 Hz, total 2H), 5.82 5.98 (each s, total 1H), 6.41 (each d, J = 46.9 Hz, total 1H11); MS (FAB) m/z 288 (M'+1). 15 A solution of ethyl (1-butoxycarbonyl-3-fluoropiperidin-4-yliden)acetate (1.32 g, 4.59 mmol) in THF (30 mL) was hydrogenated over Pd-C (TMEDA complex, 66.0 mg) at room temperature for 2 hr under hydrogen atmosphere. The catalyst was filtered off and the filtrate was concentrated to dryness. Chromatography of the residue with hexane - EtOAc (9 :1, v/v) as eluent gave ethyl 1-tert-butoxycarbonyl-3-fluoro-4-piperidinylacetate (653 mg, 73 %) as a colorless oil. 20 'H-NMR (CDCl 3 ) 8 1.26 (t, J= 7.1 Hz, 3H), 1.45 (s, 9H11), 1.53 - 1.79 (min, 2H), 1.92 - 2.09 (inm, 21H1), 2.31 (dd, J= 16.4, 6.9 Hz, 1H), 2.52 (dd, J= 16.4, 7.3 Hz, 1H), 2.61 -3.06 (mn, 2H), 4.14 (q, J = 7.1 Hz, 2H1), 4.28 - 4.77 (min, 2H); 13 C NMR (CDCl 3 ) 14.29, 25.80, 28.42, 35.99, 36.20, 60.55, 79.78, 86.72. 88.48, 154.94, 171.93; FAB-MS m/z 290 (M*+1). To a solution of ethyl 1-tert-butoxycarbonyl-3-fluoro-4-piperidinylacetate (653 mg, 2.26 25 mmol) in CH 2 C1 2 (10 mL) was added TFA (5 mL) at 0 oC. After being stirred at room temperature for 4 hr, the reaction mixture was concentrated. The residue was taken up with sat. NaHCO 3 solution and extracted with THF - EtOAc (1 : 1, v/v). The extracts were dried over MgSO 4 and concentrated to afford ethyl 3-fluoro-4-piperidinylacetate (420 mg, 98 %) as a yellow oil. 'H-NMR
(CDCL
3 ) 6 1.26 (t, J= 7.4 Hz, 3H), 1.68 - 1.80 (mn. 2H), 2.18 (mn, 1H1), 2.32 (dd, J= 16.6, 6.8 Hz, 30 1H), 2.54 (dd, J= 16.6, 7.5 Hz, 1H), 2.82 (min, 1H), 2.90, 3.00 (each d, J= 14.4 Hz, total 1H), 3.31 (min, 1H), 3.51 (min, 1H11), 4.15 (q, J= 7.4 Hz, 2H11), 4.82, 4.71 (each broad s, total 1H11). 469 WO 01/00206 PCT/US00/18079 To a solution of ethyl 3-fluoro-4-piperidinylacetate (230 mg, 1.22 mmol) and 2-(N-tert butoxy carbonyl-N-methylamino)acetaldehyde (211 mg, 1.22 mmol) in THF (5 mL) was added NaBH(OAc) 3 (386 mg, 1.82 mmol) and acetic acid (70.0 mL, 1.22 mmol) at room temperature After being stirred for 24 hr, the reaction mixture was quenched by the addition of sat. NaHCO 3 5 solution and extracted with EtOAc. The extracts were washed with brine, dried over Na 2
CO
3 , and concentrated to dryness. Chromatography of the residue with CHCl 3 - MeOH (6 : 1, v/v) as eluent gave ethyl 3-fluoro-1l-[2-(N-tert-butoxycarbonyl-N-methylamino)ethyl]-4-piperidinylacetate (236 mg, 56 %) as a reddish oil. 'H-NMR (CDC13) 8 1.25 (t, J= 7.1 Hz, 3H), 1.45 (s, 9H), 1.53 1.79 (m, 2H), 1.90 - 2.09 (m, 2H), 2.15 (dd, J= 16.4, 7.1 Hz, 1H), 2.45 -2.58 (m, 2H), 2.87 (s, 3H), 10 2.90 - 2.99 (m, 2H), 3.20 (m,1 H), 3.24 - 3.45 (m, 2H), 4.14 (q, J= 7.1 Hz, 2H), 4.61, 4.73 (each broad s, 1H); ESI-MS m/z 347. To a solution of ethyl 3-fluoro-1-[2-(N-tert-butoxycarbonyl-N-methylamino)ethyl]-4 piperidinylacetate (236 mg, 0.68 mmol) in CH 2 C 2 (10 mL) was added TFA (5 mL) at 0 oC. After being stirred at room temperature for 4 hr, the reaction mixture was concentrated. The residue 15 was taken up with sat. NaHCO 3 solution and extracted with CHCl 3 . The extracts were dried over MgSO 4 and concentrated to afford ethyl 3-fluoro-1-[2-(N-methylamino)ethyl]-4-piperidinylacetate (117 mg, 70%) as a reddish oil. 'H-NMR (CDCl 3 )6 1.26 (t, J= 7.1 Hz, 3H), 1.58 (m ,1H), 1.70 (m, 1H), 1.99 (m, 11), 2.14 (m,1H), 2.27 -2.33 (m, 1H), 2.47 (s, 3H), 2.48 - 2.56 (m, 4H), 2.72 (t,J= 6.3 Hz, 2H), 2.90 (m, 1H), 3.16 (m, 1H), 4.14 (q,J= 7.1 Hz, 2H), 4.67 (d,J = 48.3 Hz, 20 1H); ESI-MS m/z 247 (M'+1). To a solution of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (164 mg, 0.52 mmol), ethyl 3-fluoro-1-[2-(N-methylamino)ethyl]-4-piperidinylacetate (117 mg, 0.47 mmol), Et 3 N (0.10 mL, 0.71 mmol), and HOBt (13.0 mg, 0.09 mmol) in THF (5 mL) was added EDC-HCl (137 mg, 0.71 mmol). After being stirred at room temperature for 8 hr, the reaction mixture was 25 diluted with water and extracted with EtOAc. The extracts were washed with brine, dried over Na 2
SO
4 , and concentrated to dryness. Chromatography of the residue with toluene - acetone (1: 2, v/v) as eluent gave ethyl 3-fluoro-l1-[2-N-methyl-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenyl]acetamido]ethyl-4-piperidinylacetate (160 mg, 62 %) as a yellow amorphous solid. 'H NMR (CDC1 3 ) 8 1.24 - 1.28 (m, 3H), 1.51 - 2.23 (m 7H), 2.26 (s, 3H), 2.35 (s, 2H), 2.39 - 2.56 30 (m, 3H), 2.88 (m, 1H), 2.95, 3.03 (each s, total 3H), 3.11 (m, 1H), 3.44 (m, 1H), 3.56 (m, 1H1), 3.64 (s, 2H), 3.68 (s, 3H), 4.11 - 4.17 (m, 2H), 4.57, 4.69 (each s, total 1H), 6.72 - 6.82 (m, 2H), 7.10 - 7.33 (m, 5H), 7.54 (d, J= 8.1 Hz, 1H), 8.06 (d, J= 8.1 Hz, 1H); ESI-MS m/z 543 (M++1). 470 WO 01/00206 PCT/US00/18079 To a solution of ethyl 3-fluoro-1-[2-N-methyl-N-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenyl]acetamido]ethyl-4-piperidinylacetate (160 mg, 0.29 mmol) in THF (3 mL) was added 0.25N NaOH (1.30 mL, 0.32 mmol). After being stirred at room temperature for 1 hr, the reaction mixture was concentrated. The residue was diluted with water and neutralized with IN 5 HCI at 0 oC. The mixture was concentrated and purified by ion-exchanged resin (HP-20, Mitsubishi Chemical) to give 329 (110 mg, 74 %) as a yellow amorphous solid. IR (KBr) 3343, 2937, 1700, 1617, 1589, 1535, 1486, 1455, 1417 cm'; 1 H-NMR (CD 3 OD) 8 1.57 - 1.72 (m,2H), 1.98 (m, 1H), 2.26 (m, 1H), 2.28 (s, 3H), 2.37 - 2.59 (m, 3H), 2.66 - 2.89 (m, 2H), 2.95, 3.09 (each s, total 3H), 3.14 (m, 1H), 3.40 (m, 1H), 3.51 (m, 1H), 3.59 (m, 1H), 3.71 (s, 2H), 3.78 (m, 10 1H), 3.89 (s, 3H), 4.69, 4.81 (each s, total 1H11), 6.79 (dd, J= 8.1, 1.5 Hz, 1H), 6.90 (broad s, 1H), 7.01 (t, J= 7.8 Hz, 1H), 7.13 -7.19 (m, 2H), 7.58(d, J= 7.8Hz, 1H),8.00 (d,J= 8.1 Hz, 1H); ESI-MS m/z 515 (MW+1); Anal. Calcd for C 27
H
35
FN
4 0,H 2 0: C, 60.89; H, 7.00; N, 10.52. Found: C, 61.09; H, 6.80; N, 9.87. Example 280 15 4-[2-N-[2-(4-fluorophenoxy)ethyl]-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl] acetamido]ethylpiperazinyl-1-acetic acid O F N N- rN, COOH me H H (Me 330 To a solution of 2-(N-benzyl-N-tert-butoxycarbonylamino)ethanol (6.85 g, 27.3 mmol), 4 fluorophenol (3.07 g, 27.3 mmol) and PPh 3 (7.83 g, 30.0 mmol) in THF (100 mL) was added 20 DIAD (6.00 mL, 30.0 mmol) at room temperature After being stirred for 3 hr, the reaction mixture was concentrated. Chromatography of the residue with hexane - EtOAc (8 : 1, v/v) as eluent gave 1-[2-(N-benzyl-N-tert-butoxycarbonylamino)ethoxy]-4-fluorobenzene (8.19 g, 64 %) as a yellow oil. 'H-NMR (CDCl 3 )8 1.42 - 1.50 (m, 9H), 3.41 - 3.67 (m, 2H), 3.92 - 4.11 (m, 2H), 4.51 - 4.63 (m, 2H), 6.73 - 6.85 (m, 2H), 6.89 - 6.98 (m, 2H), 7.24 (m 5H); FAB-MS m/z 346 25 (MW+1). To a solution of 1-[2-(N-benzyl-N-tert-butoxycarbonylamnino)ethoxy]-4-fluorobenzene (8.19 g, 23.7 mmol) in CH 2 Cl 2 (50 mL) was added TFA (40 mL) at 0 oC. After being stirred at room temperature for 1 hr, the reaction mixture was concentrated. The residue was taken up with sat. NaHCO 3 solution and extracted with CHC1 3 . The extracts were washed with brine, dried over 30 Na 2
SO
4 , and concentrated to give 1-(2-N-benzylaminoethoxy)-4-fluorobenzene (4.38 g, 75 %) as a reddish oil. 'H-NMR (CDCl 3 )8 3.00 (t, J= 5.2 Hz, 2H), 3.87 (s, 2H), 4.04 (t, J 5.2 Hz, 2H), 6.80 471 WO 01/00206 PCT/US00/18079 - 6.85 (min, 2H), 6.92 - 6.98 (min, 2H), 7.23 - 7.36 (min, 5H); FAB-MS m/z 246 (M'+l). A mixture of 1-(2-N-benzylaminoethoxy)-4-fluorobenzene (1.08 g, 4.40 mmol), ethyl 4 (2-bromoethyl)piperazinyl-l-acetate (1.23 g, 4.40 mmol), and K 2
CO
3 (0.61 g, 17.9 mmol) in
CH
3 CN (50 mL) was heated under reflux for 8 hr. The resulting mixture was filtered and the 5 filtrate was concentrated to dryness. Chromatography of the residue with toluene - acetone (3 : 1, v/v) as eluent gave ethyl 4-[2-N-benzyl-N-[2-(4-fluorophenoxy)ethyl]amino]ethylpiperazinyl-1 acetate (1.51 g, 77 %) as a reddish oil. 'H-NMR (CDC13)8 1.27 (t, J= 7.1 Hz, 3H), 2.31 -2.66 (min, 10H), 2.75 (t, J = 6.6 Hz, 2H), 2.90 (t, J= 6.1 Hz, 2H1), 3.18 (s, 2H), 3.72 (s, 2H), 3.97 (t, J= 6.1 Hz, 2H11), 4.17 (q, J= 7.1 Hz, 2H), 6.75 - 6.79 (m, 2H), 6.91 - 6.96 (min, 2H), 7.21 - 7.35 (inm, 10 5H); FAB-MS m/z 444 (M+1). A solution of ethyl 4-[2-N-benzyl-N-[2-(4-fluorophenoxy)ethyl]amino]ethylpiperazinyl-1 acetate (1.50 g, 3.38 mmol) in EtOH (30 mL) was hydrogenated over 5 % Pd-C (53.1% wet, 0.73 g) under hydrogen atmosphere for 4 hr. The catalyst was filtered off and the filtrate was concentrated. The residue was taken up with sat. NaHCO 3 solution and extracted with CHCl 3 . 15 The extracts were washed with brine, dried over Na 2
SO
4 , and concentrated to afford ethyl 4-[2-N [2-(4-fluorophenoxy)ethyl]amino]ethylpiperazinyl-1-acetate (1.12 g, 94%) as a reddish oil. 'H NMR (CDCl 3 ) 8 1.27 (t, J= 7.1 Hz, 3H), 1.81 (broad s, 1H11), 2.47 - 2.66 (min, 12H), 3.00 (t, J= 5.4 Hz, 2H), 3.20 (s, 211), 4.03 (t, J= 5.4 Hz, 2H11), 4.18 (q, J= 7.1 Hz, 2H), 6.81 - 6.85 (mn, 2H11), 6.90 -6.99 (m, 2H); FAB-MS m/z 354 (M*+1). 20 To a solution of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (485 mg, 1.54 mmol), ethyl 4-[2-N-[2-(4-fluorophenoxy)ethyl]amino]ethylpiperazinyl-l1-acetate (545 mg, 1.54 mmol), Et 3 N (0.32 mL, 2.32 mmol), and HOBt (41.5 mg, 0.31 mmol) in THF (15 mL) was added EDC-HCI (883 mg, 2.32 mmol) at room temperature After stirring for 24 hr, the reaction mixture was diluted with water and extracted with CHC1 3 - MeOH (10 : 1, v/v). The extracts were 25 washed with brine, dried over Na 2
SO
4 , and concentrated to dryness. Chromatography of the residue with CHCl 3 - MeOH (4 : 1, v/v) as eluent to give ethyl 4-[2-N-[2-(4-fluorophenoxy)ethyl] N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl] acetamido]ethylpiperazinyl- 1-acetate (532 mg, 53 %) as a yellow amorphous solid. 'H-NMR (CDC13)8 1.26, 1.27 (each t, J= 7.1 Hz, total 3H11), 2.27 (s, 3H), 2.51 - 2.63 (min, 10H), 3.16, 3.19 (each s, total 2H), 3.51 - 3.56 (m, 2H), 3.63, 30 3.67 (each s, total 2H), 3.69 -3.81 (mn, 5H), 3.95, 4.10 (each t, J= 5.2 Hz, total 2H), 4.16, 4.18 (each q, J = 7.1 Hz, total 2H), 6.56 (d, J = 8.1 Hz, 1H11), 6.72 - 6.78 (min, 4H), 6.89 - 6.98 (min, 2H), 7.12 (t, J= 7.6 Hz, 1H), 7.20 - 7.23 (m, 3H), 7.51 (d, J= 7.6 Hz, 1H11), 8.04 (d, J= 8.1 Hz, 1H); 472 WO 01/00206 PCT/US00/18079 FAB-MS m/z 650 (M'+1). To a solution of ethyl 4-[2-N-[2-(4-fluorophenoxy)ethyl]-N-[3-methoxy-4-[N'-(2 methylphenyl) ureido]phenyl]acetamido]ethylpiperazinyl-1-acetate (532 mg, 0.82 mmol) in dioxane (8 mL) was added dropwise 0.25N NaOH (5.00 mL, 1.25 mmol) at room temperature, and 5 the reaction mixture was stirred for 1 hr. The resulting mixture was concentrated, diluted with water, and neutralized with IN HCI at 0 'C. The mixture was extracted with CHCl 3 - MeOH (4: 1, v/v). The extracts were washed with brine, dried over Na 2
SO
4 , and concentrated to dryness. Chromatography of the residue with CHCl 3 - MeOH (3 : 1, v/v) as eluent gave 330 (168 mg, 33 %) as a pale yellow amorphous solid. IR (KBr) 3338, 2938, 2829, 1635, 1533, 1506, 1454, 1415 10 cm ; 'H-NMR (DMSO-d)8 2.25 (s, 3H), 2.37 - 2.48 (m, 6H), 2.53 - 2.67 (m, 6H), 3.09 (m, 2H), 3.44 - 3.49 (m, 2H), 3.64 - 3.69 (m, 2H), 3.72 (s, 2H), 3.80, 3.84 (each s, total 3H), 4.06 - 4.09 (m, 2H), 6.73 (m, 1H), 6.85 (s, 1H), 6.91 -6.97 (m,3H), 7.07 - 7.18 (m, 4H), 7.78 (d, J= 8.1 Hz, 1H), 8.00 (dd, J= 8.1, 2.7 Hz, 1H), 8.53 (m, 1H), 8.59 (m, 1H); FAB-MS m/z 622 (M*+1); Anal. Calcd for C 33
H
4 ,oFNsO 6 -2HCI: C, 57.06; H, 6.09; N, 10.08. Found: C, 56.83; H, 6.05; N, 9.90. 15 Example 281 4-[2-N-[2-(4-acetylphenoxy)ethyl]-N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl] acetamido]ethylpiperazinyl-1-acetic acid O Ac N N NCOOH Me H H OMe 331 To a solution of 2-(N-benzyl-N-tert-butoxycarbonylamino)ethanol (5.90 g, 23.5 mmol), 4 20 hydroxyacetophenone (3.18 g, 23.5 mmol) and PPh 3 (6.74 g, 25.8 mmol) in THF (100 mL) was added DIAD (5.20 mL, 25.8 mmol) at room temperature The reaction mixture was heated under reflux for 4 hr and concentrated. Chromatography of the residue with hexane - EtOAc (5 : 1, v/v) as eluent gave 4-[2-(N-benzyl-N-tert-butoxycarbonylamino)ethoxy]acetophenone (3.64 g, 42 %) as a yellow oil. 'H-NMR (CDC13) 8 1.43 - 1.65 (m, 9H), 2.55 (s, 3H), 3.41 - 3.69 (m, 2H), 4.02 - 4.24 25 (m, 2H), 4.49 - 4.66 (m, 2H), 6.81 - 6.93 (m, 2H), 7.19 - 7.37 (m, 5H), 7.91 (d, J= 8.8 Hz, 2H); FAB-MS m/z 370 (M +1). To a solution of 4-[2-(N-benzyl-N-tert-butoxycarbonylamino)ethoxy]acetophenone (3.05 g, 8.26 mmol) in CH 2 C1 2 (30 mL) was added TFA (20 mL) at 0 'C. After being stirred at room temperature for 2 hr, the reaction mixture was concentrated. The residue was taken up with sat. 30 NaHCO 3 solution and extracted with CHCI 3 . The extracts were washed with brine, dried over 473 WO 01/00206 PCT/US00/18079 Na 2
SO
4 , and concentrated to give 4-(2-N-benzylaminoethoxy)acetophenone (2.21 g, 99 %/) as a reddish oil. 'H-NMR (CDCl 3 ) 8 2.04 (broad s, 1H), 2.55 (s, 3H), 3.05 (t, J= 5.4 Hz, 2H), 3.88 (s, 2H), 4.15 (t,J = 5.4 Hz, 2H), 6.92 (d,J= 8.9 Hz, 2H), 7.24 -7.36 (min, 5H), 7.91 (d,J= 8.9 Hz, 2H); FAB-MS m/z 270 (M t +l). 5 To a solution of ethyl 4-(2-hydroxyethyl)piperazinyl-1-acetate (11.3 g, 52.1 mmol) and CBr 4 (20.7 g, 62.5 mmol) in CH 2 C1 2 (200 mL) was added PPh 3 (19.2 g, 73.0 mmol) portionwise at 0 oC and the reaction mixture was stirred for 30 min. Hexane was added to the mixture, the precipitates were filtered off, and the filtrate was concentrated to afford ethyl 4-(2-bromoethyl) piperazinyl-1-acetate (13.7 g, 94%) as a yellow oil. 'H-NMR (CDCl 3 )8 1.27 (t, J= 7.1 Hz, 3H), 10 2.61 -2.78 (min, 8H), 2.81 (t, J= 7.6 Hz, 2H), 3.20 (s, 2H), 3.42 (t, J= 7.6Hz, 3H), 4.18 (q, J= 7.1 Hz, 2H). A mixture of 4-(2-N-benzylaminoethoxy)acetophenone (681 mg, 2.52 mmol), ethyl 4-(2 bromoethyl)piperazinyl-l-acetate (705 mg, 2.52 mmol), and K 2 C0 3 (349 g, 2.52 mmol) in CH 3 CN (10 mL) was heated under reflux for 22 hr. The resulting mixture was filtered and the filtrate was 15 concentrated to dryness. Chromatography of the residue with toluene - acetone (1 : 1, v/v) as eluent gave ethyl 4-[2-[N-benzyl-N-[2-(4-acetylphenoxy)ethyl]amino]ethylpiperazinyl- 1-acetate (920 mg, 78 %) as a reddish oil. 'H-NMR (CDCl 3 ) 8 1.27 (t, J= 7.1 Hz, 3H), 2.45 - 2.53 (inm, 10H11), 2.55 (s, 3H), 2.76 (t,J= 6.8 Hz, 2H), 2.94 (t,J = 6.1 Hz, 2H1), 3.18 (s, 2H), 3.73 (s, 2H), 4.06 (t, J = 6.1 Hz, 2H), 4.17 (q, J = 7.1 Hz, 2H), 6.85 (d, J = 7.1 Hz, 2H11), 7.22 -7.35 (min, 5H), 20 7.90 (d, J = 7.1 Hz, 2H); FAB-MS m/z 468 (M+ 1). A solution of ethyl 4-[2-[N-benzyl-N-[2-(4-acetylphenoxy)ethyl]amino]ethylpiperazinyl-1 acetate (920 mg, 1.97 mmol) in EtOH (30 mL) was hydrogenated over 5 % Pd-C (53.1% wet, 510 mg) under hydrogen atmosphere for 4 hr. The catalyst was filtered off and the filtrate was concentrated. The residue was taken up with sat. NaHCO 3 solution and extracted with CHC1 3 . 25 The extracts were washed with brine, dried over Na 2
SO
4 , and concentrated to afford ethyl 4-[2-N [2-(4-acetyl phenoxy)ethyl]amino]ethylpiperazinyl-1-acetate (690 mg, 93%) as a reddish oil. 'H NMR (CDCl 3 ) 8 1.27 (t, J= 7.1 Hz, 3H), 2.55 (s, 3H), 2.46 - 2.54 (min, 10H11), 2.78 (t, J = 6.2 Hz, 2H), 3.04 (t, J= 5.2 Hz, 2H), 3.20 (s, 2H), 4.13 (t, J= 5.2 Hz, 2H11), 4.18 (q, J= 7.1 Hz, 2H), 6.92 (d, J = 8.8 Hz, 2H), 7.92 (d, J= 8.8 Hz, 2H); FAB-MS m/z 378 (M++1). 30 To a solution of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (558 mg, 1.77 mmol), ethyl 4-[2-N-[2-(4-acetylphenoxy)ethyl]amino]ethylpiperazinyl-1 -acetate (670 mg, 474 WO 01/00206 PCT/US00/18079 1.77 mmol), Et 3 N (0.38 mL, 2.66 mmol), and HOBt (50.0 mg, 0.35 mmol) in THF (10 mL) was added EDC-HCI (883 mg, 2.32 mmol) at room temperature After stirring for 15 hr, the reaction mixture was diluted with water and extracted with CHCl 3 - MeOH (10 : 1, v/v). The extracts were washed with brine, dried over Na 2
SO
4 , and concentrated to dryness. Chromatography of the 5 residue with CHC1 3 - MeOH (4 : 1, v/v) as eluent to give ethyl 4-[2-N-[2-(4-acetylphenoxy)ethyl] N-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenyl]acetamido]ethylpiperazinyl- 1-acetate (724 mg, 61%) as a yellow amorphous solid. 1 H-NMR (CDCl 3 ) 8 1.25, 1.27 (each t, J= 7.1 Hz, total 3H), 2.29 (s, 3H), 2.45 -2.51 (m, 8H), 2.54 (s, 3H), 2.55 -2.63 (m, 2H), 3.17, 3.19 (each s, total 2H), 3.51 - 3.55 (m, 2H11), 3.60 (s, 2H11), 3.69 (s, 3H), 3.71 - 3.78 (m, 2H), 4.04 - 4.23 (m, 4H), 10 6.47 (m, 1H, J= 8.1 Hz), 6.72 - 6.76 (m, 2H), 6.85 (d, J = 8.8 Hz, 2H), 7.12 - 7.19 (m, 2H), 7.20 - 7.24 (m, 2H), 7.51 (d, 8.1 Hz, 1H), 7.88 (d, J= 8.8 Hz, 2H11), 8.04 (d, J= 8.1 Hz, 1IH); FAB-MS m/z 674 (M+1). To a solution of ethyl 4-[2-N-[2-(4-acetylphenoxy)ethyl]-N-[3-methoxy-4-[N'-(2 methylphenyl) ureido]phenyl]acetamido]ethylpiperazinyl-1-acetate (724 mg, 1.07 mmol) in THF 15 (10 miL) was added dropwise 0.25N NaOH (6.50 mL, 1.61 mmol) at room temperature, and the reaction mixture was stirred for 1 hr. The resulting mixture was concentrated, diluted with water, and neutralized with IN HCI at 0 oC. The mixture was extracted with CHCl 3 - MeOH (4: 1, v/v). The extracts were washed with brine, dried over Na 2
SO
4 , and concentrated to dryness. Chromatography of the residue with CHCl 3 - MeOH (3 : 1, v/v) as fluent gave 331 (450 mg, 65 20 %) as a pale yellow amorphous solid. IR (KBr) 3345, 2938, 2821, 1673, 1631, 1598, 1533, 1455, 1417 cm'; 'H-NMR (DMSO-d 6 ) 5 2.25 (s, 3H), 2.35 - 2.47 (m, 8H), 2.51 (s, 3H), 2.53 - 2.58 (m, 2H), 2.96 (s, 2H), 3.46 - 3.50 (m, 2H), 3.69 (s, 2H), 3.73 - 3.77 (m, 2H), 3.80, 3.83 (each s, total 3H), 4.19 -4.22 (m, 2H), 6.73 (m, 1H), 6.85 (s, 1H), 6.92 (t,J= 7.3 Hz, 1H), 7.30 (d, J= 7.3 Hz, 2H), 7.10 -7.16 (m, 2H), 7.79 (d, J= 8.3 Hz, 1H), 7.90 -7.95 (m, 2H), 8.00 (t, J= 8.3 Hz, 1H), 25 8.55 (m, 1H11), 8.61 (m, 1H); FAB-MS m/z 646 (M+1); Anal. Calcd for C 35 1H 43
N
5 0 7 -2HCl-lH 2 0: C, 57.06; H, 6.43; N, 9.51. Found: C, 59.17; H, 6.32; N, 9.61. Example 282 4-[2-N-[4-[AN'-(2-bromophenyl)ureido]3-methoxyphenyl]-N-benzylacetamido] ethyl-1 piperidinylacetic acid NI N'COOH 30 r H H (Me 332 methyl 4-[2-N-[4-[N'-(2-bromophenyl)ureido]l-3-methoxyphenyl]-N-benzylacetamido]ethyl- 1 475 WO 01/00206 PCT/US00/18079 piperidinylaceate N N N NCOOMe Br H H OMe 333 A mixture of piperidine ethanol (10.0 g, 77.4 mmol), benzyl 2-bromoacetate (17.8 g, 77.6 mmol) and K 2 C0 3 (21.4 g, 155 mmol) in CH 3 CN (200 ml) was heated under reflux with 5 stirring for 2 h. The insoluble solid was removed by filtration, and the filtrate was concentrated in vacuo. The residue was poured into ice cooled 1N-HCI and extracted with CHCl 3 . The organic layer was washed with water, drying over anhydrous Na 2
SO
4 , and concentrated in vacuo. The residue was chromatographed on silica gel with EtOAc as eluent to give benzyl 4-(2 hydroxyethyl)-l-piperidinylacetate (16.3 g, 76%) as a colorless oil. 'H-NMR (CDCl 3 )5 1.31-1.40 10 (m, 3 H), 1.52 (dd, J= 12.8, 6.3 Hz, 2H), 1.68 (brd, J= 10.0 Hz, 3 H), 2.16 (t, J= 11.6 Hz, 2 H), 2.92 (brd, J= 11.6 Hz, 2 H), 3.24 (s, 2 H), 3.69 (t,J= 6.8 Hz, 2 H), 5.16 (s, 2 H), 7.35 (m, 5 H). To a stirred solution of benzyl 4-(2-hydroxyethyl)-1-piperidinylacetate (14.9 g, 53.8 mmol) in CH 2 C1 2 (150 ml) was added Et 3 N (37.5 ml, 269 mmol) and DMSO (41.6 ml, 538 mmol). The reaction mixture was cooled to 0 oC and SO 3 Py (25.7 g, 161 mmol) was added portion wise, 15 and the resulting mixture was stirred at rt overnight. The mixture was concentrated in vacuo, and the residue was diluted with water, followed by extracted with Et 2 0. The organic layer was washed with water, dried over anhydrous Na 2
CO
3 and concentrated in vacuo. The residue was chromatographed on silica gel with EtOAc-hexane (2 : 1) as eluent to give (4-N-carbobenzyloxy methylpiperidinyl)acetaldehyde (4.63 g, 31%) as a colorless oil. 'H-NMR (CDC13) 8 1.36-1.46 (m, 20 2H), 1.69(brs, 2H), 1.72(brs, 1 H), 1.90(m, 1 H),2.21 (dt, J= 11.6,2.4Hz, 2H),2.37(dd, J= 6.8, 1.6 Hz, 2 H), 2.92 (d,J= 11.6 Hz, 2 H), 3.25 (s, 2 H), 5.16 (s, 2 H), 7.35 (m, 5 H), 9.77 (t,J= 2.0 Hz, 1 H). To a stirred solution of N-benzylamine (1.06 ml, 9.75 mmol) in MeOH (20 ml) was added AcOH (560 ml, 9.75 nmmol) and (4-N-carbobenzyloxymethylpiperidinyl)acetaldehyde (1.79 g, 25 6.50 mmol) in MeOH (5 ml) and cooled to 0 oC. NaBH 3 CN (645 mg, 9.75 mmol) was added in one portion, and the resulting mixture was stirred overnight at rt. The mixture was concentrated in vacuo, and the residue was poured into aq.NaHCO 3 , then extracted with CHC 3 . The organic layer was washed with water, dried over anhydrous Na 2
SO
4 and concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH-EtOAc (10 : 1 : 1) as eluent to give N 30 benzyl-2-[4-(N-carbobenzyloxymethylpiperidinyl)]ethylamine (872 mg, 37%) as a colorless oil. 476 WO 01/00206 PCT/US00/18079 1 H-NMR (CDCI 3 ) 5 1.36-1.46 (m, 2 H), 1.69 (brs, 2 H), 1.72 (brs, 1 H), 1.90 (m, 1 H), 2.21 (dt, J = 11.6, 2.4 Hz, 2 H), 2.37 (dd, J= 6.8, 1.6 Hz, 2 H), 2.92 (d, J= 11.6 Hz, 2 H), 3.25 (s, 2 H), 5.16 (s, 2 H), 7.35 (m, 5 H), 9.77 (t, J= 2.0 Hz, 1 H). To a stirred solution of N-benzyl-2-[4-(N-carbobenzyloxymethylpiperidinyl)]ethylamine 5 (356 mg, 0.972 mmol), 4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (369 mg, 0.972 mmol) and N,N-dimethylaminopyridine (119 mg, 0.972 mmol) in DMF (15 ml) was added EDCHCI (372 mg, 1.94 mmol) at rt, and the resulting mixture was stirred overnight. The reaction mixture was poured into water and extracted with EtOAc. The organic layer was washed with brine and dried over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was 10 chromatographed on silica gel with CHCl 3 -MeOH (10 : 1) as eluent to give benzyl 4-[2-N-[4-[N' (2-bromophenyl )ureido]3-methoxyphenyl]-N-benzylacetamido]ethyl-1-piperidinylacetate (611 mg, 86%) as a colorless oil. 'H-NMR (CDCl 3 ) mixture of rotamars 5 1.15-2.09 (series of m, 7 H), 2.06 2.14 (m, 2 H), 2.84-2.96 (m, 2 H), 3.17-3.23 (s, total 2 H), 3.21 and 3.23(s, total 2 H), 3.38-3.42 (m, 1 H), 3.65 and 3.73 (s, total 2 H), 3.83 and 3.84 (s, total 3 H),4.49 (s, 1 H), 4.60 (s, 1 H), 5.15 15 (d, J= 2.8 Hz, 2 H), 6.74-7.83 (series of m, 16 H), 7.51-7.53 (m, 1 H), 7.94-8.02 (m, 1 H), 8.18 (d, J = 8.4 Hz, 2 H); MS (FAB) m/z 727 (M), 729 (M+2). To a stirred solution of benzyl 4-[2-N-[4-[N'-(2-bromophenyl)ureido]3-methoxyphenyl] N-benzylacetamido]ethyl-1-piperidinylacetate (347 mg, 0.477 mmol) in MeOH-H 2 0 (5: 1, 6 ml) was added LiOH (13.6 mg, 0.57 mmol) at rt, and the resulting mixture was stirred for overnight. 20 The reaction mixture was poured into water and the solution was neutralized with aq. 1N-HC1, then extracted with CHC 3 . The organic layer was washed with brine and dried over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHC13-MeOH (10 : 1) as eluent to give 332 (97.3 mg, 32%) as a colorless amorphous solid, and 333 (168 mg, 54%) as a colorless amorphous solid, respectively. 332 'H-NMR (CD 3 OD), mixture of rotamars 8 25 1.29-1.90 (series of m, 7 H), 2.75-2.86 (m, 2 H), 3.28-3.51 (series of m, 6 H), 3.74 and 3.78 (s, total 2 H), 3.87 and 3.89 (s, total 3 H), 4.66 (s, 1 H), 4.85 (s, 1 H), 6.79-7.00 (series of m, 3 H), 7.16 (d, J= 7.2 Hz, 1 H), 7.23-7.37 (m, 5 H), 7.57 (dd, J= 8.4, 1.2 Hz, 1 H), 7.88-8.00 (series of m, 2 H); MS (FAB) m/z 637 (Me), 639 (M*+2). 333 'H-NMR (CDCl 3 ), mixture of rotamars 5 1.15-2.11 (series of m, 7 H), 2.89-2.97 (m, 2 H), 3.17-3.49 (series of m, 6 H), 3.17 and 3.18 (s, 30 total 3 H), 3.70 and 3.71 (s, total 3 H), 3.83 and 3.84 (s, total 2 H), 4.49, 4.60 and 4.71 (s, total 2 H), 6.75-8.16 (series of m, 14 H); MS (FAB) m/z 651 (M), 653 (M+2). Example 283 4-[2-N-[4-[N'-(2-chlorophenyl)-3-methoxyureido]phenyl]-N-benzylacetamido]ethyl-1-piperidinyl 477 WO 01/00206 PCT/US00/18079 acetic acid N N N - NCOOH C HH COMe 334 To a stirred solution of N-benzyl-2-[4-(N-carbobenzyloxymethylpiperidinyl)]ethylamine (372 mg, 1.11 mmol), 4-[AN'-(2-chlorophenyl)ureido]-3-methoxyphenyl]acetic acid (758 mg, 1.11 5 mmol) and N,N-dimethylaminopyridine (136 mg, 1.11 mmol) in DMF (15ml) was added EDCHCI (426 mg, 2.22 mmol) at rt, and the resulting mixture was stirred for 12 h. The reaction mixture was poured into water and extracted with EtOAc. The organic layer was washed with brine and dried over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (10 : 1) as eluent to give benzyl 4-[2-N-[4-[N'-(2-chlorophenyl)-3 10 methoxyureido]phenyl]-N-benzylacetamido]ethyl-l-piperidinylacetate (668 mg, 88%) as a pale yellowish oil. 'H-NMR (CDCl 3 ) mixture of rotamars 8 1.15-1.83 (series of m, 7 H), 2.04-2.13 (m, 2 H), 2.83-2.95 (m, 2 H), 2.88 and 2.99 (s, total 2 H), 3.20 and 3.23 (s, total 2 H), 3.20-3.23 (overlap, 1 H), 3.38-3.42 (m, 1 H), 3.65-3.74 (m, 3 H), 4.50 (s, 1 H), 4.61 (s, 1 H), 5.14 (d,J= 4.4 Hz, 2 H), 6.72-6.82 (series of m, 2 H), 6.94-6.99 (m, 1 H), 7.13-7.50 (series of m, 14 H), 7.94-8.02 15 (m, 1 H), 8.18 (d, J= 8.0 Hz, 2 H); MS (FAB) m/z 683 (M'+H). To a stirred solution of benzyl 4-[2-N-[4-[AN'-(2-chlorophenyl)-3-methoxyureido]phenyl] N-benzylacetamido]ethyl-1-piperidinylacetate (633 mg, 0.93 mmol) in MeOH-H 2 0 (10 : 1, 10 ml) was added LiOH (24.4 mg, 1.02 mmol) at rt, and the resulting mixture was stirred overnight. The reaction mixture was poured into water and the solution was neutrallized with aq. 1N-HC1, then 20 extracted with CHC1 3 . The organic layer was washed with brine and dried over anhydrous Na 2
SO
4 , then concentrated in vacuo. The residue was chromatographed on silica gel with CHC1 3 -MeOH (20 : 1) as eluent to give methyl 4-[2-N-[4-[AN'-(2-chlorophenyl)-3-methoxyureido]phenyl]-N benzylacetamido]ethyl-1-piperidinylacetate (504 mg, 89%) as a colorless amorphous solid. 'H NMR (CDCl 3 ), mixture of rotamars 6 1.26-1.64 (series of m, 7 H), 2.02-2.10 (m, 2 H), 2.86 (t, J= 25 10.4 Hz, 2 H), 3.17 (d, J= 8.0 Hz, 2 H), 3.16-3.22 (m, overlap, 1 H), 3.40 (t, J= 7.6 Hz, 1 H), 3.48 (s, 1 H), 3.65 and 3.73 (s, total 2 H), 3.70 and 3.71 (s, total 3 H), 3.79 and 3.80 (s, total 3 H), 4.50, 4.60 and 4.70 (s, total 2 H), 6.73-6.85 (series of m, 2 H), 6.98 (t, J= 7.6 Hz, 1 H), 7.13-7.37 (series of m, 9 H), 7.96 (m, 1 H), 8.18 (d, J= 8.0 Hz, 2 H); MS (FAB) nm/z 607 (M +H). To a stirred solution of methyl 4-[2-N-[4-[N'-(2-chlorophenyl)-3-methoxyureido]phenyl] 30 N-benzylacetamido]ethyl-1-piperidinylacetate (177 mg, 0.292 mmol) in MeOH-H 2 0 (5: 1, 6 ml) 478 WO 01/00206 PCT/US00/18079 was added LiOH (21.6 mg, 0.90 mmol), and the resulting mixture was stirred for 4 h at rt. The mixture was poured into water, and the solution was neutrallized with aq. 1N-HCI, then extracted with CHCl 3 . The organic layer was washed with brine and dried over anhydrous Na 2
SO
4 , then concentrated to yield 334 (155 mg, 92%) as a colorless amorphous solid. 'H-NMR (CD 3 OD) 5 mixture of rotamars 5 1.28-1.90 (series of m, 7 H), 2.84 (brs, 2 H), 3.30-3.52 (series of m, 6 H), 3.74 and 3.82 (s, total 2 H), 3.87 and 3.88 (s, total 3 H), 4.63 (s, 1 H), 4.66 (s, 1 H), 6.79-7.05 (series of m, 3 H), 7.16 (d, J= 7.2 Hz, 1 H), 7.24-7.40 (min, 5 H), 7.88 (s, 1 H), 7.98-8.04 (series of m, 2 H); MS (ESI) m/z 593 (M), 615 (M+Na ). It should be understood that while this invention has been described herein in terms of 10 specific embodiments set forth in detail, such embodiments are presented by way of illustration of general principles, and the invention is not necessarily limited thereto. Modifications and variations in any given material or process step will be readily apparent to those skilled in the art without departing from the true spirit and scope of the following claims, and all such modifications are included within the scope of the present invention. 479

Claims (30)

1. A compound represented by Formula I, or a salt thereof, w Wi W N N R M H H 5 wherein W is chosen from aryl group, substituted aryl group, heteroaryl group and substituted heteroaryl group; W' is chosen from arylene group, substituted arylene group, heteroarylene group and substituted heteroarylene group; 10 A is chosen from =0O, =S and =NH; R is chosen from a direct bond, alkyenylene group and -(CH 2 ).-, wherein n is chosen from 1 and 2; X is chosen from -C(0)-, -CH 2 - and S(0)2; 15 M is chosen from 3R 4 R~ R4 IN Y--Z-A R5 wherein is a divalent 4-, 5-, 6- or 7-membered heterocyclic moiety, wherein the nitrogen atom is the point of attachment to X; 20 R' , R 2 and R 3 are independently chosen from -H, -OH,-NH 2 , halogen atom, alkyl group, substituted alkyl group, aryl group, substituted aryl group, alkoxy group, substituted alkoxy group, monoalkylamino group, substituted monoalkylamino group, dialkylamino group, substituted dialkylamino group, cycloalkylanmino group, substituted cycloalkylamino group, 25 alkylsulfonylamino group, substituted alkylsulfonylamino group, 480 WO 01/00206 PCT/US00/18079 arylsulfonylamino group, substituted arylsulfonylamino group, aryloxy group, substituted aryloxy group, heteroaryloxy group, substituted heteroaryloxy group, benzyloxy group and substituted benzyloxy group, or 5 two of R 1 , R 2 and R 3 taken together may form a 3-, 4-, 5-, 6-, or 7-membered carbocyclic or heterocyclic residues optionally substituted with from 1 to 3 substituents chosen independently from -OH, halogen atom, -NH 2 , alkyl group, alkoxy group, aryl group, aryloxy group, alkylamino group, benzyloxy group and heteroaryl group; 10 R 4 is chosen from -H and lower alkyl group; Y is a direct bond or a divalent radical chosen from -C(O)-, -C(O)NH-, alkenylene group, alkynylene group and -(CH 2 )kY , wherein k is chosen from 1, 2 and 3; and 15 Y2 is a direct bond or a divalent radical chosen from -0-, -S-, -S(O), -S(O) 2 - and -NY 3 -, wherein Y3 is chosen from -H and lower alkyl group; Z is chosen from arylene group, substituted arylene 20 group, heterocyclylene group, substituted heterocyclylene group, cycloalkylene group and substituted cycloalkylene group; A ' is a direct bond or a divalent radical chosen from alkenylene group, alkynylene group, -(CH 2 ) t - and -O(CH 2 ), 25 wherein t is chosen from 1, 2 and 3; and v is chosen from 0, 1, 2, and 3; and H H Rs is chosen from -OH, lower alkoxy group, -N(H)OH, N N and H H N N4 481 WO 01/00206 PCT/US00/18079 R R 7 wherein is a divalent 4-, 5-, 6- or 7--membered heterocyclic moiety, wherein the nitrogen atom is the point of attachment to X; 5 R 6 and R 7 are independently chosen from -H, -OH, halogen atom, alkyl group and alkoxy group; Y1 is a divalent radical chosen from -0-, -S-, -S(O)-, -S(O) 2 - and -NY 4 -, wherein Y 4 is chosen from -H and lower alkyl group; 10 Z is a divalent radical chosen from arylene group, substituted arylene group, heterocyclylene group, substituted heterocyclylene group, cycloalkylene group and substituted cycloalkylene group; A 2 is a direct bond or a divalent radical chosen from alkenylene group, alkynylene group and -(CH 2 )e 15 wherein e is chosen from 1, 2 and 3; and H H R is chosen from -OH, lower alkoxy group, -N(H)OH, NI , and H H N N 1- (CHR 9 )qRo 10 20 wherein 482 WO 01/00206 PCT/US00/18079 L is 0 wherein Q. / is a divalent 4-, 5-, 6- or 7-membered heterocyclic moiety, optionally substituted with from I to 3 substitutents 5 chosen independently from alkyl group, alkoxy group, hydroxyalkyl group, -OH, benzyloxy group, -NHz, halogen atom, aryl group and heteroaryl group, said moiety may be fused to 1 or 2 additional carbocyclic or heterocyclic residues optionally substituted with from 1 to 3 substitutents chosen 10 independently from alkyl group, aryloxy group, alkoxy group, hydroxyalkyl group, -OH, benzyloxy group, -NH 2 ,, halogen atom, aryl group and heteroaryl group; m and q are independently chosen from 0, 1, 2 and 3; Xi is chosen from -CH= and -N=; 15 R 9 is chosen from -H and lower alkyl group; H H R 1 is chosen from -COOH, lower alkoxycarbonyl group, O H H H N- and N- ; and Z 2 is chosen from -H, COOH and lower alkoxycarbonyl group; and -R 1 -_Z3-Q 2 -L 1 20 wherein 11 N- N-)1 R 1 is chosen from -0-, K " -N and -NR 12 wherein 483 WO 01/00206 PCT/US00/18079 R 1 2 is chosen from -H, alkyl group, substituted alkyl group, cycloalkyl group, substituted cycloalkyl group, aryl group, substituted aryl group, benzyl group, substituted benzyl group, lower alkenyl group, substituted lower alkenyl group and lower 5 alkynyl group the left hand bond is the point of attachment to -X- and the right hand bond is the point of attachment to -Z3; Z 3 is chosen from a direct bond, a divalent aliphatic hydrocarbon moiety having 1 to 12 carbon atoms, 10 wherein one or more carbon atoms may be replaced with -0- or -NR 13 wherein R 13 is chosen from -H and lower alkyl group, and 15 one or more hydrogen atoms attached to an aliphatic carbon atom may be replaced with lower alkyl group; and wherein x is chosen from 0 and 1; 20 y is chosen from 1, 2, and 3; and R is chosen from -H, -OH and halogen atom, , , and, when R i l 1 is -NR12 , I-Z 4 -1 wherein 14a 25 Z 4 is chosen from wherein Rl 4 a is chosen from -H, -OH, lower alkyl group 484 WO 01/00206 PCT/US00/18079 and halogen atom; / - and , wherein the left hand bond is the pointof attachment to R" 1 and 5 the right hand bond is the point of attachment to Q2; Q2 is a divalent radical chosen from arylene group, substituted arylene group, heterocyclylene group, substituted heterocyclylene group, cycloalkylene group, substituted cycloalkylene group, R156 wherein R 15 and R 16 are independently chosen 10 from -H, halogen atom and lower alkyl group; and K7 18 17 1 1 R wherein R and R1 are independently chosen from -H, lower alkyl group, substituted lower alkyl group and lower alkenyl group; and LI is chosen from -COOH and -COOR 19 15 wherein R is a lower alkyl group.
2. A compound according to claim 1, or a salt thereof, wherein M is R 3 -N Y-Z-A R 5
3. A compound according to claim 1, or a salt thereof, wherein M is > Z tA2LR8 20 485 WO 01/00206 PCT/US00/18079
4. A compound according to claim 1, or a salt thereof, wherein M is I(CHR 9 )qR 10
5. A compound according to claim 1, or a salt thererof, wherein M is --R11_ZQ2-L1 5
6. A compound according to claim 2, or a salt thereof, wherein at least one radical of R 1 , R 2 and R 3 is -OH or halogen atom.
7. A compound according to claim 6, or a salt thereof, wherein A is =0O, R is -(CH 2 )n and X is -C(O)-.
8. A compound according to claim 7, or a salt thereof, wherein Y is chosen from 10 alkenylene group, alkynylene group and -(CH 2 )kY 2 ; Y 2 is chosen from a direct bond, -0-, -S(O) and -NY 3 -; and Y 3 is -H.
9. A compound according to claim 8, or a salt thereof, wherein Y is chosen from -0- and -NY 3 .
10. A compound according to claim 2, or a salt thereof, wherein W is unsubstituted 15 phenyl group or phenyl group having one or two substituents chosen from lower alkyl group and halogen atom at the ortho positions thereof.
11. A compound according to claim 10, or a salt thereof, wherein W 1 is unsubstituted phenylene group or phenylene group having a substituent chosen from methoxy group, lower alkyl group and halogen atom at the ortho position to the -NH- thereof. 20
12. A compound according to claim 2, or a salt thereof, wherein W 1 is phenylene group having a substituent chosen from methoxy group, lower alkyl group and halogen atom at the ortho position to the -NH- thereof and having 1 to 3 substituents chosen from lower alkyl group and halogen atom. 486 WO 01/00206 PCT/US00/18079
13. A compound according to claim 2, or a salt thereof, wherein A is a direct bond or -(CH2)t- .
14. A compound according to claim 13, or a salt thereof, wherein A 1 is a direct bond.
15. A compound according to claim 14, or a salt thereof, wherein R 5 is -OH. 5
16. A compound according to claim 2, or a salt thereof, wherein W is unsubstituted phenyl group or phenyl group having one or two substituents chosen from lower alkyl group and halogen atom at the ortho positions thereof; W 1 is unsubstituted phenylene group or phenylene group having a substituent chosen from methoxy group, lower alkyl group and halogen atom at the ortho position to the -NH- thereof; R is -CH 2 -; X is -C(O)- and R 5 is -OH. 10
17. A compound according to claim 16, or a salt thereof, chosen from the group consisting of O WH 0 / / 0 H H O H O H O NH OH NH OH N OH N O0 H - " H MeO H O N H ON H N H H 0 0 COOH Nk N O CO 2 H NR O02 H H 487 WO 01/00206 PCT/US00/18079 0~%N N NR -aC2H H H . 0 lH H \/ O H0 2 H 0 N N H H 0 0 2 H CO0 2 H NN N N jNN ,H H H H~r 0 2 H C0 2 H ,?- C02H 0 NN o C 2 '-- 'C 2 N N ?H HHH 5H H H H 00 NNN -Q- 0 0\H HD C0 2 H 0'q ON -pIl- 0 2 Hj 0:c N C02 H H qN N 0 C0 ci H H 0 ci 0 Z NY~ NN 11,N 0o 0 H H CO0 H NfN N~I C02H H Hj aC2 0 2 N H1 488 WO 01/00206 PCT/US00/18079 ~ ~ N F 0 O 0 F H H 0O CO2 H H H O HO2C 00 -o SN NJ N o2 0 02H CO 2 S 0 N CO2HCO 2 H , -H H HNZ 0 F N MeN N H H N llNq HN C0 2 HY O&C OBn N -C ,? H H /.O \OC0 2 H 5 0 2 H 02 PH N OH H- N O :OCO 2 H N N O N 2 H H 0 e H H CO 2 H F N N 0 V 0 \/ NCO 2 H H H OH C 489 WO 01/00206 PCT/US00/18079 ci OH 2HNN COD J N ,CNQ2H ,?- H~ N 'I N~N ~ 'NK9-C H H 0 O 2 H M H H N 1N N ?NN-\j~CO 2 H NC2 F HMI H N~K N!&-' 1 CO 2 HH N N'J:; ( N'NN- C2 I H'4-jrC 2 H 1 ycH H 0 MeNMe C2 N H2 N -NN'N NNN- C2 Me H- N 2 (IY Mel' I 2 02 Y'llq Q-0H 00 H - Me ~H Me N NN I IH2 -?eNNN H Me N2 ?H H Me2 490 WO 01/00206 PCT/US00/18079 F COOH N NN COOH N N O H COOH N O N OH Me OMe Me OMe NQN OOH ~NN OH 5 OH OOH H OMe H H OMe I COOH COOH I '?MN5OH NF 5 r 1 e~ Me ~e H 6Me F N 0 -QCOOH H H MeMe~O HM He Me LMeOH N~~O C O N N ICOOH N OOH MeH H Me I Me Q O49 (NXN 0' OOH N~~&o H H H Me 'N:UUH MeH H 491 WO 01/00206 PCT/US00/18079 OOH NXN H Q OOH H(M H H Me NH Me QCOOH N ND OH H2 NH N N OOH JNxq)o-O COOH OOH N OOHMe FF N OOH N N N COOH I H H Me I H H Me OOH MON N N2OFOH JN )o OOH (NIN a OOH H Me M eF M ecXC H O H Me H H Me N N OOH lLNIN OOH H1MeNe H OMe 492 WO 01/00206 PCT/US00/18079 F ' N re OOH OOH OOHQ OOH Q H HOOMe H H H He OOH OH Me ?OMeHOOH II o COOH - COOH AN / COOH CO OH 5O OHCMe MO H Oe 0 M OOH OOH N N4 O O H 9 N 3 O O eN/I N Me NCH e Me NO O H -O COH -QCH N N O N ) O l / COACH l oe\ /COCH N J TN HIr N CO HfN% H \ H H CC M~e H Me' H Me 9 ~ N LN M~ / c O O H N1 HNN MC / O O H NN/ COOH / OOH rMeH H me H H Me 493 WO 01/00206 PCT/US00/18079 N I OOH N N'1: NOOH CC ~ OOO H O 5OOH OOH Me H H Me F I N OOH N OOH clMe IMe I'I - OO NC_ COOH N N COOH No N'N COOH \NNCOOH r H H me Ol Oe 5 BrH H OMe Ore H COH e HN OMe 00 N N I Nj CO /I 'aCOOH IHH Me MerHH Me O~e 494 WO 01/00206 PCT/US00/18079 t-ObB_/ OOH q~~X YOCOOH CH HOMe ~ 1 H H Me Me K M rHH Me COO 4 3 COOH r--C O c~HH M Me / M N K~HH le -(QCOOH NJN~ aoKDCOOH MH H M eM I lo&COOH 4:a~-QCOOH MeH Me MHHMe Me M I >O-OCOOHOH Me HOHOM 495 WO 01/00206 PCT/US00/18079 OO OOH -H H e r H H Me /m M ?H MeN O-Q-COOH "N' I/ OOH ..-- HMe H H9 e Y me Me N'J K-COH J oQCOOH NOON MeH I Me H H Me F M N NH Me NI'- Ia COOH N Nle aCOOH ?Me H HI H Me N F Nlo~aOOH O ?r HIMe H H-9 Me C0 ~HHMeN~ HO rOOHZ N N ~ YN rI KFI H H H - m49M WO 01/00206 PCT/US00/18079 FF F F/ NJ '?N H H ~ 1 HHMe F NF NI O-Q-COOH Q a CH ?~~ Me H H HH FM H F O- C O L N '- N O ~ COOH H Me H H I ~N~ Q a-COOH KNqNLN OOH ~rH H H H r e F H I r-JC O HI 497 WO 01/00206 PCT/US00/18079
18. A compound according to claim 17, or a salt thereof, chosen from the group consisting of N O H / N N\ Me H 0 HN-4 O C N 00 H HN N 0 2 H 0 N N N OOC (H H H H 5 CO 2 H CO 2 H ClI C02 02 498 0 N?-H 0 2 H NN 0 H H HH, N H H 1:0 N 2 2 0 0QHC 2 H H H0 2 02HN N Njp YO 498 WO 01/00206 PCT/US00/18079 KNY NQ2 -02H H HH H H 2 N H0 2 C N 'I N- " 00 Me H H 0 0? O e C0 2 HC02 NkN rN'N% H~ H 0 2 H N H H- A F ~NNN N r a H 2 N 0 NNN H H H H 50 H 02 oC2 9 H H H 0 2 H H( N oN C0 2 H H MeH H 0 N NN 0\C 2 H NIHJ )ZC 2 H HfjY H P 499 WO 01/00206 PCT/US00/18079 0 NY?" H NH/ H~ HH N CO 2H N NC r H HH N _ - NCO2H N'N 02H ( .1H H H H*er F N N N I0 2 H H 02 NH 2 N N 0 H Me NH 2 0 2 H NNMe N. 0N' rO2H Me Me 5 1 N ~ - c HNL. e N'O H C O O H 5 e N ,H 2 e 0 N" H Me O Q 7 0 COOH MCOOH OH COOH ON, OH A H O~N H2HM IMee H H (Me Q " L " O NN 0 QQ C00 NXNQOOH Me H OMe CI H OMe N -OH F -N C O O H e Br H H OMe 500 WO 01/00206 PCT/US00/18079 YN IN 0O ?NN[ ~ NZ _CO HO O OHH M eN NO O N NI NCH N ZN 0 -COOH Y # K eOQ-COOH 9N 9CJCOOH 5 -H H KVCOOH N NCOOH N I N COOH NNjYQ& COOH NN r<NlDNo H 2 "y N CIeH H 6Me _XOH Ij H H 6Me _CO bH Me 0 COOH N-I% M N OOH 5 eMe 6rMe N NO N N OOH NCOOH COOH e Me MeHH501 50 WO 01/00206 PCT/US00/18079 H OMe _COOH M H H MeOOH N~H H Me )OOH I OOH 5f ~ No OOH N N' N OO N N OOH H H Me N N OOH N OOH H H Me Me H OMe N NNOOH NN &HCOOH IH H OMe IH H Me H OOH 9Th N'N 1 0N-Q:-COOH N5N rN-9-COOH 5 eH H Me I H H Me N N No" Q' NXN-Q No H Me \ 2 OOH N COOH Q1X N 15 OH ~r H Me LI3COOH ieH H Me H -- OH OO N-Nyr OOOH eO HQH ~e H Me MeH OMe H OOH IOOH CO clH H OMe Me H H 502 WO 01/00206 PCT/US00/18079 lo-Q-COOH N , CO (KN'N \ /COOH 'Y eMe H H Me Y:.H H Me ,NX NN S I /COOH \ / O 6Me MeH Me N!H N' Me ~eOOH 0 I XN I\/ OOH MeH H Me -l HOH MeOH N N a C (>0H H H-? HOO H(Nk .C 5 Me Br H H Me lo-Q-COOH ? NH H Me N COOH A NI~r~% M OOH Br H H 6Me rHH Me e N0 Me e N OOH ANCO Me MeH 503 WO 01/00206 PCT/US00/18079 Me Me Me NO -GCOOH 0N N N COOH IN QJ H M M N, H ?#N N- COOH NXN /COOH N N COOH H Me H Me r 1 Me LAN'N N .OQ_ -COOH N N kqo-CCOOH eMe Me Nl N o a OO - C NM O-C COOH ? N' O- C- COOH H Me H Me N I NI aC 'OH NNN 4 -&~COOH M~HH Me riH H Me NNI NI&OO (?CAJ 4:aCe Me M 504 WO 01/00206 PCT/US00/18079 v' N T IN -&OOH I' O MeMe MeMe o Q cl H ~~H Me eHHm rII~H Me F~j -C~C 2 0 OOH ? NJ 0Me OOH Me Hr e me 0 0 N' OaOOH N'N" NoaOOH ciH me M~e H H 0M N Nf~p" I -aCOOHN I N?" NO0HOO 5 H H0I H H e ?Me e N Me~ M M e505 WO 01/00206 PCT/US00/18079 N CO NH HOO F F N O COOH N N II O-C COOH F F r He H H I. F F N -NO C COOH N N COOH C OHN N C O O H H H CO HH I F o-CJ-a COOH H H H F F 10
19. A compound according to claim 2, or a salt thereof, wherein R 5 is lower alkoxy group. 506 WO 01/00206 PCT/US00/18079 AMENDED CLAIMS [received by the International Bureau on 24 November 2000 (24.11.00); new claims 20-30 added; other claims unchanged (1 page)]
20. A method of inhibiting cell adhesion in a mammal, said method comprising administering to said mammal an effective amount of a compound according to any of claims 1 to 19.
21. A method of treating a condition associated with VLA-4 mediated cell adhesion in a mammal, aid method comprising administering to said mammal an effective amount of a compound according to any of claims 1 to 19.
22. A method according to claim 21, wherein the condition associated with VLA-4 mediated cell adhesion is chosen from inflammatory responses, autoimmune responses and tumor metastasis.
23. A method according to claim 21, wherein the condition associated with VLA-4 mediated cell adhesion is chosen from asthma, diabetes, arthritis, psoriasis, multiple sclerosis, inflammatory bowel disease and transplantation rejection.
24. Use of a compound according to any of claims 1-19 for therapy.
25. Use of a compound according to any of claims 1-19 in the manufacture of a medicament for the treatment of a condition associated with VLA-4 mediated cell adhesion.
26. A pharmaceutical composition comprising as a therapeutic agent, a compound according to any of claims 1 to 19 and a pharmaceutically acceptable carrier or excipient.
27. A pharmaceutical composition according to claim 26, further comprising one or more additional therapeutic agents.
28. A pharmaceutical composition according to claim 27, wherein said one or more additional therapeutic agents are chosen from the group consisting of antiinflammatory, antirheumatic, corticosteroid, immunosuppressive, antipsoriatic, bronchodilator, antiasthmatic and antidiabetic agents.
29. A pharmaceutical composition according to claim 28, wherein one of said one or more additional therapeutic agents is an antiinflammatory agent.
30. A pharmaceutical composition according to claim 29, wherein said antiinflammatory agent is chosen from a steroid and an NSAID. 507
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