AU781438B2 - VLA-4 inhibitor compounds - Google Patents

VLA-4 inhibitor compounds Download PDF

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Publication number
AU781438B2
AU781438B2 AU59031/00A AU5903100A AU781438B2 AU 781438 B2 AU781438 B2 AU 781438B2 AU 59031/00 A AU59031/00 A AU 59031/00A AU 5903100 A AU5903100 A AU 5903100A AU 781438 B2 AU781438 B2 AU 781438B2
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Australia
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ooh
mixture
group
chosen
mmol
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AU5903100A (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 Inc
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Publication of AU781438B2 publication Critical patent/AU781438B2/en
Assigned to PHARMACOPEIA DRUG DISCOVERY, INC., DAIICHI PHARMACEUTICAL CO., LTD. reassignment PHARMACOPEIA DRUG DISCOVERY, INC. Alteration of Name(s) of Applicant(s) under S113 Assignors: DAIICHI PHARMACEUTICAL CO., LTD., PHARMACOPEIA, INC.
Anticipated expiration legal-status Critical
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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
    • C07D207/08—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 with hydrocarbon radicals, substituted by hetero atoms, attached to ring carbon atoms
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    • C07D207/18—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 one double bond between ring members or between a ring member and a non-ring member
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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
    • C—CHEMISTRY; METALLURGY
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    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
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    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
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    • C07D401/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a chain containing hetero atoms as chain links
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    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
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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, oa43 1 integrin, also known as VLA-4. Compounds of the present 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 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, 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 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.
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.
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, Cell, 69:11 (1992)). There are at least 16 different a subunits (aI-ca, aL, a, a aX. aIb, c v) and at least 9 different P (P3IP9) WO 01/00206 PCT/US00/18079 subunits. Integrins are divided into sub-families, based upon the P subunit. Leukocytes express a number of different integrins, including caP3 1 asp, aP a07, LP 2, axP 2 and aVP3.
a4P3 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 various inflammatory disorders (Helmer, Ann. Rev. Immunol., 8:365 (1990)). a4,3 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 a4Pi/VCAM-1 pathway (Lobb et al., J. Clin. Invest., 94:1722-28 (1994)). In a 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. Exy.
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 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 (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 IgGI 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)).
Another approach has used three-dimensional crystallographic structures of VCAM-1 fragments to synthesize cyclic peptide antagonists that closely mimicked the ax, integrin binding loop in domain 1 of VCAM-1. Synthetic VCAM-I peptide CQIDSPC, was able to inhibit the adhesion of VLA-4expressing cells to purified VCAM-I (Wang et al., Proc. Natl. Acad. Sci. USA, 92:5714 (1995)).
An additional strategy is to block the binding of a4Pi to its other counter receptor, that is, an alternatively spliced region of fibronectin containing the connecting segment-I (CS-1) motif 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 s allografts (Molossi et al., J. Clin. Invest., 95:2601 (1995)). Each of these studies provide evidence that selective inhibition of c 4 f3/VCAM-1 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 io exhibiting VLA-4 inhibitory activity in in vitro binding assays, none of the described compounds have exhibited efficacy in oral 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 S 15 pathologies associated with leukocyte migration and adhesion.
SUMMARY OF THE INVENTION SThe compounds of the present invention selectively inhibit the binding of ligands to c4o3 and therefore, are useful for inhibition, prevention and suppression of VLA-4mediated cell adhesion and the pathologies associated with that adhesion, such as, for example, inflammation, asthma, arthritis, diabetes, autoimmune responses, multiple sclerosis, psoriasis, transplantation rejection, and tumor metastasis.
According to a first aspect, there is provided a compound represented by Formula I, or a salt thereof,
C
C
A
H H wherein W is a substituted or unsubstituted phenyl W' is a substituted or unsubstituted divalent group of phenyl, pyridine, pyrrolidine or thiazole, A is =S or =NH; R is -(CH 2 wherein n is 1 or 2; X is M is chosen from the following groups
R
2
R
3 R R 4 QR O NY-Z-A
R
N
wherein
Q_
is a divalent 5- or 6- membered heterocyclic radical, 10 with the nitrogen atom being in the position of its attachment to X; :wherein Q represents -CH 2 or S2 3
R
2 and R 3 are independently chosen from the group consisting of-H, -OH, quinolinyloxy, -NH 2 mono- or dialkylamino, alkylsulfonylamino, arylsulfonylamino, naphthalylsulfonylamino, dialkylamino substituted naphthalylsulfonylamino, Ci-C 6 alkyl, benzyloxymethyl, halogen, phenyl, CI-C 4 alkoxy, phenyloxy, naphthyloxy, and phenyloxy substituted with COOH or halogen; or two of adjacent R R 2 and R 3 taken together may form an alkylene group having 1 to 5 carbon atoms or alkylenedioxy group having 1 to 3 carbon atoms optionally substituted with from 1 to 3 alkyl group(s); 20 R 4 represents hydrogen or lower alkyl; Y is chosen from a bond, a C 2
-C
8 alkenylene group, alkynylene group, and -(CH 2 )k-Y 2 wherein k is chosen from 0, 1, 2 and 3; and Y2 represents a direct bond or a divalent radical chosen from
-S(O)
2 and -NY wherein
Y
3 is independently chosen from hydrogen and lower alkyl; Z represents C 3
-C
8 cycloalkylene, phenylene, pyridylene optionally substituted with halogen, piperidine, piperazine, phenyl, or phenyl independently substituted with one or two group(s), chosen from alkyl, alkoxy, halogen, [R:\LIBFF]12694.doc:HJG 3b amino, mono- and dialkylsubstituted amino, acylamino, nitro, carboxy; A' is a direct bond or represents bond, (CH 2 or alkynyl, wherein t is chosen from 1, 2, 3; NH NH NH NH N N or N-N
R
5 represents OH or lower alkoxy; and -Rl 3 Q2_ L 1 wherein
N
V N \N-4 R is chosen from and -NR 12 wherein S. R 12 is chosen from lower alkyl optionally substituted with alkyl, 3-6cycloalkyl, hydroxy, phenyl (optionally substituted with amino or nitro), amino, cyclopropylamino, dialkylamino, diallylamino, methoxymethylamino, 15 morpholino, dihalogen substituted pyrrolidino or 4-alkylpiperidino; lower alkenyl; lower alkynyl and phenyl; S* Z 3 is chosen from a divalent aliphatic hydrocarbon moiety having 1 to 12 carbon atom,
R
wherein x is 0 or 1; y is 1,2 or 3;
R
1 4 is -H or-OH; I R:\LIBFF] I 2694doc:HJG
N
Ny and {4 provided that Z 4 is chosen from the following three formulated divalent groups in the limited case of R 1 is -NR 1 2 14 Ra
N
wherein 4 Ra is -H or halogen; N Q2 represents
R
7 R and R' are independently -H or a lower alkyl; 0 I0s or phenylene which may be substituted with halogen, alkoxy, amino, *mono or dialkylamino, acylamino, piperidino, nitro or carboxy; R F 1 9 SL' is chosen from -COOH and -COOR' 9 wherein
R'
9 is a lower alkyl.
According to a second aspect, there is provided a method of inhibiting cell adhesion in a mammal, said method comprising administering to said mammal an effective amount of a compound according to the first aspect.
According to a third aspect there is provided a method of treating a condition associated with VLA-4 mediated cell adhesion in a mammal, said method comprising administering to said mammal an effective amount of a compound according to the first aspect.
[R:\LIBFF] 2694.doc:HJG According to a fourth aspect there is provided use of a compound according to the first aspect for therapy.
According to a fifth aspect there is provided use of a compound according to the s first aspect in the manufacture of a medicament for the treatment of a condition associated with VLA-4 mediated cell adhesion.
According to a sixth aspect there is provided a pharmaceutical composition comprising as a therapeutic agent, a compound according to the first aspect and a pharmaceutically acceptable carrier or excipient.
There is disclosed herein a compound represented by Formula I, or a salt thereof,
A
N NW1R X M H H wherein S 15 W is chosen from aryl group, substituted aryl group, heteroaryl group and substituted heteroaryl group; S" W 1 is chosen from arylene group, substituted arylene group, heteroarylene group and substituted heteroarylene group; A is chosen from =S and =NH; WO 01/00206 PCT/US00/18079 R is chosen from a direct bond, alkyenylene group and -(CH 2 wherein n is chosen from 1 and 2; X is chosen from -CH 2 and S(O) 2 M is chosen from 2 3 -N Y-Z- A R wherein
Q-X
is a divalent 6- or 7-membered heterocyclic moiety, wherein the nitrogen atom is the point of attachment to X;
R
1 R and R 3 are independently chosen from -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, 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 two of R 1
R
2 and R 3 taken together may form a or 7-membered carbocyclic or heterocyclic residues optionally substituted with from I 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;
R
4 is chosen from -H and lower alkyl group; Y is a direct bond or a divalent radical chosen from -C(O)NH-, alkenylene group, alkynylene group and -(CH)kY 2 wherein k is chosen from 1, 2 and 3; and
Y
2 is a direct bond or a divalent radical chosen from
-S(O)
2 and -NY 3 WO 01/00206 PCT/US00/18079 wherein Y is chosen from -H and lower alkyl group; Z is chosen from arylene group, substituted arylene 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 and -O(CH 2 wherein t is chosen from 1, 2 and 3; and v is chosen from 0, 1, 2, and 3; and H
H
R
5 is chosen from -OH, lower alkoxy group, -N(H)OH, N-N and H H wherein is a divalent 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 -OH, halogen atom, alkyl group and alkoxy group; Y' is a divalent radical chosen from -S(0) 2 and -NY 4 wherein
Y
4 is chosen from -H and lower alkyl group; 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, WO 01/00206 PCT/US00/18079 alkynylene group and -(CH 2
)C
wherein e is chosen from 1, 2 and 3; and H H UN Nand R is chosen from -OH, lower alkoxy group, -N(H)OH, 1,(CHR 9 )qR' L is wherein 1<7" is a divalent 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, -NH2, 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 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; X' is chosen from -CH= and
R
9 is chosen from -H and lower alkyl group; H H 0" is chosen from -COOH, lower alkoxycarbonyl group, H H H N and U and WO 01/00206 PCT/USOO/18079
Z
2 is chosen from COOH and lower alkoxycarbonyl group; and -R 11._Z3-Q2_t1 wherein R II is chosen from V and -NR' 2 wherein
R
1 2 is chosen from 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 alkynyl group the left hand bond is the point of attachment to 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, wherein one or more carbon atoms may be replaced with or -NR 13 wherein
R
13 is chosen from -H and lower alkyl group, and 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; y is chosen from 1, 2, and 3; and
R
14 is chosen from -OH and halogen atom, 0 Se' and, when WO 01/00206 PCT/US00/18079 R" is -NR' 1 4 wherein 1-z 4 -1 wherein 14a Z4 is chosen from wherein
R'
4 a is chosen from -OH, lower alkyl group and halogen atom; and wherein the left hand bond is the point of attachment to R" 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, cycloalkylene group, substituted cycloalkylene group,
R
1 S R 16 wherein R i and R 6 are independently chosen from halogen atom and lower alkyl group; and SR wherein R' 7 and R 1 8 are independently chosen from lower alkyl group, substituted lower alkyl group and lower alkenyl group; and L' is chosen from -COOH and -COOR 19 wherein
R
9 is a lower alkyl group.
WO 01/00206 PCT/USOO/18079 R R-Z-A RS In a preferred embodiment of Formula I, M is In this embodiment, more preferred compounds are those wherein A is R is -(CH 2 and X is Y is preferably chosen from alkenylene group, alkynylene group, -(CHAY 2
-CH
2 and and more preferably, Y is 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-.
In preferred compounds of this embodiment, A is preferably =0 and A' is a direct bond or
-(CH
2 More preferred compounds are those wherein A 1 is a direct bond and R 5 is -OH.
R
2 3 Rt jR 4 Preferred compounds of Formula I, wherein M is R5and A is =0 are represented in Table 1. With respect to the representation of the lower bond is the point of attachment to -NH- and the upper bond is the point of attachment to The entry entitled
R--R
5 depicts that portion of the particular compound represented by IR-LX NA Y R' WO 01/00206 WO 0100206PCT/USOO/18079 TABLE1 .:Spectrum
WI
N
0,11 C0 2
H
WO 01/00206 PCT/USOO/18079 WO 01/00206 WO 0100206PCT/USOO/18079 11 0% C0 2
H
0
CO
2
H
l?- WO 01/00206 WO 0100206PCT/USOO/18079 ABLE.1 I Spectrum IR R: W 512 0 C0 2
H
526 532 0 O~p CO 2
H
H0 2
C
514 0 C0 2
H
500 0 C0 2
H
516 0 CO0 2
H
534 C 2 0 WO 01/00206 WO 0100206PCTIUSOO/18079 TBLE1 566 C 2 002
I-
0aCO 2
H
0-0 0K' C2 1*- Co 2
H
548 .N 0 2
H
110 562 N.N 0 C0 2
H
552 N 0 2
H
A 0 WO 01/00206 WO 0100206PCT/USOO/18079 TABLE'1 Mass.., Spectrum -W- 0 IlOCOH A 0 2
N
H
2
N
516 532 562 F--y 0 qo C0 2
H
H-0 2
C
WO 01/00206 PCT/USOO/18079 TABLE1 Spectirm 0~
CO
2
H
WO 01/00206 WO 0100206PCT/USOO/18079 TABLE.
.I.
.M Spectirum:: 566 V- F 0~ R- 2 575 V o N C 2
H
HN
553r
CO
2
H
523 0 Q 0 2
H
F 11 524 ~~0 517N 535 F oe
O
521
F
WO 01/00206 WO 0100206PCT/USOO/18079 TABLE 1 Mss Spectrum: R 0-p 0Nr: COOH -0 WO 01/00206 WO (i100206PCTIISOOII8079 q Br Me OMe WO 01/00206 WO 0100206PCTUSOO/18079 I TTBLE::: I 629 9i Bn O M e N C
O
COOH
538 cl ~OMe 0 OJD 554 F OlMe /r OOH 538 Yl
OOH
518 R* OMe 0 OOH 601 F p OMe
-O-Q-COOHC
WO 01/00206 WO 0100206PCT/usooI 18079 MNass:Spec~j V, W- J-- 516 556 OMe 0N '-O--G-OOH 556 OMe 561g NZ
_-COH
WO 01/00206 WO 0100206PCT/JSOO/18079
TABLE
Mass Spe 572 q y p 6 5 2 9 xM 1
KN,COOH
555 QF cI 6 Me 0
COOH
487H 9-z -\-COoH 600ry Br Me 536 9yx Me <1 gN O-CJ--COOH 554 9 H 2 Me 4 N O-Cj-COOH WO 01/00206 WO 0100206PCT/USOO/18079 TABLE-' :Mass Spec.j. i -R-R 534 9H dMe
C-O-COOH
0 2 M e~ C O
O
7 4 9 1 M e
I
044 OOH 580 FNN-cO Cl OMe X- 581Me\ 533
NH
2 I OMe \-yNQ -OO 0 OOH 548 PMe 0 OOH 552 PMe 0 OOH WO 01/00206 539 584 523 568 61 PCT/JSOO/18079 WO 01/00206 WO 0100206PCT/USOO/18079 Q~ Br OMe
Y?-X
OMe q x Br OMe Y1 /X Me WO 01/00206 WO 0100206PCT/JSOO/18079 TABLE-V 1 MassSpec. R. .7.RW <yr <DCOOH 648 OMe O
',CO-COOH
581 clOMe I o-
COOH
589 Y
O
OMe 537 I~
COOH
CIOMe0 Me -N..-COOH WO 01/00206 WO 0100206PCT/USOO/18079 I TABLE II Imass Spec! W -W 681 B r OMe
-~COOH
522 FOMe X1 00-OOH 9 0 e
\~~-CJ--OOH
624 OMe
N
OOH
534 ~~OMe q :O
H
4 9 4 9 x x- C J C O O H 550
QH
570
QH
624 -COOH
Q/COOH
WO 01/00206 WO 0100206PCTLJSOOI18079 9/
COOH
9/ Me WO 01/00206 520 54 598 643 585 601 71 PCTUSOO/18079 WO 01/00206 686 731 573 617 49 PCT/USOO/18079 WO 01/00206 Mass Sl 743 548 568 613 506 PCT/JSOO/18079 WO 01/00206 CIS0I87 PCT/USOO/18079
TABLEI
Ma ss- Spec
R:
~9r HN' 11 Br OMe 0 O 524F
-OOH
clMe
OOH
689 cI OMe
OOH
594 qzCOOH 681- 04C OOH 726 ?rh Me 590 rcI N~9 F OMe "N WO 01/00206 WO 0100206PCT/USOO/18079 II Mss.Spec. WI -WI
R
OMe- O OOH 6 4OMe OOH 659
OOH
534 He ~Me OOH 554 O H ~~e 0 OOH 599 OH Mr~ e V,
OOH
571F
\OH
648 cl V-6OOH WO 01/00206 WO 0100206PCT[USOO/18079 -TABLEil1.
Mr~assSpec.- I_ 643 Br OMeO
H
532 e 552 OMe 5y7 OMe 0.
62 ClNOMe OMe 0 F_ O 532 y N- y OMe 0~ OOH 552 0 OOH WO 01/00206 PCT/USOO/18079
F
V-XNJOO
H
F
hr- Nr-LO
H
WO 01/00206 WO 0100206PCT/USOO/18079 I._____TABLE 1.
560F 605 9>,F 562
OOH
582 O
H
627 V q 0 508 5 8 V 1 0) OOH 522 9>,OO 522F Br~
O
WO 01/00206 WO 0100206PCT/USOO/18079
TABLET.
526 c 591 9 M j OMe
N
00 37F WO 01/00206 WO 0100206PCT/USOO/18079 Mass:S''peC 1 W- 1 589 x,-I F0 526 Fio(j2-~ 632 F cl OMe N OOH 616 -l F VVI
OOH
54CF 3 0 0 524 FqN~ 0 O 541F WO 01/00206 WO 0100206PCTIUSOO/18079
.:TABLEXI
542 yF\F O 4 2 Ic
F
607 QLF Sr 571 Fy 619 Br 560 F V<yNN H- WO 01/00206 PCT/US00/18079 Q
R
7 R8 In another preferred embodiment of Formula I, M is In this embodiment, more preferred compounds are those wherein A is R is -(CH 2 n and X is Y is preferably chosen from -S(0) 2 and -NY,and more preferably, is 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-.
In this embodiment of Formula I, A is preferably =0 and A 2 is a direct bond or
-(CH
2 More preferred compounds are those wherein A 2 is a direct bond and R 8 is -OH.
R Z-A 1 Preferred compounds of Formula I, wherein M is i and A is 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 The entry entitled -R-R 8 depicts that portion of the particular compound represented by S1Z'A WO 01/00206 WO 0100206PCT/USOOI 18079 irl-Y 1(CHR 9 )qR 10 In another preferred embodiment of Formula 1, M is .In this embodiment, preferred compounds are those wherein A is R' 0 is -CO 2 H and q is 0 or 1. More preferred are compounds wherein R' 0 is -CO 2 H, q is 0 or 1, most preferably 0, and m is 2.
When A is L is preferably chosen from (N A HO,, NH2 N
(CN
HO OH IH 2 N 0C N N N N 0 0
N
-,p-CIPh WO 01/00206 WO 0100206PCT/USOO/1 8079 PhO Ph,
N
0 pMOhBnO
N
N
1 0a SN S N2Y 'N Y\N
N
K- 0 9N YN
H
2 N HN~ N N N 0a 0 andw O
H
More preferably, L is chosen from HO OH N N 1 01 I Y\ eNL N.
~,p-CIPh 0 PhO
NN
I Io ,p-MeOPh 0 B n o
NN
-o Io Ia Ph S2 NI N
NN
0 an LI 0 WO 01/00206 PCT/USOO/18079
HO,
Most preferably, L is chosen from HO OH N
-N
0 0 p-CIPh 1 2 N
O
N0 0O
H
p-MeOPh S N KY N I 0 o 1L o S and O Preferred compounds of Formula I, wherein R is -CH 2 and X is 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-.
(CHR
9 )qR 10 Preferred compounds of Formula I, wherein M is L-Z2 ,A is R is -CH 2 and X is are represented in Table 3. With respect to the representation of-W', the lower bond is the point of attachment to -NH- and the upper bond is the point of attachment to The entry entitled -N--Z 2 depicts that portion of the particular compound represented by
CHR
9 )qRo 1 WO 01/00206 WO 01/11206PCT/USOO/1 8079 T ABLE 3.
pec W- I -WI- I--N Z 554.7
HO,,
493.6 540.6 523.7 555.7 583.8 WO 01/00206 524.7 509.7 538.7 554.7 569.7 PCT/USOO/1 8079 WO 01/00206 567.8 570.4 552.7 619.5 575.1 PCTUSOOI18079 TABLE 3 -Wy.! I L 2rC0 2
H
N
H0,, N rCO 2
H
N
CNI
C0 2
H
(N)O2 WO 01/00206 PCT/USOO/18079 r C2
N
I)
WO 01/00206 WO 0100206PCTIUSOO/18079
TABLE
Mass
I.
555.6 1 I HO, IC, 556.6 570.7 570.7 673.8
H
3 600.7 WO 01/00206 PCT/USOO/18079 WO 01/00206 PCT/US00/18079 Spec W WL- N Z 2 562.6 H
CO
2
H
"u y v Yet another preferred embodiment of Formula I includes compounds wherein M is -R -Z Q L. Preferably, A is R is -CH 2 and X is Preferably 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-.
In compounds wherein Q 2 is R 1
R
18 and Z 3 is a divalent aliphatic hydrocarbon moiety, A 2 preferred compounds are those wherein R 1 is or -NR 2 more preferably NR 12 wherein
R
12 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 R1 R and Z 3 is R R" is preferably
-NR'
2 In these compounds, x and y are preferably 1. Preferred choices for R' 4 include H.
-OH and 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 to -NH-.
WO 01/00206 PCT/USO0/18079 In compounds wherein Q 2 is R 1
R'
8 and Z 3 is o R 1 is preferably chosen from and -NR' 2 preferably wherein R 12 is chosen from -H and lower alkyl group.
Preferably, R 17 and R 1 8 are each 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 R 17 8 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 1 8 is lower alkyl group or substituted lower alkyl group.
In compounds wherein Q 2 is R 18 and Z 3 is R 1 is preferably -NH- and R 1 7 and R' 8 are each preferably 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 Q2 is chosen from aryl group, substituted aryl group and
R
1 S, and more preferably from phenyl group and phenyl group substituted at the point of attachment to Z 3
,Z
3 is preferably a divalent aliphatic hydrocarbon moiety.
Yet another embodiment of the invention is a compound represented by Formula I, 0 Z3 W N/ N R11 L3 H H H WO 01/00206 PCT/US00/18079 wherein the substituents W, R" and Z 3 are defined as in Formula I, and L 3 is chosen from u 4 COOR20, wherein R 20 is preferably chosen from -H and lower alkyl, CH 3 J and Still another embodiment of the invention is a compound represented by Formula m, WN wR 11 d (CH 2 f H H I WR M wherein the substituents W, and R" 1 are defined as in Formula I, and d is chosen from 0 and 1, and f is chosen from I and 2.
11-Z3-Q2- 1 Preferred compounds of Formula I, wherein M is A is R is
-CH
2 and X is are represented in Table 4. With respect to the representation of-W', the lower bond is the point of attachment to -NH- and the upper bond is the point of attachment to The entry entitled L' depicts that portion of the particular compound represented by H-Z 4 -L 1 2 WO 01/00206 PCT/USOO/18079 TABLE 4, Maiss Sle I J -R 1 466.22 H KI JO2H
CH
3 0 496.22 02H
CH
3
OCH
3 0 445.19 -'IC02H2
CH
3 475.20 FlyJ C0 2
H
OH
3
OCH
3 497.22 0 O 2
H
OH
3 OCH 3 u 510.42 N ND--/'C2H
OH
3
OCH
3 517.62 IN
I
CH
3
OCH
3 473.56 C-0ii. CO 2
H
CH
3 504.1 HI CC o2H
OH
3
OCH
3 1 529.17 9HQJ
OH
3
OCH
3 53 WO 01/00206 WO 0100206PCT/USOO/18079 501.16 HOX?
CH
3
OCH
3 C 2 427.20 1
CH
3 OCH 3 441.22 0 2
H
CH
3 OCH 3 468.1 C 2
CH
3 OCH 3 532.2 vp
CH
3 OCH 3 624.2 9<
CH
3
OCH
3 I N C0 2
H
455.16 C 2
H
CH
3 OCH 3 483.70 N l C 2
CH
3 OCH 3 959.54 C0 2
H
CH
3 54 WO 01/00206 WO 0100206PCTIIJSOO/18079 923C WO 01/00206 WO 01/024)6PCT/USOO/18079 TABLE:4' 507.55 jJNC0 2
H
CH
3 OCH 3 521.58 C 2
CH
3 OCH 3
CO
2
H
CH
3 OCH 3 HNI-'CO2H
CH
3 OCH 3 414.10HN- 2
CH
3 00H 3 441.2 .2H C2
CH
3 OCH 3 511.47yJ rNNNN,,C
CH
3 OCH 3 459.11 H HN 0
O-CO
2
H
CH
3 OCH 3 504.6 H
CH
3 ~N Y CO 2
H
CH3
OCH
3 500.2
CH
3 WO 01/00206 PCT/USOO18079 TABL.E 4:: MS. 530.2 0
CH
3 OCH 3 516.4 0
CH
3 OCH 3 H 486.2 0 CH3 544.2 qK VpqA
CH
3
OCH
3 500.2 c I OH 506.2
CH-
3 V Cl- 3
OH
OH
516.2 9
CH
3
OCH
3 498.2 9-OA
CH
3
CH
3
H
47539~AN
OH
CH
3 WO 01/00206 WO 0100206PCT/1iS00/18079 502.2 HO0
CH
3 506.3 N
O
CH
3 OCH 3 IO 518.1 0 A HO
CH
3 OCH 3 II 530.1
H
CH
3
OCH
3 N 526 CH3 OH3 LN 3
N
602 IK rf~
CH
3
OCH
3 x
COOH
573 '4 %C
O
CH
3 OCH 3 OH 622 y
CH
3 HN~
CC
WO 01/00206 WO 0100206PCT/USOO/18079 6460 C H 3 O C H 3 N C O 518
N
CI
OCH
3 CCO 563 y
N
OCH
3
LQCOOH
cl \,7_Xl X..COOH CI
OCH
3 638
OCH
3
COOH
639
OCH
3 'S C CI
OCH
3 FN COOH 593
\Q
CI OC3_4!
COOH
513 9 \9
CH
3 0OCH 3 L COOH WO 01/00206 PCTIIJS0OI18079
T.ABLE
Mass.Specf: 1 7W.7 WO 01/00206 516 486 521.56 535.55 507.54 535.59 522.51 539.55 502.56 515 PCTIJSOOI18079 WO 01/00206 PTUO/87 PCT/USOO/18079 547.6
C
2 0
CH
3 OCH 3 H 620.69
CH
3 OCH 3 OC H 3 633.73 Q., 1 CH 3
CH"'CH
3
OH
590.67 y q
N
CH
3 OCH 3
OCH
F OCH3 F OCH 3 c 52994CH3
OH
FH
OCH
3
C
CH3
OHH
CH
3
OCH
3
CI
489.56 I~KCH 3 o
CH
3 OCH 3 WO 01/00206 Mass St 461.51 495.95 491.54 461.51 459 PCT/USOO/18079 WO 01/00206 WO 0100206PCT/USOO/18079 552.590
FOCH
3
H
3
C{NH
523.55 0 CH3 H 3 C CH 3
OH
OCH
3 529.94 0 H H l O F
OCH
3 6H 3
CI
490.55 9
CH
3
OCH
3
H
543.97 0 yHiy H OH F OCH 3
CH
3
C
540.01 0
CH
3
OCH
3
CH
3
CI
548.63 0 LII H3~y-OH
CH
3
OCH
3
H
3 C NH %H 3 504.58 0 Yy VCH3 OH
CH
3
OCH
3 CH 3 620.07 09- F OCH 3
(ZH
3
C
WO 01/00206 WO 0100206PCT/USOO/18079 MassSpecJ.. W J 562.66 9K\XI 0 C3OCH 3 H 3 C yN-CH 3
CH
3 578.63 C3 0
FOCH
3 0 506.55 0 CH3 j OH
CH
3
OCH
3 NH 2 574.67 P V AC3 0 O
CH
3 OCH 3 0 510.51 0 F
OCH
3 H 506.55 0
CH
3
OCH
3 6H 3
NH
2 505.56 H OH 9Kv
CH
3 OCH 3 H 3 C CH 3 556.41 0
\HO,()-OH
BrOCH 3
CH
3 596.67 92N
O
CH
3 OCH 3
CH
3 WO 01/00206 WO 0100206PCT/USOO/18079 Mvass SpecJ]-I i 532.63 0H
CH
3
OCH
3
CH
3 691.53 02N &~cOH BrH 3 CH 3 541.00
HO
c I
OCH
3 H 3 C CH 3
NH
2 6070 rco COOH Me V
NH
2 497 HCO 525
(COOH
cI OMe V N 527
~COOH
cl M eN H 2 542 :~1IIIJ ~fH OMe "<fCO 498H 543Ic COOH OMe WO 01/00206 WO 0100206PCT/JSOOI 18079 TABL 4...7 611 QL
COOH
CI OMe V 512 <~COOH cl ~OMeV 518 )zCO 569
COOH
569 OMe V C:f00H 591 591 iiC0 N CO OH MI~ e N-
COOH
c I OMe 583 Q
CO)
B r Me L WO 01/00206 WO 0100206PCTIJSOOI18079 561 (cl)pJCOOH CIOMeNH 549 CO OH 604 O~e _N,_ryCOOH 589 1 2 C H OMe
CO
506 9-zry JyCOOH ~Me 607\- C YCOOH OMe 589 575 O~e Ot~J~h~COOH 601 OMe hN y
OOH
603 Cy IN- yCOOH 9-Z ~OMe N-a WO 01/00206 WO 0100206PCTfUSOO/18079 617 Q- M
COOH
625 9 \9 OMe
V
6119 z OMe N&cCOOH 522 HO-) 0
JCXO
565 NK MeO
/N~~QCOOH
593 ):)--lCOOH 518 9x ~T COOH OMe 538 ~T~COOH 627 SJINV ,.yCOOH 585 MeO lp, Pc)COOH WO 01/00206 WO 0100206PCT/USO0/18079 624 -N COOH 609 639)
COOH
OMe jT 6379 CO LNZ ~JCOOH 621 581)
COOH
6 3 7 M e
M
Me ~COOH WO 01/00206 WO 01/(1206PCT/USOO/18079 611 Fj~~CO F- Q~COOH 488 0 7
COOH
0 4 C O O H 522 NOIc 640
XCOOH
676 i;i
~COOH
548 XCO 613 ~COH 666 Br Me ,/VNhjyC00 WO 01/00206 WO 0100206PCT/USO0/1 8079 o x d 1 616
COOH
6 Br MeCOOH 502 Z
COOH
Me
V
514 COOHr 484 9-z- jfCOOH 72 WO 01/00206 WO 0100206PCT/USOO/18079 TAlm .4 s pec! 585I (c,~fOOH 512 9.,x Me
COOH
482 0 c~co 532 -x O~e
COOH
560
K
Me K>
COOH
6250 Br Me 583 ~~CO 568 534 CQOH ~~Me N-y 534 Me
~COOH
536 ~JfCOOH WO 01/00206 PCT/US0O/18079 Mass Spec 581 i 0
COOH
522 MOe COOH 520 0 j ,COOH 520r^ ^co P C61OHo The principles of the present invention also encompass prodrugs in the scope of Formula I, and compounds representative thereof include those wherein Rs or R 8 is a lower alkoxy group, and those wherein R 1 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 a4pl receptor sites in a mammal, wherein the method comprises administering an effective amount of a compound represented by Formula I. As 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 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.
WO 01/00206 PCT/USOf/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.
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 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 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 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 made from chloroprocaine, choline, N,M-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 acids. Such acids include acetic, benzenesulfonic, benzoic, camphorsulfonic, citric, WO 01/00206 WO 0100206PCT/USOO/18079 ethanesulfonic, methanesulfonic, fiirnanic, glucomic, glutamnic, hydrobromic, hydrochloric, lactic, maleic, malic, niandelic, nitric, pamoic, pantothenic, phosphoric, succinic, sulfuric, tartaric, ptoluenesulfonic acids, and the like. Particularly preferred are citric, hydrochloric, nialeic, fuimaric, phosphoric, sulfuric, tartaric and p-toluenesulfonic acids. Compounds of the invention may also be in the form of hydrates.
DETAILED DESCRIPTON OF THE INVENTION Abbreviations Definitions The following terrns and abbreviations have the indicated meaning throughout this disclosure.
293E HEK cells 293E human embryonic kidney cells Ac acetyl anti-a4-PE conjugated monoclonal antibody against integrin cx4 subunit, phycoerythrin conjugated anti-3 I-FITC conjugated monoclonal antibedy against integrin 131 subunit, fluorescein conjugated ax5f31 integrin cx5 P 1, fibronectin receptor, axvP3 integrin cxv133, vitronectin receptor (x4037 integrin (4037 Bn benzyl Bc t-butoxycarbonyl BSA bovine serum albumin c- cyclocDNA complementary DNA CHO cells Chinese Hamster Ovary cells p-ClPh para-chlorophenyl CMV promoter cytomegalovirus promoter m-CPBA 3-chloroperoxybenzoic acid DAST diethylarninosulfuir trifluoride DCM dichioroniethane methylene chloride =CH 2
CI
2 DELFIA dissociation enhanced lanthanide fluor-immune assay DIAD diisopropyl azodicarboxylate DIC diisopropylcarbodiimide DlEA NN-diisopropylethylamine WO 01/00206 WO 0100206PCT/JSO0I 18079
DMAP
DMEM
DMF
DTPA
EDC
Et 2
O
FACS
Emoc GPIlb/lla
HEPES
HMDS
HOAc HOBt human IgG I
ICAM
LDV
LEA- I and Mac- I LiH-MDS Me p-MeOPh nM
PBS
PEG
Ph PhOH PyBroP RPMI medium
TEA
TEAA
THE
TLC
TMS
Ts VCAM- I (D ID7) 4-N,N-dimethylaminopyridine Dulbecco's Modified Eagle's Medium NN-dimethylformamide Diethylenetriaminepentaacetic acid I -ethyl-3-(3-dimethylaminopropyl)carbodiimide ethyl ether fluorescence cell sorting 9-fluorenylmethoxycarbonyl integrin cxllbf33, fibrinogen receptor N-(2-hydroxyethyl)piperazine-N'-(2-ethanesulfonic acid) 1, 1, 1,3,3,3-hexamethyldisilzane acetic acid 1-hydroxybenzotriazole human immunoglobulin G 1 intracellular adhesion molecule Leu-Asp-Val Lymphocyte function-related antigen lithium 1,1,1 ,3,3,3-hexamethyldisilazane methyl para-methoxyphenyl nanomolar phosphate buffered saline polyethylene glycol phenyl phenol bromo-tris-pyrrolidinphosphonium hexafluorophosphate Russell Park Memorial Institute medium trifluoroacetic acid trifluoroacetic acid anhydride tetrahydrofuran thin-layer chromatography trimethylsilyl toluenesulfonyl vascular cell adhesion molecule (containing one to seven 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 IgG molecule "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.
"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 carbon atoms. The rings may optionally be substituted with 1-3 substituents selected from alkyl, 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.
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.
"Cycloalkylene group" means a divalent radical formed by removing a hydrogen atom from a "cycloalkyl group." WO 01/00206 PCT/US00/18079 "Divalent C, to C2 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 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, 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 1 to 12 carbon atoms, preferably 1 to 6 carbon atoms, and from 1 to 4 heteroatoms chosen from O, N and S; or a bicyclic 9- or 10-membered heteroaromatic ring system containing 14 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, 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 I to 20 carbon atoms, preferably from 1 to 8 carbon atoms, 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, secbutoxy, tert-butoxy, cyclopropoxy, cyclohexyloxy groups and the like. "Lower alkoxy group" refers to alkoxy groups having from 1 to 4 carbon atoms.
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-l-yl, 2-methylbuten-1-yl, 3-methylbuten-I-yl, hexen-1-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." "Alynyl group" refers to an unsaturated acyclic hydrocarbon radical containing at least 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, alkoxy, hydroxy, aryl, cyano, carboxy, alkoxycarbonyl, monoallcylamino, alkyloxy, cyanoalkyl, cycloallcyl, 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.
"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 alcyl, 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 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, cycloallcyl, WO 01/00206 WO 0100206PCT/USOO/18079 alkylthio, alkylsulfinyl, alkylsulfonyl, arylthio, carboxyalkyl, alkoxycarbonylalkyl, haloalkyl, acylamino, dialkylamino, cyclicamidno 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, alkoxy, hydroxy, aryl, cyano, carboxy, alkoxycarbonyl, monoallcylamnino, alkyloxy, cyanoalkyl, cycloalkyl, alkylthio, alkylsulfinyl, alkylsulfonyl, arylthio, carboxyalkyl, alkoxycarboniylalkyl, haloalkyl, acylanuno, dialkylaxnino, 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, hydroxy, aryL, cyano, carboxy, alkoxycarbonyl, monoalicylamino, alkyloxy, cyanoalcyl, cycloalkyl, alkylthio, alkylsullinyl, 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, hydroxy, aryl, cyano, carboxy, alkoxycarbonyl, monoalkylamino, ailkyloxy, cyanoalkyl, cycloalkyl, alkylthio, ailkylsulfinyl, alkylsulfonyl, arylthio, carboxyalkyl, alkoxycarbonylalkyl, haloalkyl, acylamino, dialkylamidno, cyclicanino 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 substituent such as alkyl, amino, alkoxy, hydroxy, aryl, cyano, carboxy, alkoxycarbonyl, monoalkylamino, alkyloxy, cyanoalkyl, cycloalkyl, alkylthio, alkylsulfinyl, alkylsulfonyl, arylthio, carboxyalcyl, alkoxycarbonylalkyl, haloalkyl, acylamino, dialkylamino, cyclicamino groups, halogen atom and nitro.
"Substituted heteroarylene group" or "substituted heterocyclylene group" means a heteroarylene or heterocyclylene group wherein at least one hydrogen atom attached to a ring thereof is replaced with a substituent such as ailcyl, amino, alkoxy, hydroxy, aryl, cyano, carboxy, alkoxycarbonyl, monoalkylamino, ailcyloxy, cyanoalkyl, cycloalkyl, alkylthio, ailcylsulfinyl, alkylsulfonyl, arylthio, carboxyalkyl, alkoxycarbonylalkyl, haloalkyl, acylamino, dialicylainino, cyclicaniino groups, halogen atom and nitro.
WO 01/00206 PCT/USOO/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.
"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, 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 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 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 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 "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: beads, pellets, disks, 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- WO 01/00206 PCT/US00/18079 acrylamide beads, latex beads, dimethylacrylamide beads optionally cross-linked with N,N'-bisacryloyl ethylene diamine, glass particles coated with hydrophobic polymer, etc., material having a rigid or semi-rigid surface; and soluble supports such as polyethylene glycol or low molecular weight, non-cross-linked polystyrene. The solid supports may, and usually do, have functional groups such as amino, hydroxy, carboxy, or halo groups; where amino groups are the most common.
Tentagel
T
m NH 2 (Rapp Polymere, Tubingen, Germany) is a preferred amine functionalized polyethylene glycol- grafted polystyrene resin. Tentagel'"-S-PHB resin has a parahydroxy benzyl linker which can be cleaved by the use of 90% trifluoroacetic acid in 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 W095/30642, the disclosure of which is incorporated herein by reference. In the synthesis described in WO95/30642, the linker is a 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 or or as or for amino acids.
The present invention is meant to include all such possible diastereomers as well as their racemic and optically pure forms. Optically active and or and 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 and geometric 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 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.
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 (ICso) and the VLA-4 selectivity of test compounds for an a4p 1 receptor using in vitro assays are determined in direct binding assays and competitive assays with other integrin receptors such as 32 (LFA-1 and Mac-1), P3 (GPIIb/IIa and avp3) and p1 (a4p7). Compounds of the present invention have K, values 1 M. Preferred compounds of the invention are those having KI 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 examples are provided by way of illustration only, and are not intended to limit the invention thereto.
H H H HH H H O H Q H H
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H H 0 2
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WO 01/00206 WO 0100206PCTI USOOI 18079 N"N'y0 H H 0 2
H
N WQ-- 0 2
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NI Njir4 N _O 'C 2
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F-'N
Me N N0 11 2H2 HHH W H 0 2 H
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WO 01/00206 WO 0100206PCT/USOO/18079 HtN H2
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2
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7H H (?'MeN 2 %H
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NH H 0 2 H ?~~C2 HH M e c NH e H
OH
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OHH
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WO 01/00206 WO 0100206PCTIIJS0OI18079
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WO 01/00206 WO 0100206PCT/USOO/18079 F F OOH COOH <aCOCH OOH Me OOH l OCH
M
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c 0 H,6 H -e H_6 WO 01/00206 WO 01/02116PCT/USOO/18079 0 HiH 2-toH 0
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3
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3H 3 H NOOH WO 01/00206PT/S I189 PCT/USOO/18079 JF <JyJMe N rN eK M N-COOH Me HNHCOOH y1e y1e 6iHHMe rOJNO ~rH H We TO&.COOH C IH H% 2 6M TNOO y1e Me H H 6Me C CC 'N N N x Me H H e, yCOOH t~4
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3
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3
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3 H 3 WO 01/00206 YyN PCT/USOO/18079 WO 01/00206 WO 0100206PCT/USOO/18079 pj<jfyCOOH 0 COOH N M% 7 NH2 Me H H
CO
CI1H H M H ~/COQH Me H H OMe a ti m ome Me Me-T r-COOH Me H H 6M7 M"-Y7J-> 0
COOH
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M eQ mre 'HH 6yMeo MHHMe Y COOH Me' H 6e WO 01/00206 WO 0100206PCT/USOO/18079 S~J~c,~2COOH Gla,
COOH
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M
6Nye eg COOH e H Me HM~ Me HHI 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 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 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 a4 and 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 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 DID7-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 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 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 ca4 1 of each test compound among other integrin receptors, namely, P2 (LFA-I and Mac-l), P3 (GPIIbIIIa and avp3), P1 (aSpl) and P7 (ca47) was examined. LFA-1 binds to ICAM-1 and mediates the emigration ofleukocytes into inflammatory sites. Mac-1 binds to a number ofligands, including ICAM-1 and fibrinogen, and plays an important role in 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 ofextracellular 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 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 five immunoglobulin domains of human ICAM-1 (D1DS) attached above the hinge region of an IgG1 molecule, was used. In the assays of GPIIb/Ia 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 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.
Eu'-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°C, 5% CO, for 48 hours and then washed twice with washing buffer, then blot dried. 50 j1 of the inhibitor solution diluted with assay buffer DMSO final) is added to each well, followed by 50 /l of Eu'-VCAM-l IgG diluted with assay buffer at 2 nM. The plate is incubated at room temperature for at least 30 min. Each well is then washed four times with washing buffer and blot dried. 100 ul 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 DELFIA Fluorometer 1234, Wallace, Inc., USA). In this assay, the washing buffer comprises 25 mM HEPES (pH 150 mM NaCI, 1 mM CaCI,, 1 mM MgCl 2 ,and 4 mM MnCI,; the assay buffer comprises 25 mM HEPES (pH 150 mM NaCI, 1 mM CaCI 2 1 mM MgCI,, 4 mM MnCI 2 1% BSA, and 20 ,uM DTPA.
In Vivo Assays 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 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 of Ascaris-induced bronchial inflammation, one of the representative compounds (example number 32) inhibited eosinophil infiltration by 49% at an oral dosage of 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 at an oral dosage of 50 mg/kg.
Other representative compounds were also tested in mice. The dosage, route of 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 TABLE Dosage/Regimen Route Inhibition of Eosinophil Infiltration Compound 79 30 mg/kg b.i.d' x 2days i.v. 35.9 Compound 90 10 mg/kg t.i.d 2 x2 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.dx 2days p.o. 18.9 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 1 two times a day (0 and 8 hrs) 2 three times a day 8 and 16 hrs) 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 /g 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 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 DosagelRegimen Route Inhibition of Eosinophil Infiltration Compound 90 3 mg/kg 2days i.v. 25.4 mg/kg 2 days i.v. 46.7 mg/kg 2days i.v. 83.7 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-Assav Method 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 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 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- Administration Serum Concentration (ng/ml) Compound 58 50 mg/kg 30 3614 Compound 68 10 mg/kg 15 261 Compound 78 10 mg/kg 15 368 Compound 79 10 mg/kg 15 618 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 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 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 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 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 TABLE 8 MOUSE Dosage AUC' (ng-h/mL) MRT2 (hr) CL 3 (mL/min/kg) Compound 68 10 mg/kg 1595 (0-6 hr) 0.2 104.5 1595 0.2 104.5 mg/kg 1307 (0-6 hr) 1.6 637.6 WO 01/00206 PCT/US00/18079 1751(0-) 4.1 476.0 Compound 90 10 mg/kg 8995 (0-6 hr) 0.5 18.53 9219(0-) 0.7 18.08 mg/kg 2540 (0-6 hr) 1.2 65.62 2950 2.5 56.50 Compound 80 10 mg/kg 5259 (0-6 hr) 0.4 31.69 5389 0.6 30.93 mg/kg 2190 (0-6 hr) 2.3 152.24 3615 6.5 92.21 total area under the plasma concentration (measured by LC/MS/MS method) versus time curve 2 mean residence time apparent plasma clearance TABLE 9 RAT Dosage AUC' (ng-h/mL) (hr) CL (ni/mirift) Compound 90 10 mg/kg 31915 (0-6 hr) 0.8 5.23 33488(0-) 1.1 4.98 mg/kg 11454 (0-6 hr) 1.7 17.85 19968 8.8 9.99 Compound 79 10 mg/kg 5259 (0-6 hr) 0.4 31.69 5389 0.6 30.93 mg/kg 22119 (0-6 hr) 0.5 24867 0.7 6.7 Compound 68 10 mg/kg 6345 (0-6 hr) 0.63 26.45 6405 0.67 26.20 mg/kg 1867 (0-6 hr) 1.1 181.1 2086 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 Pharmacokinetic parameters and the time course of the serum concentration of a single 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 MONKEY AUC' (ng.h/mL) Cltot 2 (mlJmin/kg)
F
Compound 195 8405 ATENOLOL 5021 6.7 total area under the plasma concentration (measured by the LC/MS/MS method) versus time curve 2 apparent plasma clearance TABLE 11 TIME 5 min 30 min 1 hr 2 hr 4 hr 8 hr Serum cone' (ng/ml) Compound 195 65447 3900 2225 772 194 28 Serum conc' (ng/ml) ATENOLOL 4057 1189 771 479 348 137 Smeasured by the LC/MS/MS method Binding assay of VCAM-1 to VLA-4 expressing cells Preparation of VCAM IgG fusion protein A VCAM IgG fusion protein containing the one to seven immunoglobulin domains of VCAM-I (D1D7) ligated to the hinge CH2 and CH3 regions of human IgGI was used in the binding assay.
Construction of a stable cell line expressing DID7-VCAM IgG fusion protein An Epstein-Barr virus based, episomal plasmid containing a D1D7-VCAM IgG fusion gene under transcriptional control of the CMV promoter, was transfected into 293E human embryonic kidney cells. Stably transfected cells were selected using 250 ,g/mL hygromycin in DMEM with 10% fetal calf serum. The cells secreted D1D7 VCAM IgG fusion protein into the medium cumulatively for up to 9 days.
Purification of DID7 VCAM IgG fusion protein The cells were cultured in DMEM with 10% fetal calf serum for 2 days, then changed to medium and cultured for a further 10 days. The medium was centrifuged, filtered and then WO 01/00206 PCT/US00/18079 incubated overnight with Protein A Sepharose 4. The Protein A Sepharose was washed extensively and the D ID7 VCAM IgG fusion protein bound was eluted using 100 mM citric acid, pH 3.
Preparation of europium labeled-DID7 VCAM IgG fusion protein The DID7-VCAM IgG fusion protein, at 1 mg/mL, was dialyzed against 50 mM NaHCO,, 0.9% NaCI, 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.
Construction of cell line expressing VLA-4 (CHO/VLA-4) A CHO cell line stably transfected with the cDNA of a4 and 31 was used in the binding assay. The gene for human a4 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 31 gene was amplified by PCR from human peripheral leukocyte cDNA and 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-KI cells were stably co-transfected with plasmids encoding the a4 and p1 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 Diego, CA) and anti-p1 -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, 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 370C, 5% CO, for 48 hours until confluent. On the day of assay, the wells were emptied and washed twice with 350 ,1 of a washing buffer containing 25 mM HEPES, pH 7.5, 150 mM NaCI, 1 mM MgCl 2 WO 01/00206 PCT/US00/18079 1 mM CaCI, 2 mM MnC12. 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 pM DTPA and 1% dimethysulfoxide), in the presence of 2 nM of europium-labeled D1D7-VCAM IgG fusion protein. Final concentrations used ranged from 0.1 gM. 50 ll 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-a4 monoclonal antibody (L25.3, Becton Dickinson, Bedford, MA).
The cells were allowed to incubate with the test compound mixture, in the presence of europium-labeled DID7-VCAM IgG fusion protein at room temperature for at least 30 minutes.
The cells were then washed three times with 350 pl of washing buffer, using a Skatron plate washer and blot dry. An 100 1 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 (Model: Victor", 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 DID7-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 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.
Preparation of VCAM coated plate A 50 kl aliquot of the DID7-VCAM IgG fusion protein (1.5 klg/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 4°C. On the day of assay, the wells were emptied and washed twice with 350 gl of PBS. The plate was then blocked with 100 ml 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 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 pM of calcein- AM (Molecular Probe, Eugene, OR, catalog no. C1430) in RPMI medium, at room temperature for min in the dark. Following labeling, cells were washed twice with RPMI medium and resupended at 1 x 106 cells/mL.
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 pl 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 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 reader set at 485 nM excitation and 530 nM emission. The IC 50 (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 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.
Modifications and variations in any given material or process step will be readily apparent to one WO 01/00206 PCT/US0O/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.
Example 1 Boc'
A
A three-necked 500 mL round-bottomed flask was charged with 200 mL of THF and NaH g, 62.9 mmol). A solution of 1-vinyl-2-pyrrolidinone (6.9 g, 62.9 mmol) and methyl 3iodobenzoate (15.0 g, 57.3 mmol) in THF (100 mL) was added dropwise to the flask over 15 min.
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 NaHC03 until pH 9 and then the mixture was extracted 3x with EtOAc. The combined 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 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 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 KCO, 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, and concentrated in vacuo to afford a yellow viscous oil. The oil was chromatographed (25% EtOAc:hexanes) to afford 7.0 g A.
Br Boc A Oc A' Hydrochloric acid (gas) was bubbled through 15.8 g (73.5 mmol) 2-bromophenylacetic 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 CHC1. 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 in a 250 mL round-bottomed flask.
To this flask was added 3.5 g (147 (mmol) sodium hydride 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 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 sodium hydroxide, extracted with 3 x 125 mL CH 2
CI
2 and the combined organic layers were washed with a saturated aqueous solution of sodium chloride, dried over magnesium sulfate, and the solvent was removed under educed pressure to give 15.0 g (63.0 mmol, 86%) of 2-(2bromobenzyl)-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 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 CI, 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.
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-tbutyl-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 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'.
WO 01/00206 PCT/US00/18079
H
Fmo 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) 3 (12 mg, 0.3 nmmol), methyl acrylate (0.47 g, 5.5 mmol), and NaOAc (0.5 g, 5.5 mmol). This mixture was then heated to 80 0 C 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 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 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 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
TFA/CH
2
CI
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 dioxane/water, adding K 2
CO
3 (1.2 and Fmoc-Cl (1.08 g, 4.0 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,, and then concentrated in vacuo to afford 495 mg 1 as a white crystalline solid.
WO 01/00206 PCT/US00/18079 o 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 were drained and the resin was washed 3x with DMF, MeOH, and CH 2 CIl. 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 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 EtO until a precipitate formed. This precipitate was collected and dried in vacuo to afford 67 mg 1' as a white crystalline solid.
Example 2
TMS
\N
Bo 2 A (4.2 g, 11.3 mmol) was placed in a 100 mL round-bottomed flask and dissolved in NEt 3 mL). To this solution was added Pd(PPh 3 2 Cl (0.16 g, 0.23 mmol), Cul (21 mg, 0.12 mmol), and trimethylsilylacetylene (1.38 g, 13.5 mmol). This mixture was stirred at room temp for 14 hr.
The reaction mixture was quenched by the addition of I N HCI (100 mL). The solution was then extracted 3x with EtOAc, dried over MgSO,, 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
N
Fmoc 3 WO 01/00206 PCT/US00/18079 A solution of dicyclohexylborane was generated by the addition of borane-THF (12.0 mL, 12 mmol), at 0°C to a solution of cyclohexene (2.3 mL) in 6 mL of anhydrous THF. This solution was stirred for an additional 1 hr at 0°C. The acetylene (2.0 g, 5.84 mmol) was then added dropwise over 15 min at 0°C and then allowed to warm to room temp over 1 hr. The reaction mixture was then diluted with MeOH (20 mL) and then recooled to 0°C. A solution of 2 N NaOH (6 mL) and 30% H 2 0, (3.5 mL) was then added dropwise. The reaction mixture was then stirred at 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 extracts were then acidified with 1 N HCI and extracted with EtOAc dried over MgSO,, 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 C 1, 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 (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
HHN-
0 3' 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, 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
CI
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,.
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
CI
2 The compound was then cleaved from the resin and the filtrate was collected and then 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 EtO 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
H
Fmoc 4 The acetylene was deprotected by placing 2 (0.75 g, 2.1 mmol) in MeOH mL) and adding to this solution KOH (1.4 The resulting solution was stirred at room temp for 1 hr. The reaction mixture was then concentrated in vacuo and acidified with 1 N HCI. The resulting aqueous solution was extracted 3x EtOAc, the combined organics were dried over 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°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 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 1 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 and Fmoc-CI (1.29 g, 4.9 mmol). This mixture was stirred at room temp for 14 hr and then poured in 1 N HCI (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 MeOH/ dichloromethane to afford 110 mg of the desired compound.
HO
02O 0 4' 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 Cl,. 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
C,.
To the above resin was added 9 mL of DMF, 4-[N'-(o-Tolylurea)]-phenylacetic acid (132 mg, 0.42 mmol), PyBroP mg, 0.42 mmol), and DIEA (107mg, 0.82 mmol). The contents were shaken for 14 hr at room temp and then drained and washed 3x with DMF, MeOH, and
CH
2
CI
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 O until a precipitate formed. This precipitate was collected and dried in vacuo to afford 27 mg 4' as a white crystalline material.
Example 4 Hco Fmoc The iodide (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, was bubbled through for 10 min. The reaction mixture was poured onto 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
TFA/CH
2
CI
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 dioxane/water, adding K 2 CO (15 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.
WO 01/00206 PCT/US00/18079 O 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 contents were drained and the resin was washed 3x with DMF, MeOH, and CH,CI,. 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, 0.42 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 CHCl,. 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 O until a precipitate formed. This precipitate was collected and dried in vacuo to afford 51 mg 5' as a white crystalline material.
Example (E)-4-[2-[1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2-pyrrolidinyl]ethenyl] benzoic acid
SCO
2
H
6 To a cold (minus 78 OC), stirred solution of triethyl 4-phosphonomethylbenzoate (904 mg, 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 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 chromatographed on silica-gel with n-hexane-EtOAc 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 OC; IR (KBr) 1710, 1697, 1681 'H-NMR (CDCI 3 51.39 (12 H, series of m), 1.77-1.93 (3 H, 2.11 (1 H, 3.47 (2 H, 4.34-4.54 (total 3 H, 6.22 (1 H, 6.43 (1 WO 01/00206 WO 0100206PCT/USOO/18079 H, d, J 14.2 Hz), 7.39 (2 H, J 8.3 Hz), 7.97 (2 H, d, J 8.3 Hz); MIS (FAB) ,n/z 346 (MW+1); Anal Calcd for C 20
H
2 7N0 4 C, 69.54; HK 7.88; N, 4.05. Found: C, 69.52; H, 8.08; N, 4.07.
To a stirred solution of ethyl -Qert-butoxycarbonyl)-2-pyrrolidinyllethenylJ benzoate (700 mg, 2.03 mmol) in CH 2 Cl 2 (3 niL) was added TFA (3 xnL) and the resulting mixture was stirred for 3 hr. The mixture was concentrated and the residue was made basic by the addition of sat. NaHCO 3 The n-dxture was extracted with CHCl 3 (2 x 100 mL). The combined extracts were dried over Na 2 CO3 and concentrated in vacuo to give 434 mg ethyl pyrrolidinyl) ethenyl]benzoate as a brown oil. 'H-NMvR (CDCl 3 8 1.39 (3 H, t, J =7.3 Hz), 1.52- 2.06 (4 K, series of in), 2.93-2.99 (1 H, mn), 3.07-3.13 (1 H, in), 3.74 (1 H, q, J =7.3 Hz), 4.37 (2 H, q, J= 7.3 Hz), 6.34 (1 H, dd, J= 15.6, 7.3 Hz), 6.54( HK 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 mng, 1.77 minol), pentafluorophenyl -(2-methylphenyl)ureidolphenylacetate (797 mg, 1.77 minol), EtN (0.37 inL, 2.66 minol) in DMF (15 inL) was stirred for 15 hr. The mixture was diluted with EtOAc (300 inL). The solution was washed with brine (2 x 200 mL), dried over MgS504, and evaporated off in vacuo. The residue was chromatographed on silica-gel with CHCI,-EtOAc 1) as eluent to give 906 mg ethyl 1-[4-[N'-(2-inethylphenyl)ureidolphenylacetyl]-2-pyrrwlidinyl] ethenyljbenzoate as a brown oil. 'H-NMR (CDC1 3 8 1.39 (3 H, t, J 7.3 Hz), 1.83-2.20 (4 H, series of in), 2.24 (3 H, d, J =4.9 Hz), 3.63 (4 H, 4.36 (2 H, q, J 7.3 Hz), 4.62 and 4.84 (total 1 H, in), 6.18-6.47 (2 H, in), 7.03-8.02 (14 HK series of in).
A stirred mixture of ethyl (E)-4-[2-[Il-[4-[N'-(2-inethylphenyl)ureido] phenylacetylj-2pyrrolidinyllethenyl~benzoate (906 mg, 1.77 inmol) in 0.25 N NaOH (14 inL) and THF (14 niL) was heated under reflux for 3 days. The mixture was poured into ice-IN HCI (200 mQL and the precipitate was collected with suction. The solid was recrystallized from CHCl 3 -MeOH-n-hexane to give 453 mng 6 as a light yellow crystalline powder. mp 165-168 IR (KBr) 3282, 2974, 2663, 2537, 1700, 1685 'H-NMR (DMS04d) 8 1.74-2.12 (4 H, in), 2.24 (3 H, d, J= 4.9 Hz), 3.35-3.66 (4 H, in), 4.67-4.74 (1 H, in), 6.25-6.41 (1 H, in), 6.53 (1 H, 6.93 (1 H, t, J= 7.3 Hz), 7.08-7.92 (12 H, series of in), 9.00 (1 H, in), 12.87 (1 H, br MIS (FAB) m/z 484 AnaL Calcd for C2 9 H42N 3 0 4 -0.5H 2 O0: C, 70.71; H, 6.14; N, 8.39. Found: C, 70.46; H, 6.07; N, 8.39.
Example-6 WO 01/00206 PCT/USOO/18079 1-14-[N' -(2-methylphenyl)ureidojphenylacetylj-2-pyrrolidinyl~ethyl]benzoic acid NYN Y02H A mixture of 1-[4-[N'-(2-methylphenyl)ureidojphenylacetyll-2-pyrolidinylJ ethenylJbenzoate (200 mg, 0.414 mmol) and 5% Pd/C (200 mg) in MeOH (20 mL) was 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 CHCI,-MeOH as eluent to give 201 mg 7 as a colorless crystalline powder. mp 180-190 IR (KBr) 3345, 3124, 3060, 3027, 2960, 2927, 2875, 1706, 1672 'H-NvR (DMSOd)5 1.04-3.96 (total 16 H series of 6.91-7.41 (9 H, 7.79-7.90 (3 8.23 (1 H, br 9.31 (1 H, br MS (FAB) ni/z 486 (N
T
Anal. Calcd for Cn-1 31
N
3 042.25H 2 0: C, 66.21; H, 6.80; N, 7.99. Found: C, 65.97; H, 6.20; N, 7.72.
Example 7 1 [4-[AP(2-methyphenyl)ureidophenylacetyl]pyrrolidinylJmethoxylbenzoic acid -(J-CO 2 H 8 8 (S)-4-[2-[1-[3-methoxy-4-(M"-phenylureido)phenylacetyllpyrrolidinyllmethoxylbenzoic acid aNY'N- _0 02H 9 (S)-4-[2-[1-[4-[M,"-(2-chlorophenyl)ureidolphenylacetyljpyrrolidinylmethoxybenzoic acid y~ 0^8r~cC02H 8, 9 and 10 To a stirred mixture of Boc-prolinol (3.00 g, 14.9 nunol), ethyl phydroxybenzoate (2.40 g, 14.5 mnol), and triphenylphosphine (3.91 g, 14.9 mol) in THF (80 nL) 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.
The residue was purified by column chromatography on silica gel with EtOAc-n- hexane vlv) as eluent to give 4.88 g (93 O/o) ethyl (S)-4-(1-terf-butoxycarbonyl-2-pyrrolidinyl)methoxybenzoate 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 solution. The aqueous solution was washed with EtO2 (x2) and then acidified by the addition of 1 N HCI. 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-(l-tert-butoxycarbonyl-2-pyrrolidinyl)methoxybenzoic acid was added
CH
2 ClI (10 mL) and TFA (10 mL). The mixture was stirred at room temp for 1 hr EtO2 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 Fmoc-Cl (3.34 g, 12.9 mmol) was added to the solution, and the resulting mixture was stirred for 20 hr at room temp. The mixture was washed with EtO2 (x2) and aqueous layer was separated. The layer was acidified by the addition of 1 N HCI. The 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 (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-(l-Fmoc-2pyrrolidinyl) methoxybenzoic acid (500 mg, 1.13 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
CI
2 (7 mL). The mixture was shaken for 20 hr and drained. The resin was washed with DMF MeOH (x3),
CH
2 CIl (x3) and dried under a reduced pressure to give 522 mg of resin, which was used to prepare 8, 9 and 8 To the above resin (115 mg) was added a solution of piperidine-DMF (50 v/v, 4 mL) and the mixture was shaken for 1 hr. The resin was washed with DMF (x3), MeOH CHCI 2 To the resin was added DMF (4 mL), CH 2 CI, (2 mL), 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 MeOH CH,C1 2 To the resin was added a solution of TFA in
CH
2 CI, (50 v/v, 4 mL) and the mixture was shaken for 2 hr. The mixture was filtered and the 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 O 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 9 To the above resin (60 mg) was added a solution of piperidine in DMF (50 v/v, 3 mL) and the mixture was shaken for 2 hr. The resin was washed with DMF MeOH CHCIl To the resin was added DMF (2 mL), CH 2
CI
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 MeOH CH 2 CI, To the resin was added a solution of TFA in CH 2
CI
2 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 To the above resin (637 mg) was added a solution of piperidine in DMF (50 v/v, 20 mL) and the mixture was shaken for 4 hr. The resin was washed with DMF MeOH
CHCI
2 To the resin was added DMF (12 mL), CH 2 CI, (8 mL), 4-(Fmocamino)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 CH 2
CI
2 (x3) and dried under a reduced pressure to afford 617 mg of the resin. 57 mg 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 MeOH CH 2 CIl 2-chlorophenyl isocyanate (0.050 mL, 0.41 mmol) was added to a suspension of resin in THF (1 mL) and CH 2 ClI (1 mL). The mixture was shaken for 20 hr and drained. The resin was washed with DMF MeOH (x3),
CH
2
CI
2 To the resin was added a solution of TFA in CH 2 CI, (25 v/v, 2 mL) and the 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 l) Example 8 (S)-3-[2-[1-[3-methoxy-4-(N'-phenylureido)phenylacetyl]pyrrolidinyl]methoxy] phenylacetic acid WO 01/00206 PCT/US00/18079 0 2
H
H H 11 (S)-3-[2-[1-[4-[N'-(2-methylphenyl)ureido]phenylacetyl]pyrrolidinyl]methoxy] phenylacetic acid H H 12 11 and 12 To a stirred mixture of Boc-prolinol (3.51 g, 17.5 mmol), methyl m-hydroxyphenylacetate (2.90 g, 17.5 mmol), triphenylphosphine (4.60 g, 17.6 mol) in THF 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 by column chromatography on silica-gel with EtOAc- hexane v/v) as eluent to give 5.49 g methyl (S)-3-(l-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 mixture was extracted with EtO and the aqueous layer was acidified by the addition of 1 N HCI. The mixture was extracted with EtOAc. The extract was washed with water, brine, dried over MgSO, and then concentrated in vacuo to afford 4.43 g (88 (S)-3-(l-tert-butoxycarbonyl -2pyrrolidinyl)methoxy phenylacetic acid as a viscous oil.
A mixture of the above (S)-3-(l-tert-butoxycarbonyl-2-pyrrolidinyl) methoxyphenyl acetic acid in CH 2
CI
2 (10 mL) and TFA (10 mL) was stirred for 1 hr at room temp. EtO2 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 EtO2 and the aqueous layer was acidified by the addition of I N HCI. The mixture was extracted 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-(l-Fmoc-2-pyrrolidinyl)methoxyphenylacetic acid as a viscous oil.
WO 01/00206 PCT/US00/18079 Wang resin (0.71 mmol/g, 400 mg) was suspended in a solution of(S)-3-(1-Fmoc-2pyrrolidinyl) methoxyphenylacetic acid (520 mg, 1.14 mol), DMAP (35 mg, 0.29 mmol), HOBt mg, 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 MeOH
CH
2 CI, (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 MeOH CH 2
CI
2 To the resin was added DMF (1.5 mL), CH 2 CI, (1.5 mL) 3-methoxy-4-(N'-phenylureido) phenylacetic acid (42 mg, 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 MeOH CH 2
CI
2 To the resin was added a solution of TFA in CH 2 CIl (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, EtO was added to the residue and the solid was collected to afford 8 mg 11 as a crystalline solid. MS (FAB) m/z 518 (MI+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 MeOH CH 2 CIl To the resin was added DMF (1.5 mL), CH 2
CI
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 shaken for 15 hr and drained. The resin was washed with DMF MeOH CH 2 Cl, A mixture of the resin in TFA-CH 2
CI
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 O was added to the residue and the solid was collected to afford 11 mg 12 as a crystalline solid. MS (FAB) m/z 502 Example 9 4-[2-[1-[3-methoxy-4-(N' -phenylureido)phenylacetyl]-2-pyrrolidinyl]ethynyl] benzoic acid OOH 13 WO 01/00206 WO 0100206PCT/USOO/18079 To a stirred cold (minus 50 solution of N-boc-prolinal (5.98 g, 30 nimol) and PPh 3 (62.95 g, 240 mmnol) in CH 2
CI
2 (200 niL was slowly added a solution of CBr 4 (39.80 g, 120 nimol) in CH 2
CI
2 (50 niL), and the stirring was continued for 1 hr at 0 To this mixture was added sat.
NaHCO 3 and the mixture was extracted with CHCI 3 The extract was washed with H 2 0, dried over MgSO 4 and evaporated. The residue was chromatographed on silica-gel with CHC1 3 and nhexane-AcOEt 1, vtv) as eluent to give 7.84 g 1-(tert-butoxycarbonyl)-2-(2,2dibromoethenyl) pyrrolidine as colorless plates. nip 61-63; 111 (KBr) 1693 'H-NMR (CDC 3 8 1.46 (9 H, 1.72-2.19 (4HK in), 3.35-3.45 (2K-1 m),4.35 (lH, br 6.36 (lH, br MS (FAB) m/z 352, 354, 356, 358; Anal. Calcd for C 11
H,,NO
2 Br 2 C, 37.2 1; H, 4.83; N, 3.94. Found: C, 37.14; H, 4.83; N, 4.00.
To a stirred cold (minus 78 OC) solution of 1-Qtert-butoxycarbonyl)-2-(2,2dibromoethenyl) pyrrolidine (7.81 g, 22 nimol) in THE (200 niL) was added n-BuLi (1.59 M in hexane, 28 niL, 44 nimol) 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 CI and extracted with EtOAc. The extract was washed with brine, dried over MgSO 4 and evaporated. The residue was chronatographed on silica-gel with n-hexane-AcOEt (10: 1, v/v) as eluent to give 4.15 g l-(tert-butoxycarbonyl)- 2-ethynyl pyrrolidine as a light yellow oil. 'H-NMR (CDCI,) 8 1.48 (9 H, 1.82-2.21 (4 H, in), 3.30-3.45 (2 H, in), 4.41-4.52 (1 H, in).
A suspension of ethyl 4-iedobenzoate (1.7 niL, 10 nimol), Pd(PPh 34 (578 mng, 0.5 nimol), and Ciii (190 minol, 1 nimol) in i-Pr 2 NH (20 niL) was stirred for 0.5 hr under N 2 To this mixture was added a solution of 1 -Qert-butoxycarbonyl)-2-etliynylpyrrolidine (1.95 g, 10 mmnol) in i-Pr 2
NH
niL) for over 10 min. After stirring for 3 hr at room temip, 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 chromatographed on si lica-gel with n-hexane-AcOEt (10: 1, v/v) as eluent to give 2.77 g l-Qtert-butyloxycarbonyl)-2-(2-(4-cthoxycarbonylphenyl)ethynyl) pyrrolidine as a colorless oil. IH-NMvR (CDCI,) 8 1.37 (3 H, L, J=6.8 Hz), 1.49 (9 H, 1.85-2.12 (4 H, mn), 3.37- 3.51 (2 H, in), 4.37 (2 H, q, J=6.8 Hz), 4.54-4.77 (1 H, in), 7.44 (2 H, d, J=7.8 Hz), 7.96 (2 H, d, J=7.8 Hz) To a stirred solution of l-(terI-butoxycarbonyl)-2-[2-(4-ethoxycarbonylphenyl) ethynyll pyrrolidine (2.75 g, 8 nimol) in CH 2
CI
2 (5 niL) was added TEA (5 niL, and the resulting mixture was stirred overnight. The mixture was concentrated in vacuo and made basic with sat. NaHCO 3 WO 01/00206 WO 0100206PCT/USOO/18079 and extracted with CHC 3 The extract was washed with brine, dried over MgSO 4 evaporated to give 1.95 g 2-f 2-(4-ethoxycarbonylphenyl) ethynylipyrrolidine as a light yellow oil. 'H-NMR
(CDC
3 5 1.38 (3 H, t, J=6.8 Hz), 1.82-2.16 (4 H, in), 3.01-3.48 (2 H, in), 4.00-4.11(1 H, in), 4.37 (2H, q, J=6.8 Hz), 4.54-4.77 (1 H, in), 7.44-7.46 (2 H, in), 7.95-7.97 (2 H, mn).
A mixture of 3-methoxy-4-(N'-phenylureido)phenylacetic acid (180 mg, 0.6 mmnol), 2-(2- (4-ethoxycarbonylphenyl)ethynyl)pyrrolidine (146 mg, 0.6 mnmol), EDC (173 mg, 0.9 rnuol), DMAP (73 mg, 0.6 mmol), and cat. HOBt in DMF (10 mL) was stirred overnight. The mixture was poured into 1 N HCl and the solid was collected with suction. The residue was dissolved in CHCl 3 and dried over MgSO,. After removal of the solvent, the residue was chromatographed on silica-gel with CHCl 3 -MeOH (10: 1, v/v) as eluent to give 192 mg ethyl 4-[2-[l-[3-methoxy- 4-(N'-phenylureido)phenylacetylJ-2-pyfrolidinyl] ethyniyl]benzoate as a light yellow amorphous solid. 'H-NMvR (CDCl 3 8 1.38 (3 H, t, J=6.8 Hz), 1.98-2.24 (4 H, in), 3.48-3.89 (2 H, in), 3.53 (2 H, 3.62 (3 H, 4.33-4.40 (2 H, in), 4.78-5.04 (1 H, in), 6.77-8.00 (14 H, in).
To a stirred solution of ethyl -[3-methoxy-4-(N'-phenylureido) phenylacetylJ-2pyrrolidinyl] ethynyllbenzoate (184 ing, 0.35 nol) in THF (5 mL) was added 0.25 N NaOH (4 mQL. The resulting mixture was stirred overnight. The mixture was poured into H,0 and made acidic by the addition of 1 N HCl (1 inL). The solid was collected with suction and dissolved in CHCI,. The solution was dried over MgSO 4 and evaporated. The residue was recrystallized from
CHCI
3 -n-hexane to give 65 mng 13 as a white crystalline powder. mp 154-157; IR (KBr) 3346, 2952, 2615, 1712, 1693cm-'; 'H-NMvR (CDC1 3 8 1.92-2.29 (4 H, in), 3.32-3.82 (2 H, in), 3.78 (2 H, 3.80 (3 H, 4.87-5.11(1 H, in), 6.77-9.26 (14 H, in), 13. 10 (1 H, br MS (FAB) ,n/z 498 1).
Example 4-[2-f 1 -[3-inethoxy-4-[N'-(2-methylphenyl)ureido~phenylacetylJ-2-pyrrolidinylethynyl benzoic acid COCH 14 A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetic acid (1.45 g, 4.6 rnmiol), 2-(2-(4-ethoxycarbonylphenyl)ethynyl)pyrrolidine 12 g, 4.6 iniol), EDC 1.32 g (6.9 WO 01/00206 WO 0100206PCT/USOO/1 8079 mmol), DMAP (562 mg, 4.6 inmol) 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 CHCI, and dried over MgSO 4 After removal of the solvent, the residue was chromatographed on silicagel with CHCI,-MeOH (100: 1, v/v) as eluent to give 2.20 g ethyl 4-[2-[1-[3-methoxy-4-(N'- (2-methylphenyl)ureidoj phenylacetylJ-2-pyrrolidinyllethynylJbenzoate as a white amorphous solid.
IH-NMvR (CDC1 3 5 1.37-1.41 (3 H, in), 1.94-2.22 (4 H, in), 2.29 (3 H, 3.41-3.89 (2 H, in), 3.62 (3 H, 3.69 (2 HL 4.34-4.40 (2 H, in), 4.72-5.01 (1 H, mn), 6.33 (1 H, br 6.80-8.06 (12 H, in).
To a stirred solution of ethyl l-[3-methoxy-4-(N' -(2-methylphenyl)ureido~phenyl acetylJ-2-pyrrolidinyl~ethynyl]benzoate (2.16 g, 4 minol) in THF (30 inL) 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,. The solution was dried over MgSO, and evaporated. The residue was recrystallized from CHCI 3 -n-hexane to give 555 mg 14 as a white crystalline powder. mp 16 1-164; IR (KBr) 3338, 2954, 2875, 1707, 1691 cm-; 'H-NMR (CDC 3 8 1.96-2.10 (4 H, in), 2.24 (3 H, 3.32-3.81 (2 H, in), 3.62 (2 H, 3.81 (3 H, 4.87-5. 10 (1 H, in), 6.76-8.58 (13 H, in); MS (FAB) m/z 5 12 (NC+ 1) Example 11 1-[3-inethoxy-4-[N'-(2-methylphenyl)ureido~phenylacetylJ-2-pyrrolidinyl]ethynyl] phenylacetic acid H. HCOOH is A mixture of 3-inethoxy-4fN'-(2-methylphenyl)ureidolphenylacetic acid (141 ing, 0.45 inmol), 2-[2-(3-ethoxycarbonylinethylphenyl)ethynyllpyrrolidine (116 ing, 0.45 iniol), EDC (130 mng, 0.68 inmol), DMAP (55 mg, 0.45 ninol), cat. HOBt in DMF (10 niL was stirred overnight.
The mixture was poured into 1 N HCl 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 mL). 0.25 N NaOH (4 mL) was added to this solution with stirring. After stirring overnight, the mixture was poured into IN HCI (2OmQ). The resulting precipitate was collected with suction and dissolved in CHC1 3 The solution was dried over MgSO 4 and evaporated. The residue was WO 01/00206 WO 0100206PCT/USOO/18079 chromatographed on silica-gel with CHCI 3 -MeOH 1, v/v) as eluent to give 92 mg 15 as a white amorphous solid. 'H-NMR (CDCI 3 8 1.96-2.18 (7 H, in), 3.50-3.88 (9 H, in), 4.78-4.98 (2 HK 6.72-7.99 (14 H, in); MS (FAB) m/z 526 1).
Example 12 4-fl1-[4-[N'-(2-methylphenyl)ureidolphenylacetylJ-2-pyrrolidinylinethoxy]isophthalic acid OOH 16 To a stirred solution of pentafluorophenyl ester of 4-[N'-(2-methylphenyl)ureidoJ phenylacetic acid (2.32 g, 5.15 mniol), diinethyl (S)-4-(2-pyrrolidinylmethoxy)isophthalate (1.51 g, 5.15 nimol) in DMF (20 niL) was added Et 3 N (1.0 mL, 6.65 mmol), and the mixture was stirred 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 CHCI 3 -MeOH (10: 1, v/v) as eluent to give 1.58 g (550%) dimethyl 1-[4-[N'-(2-methylphenyl)ureidoJ phenylacetylJ-2-pyrrolidinylinethoxy~isophthalate as a yellow crystalline solid. 'H-NMR (CDCI 3 8 1.87-2.25 (mn, total 7 3.50-3.65 (in, 4 3.85 3 3.89 3 4.18-4.31 (in, 2 4.44 (in, 1 6.95-8.45 (in, total 13 H).
To a stirred solution of dimethyl 4-[l-[4-[N'-(2-methylphenyl)ureido] phenylacetyl] pyrrolidinylmethoxy]isophthalate (1.56 g, 2.79 mmnol) in THE (30 mL) was added 0.25 N NaOH mL), and the reaction mixture was heated under reflux overnight. The resulting mixture was poured into I N HCI, and solid was collected. The crude solid was washed with EtO to give 574 mg 16 as ayellow amorphous solid. LR(KBr) 1710cm-'; 'H-NMR(DMSO-diJ)8 1.83-2.18 (in, 4 2.24 3 3.36-4.28 (mn, 8 H) 6.91-9.02 (series of in, total 13 12.89 (hr s, 1 H); MS (FAB) m/z 532 (Mr+ 1).
Example 13 44[24 l-(3-methoxy-4['-(2-nethylphenyl)ueidojphenylacetyl-2-pyrrolidinyl] ethenylJ benzoic acid OOH 17 WO 01/00206 WO 0100206PCT/USOO/18079 To a stirred solution of the 4-[2-[2-(NV-tert-butoxycarbonyl)pyrrolydinylethenyl benzonitrile (2.26g, 7.57mmol) in CH 2
CI
2 (23 niL was added dropwise a 1.5M solution of diisopropylaluininum hydride (toluene solution) (6.O6niL, 9.O9mmol) at 0 0 C for over 15 min.
The resulting solution was stirred for3 hr at 0 0 C. The solution was quenched by the addition of sat.NH 4 CI. The resulting mixture was filtered through Celite, and the filtrate was extracted with EtOAc. The filtrate was washed with brine, dried over NaSO 4 and evaporated in vacuo to afford 1.89 g 4-[2-[2-(N-Iert-butoxy carbonyl)pyrrolidiniyljethenylJ 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 of AgNO 3 (2.13g, 12.5mmol) in CH3CN (10 mL) at 0 0 C for over 0.5 hr. To the stirred above mixture was added dropwise a solution of 4-12-[2-(N-Ierl- butoxycarbonyl) pyrrolydinyijethenyl] benzaldehyde (1 .89g, 6.27mmol) in CH 3 CN (lOmL) 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 acidified by carefully adding 1 N HC1, 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) pyrrolidinyllethenyl] 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 immol) in MeOH-benzene(l vfv, 3OmQ) was added dropwise a 2 M-n-hexane solution of TMSCHN 2 (1.32mL, 2.65nminol) 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(l v/v) as eluent to afford 0.64g (88%) methyl 4-[2-[2-(N-teri-butoxycarboriyl) pyrrolidinyll ethenyl]benzoate as a pale yellow crystalline material.
To a stirred solution of methyl 4-[2-[2-(N-tert-butoxycarbonyl)pyrrolidinyl ethenyl] benzoate (0.64g, 1. 93mmol) in CH 2 Cl 2 (5mQL 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 CHIM 3 The extract was dried over Na 2
SQ
4 and evaporated in vacuo to afford 0.45g methyl 4-[2-(2-pyrrolidinyl)ethenyllbenzoate as a yellow crystalline material.
WO 01/00206 WO 0100206PCT/1JSOO/18079 To a stirred mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetic acid (285mg, 0.9O6mmol), methyl 4-[2-(2-pyrrolydinyl)ethenyl~benzoate (2 10mg, 0.9O6mmol) in DMF (4mL was added 1-ethyl-3-(3-dmethylaminopropyl)carbodiimide (EDC) (209mg, 1.O9mmoI), 1hydroxybenzotriazole (HOBt) (147mg, 1.O9mmol) and 4-dimethylanuinopyridine (DMAP) (11lmg, 0.0906mmol) at room temp. After the resulting mixture was stirred for 48 hr at room temp, the mixture was poured into ice- I N HCI and extracted with EtOAc. The extract was dried over Na 2 SO, and evaporated in vacuo. The residue was chromatographed on silica-gel with acetonetoluene (1:4 to 1: 1, v/v) as eluent to afford 0.47g methyl 4-[2-[1-[3-methoxy-4-IN'-(2methylphenyl)ureido] phenylacetyl]-2-pyrrolidinyllethenyl]benzoate as a white crystalline material.
To a stirred solution of methyl 1 [-[-[3-methoxy-4-[N' -(2-methylphenyl) ureidoiphenyl acetylJ-2-pyrrolidinyllethenyljbenzoate (0.47g 0.89 immol) in THF (5mL) was added 0.25 N NaOH (5.36miL). The resulting mixture was stirred at room temp for 20 hr. The mixture was acidified with IN HCL The mixture was extracted with EtOAc. The extract was washed with brine, dried over Na 2
SQ
4 and evaporated in vacuo to afford 430mg 17 as a white crystalline material. 'H-NMvR (400MHz, DMSO-d~j 8 1.78-2.13 (4H, in), 2.50 (311, 3..44 -3.68 (4H, in), 3.75 and 3.82 (3H, 4.71 (11H, in), 6.26-8.59 (15H, in).
Example 14 4-12-[ 1-[3-methoxy-4-(N'-phenylureido)phenylacetylJ-2-pyrrolidinyl]ethenyl benzoic acid OOH i To a stirred solution of 3-methoxy-4-(N'-phenylureido)phenylacetic acid (305mg, l.Oimmol), methyl 4-[2-(2-pyrolydinyl)ethenylJbenzoate (235mg, l.Olmmol) in DMF (4mL) was added 1 -ethyl-3-(3-dimethylaminopropyl)carbodiimide (232mg, 1.21 mmol), 1hydroxybenzotniazole (HQBt) (164mg, 1.2 lmmol), and 4-dimethylaminopyridine (DMAP) (12mg, 0. l0imniol) at room temp. After the mixture was stirred for 48 hr, the mixture was acidified by the addition of 1 N HCI. 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 methyl 1-[3-methoxy-4 -(NV'-phenylureido) phenylacetylJ-2-pyrrwlidinyl]ethenylj benzoate acid as a white crystalline material.
WO 01/00206 WO 0100206PCT/LJSOOII 8079 To a stirred solution of methyl 1-[3-methoxy-4-(N' -phenylureido) phenylacetylJ-2pyrrolidinyllethenyllbenzoate 0.43g, .837ntmoI) in THF (5mL) was added a solution of 0.25 N NaQH (5.04mL) at roam temp. After the resulting mixture was stirred for 20 hr, the mixture was acidified by the carefully addition of 1 N HCI. The mixture was extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 and evaporated in vacuo to afford 3 97mg 18 as a white crystalline material. 'H-NMR (400MHz, DMSO-d 6 8 1.78-2.13 (4H, in), 3.17-3.68 (4H, in), 3.74, 3.82 (3H, 4.71 (1H, in), 6.27-9.28 (16H, in).
Example 4-[2-f l-[3-methoxy-4-[f'-(2-methylphenyl)ureidoJphenylacetylJ-2-pyrrolidinyllethyllbenzoic acid OOH 19 A mixture of 4-[2-f 1-[3-inethoxy-4-[N'(2-methylphenyl)ureidolphenylacetyl]-2pyrrolidinyl] ethenylibenzoic: acid (184mg, 0.358nunol) and 5% Pd-C(368mg) in MeOH was hydrogenated in an atmospheric pressure at roam temp. After the mixture was stirred for 21 hr at roam temp, insoluble catalyst was filtered off and the filtrate was evaporated in vacuo. The residue was chromnatographed on silica-gel with MeOH-CHCI 3 (1:4 to 1:3, v/v) as eluent to afford 123mg 19 as a white crystalline material. 'H-NMR (DMSO-d4) 8 1.55-2.03 (in, 6H), 2.24 3H4), 2.60 (in, 2H), 3.17-3.59 (in, 4H), 3.83 3H), 3.97 (in, 114), 6.6 1-8.57 (in, 13H); MS (FAB) mfz 516 1).
Example 16 3-fl -[3-inethoxy-4-[N '-(2-methylphenyl)ureido]phenylacetylJ-2-pyrrolidiny] methylthiobenzoic: acid H H To a solution of m-iodophenol (20.0 g, 90.9 mmol) in DMFW (200inL was added 1,4diazabicyclo [2,2,2joctane (20.4g, 181.8 inmol) and dimethylthiocarbamoyl chloride (16.9 g, 136.4 minol). The resulting cloudy solution was stirred for 0.5 hr at 35*C and then heated at 75'C for hr. After coaling, 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 O-m-iodophenyl diinethylthio carbamate as a pale yellow crystalline powder. JR (KBr) 1540, 1463, 1278, 1193, 1166, ll24cm"; IH-Nl'vR (400 M~z, CDCI,) 8 3.3 3 3H), 3.45 7.05 7.14 (in, 2Hf), 7.43 J 1.9 Hz, WO 01/00206 WO 0100206PCT/US018079 7.58 (dd, J 1.0, 7.8 Hz, 1H), MS (FAB) m/z 307 Anal. Calcd for C 9 Hj 0 1NOS: 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 nunol) in Ph 2
O
was heated at 230*C for 10 hr. After cooling, the reaction mixture was chromatographed on silica-gel with n-hexane-EtOAc 1, v/v)as eluent to give 9.31 g S-m-iodophenyl dimetliylthio carbamnate as a pale yellow oil. 'H-NMR (400 MIRz, CDCI 3 5 3.08 (br s, 6H), 7.11 (t, J 7.8 Hz, 1H), 7.4.6 J1= 7.3 Hz, 1H), 7.71 J =7.3 Hz, 1H), 7.85 IH); MS (FAD) m/z 307 To a solution of S-m-iodophenyl dimetliylthiocarbamate (5.01 g, 16.31 minol) in MeOH (20 mL) was added 28 0 /o-MeONa in MeOH (3.46 inL, 17.94mmol). The resulting mixture was stirred at room temp for 3.5 hr and then heated at 70'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
SO
4 The organic layer was concentrated under a reduced pressure. The residue was chromatographed on silica-gel with nhexane-AcOEt (10: 1, v/v) as eluent to afford 3.42 g m-iodothiophenol as an oil. 'H-NVR (400 M1Hz, CDCI,) 8 3.45 1H), 6.95 J 7.8 Hz,H),7.23 (dJ 7.8 Hz 1H), 7.48 J 7.3 Hz,lIH), 7.64 J =1.5 Hz,LIH); 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 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 1, v/v) as eluent to afford 5.76 g (81 N-Qtert-butoxycarbonyl)-2pyrrolidinylmethyl p-toluenesulfonate as a colorless oil. IH-NMR (400 MHz, CDCI,) 8 1.36 and 1.41 (s each, total 9H), 1.82 (br m, 2H), 1.96 (br m, 2H), 2.44 3H), 3.30 (br m, 2H), 3.89 (br s, 1H), 3.96 (br s, 1H), 4.09 (br m, lH), 7.34 (br s, 2H), 7.77 J= 8.3 Hz, 2H); MS (FAD) m/z 356 1).
To a stiffed mixture of m-iodothiophenol (2.67 g, 11.31 mmol) and N-QIert-butoxy carbonyl)-2- pyrrolidinylmethyl p-toluenesulfonate (3.34 g, 9.43 mmol) in pyridine (9.4 mL) was added 8N KOH (1.77 mL). The resulting midxture was stirred at room temp overnight. The reaction WO 01/00206 WO 0100206PCTUSOOI18079 mixture was diluted with EtOAc. The solution was washed with H120, sat. NHCI 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 1, vtv) as eluent to afford 1.79 g (451/) [N-QIerz-butoxycarbanyl)-2-pyrrolidinyllmethyl 3-iodophenyl sulfide as an oil. IH-NMR (400 MI-Iz, CDCI,) 8 1.45 911), 1.78 2.01 (br m, 4M1, 2.71 (dt, 114I), 3.32 3.49 (br m, 314), 3.90 4.02 (br m, 114), 7.12 J= 7.8 Hz, 1H), 7.18 7.8 Hz, 111), 7.57 (dd, J= 2.0, 8.3 Hz, 2H); MS (FAB) m/z 420 1).
To a stirred solution of [1-(tert-butoxycarbonyl)-2-pyrrolidinyllmethyl 3-iodophenyl sulfide (1.76 g, 4.20 mmol) in DMSO (20 mL) and MeOH (16 mL) was added Et 3 N (1.28 mL, 9.24 inmol), Pd(OAc) 2 (47.1 mg, 0.21 mmol), and l,3-bis(diphenylphosphino)propane (86.6 mg, 0.2 immol), 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 1, v/v) as eluent to afford 1.28 g (87 methyl 3-[I1.Qert-butoxycarbonyl)-2-pyrrolidinyllmethylthiobenzoate as an oil. 'H- NMvR (400 MHz, CDCl 3 5 1.42 and 1.45 (each s, 9H), 1.79 2.05 (br m, 4H) 2.83 (dt, J 10.8, 30.3 Hz, 114), 3.34 3.54 (br m, 314), 3.92 3H1), 3.92 and 4.05 J 7.8 Hz, 114), 7.36 J 7.8 Hz, 114), 7.63 (br d, J 14.7 Hz, 11H), 7.83 (br d, J 12.7 Hz, 111), 8.04 111); MS (FAB) ,n/z 352 To a stirred solution of methyl 3-[1-(tert-butoxycarbonyl)-2-pyrrolidinylI methyltbio benzoate (1.46 g, 4.16 mmol) in CH 2
CI
2 (30 mL) was added TEA (15 niL) at 0 0 C. 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 IN NaOH and extracted with CHCI,. The extract was washed with brine, dried over NaSO,, and concentrated under a reduced pressure to afford 947 mg (9 methyl 3-(2-pyrrolidinyl) methylthio benzoate as a brown oil. 'H-NMR (400 MHz, CDCI 3 5 1.45 1.54 (in, 111), 1.72 2.00 (in, 411) 2.88 3. 10 (in, 411), 3.30 (in, 111), 3.92 311), 7.34 J =7.8 Hz, 1H), 7.52 (in, 111), 7.84 (in, 111), 8.01 J= 2.0 Hz, 111); MS (FAR) m/z 252 The mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetic acid (1.18 g, 3.77 minol), EDC (1.08 g, 5.65 inmol), DMAP (23 mg, 0. 19 nunol) and HOBt (25 mg, 0. 19 minol) in DMF (5 inL) was stirred at room temp for lhr methyl 3-(2-pyrrolidinyl)methylthio benzoate (947 mg, 3.77 rnmol) was added to the mixture and the resulting mixture was stirred overnight. After WO 01/00206 WO 0100206PCTIIJSOO/18079 DMAP (460 mg, 3.77 mmol) and HOBt (835 mg, 6.18 mmol) was added and stirred for a further hr. The reaction mixture was diluted with EtOAc. The solution was washed with brine and dried over Na 2
SQ
4 The solvent was removed under a reduced pressure. The residue was chromatographed on silica-gel with n-hexane-EtOAc v/v) a eluent to afford 294.3 mg methyl 3-[1 -[3-methoxy-4-[N -(2-methylphenyl)ureido Jphenylacetyl]-2-pyrrolidinyllmethylthio benzoate as a pale yellowish amorphous. IR (KBr) 2875, 1724, 1620, 1284, 1 l82cm- 'H-NMR (400 M1Hz, CDCI 3 8 1.86 -2.05 (in, 4H), 2.31I(s, 314), 2.84 (dd, J= 9.3, 13.7 Hz, 11H), 3.43 3.59 (m,514), 3.73 314), 3.92 3H), 4.33 (in, 11H), 6.16 1H), 6.77 6.80 (mn, 2H), 7.04 lH), 7.16 J= 8.3 Hz, lH), 7.38 J1=7.8 Hz, 7.49 J= 7.8 Hz, 1H), 7.73 (dt, J= 1.0, 7.8 Hz, 1H4), 7.79 (dd, J 2.0, 6.8 Hz, lH), 8.01 J 2.0 Hz, 1H), 8.05 (dd, J 2.4, 7.8 Hz, 1H); MS (FAB) m/lz 548 1).
To a stirred solution of methyl 3-[I-[3-methoxy-4-[N'-(2-methylphenyl)ureidoj phenylacetyl]-2-pyrrwlidinyl]methylthiobenzoate (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 inmol), and the resulting mixture was stirred for 9 hr at room temp. The mixture was treated with IN HCI and extracted with CHCI,. 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 CHCI 3 -MeOH (10: 1, vlv), and crystallized from n-hexane-EtQAc to afford 89.7mg 20 as a white powder. IR (KBr) 2960, 1708cm'; 'H-NMvR (400 M4Hz, DMSO-d 6 8 1.82 2.01 (mn, 4H), 2.24 311), 2.93 (dd, J 9.3, 12.7 Hz, 111), 3.40 -3.54 (in, 5H), 3.86 3H), 4.13 (br m, 1H), 6.74 J= 8.3 Hz, 1H), 6.87 (d, J1= 1.5 Hz, 1H), 6.94 J= 7.8 Hz, 114), 7.10 7.17 (in, 2H), 7.42 J 7.8 Hz, 1H), 7.70 J 7.8 Hz, 1H), 7.74 J 8.3 Hz, 1H), 7.79 J 7.8 Hz, IN), 7.84 IH), 8.00 J1= 8.3 Hz, 1H), 8.48 1H), 8.57 IH); MS (FAB) m/z 534 Example 17 3-[I -[3-methoxy-4-[N '-(2-methylphenyl)ureidoJphenylacetyll-2-pyrrolidinyljmethylsulfonylI benzoic acid
~N()COOH
H H 21 To a stirred solution of methyl 3-[Il-[3-methoxy-4-[N'-(2-methyl phenyl)ureidojphenyl acetyll-2-pyrrolidinyllmethylthio~benzoate (131.8 mg, 0.241 inmol) in CH 2
CI
2 (3 inL) was added m-CPBA (130.5 mg, 0.529 mmol) at 0 0 C. The reaction mixture was stirred at room temp for hr. The mixture was diluted with CHC1 3 and quenched with sat. Na 2
S
2
O
3 solution The organic WO 01/00206 WO 0100206PCT/USOO/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-[[I-[3-methoxy-4-[N'-(2-methylphenyl) ureidolphenyl acetyll-2-pyrrolidinylJ-methylsulfornyllbenzoate as an amorphous solid. To a stirred solution of this crude compound in THF (7.4 mL) and H420 (1.8 mL) was added LIOH (17.3 mg, 0.723 mmol), and the stirring was continued overnight at room temp. The reaction mixture was diluted with CHC1 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 CHCI 3 MeOH (10: 1, v/v) as eluent, and the crude solid was recrystallized from n-hexane-EtOAc to afford 69.9 mg (5 21 as a white crystalline powder. mp 243 -245; IR (KBr) 3354, 2974, 1533cm-f; 'H-NMR (400 DMSO-d) 8 1.80 2.00 (in, 414), 2.24 Cs, 314), 3.19 3.62 (in, 614), 3.82 (s, 3H), 4.18 (in, 111), 6.67 J =8.8 Hz, 114), 6.80 J= 1.0 Hz, 114), 6.93 J1= 7.3 Hz, 114), 7. 7.17 (in, 214), 7.66 J =7.8 Hz, 114), 7.78 J 8.3 Hz, 114), 7.91 7.99 (in, 214), 8.20 J 7.3 Hz, 114), 8.34 114), 8.48 114), 8.56 114); MIS (FAB) m/z 566 (NV+ Anal Calcd for C29I4 31
N
3
O
7 SdHCl1IH 2 O: C, 56.17; H, 5.53; N, 6.78. Found: C, 55.92; H,5.58; N, 6.71.
Exampl~e 18 4-1(1 -methoxy-4-[N '-(2-methylphenyl)ureidolphenylacetylJ-2-pyrrolidinyl] methylsulfonyl] benzoic acid PHHN OOH 2 To a stirred solution of methyl 4-[[1-[3-methoxy-4-IN'-(2-methyl phenyl)ureidoJ phenylacetyl]-2-pyrrolidinyllmethylthio~benzoate (300 mg, 0.548 mmol) in CHC1 2 (6 inL) was added m-CPBA (297 mg, 1.206 nimol) at 0 0 C, 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-[I1-13-methoXY4-[N'-(2methylpheryl)ureido]phenylacetyl]-2-pyrrolidinylJ-methylsulfonyljbenzoate as a crude yellow oil.
To a stirred solution of this crude compound in THE C4.4 mL) and 1420(1. IlmL) was added LiQH (39.4 mg, 1.643 inmol), 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 eluting with CHCI 3 -MeOH (10: 1, and crystallized from n-hexane-EtOAc to afford 128.0 mng 22 as a white powder. IR (KBr) 3388, 2974, 1537, 1298, 1 155cm'; 'H-NMR (400 Mflz, DMSO-Q~ 8 1.80 1.98 (Cm, 414), 2.24 314), 2.54 114), 3.20 3.70 Cm, 514), 3.82 3H4), 4.16 WO 01/00206 WO 0100206PCT/USOO/18079 (brm, I1H), 6.67 (dd,j= 1.5, 8.3 Hz, 1H), 6.80 1.5Hz, 1H), 6.93 7.3Hz, LH), 7.10 7.16(in, 2H), 7.78 J =7.3 Hz, 1H), 7.95 J= 8.3 Hz, 2H4), 7.98 J=8.3 Hz, 1H), 8.14 (d, J 8.3 Hz, 211), 8.49 1H), 8.57 lH); MS (FAB) ni/z 566 (M*+l);AnaL. Calcd for
C
29
H
31
N
3
O
7 S-3H 2 O: C,56.21; H, 6.02; N, 6.78. Found: C, 56.76; H,5.37; N, 6.70.
Example 19 1-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetyl] -2-pyrrolidinyl] methyithiol benzoic acid
NN
23 To a stirred solution of p-iodophenol (20.0 g, 90.9 mmol) in DME (200 mL) was added 1,4-diazabicyclo 12,2,2loctane (20.4g, 181.8 inmol) and dimethyithiocarbamoyl chloride (16.9 g, 136.4 mniol). The resulting solution was stirred for 3.5 hr at 75'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
SQ
4 and evaporated under a reduced pressure. The crude solid was recrystallized from H 2 0 to give the 26.75 g O-p-iodophenyl dimethylthiocarbaniate as a pale a yellow crystalline powder. IR (KBr) 1479, 1207, 827cm-1; IH- NN~IR (400 MfHz, CDCI,) 8 3.37 3H), 3.45 3H4), 6.83 J 8.8 Hz, 2H), 7.69 J 8.3 Hz, 2H); MS (FAB) m/z 307 Anal. Caled for C 9
H,
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 dimethylthiocarbaxnate (10.0 g, 32.6 inmol) in Ph 2
O
(25mL) was heated at 230*C for 5.5 hr. After cooling, the reaction mixture was chromatographed on silica-gel with n-hexane-EtOAc 1, v/v) as eluent to give 2.55 g S-p-iodophenyl dimethylthio carbamate as awhite crystalline powder. IR (KBr) 3299, 1651, 1469, 137lcm-'; 'H- NMR (400 MHz, CDCI,) 8 3.03 (br s, 3H), 3.08 (br s, 3H), 7.21 J 8.3 Hz, 2H), 7.70 J= 8.3 Hz, 2H); MS (FAB) m/lz 308 Anal. Calcd for C 9 H4, 0 INOS: C,35.19; H,3.28; N,4.56.
Found: C,35.49; H,3.28; N, 4.43.
To a solution of S-p-iodophenyl dimethylthiocarbamate (2.55 g, 8.31 mmnol) in MeOH mQL was added MeONa (495 mng, 9. l4mmol), and the resulting mixture was stirred at 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,0, brine, and dried over Na 2
SO
4 The organic layer was concentrated under a reduced pressure and the residue was WO 01/00206 WO 0100206PCT/USOO/18079 chromatographed on silica-gel with n-hexane EtOAc 1, vlv) as eluent to afford 1.75 g 89 piadothiophenol as a pale yellow crystalline solid. IR (KBr) 2559, 1097, 1002, 806cm'; 'H-NMR (400 MHz, CDCI 3 5 3.43 I1H), 7. 10 J 8.3 Hz, 2H), 7.53 J 8.3 Hz, 2H); MS (FAB) m/z 236 AnaI. Calcd for C 6
H
5 IS: C, 30.53; H, 2.13. Found: C, 30.57; H, 2.15.
To a stirred mixture of p-iodothiophenol (1.75 g, 7.43 mnzol) and N-Qterlbutoxycarbonyl)-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. NHICI, brine, and dried over Na 2
SO
4 The organic layer was concentrated under a reduced pressure. The residue was chromatographed on silica-gel with n-hexane-EtOAc 1, vlv) as eluent to afford 1.49 g (531/) [N-QIert-butoxycarbonyl)-2-pyrrolidinyllmethyl 4-iodophenyl sulfide as a pale yellowish oil. 'H-NMR (400 MHz, CDCl 3 8 9H), 1.78 2.01(br m, 4H), 2.71 (dt, 1H), 3.3 2 3.49 (br mn, 3H), 3.90 4.02 (br m, 1H), 7.12 J 7.8 Hz,L1H), 7.18 (dJ= 7.8Hz, 57(dd,J 2 .0,8.3Hz,2H);MS(FAB)m/z 420(M*+1).
To a stirred solution of [1-QIert-butoxycarbonyl)-2-pyrrolidinylJ methyl 4-iodophenyl sulfide (1.49 g, 3.56 nunol) in DMSO (16 mL) and MeOH (13mb) 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 muin, and the miixture was stirred at 70*C overnight. After cooling, the mixture was concentrated to a small volume. 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 the residue was chromatographed on silica-gel with n-hexane-EtOAc 1, v/v) as eluent to afford 1. 16 g methyl 4-f l-(tert-butoxycarbonyI)-2pyrrolidinyllmethylthiobenzoate as an oil. 'H-NMR (400 MlHz, CDCI 3 5 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 (in, 3H), 3.89 3H), 4.03 (br d, J 27.3 Hz, 1H), 7.38 J 7.3 Hz, IH), 7.47 J 7.3 Hz, 111), 7.92 (br s, 2H); MS (FAB) m/z 352 1).
To a stirred solution of methyl 4-[1-Qtert-butoxycarbonyl)-2-pyrrolidinyl~methylthio benzoate 16 g, 3.32 mmol) in CH 2 Cl 2 (20 rub) 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 was treated with IN NaOH. The mixture was extracted with CHCl 3 The extract was washed with brine, dried over KOH, and concentrated under a reduced pressure to afford 767 mg methyl WO 01/00206 WO 0100206PCT/USOO/18079 4-(2-pyrrolidinyl) methylthio benzoate as a yellow oil. 'H-NMR (400 MIlz, CDCl 3 8 (dt, J 3.9, 12.7 Hz, 111), 1.85 2.09 (in, 2H) 2.13 (in, 114), 3.11 3.27 (mn, 3H), 3.40 (dd, J 6.8, 13.2 Hz, 114), 3.54 (dd, J= 7.3, 15.1 Hz, 114), 3.89 311), 5.07 (br, 111), 7.38 J =8.3 Hz, 2H), 7.91 (d, J 8.3 Hz, 2H1); MS (FAB) m/z 252 To a stirred mixture of 3-methoxy-4-IN'-(2-methylpheniyl)ureidolphenylacetic acid (1.30 g, 4.136 mmol), Et 3 N (0.63 mL, 4.549 mniol) in DMF (20 inL) 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 inL) and precipitate was collected with suction. The crude solid was washed with 0.1IN HC1, H20, n-hexane, and dried at 40*C to afford 1.91 g pentafluorophenyl 3methoxy-4-[N'-(2-methylphenyl) ureidoiphenylacetate as a pale brownish crystalline powder. IR (KBr) 1785, 1224, 1216cm-'; 'H-NMR (400 MHz, CDCI 3 82.29 3H), 3.76 (s,314), 3.90 2H), 6.49 IH), 6.81 J 1.5 Hz, 1H), 6.91 (dd, J 8.3 Hz, 114), 7.15 J =7.3 Hz, 3H1), 7.24 (in, 111), 7.50 J= 7.8 Hz, 11H), 8.17 (d,J =7.8 Hz, IH); MS (FAB)m/tz 481 AnaL.
Calcd for CH3N 3 0 5 Sd /4H,0: C, 57.5 1; H, 3.57; N, 5.83. Found: C, 57.40; H,3.75; N, 5.68.
A mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureidoJ phenylacetate (1.47 g, 3.05 mmol), methyl 4-(2-pyrrolidinyl)methylthiobenzoate (767 mg, 3.05 inmol), Et 3
N
(0.51 mL, 3.66 mmol) in DMIF (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 v/v) as eluent to afford 1.366 g methyl 4-[[1-[3-methoxy-4-[N'-(2methylphenyl)ureido~phenylacetylJ-2-pyrrolidinylJ methylthiolbenzoate as a white crystalline powder. IR (KBr) 1785,1224, 1216cm'; 'H-NMR (400 MHz, CDC1 3 8 1.88 1.99 (in, 411), 2.30 311), 2.75 (dd, J 13.2 Hz, 111), 3.43 -3.55 3.56 2H), 3.64 (dd, J= 1. 1, 14.2 Hz, 111), 3.73 311), 3.88 311), 4.33 (mn, 111), 6.29 111), 6.78 6.81 (in, 211), 7.11 7.26 (mn, 511), 7.50 J 8.3 Hz, 311), 7.93 J 8.8 Hz, 211), 8.07 J 7.8 Hz, 111); MS (FAB) Mn/z 548 AnaL. Calcd for C3DH 33
N
3 0 5 SdI/4H 2 0: C, 65.26; HL 6.12; N, 7.61. Found: C, 65.48; H,6.20; N, 7.47.
To a stirred solution of methyl 4-[[l-[3-methoxy-4-[N'-(2-methylphenyl)ureido phenylacetylJ.2-pyrrolidinyllmethylthiolbenzoate (300 mg, 0.548 innol) in THE (5.5 inL) and 1120 1 mL) was added LiOH (39.4mg, 1.643 innol), and the reaction mixture was stirred at room temp overnight and at 50*C for 9 hr. The mixture was diluted with CHCI 3 The solution was WO 01/00206 WO 0100206PCT/USOO/18079 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-EtOAc-MeOH to afford 218.6 mg 23 as a white crystalline powder. IR (KBr) 33 18,2952, 1596, 1536, 1299, 1155cm'; 'H-NMR (400 M4Hz, DMSO-dj) 5 1.82 2.05 (in, 4H), 2.25 3H), 2.91 (dd, J =9.8, 13.2 Hz, 1H), 3.47 3.52 (in, 3H), 3.57 211), 3.87 31H), 4.14 (br mn, 1H), 6.76 J 1.5, 8.3 Hzd, 114), 6.89 J= 1.5 Hz, 1H), 6.94 J =7.3 Hz, 1H), 7.11 -7.19 (mn, 21H), 7.57 J =8.3 Hz, 211), 7.80 J =8.3 Hz, 111), 7.83 J =8.3 Hz, 211), 8.02 J =8.3 Hz, 1H), 8.49 1H), 8.58 1H); MS (FAB) m/z 534 Anal. Cajcd for C2,H- 31
N
3
Q
5 S-5/4H- 2 O: 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.
Example 4-[[I-13-inethoxy-4-[A'-(2-inethylphenyl)ureido~phenylacetyl]-2-pyrrolidinyl methylsulfinyl] benzoic acid H9H OOH 2 To a stirred solution of methyl 4-[Il-[3-methoxy-4-[N'-(2-inethyI phenyl)ureidoj phenylacetylJ-2-pyrrolidinyllinethyltbiojbenzoate (264 ing, 0.482 inmol) in CH.Cl, (5.2 mL) was added mn-CPBA (118.8 mg, 0.482 minol) at 0 0 C, and the mixture was stirred at room temp for I hr. The mixture was diluted with CHCl 3 and quenched with sat NaS%0 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-inethylphenyl)ureido] phenylacetyl]-2-pyrrolidinyl~inethylsulfinylJ benzoate as a crude amorphous solid. To a stirred solution of this crude compound in THF (4 mL) and H 2 0 (lmL) 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 HCI, brine, and dried over NaSO,. The solvent was removed under a reduced pressure, and the obtained crude solid was recrystallized from n-hexane-CHCI 3 -MeOH to afford 193.2 mg 24 as a white crystalline powder. JR (KBr) 3338, 2956, 1708, 1529, 1299, 1207, 1 155cm-'; 'H-NM (400 M[Rz, DMSO-d) 8 1.70 2.06 (mn, 4H), 2.24 3H), 2.90 (dd, J1 8.3, 13.2 Hz, 111), 3.02 3.08 (in, 111), 3.16 3.25 (in, 11H), 3.41 3.60 (in, 311), 3.84 311), 4.40 (br s, 1H), 6.74 J= 7.8 Hz, lH), 6.87 IH), 6.94 J= 7.3 Hz, 111), 7.11 -7.17 (mn, 211), 7.75 7.81 (in, 3H), 7.98 8.05 (mn, 111), 8. 10 J 8.3 Hz, 211), 8.46 lH), 8.56 111); MS (FAB) m/lz 550 572 Anal. Calcd for C 2
H
31
N
3
O
6 S-3/2H- 2 O: C, 60.40; H, 5.94; N, 7.29. Found: C, 60.15; H,5.82; N, 6.90.
WO 01/00206 WO 0100206PCT/USOO/1 8079 Example 21 1-[3-methoxy-4-[AN'-(2-methylphenyl)ureido]pheniylacetiyl]-2-pyrrolidinylmethoxy benzoic acid.
IH ~OOH 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 nunol) in THF (40 mL) was added DIAD 10 mL, 15.74 mmol). The resulting mixture was heated under reflux for 14 hr. The mixture was evaporated off in vacua and the residue was purified by column chromatography on silica-gel with n-hexane-EtOAc 1, v/v) as eluent to give 3.34 g methyl (S)-4-[1I-(tert-butoxycarbonyl)-2pyrrolidinylmethoxy] benzoate as an oil. 'H-NMR (CDC 3 5 1.48 9 1.67 J=9.3 Hz, I 1.87-2.03 (in, 3 3.36-3.43 3.87-4.09 (m,1IH), 4.13-4.20 (m,2 H, 6.94 J=8.3 Hz, 211), 7.98 J=8.3 Hz, 2H1).
A mixture of methyl (S)-4-[1-Qtert-butoxycarbonyl)-2-pyrrolidinylmethoxyj benzoate (3.34 g, 9.96 mnnol) in TEA (20 mL) and CH 2
CI
2 (35 mL) was stirred at room temp for 15 hr. The mixture was concentrated in vacua and made basic with sat. NaHCO 3 The mixture was extracted with CHCI 3 washed with brine, and dried over NaCQ 3 The organic layer was evaporated to give 1.70 g methyl (S')-4-(2-pyrrolidinylmethoxy)benzoate as a yellow oil. IH-NMR (CDC1 3 1.54-1.61 (in, I 1.77-1.86 (in, 2 1.87-1.97 (in, 1 2.00 (bs, 1 2.93-3.06 (in, 2 H), 3.52-3.57 (in, 1 3.88 3 3.90-3.99 (in, 2 6.92 J=9.0 Hz, 2 7.98 J=9.0 Hz, 2
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 mng, 1.40 inmol), EDC (312 mg, 1.63 mng), HOBt (220 mg, 1.63 nol), and a catalytic amount of DMAP in DMF (15 mL) was stiffed for 6 hr. The resulting mixture was diluted with EtOAc, washed with 0.5 N HCl, sat. NaHCO 3 brine, and dried over MgSO 4 The solvent was evaporated off in vacua to give an oily residue, which was purified by column chromatography on silica-gel with CHCI 3 -MeOH (50: 1, vlv) as eluent to give 540 mng methyl (S)-4-[1-[3-methoxy-4-[N'-(2methylpheniyl)ureidojphenylacetylj-2-pyrrolidinylmethoxy] benzoate as an oil. 'H-NMR (CDCI 3 8 1.81-2.12 (in, 4 H) 2.88 (bs, 3 3.48-3.61 (mn, total 7 3.88 3 4.104.21 (in, 2 4.42- 4.46 (in, 1 6.75-8.08 (series of in, total 13 H).
WO 01/00206 WO 0100206PCT/USOO/18079 To a stirred solution of methyl (S)-4-[I-[3-methoxy-4-[IN'-(2-methylphenyl) ureido] phenylacetylj-2-pyrrolidinylmethoxylbenzoate (540 mg, 1.02 mmol) in THF (10 mL) was added 0.25 N NaOH (10 mL). The resulting mixture was heated under refiux for 16 hr. The mixture was poured into I N HCI and the solid was collected. The crude solid was washed with EtO to give 278 mg 25 as a white amorphous solid. IR (KBr) 1708 'H-NMR (DMSO-d 6 5 1.83- 2.14 (in, 4 2.21 3 2.46 2 3.78 3 H4), 3.95-4.02 (in, 1 4.13-4.16 (in, 1 H), 4.24 (bs, 1 6.5 1-7.98 (series of m, 12 8.43 1 8.53 1 12.57 (bs, 1 MIS (FAB) m/z 517 (1W).
Example 22 1-11 -[3-methoxy-4-[N' -(2-methylphenyl)ureido]benzoylJ-L-prolyll-4-piperidinylacetic acid OOH 26 A mixture of 3-methoxy-4-nitrobenzoic acid (229mg, 1. l6nunol), tert-butyl 4-(1prolylpiperidinyl)acetate (344mg, 1.16inmoI), HOBt (1 88mg, 1.39minol), DMAP (14.2mg, 0. 1 l6mmol), and EDC (267mg, 1 .39nunol) in DMF (7mL) was stirred for 22 hr at room temp.
The mixture was diluted with EtOAc (5OmL) and washed successively with 1 N HCI, sat. NaHCO 3 and H,0. The organic layer was dried over Na 2 SO, and evaporated in vacua. The residue was chromatographed on silica-gel with MeOH:CHCI 3 (1:30, v/v) as eluent to afford 520mg tertbuty 1-(3-inethoxy-4-nitrobenzoyl)prolyl-4-(1-piperidinyl)acetate as a white crystalline material.
'H-NMR (CDCI 3 5 1.12-1.33 and 1.62-2.23 (each m,911), 1.44 2.65,3.13,3.47, 3.67, 4.44, and 4.61 (each m, 8H), 3.99 3H), 5.05 (in, lH), 7.21 J8.3Hz, lH), 7.31 1H), 7.86 (d, J=8.3Hz, 1H).
A stirred mixture of fert-buty 1-(3-methoxy-4-nitrobenzoyl)prolyl-4-(1-piperidinyl) acetate (0.52g, 1 .O9mmol) and 5% Pd-C (2.08g) in MeOH (liL) was hydrogenated under an atmospheric pressure for 94 hr at room temp. Insoluble catalyst was removed, and the filtrate was evaporated in vacua. The residue was chromatographed on silica-gel with MeOH:CHC1 3 (1:40 to 1:6 vlv) as eluent to afford 279mg tert-buty 1 -(4-amnino-3 -methoxybenzoyl)prolyl-4-( 1piperidinyl)acetate as a white crystalline material. 'H-NMR (CDC 3 8 1. 16-2.17, 2.69, 3.06, 3.67, 4.12, and 4.5 9 (each mn, 17H), 3.86 3H), 5. 10 (mn, I1H), 6.64 (in, I1H), 7.12 (each mn, 211).
WO 01/00206 WO 0100206PCT/USOO/18079 To a stirred solution of tert-buty 1-(4-amiino-3-methoxybenzoyl)-L-prolyl-4-(1-piperidinyl) acetate (279mg, 0.627mmo1) and EtN (0.0876mL, 0.627mmo1) in THE (4mL) was added dropwise o-tolyl isocyanate (0.0777mL, 0.627mmo1) 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 was collected with suction. The crude solid was purified by silica-gel column chromatography with MeOH:CHCI, (1:40, vfv) as eluent to afford 254mg tert-butyl 1-[1-[3-methoxy-4-[N'-(2methylphenyl)ureidoj benzoylJ-L-prolylj-4-(1-piperidinyl)acetate as a crystalline solid. 'H-NMR
(CDCI
3 8 1.43 9H), 1. 13-1.25 and 1.76-2. 14 (each mn, 9H), 2.60, 3.18, 3.71, 4.06, and 4.57 (each mn, 8H), 3.67 3H), 5.06 (in, 1H), 6.63, and 6.90 2H), 6.98-7.23, and 7.64 (each m, 7.56 J=7.8Hz, 1H), 8.21 J=8.8Hz, IH).
A solution of tert-butyl 1-11 -[3-methoxy-4-[NV'-(2-methylphenyl)ureidoj benzoyl]prolyl]- 4-(1-piperidinyl)acetate (254mg, 0.44Ominol) in CH 2
CI
2 (6mL) and TEA (6mL) was stirred for 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 26 as a white crystalline solid. 'H-NMR( DMSO-d) 8 0.47, 1.05, 1.44, and 1.62-1.99 (each m, 911), 2.49 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 3H), 4.92(m, 1H), 6.82, 6.95, 7.11, 7.77, 8.20, 8.57, and 8.75 (in, 9H); MIS (FAB) m/~z 523 Anal. Calcd for C 2 gH 3
,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 (S)-3-methoxy-4-[ 1-[3-methoxy-4-[N' -(2-methylphenyl)ureido~phenylacetylj-2-pyrrolidinyl methoxylbenzoic acid 9H H N~ qOOH 27 To a stirred solution of ethyl 4-hydroxy-3-methoxybenzoate (3.00 g, 15.29 mmol), Boc-prolinol (3.08 g, 15.30 mmnol), Ph 3 P (4.81 g, 18.34 nunol) in THF (50 mL) was added DIAD (3.61 mL, 18.33 inmol) 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 CHC1 3 -MeOH (50: 1, v/v) as eluent to give ethyl (S)-3-methoxy-4-[1-Qertbutoxycarbonyl)-2-pyrrolidinyl methoxy] benzoate as a gum. The above ethyl (S)-3-methoxy-4-[ 1- (teri-butoxycarbonyl)-2-pyfrolidinylmethoxy] benzoate was dissolved in CH 2
CI
2 (50 mL) and TEA (45 mL). The mixture was stirred for 2 days at room temp. The resulting mixture was WO 01/00206 WO 0100206PCT11JSO0118079 concentrated in vacuo and made basic with sat. NaHCO 3 The mixture was extracted with CHCI 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.
'H-NMR (CDC1 3 8 1.39 3 H, J=7.1 Hz), 1.52-1.59 (in, 1 1.76-1.88 (in, 2 1.92-2.01 (in, 1H), 2.92-3.06 (in, 2 3.56-3.63 (in, 1 3.90 3 3.91-4.02 (mn, 2 4.35 2 H, J=7. 1 Hz), 6.89 1 H, J=8.3 Hz), 7.54 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 mng, 1.52 inmol) in DMF (8 mL) was added pentafluorphenyl ester of 3-methoxy-4-[N'-(2inethylphenyl) ureidoiphenylacetic acid (728 mg, 1.52 mmol) and Et 3 N (0.26 mL, 1.87 inmol).
And the resulting mixture was stirred at room temp overnight. The mixture was diluted with EtOAc, washed with 1 N HCI, sat. NaHCO 3 brine, and dried over MgSO,. The solvent was evaporated off in vacua 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 ethyl (S)-3-methoxy-4-[l-[3-methoxy-4- [N'-(2-methylphenyl)ureido]phenyl acetylj-2-pyrrolidinyl methoxylbenzoate as an amorphous solid. 'H-NMR (CDCI 3 8 1.38 3 H, J=7.3 Hz), 1.88-2.20 (in, 4 H, in), 2.24 (in, 3 3.44-3.50 (in, I 3.53-3.58 (in, 7 3.82 3 4.09-4.17 (in, 1 4.22-4.25 (in, 1 4.35 2 H, J=7.3 Hz), 4.38-4.49 (in, 1 6.71-6.78 (mn, 1 6.99 1 H, J=8.3 Hz), 7.04-7.07 (in, 1H), 7. 16-7.19 (in, 2 7.49-7.66 (in, 3 8.06 1 H, J=8.3 Hz).
To a stirred solution of ethyl (S)-3-methoxy-4-[Il-[3-inethoxy-4-[N'-(2-methylphenyl) ureidolphenylacetyl]-2-pyrrolidinyllmethoxylbenzoate (760 mg, 1.32 mmol) in TJHE (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 rnL) and the solid was collected.
The crude solid was washed with EtO to give 429 mng 27 as a yellow amorphous solid. mp 132-135; IR (KBr) 1707 'H-NMR (DMSO-d 6 8 1. 84-2. 18 (in, 4 2.25 3 2.49-2.5 1 (in, 2 3.29-3.59 (in, 4 3.80 3 3.82 3 4.00-4.05 (in, 1 6.53-8.01 (in, 10 H), 8.45 1 8.54 1 12.63 (bs, 1 MS (FAB) m/z 548 Example 24 -methoxy-4-[N' -(2-inethylphenyl)ureidojphenylacetyl]-2-pyrrolidinylnethoxyJ phthalic acid WO 01/00206 PTUO/87 PCT/USOO/18079 (AX)H 28 To a stirred solution of dimethyl 4-hydroxyphthalate (3.00 g, 14.27 mmol), N-Bocprolinol (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 resulting mixture was evaporated and the residue was purified by column chromatography on silica-gel with n-hexane-EtQAc 1, vA') as eluent to give 5.75 g dimethyl (S)-4-[lI-Qtertbutoxycarbonyl)-2-pyrrolidinyl methoxy] phithalate as an oil. 'H-NMR (CDCI 3 5 1.47 9 H), 1.86-2.05 (in, 4 3.36-3.40 (in, 2 3.87 (in, 3 3.91 3 3.96-4.19 (in, 3 7.03-7.24 (in, 2 7.80 (in, I1H).
To a solution of dimethyl (S)-4-[1-QIert-butoxycarbonyl)-2-pyrrolidinylmethoxy] phithalate (5.75 g, 14.62 inmol) in CHC1 2 (25 inL) was added TFA (20 niL), and the resulting mixture was stirred for 50 mmn at room temp. The resulting mixture was concentrated in vacuo and made basic with sat. NaHCO 3 The mixture was extracted with CH 2
CI
2 washed with brine, dried over MgSO 4 and evaporated off in vacuo. The residue was purified by column chromatography on silica-gel with CHCI 3 -MeOH (50: 1, v/v) as eluent to give 790 ing diinethyl pyrrolidinylinethoxy) phithalate as a brown oil. 'H-NMR (CDC 3 8 1.48-1.57 (in, 1 1.72-1.84 (in, 2 1.89-1.98 (in, 2 2.91-3.03 (mn, 2 3.48-3.54 (in, I 3.82-3.97 (in, total 8 H), 6.98 (dd, 1 H, J=2.4, 8.8 Hz), 7.06 I H, J=2.4 Hz), 7.78 1 H, J=8.8 Hz).
To a stirred solution of dimethyl (S)-4-(2-pyrrolidinylmethoxy)phthalate (212 mng, 0.72 minol) in DMF (8 inL) was added pentafluorophenyl ester of the 3-inethoxy-4-[N'-(2inethylphenyl)ureido] phenylacetic acid (346 mng, 0.72 nunol) and Et 3 N (120 ml], 0.86 iniol), and the mixture was stirred overnight. The resulting mixture was diluted with EtOAc, washed with 1 N HCI, sat. NaHCO 3 brine, and dried over MgSQ 4 The solvent was evaporated off in vacuo to give 413 ing diinethyl -[3-inethoxy-4-[N'-(2-inethylphenyl)ureidolphenylacetyl]-2pyrrolidinylinethoxy]phthalate as an oil. 'H-NM4R (CDCI,) 5 1.92-2.12 (in, 4 2.29 (br s, 3 H), 3.51-3.64 (mn, 7 3.87 3 3.89 3 4.10-4.19 (in, 2 4.44 (in, 1 6.73-8.02 (series of in, total 12 H).
To a stirred solution of diinethyl 1-[3-methoxy-4-[N' -iethylpheriyl)ureido] WO 01/00206 WO 0100206PCT/JSOO/18079 phenylacetyl]-2-pyrrolidinylmethoxylphthalate (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 rorn 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 EtO to give 3 10 mg 28 as a yellow amorphous solid. 1k (KBr) 1701 cm-1; 'H-NMR (DMSO-d) 8 1.87-2.18 (in, 4 2.25 3 2.50 2 3.38-3.60 (in, 4 3.83 3 4.00-4.14 (in, 1 6.74-8.02 (series of m, 10 8.46 1 8.54 1 MS (FAB) ni/z 562 Example 3-chloro-4-II1-[3-methoxy-4-[N'-(2-methylphenyl)ureidojphenylacetylj-2-pyrrolidinyllmethoxy] benzoic acid CI~> OOH 29 To a stirred solution of methyl 3-chloro-4-hydroxybenzoate (600 mng, 3,21l5nnol), N-tertbutoxycalbonylprolinol (647,1 mng, 3,215 mmol), and Ph 3 P (1.0 1 g, 3.858 inmol) in THE (10 mL) was added dropwise diisopropyl azodicarboxylate (DIAD) (0.8 mL, 3.890 mmol) at room temp and the mixture was stirred for 3days at room temp, and for 18 hr at 70*C. The reaction mixture was evaporated off in vacua, and the residue was chromatographed on silica-gel with n-hexane:EtQAc 1, v/v) as eluent to give 1. 147g methyl 3-chloro- 1-QIert-butoxycarbonyl)-2pyrrolidinylimethoxy benzoate as an oil. 'H-NMR (400 MfHz, CDCI 3 8 1.46, 1.48 (s each, 9H), 1.59 1.63 (br, 1H), 1.88 (br s, 1H), 2.05 lH), 2.05 2.21 (in, 2H), 3.34 -3.45 (br m, 3.89 3H), 3.97(br mn, 0.5H), 4.21 (br s, 2H), 7.05 J= 8.8 Hz, 1H), 7.90 (dcl, J 8.8 Hz, I1H), 8.04 J 2.0 Hz, I1H); MS (FAB) n/z 370 1).
To a stirred solution of methyl 3-chloro-[l-(tert-butoxycarboniyl)-2-pyrrolidiniyljmethoxy benzoate (1.14 g, 3. 10 inmol) in CH 2 Cl 2 (20 mL) was added TEA (5 mL) at 0 and 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 mixture was extracted with CHC1 3 The extact was washed with brine, dried over KOH, and concentrated under a reduced pressure to afford 741 mg methyl 3-chloro-4-(2-pyrrolidinyl)methoxybenzoate as a yellow oil. 'H-NMR (400 MHz,
CDCI
3 5 1.60 1.67 (in, 1H), 1.78 2.02 (in, 3H), 2.93 2.98 (in, lH), 3.03 3.09 (mn, III), 3.59 (dt, J 9.3 Hz, IH), 3.89(s, 3H), 3.98(dd, J= 6.3, 8.8 Hz, IH), 4.05 (dd, J= 4.9, 9.3 Hz, IN), 6.93 J= 8.8 Hz, 1H), 7.90 (dd, J 2.0, 8.8 Hz, IN), 8.04 J 2.0 Hz, IN); MS (FAB) m/z 270 1).
WO 01/00206 WO 0100206PCT/USOO/18079 The midxture of pentafluorophenyl 3-methoxy-4-[N'"-(2-methylpheniyl)ureido phenylacetate (500 mg, 1 .O4mmoI), methyl 3-chloro-4-(2-pyrrolidinyl)methoxybenzoate (281 mg, 1.04 nunol), Et 3 N 17 mL, 1.25 nunol) 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
SQ
4 The solvent was removed under a reduced pressure, and the residue was chromatographed on silica-gel with nhexane EtOAc v/v) as eluent to afford methyl 3-chloro-4-[[I-[3-methoxy-4-jN'-(2methylphenyl)ureido]phenylacetylj-2-pyrrolidiryl] methoxy]benzoate (620 mg, 1.04 mxnol) as a white crystalline solid. To a stirred solution of this compound in THE (8 mL) and H20 (2 mL) was added LiOH (74.9mg, 3.126 mniol), and the mixture was stirred at room temp for 2 days. The mixture was diluted with CHCI 3 and treated with IN HCI. The solution was washed with brine, dried over NaSO 4 and evaporated in vacua. The crude solid was recrystallized from n-hexane- EtOAc-CHCI 3 to afford 561.2 mg 29 as a white crystalline material. IR (KBr) 1676, 1599, 1487, 1267, 758, 754cm-'; 'H-NMR (400 M4Hz, DMSO-d) 8 1.82 2.24 4H), 2.25 3H), 3.48 -3.60 (in, 4H), 3.78 3H), 4.18 (in, 2H), 4.31 (in, 1H), 6.74 (dd, J 1.5, 8.3 Hz, lH), 6.84 J 2.0 Hz, IH), 6.91 6.95 (in, 1H), 7.11 7.17 (in, 3H), 7.79 (dd, J 2.0, 8.3 Hz, 2H), 7.85 J 2.0 Hz, I1H), 7.98 J1= 8.3 Hz, 111), 8.53(s, 1ff), 8.58 I MS (FAD) m/z 552 Example 26 3,5-dichloro-4-[[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureidojphenylacetyl]-2-pyrolidinylj methoxy] benzoic acid COOH H OH 3 To a stirred solution of methyl 3,5-dichloro-4-hydroxybenzoate (600mg, 2.714 inmol), Ntert-butoxycarbonylprolinol (546mg, 2.7 l4mmoI), and Ph 3 P (854 mg, 3.257 nunol) in THE (l0m.L) was added dropwise DIAD (0.68mL, 3.283 nunol) at room temp and the mixture was stirred for 3days at room temp, and for 18 hr at 70*C. The reaction mixture was concentrated and the residue was chromatographed on silica-gel with n-hexane EtOAc 1, v/v) as eluent to give 988.8 mng methyl 1-(tert-butoxycarbonyl)-2-pyrrolidinyl]inethoxy-3,5-dichlorobenzoate a pale yellowish oil. 'H-NNvI (400 N11-z, CDCI 3 5 1.44 9H), 1.88 -2.15 (br m, 3H-I), 2.34 (br s, IM), 3.40 3.44 (in, 2H), 3.92 3H), 3.92, 4.14 (in, 11H), 4.18 (br s, 2H), 7.98 2H); MS (FAB) m/z 404 I).
To a stirred solution of methyl 4-[1-Qtert-butoxycarbonyl)-2-pyrrolidinyljinethoxy-3,5dichlorobefizoate (988 mng, 3.248 inmol) in CH 2
CI
2 (20 inL) was added TEA (5 mL) at 0 0 C, and WO 01/00206 WO 0100206PCT/USOO/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 CHC 3 The extract was washed with brine, dried over Na 2
SQ
4 and concentrated under a reduced pressure to afford 672 mg methyl 3,5-dichloro-4-(2-pyrrolidinyl)methoxybenzoate as a pale yellowish oil. 'H-NNM (400 MU-z, CDCI 3 8 1.62 1.69 (in, 1H), 1.78 1.86 (in, 2H), 1.89 1.99 (in, 1H), 2.92 2.98 (in, 1H), 3.04 3.09 (mn, 1H), 3.55 3.60 (mn, lH), 3.91 3H), 4.01 (dd, J 6.8, 8.8 Hz, 1H), 4.08 (dd, J 4.9, 8.8 Hz, I1H), 7.97 2H); MS (FAB) ni/z 304 1).
A mixture of pentafluorophenyl 3-inethoxy-4-[N'-(2-methylphenyl)ureido] phenylacetate (385.8 mg, 0.803 mmol), methyl 3,5-dichloro-4-(2-pyrrolidiniyl)methoxybenzoate (244.3 mg, 0.803 minol), Et 3 N 13 mL, 0. 964 inmol) in DMF (4 inL) 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 nhexane:EtQAc v/v) as eluent to afford methyl 3,5-dichloro-4-[[l-[3-inethoxy-4-[N'P-(2methylphenyl)ureidolphenytacetylJ-2-pyrrolidinylJ methoxy~benzoate as an oil. To a stirred solution of this compound in THE (8 mL) and H 2 0 (2 inL was added LiOH (57.7 mg, 2.409 mmiol), and the mixture was stirred at room temp overnight. The mixture was concentrated in vacua, and the residue was diluted with CHCI 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-CHC 3 to afford 428.2 mg (9 30 as a white crystalline powder. JR (KBr) 1618, 1535, 1454, 1257, 754cin-'; 'H-NNvIR (400 Mffz, DMS0-l 6 8 1.83 2.24 (mn, 4H), 2.24 3H), 3.50 3.58 (mn, 4H), 3.84 3H), 3.98 4.05 (mn, IH), 4.15 (dd, J 2.9, 8.7 Hz, 4.29 (br mn, IH), 6.74 J =8.3 Hz, lH), 6.87 IH), 6.93 J= 7.3 Hz, 114), 7. 11 J 7.8 Hz, 7.16 J 8.3 Hz, 1H4), 7.79 J 8.3 Hz, IH), 7.86 LH), 7.87 J 9.8 Hz, 1H), 7.99 J 8.3 Hz, 1141), 8.49 IH), 8.58 IN); MS (FAB) ,n/z 586 Example 27 4-ti -[3-inethoxy-4-[N'-(2-methylphenyl)ureidoJphenylacetyl]-2-pyrrolidinylnethoxy]-3 nitrobenzoic acid IH H "IN OOH 3 To a stirred solution of 4-hydroxy-3-nitrobenzoic acid (3.00g, 0.0 164ino1) in MeONbenzene v/v) was added dropwise 2.0 M-n-hexane solution of TMSCHN 2 (8.2mL, 0.0 164ino1) at room temp. After the resulting solution was stirred for 4 hr at room temp, the mixture was WO 01/00206 WO 0100206PCT/USOO/18079 evaporated off in vacua. The oily residue was chromatographed on silica-gel with CHC1 3 as eluent to afford 4.23g methyl 4-hydoroxy-3-nitrobenzoate as a pale yellow crystalline material.
To a stirred mixture of N-tert-butoxycarbonylprolinol (1 .02g, 5.O7mmol), methyl 4hydoroxy-3-nitrobenzoate (1.00g, 5.O7mmol), and Ph 3 P (1.46g, 5.58mmol) in THF (lOmL) was added dropwise diisopropyl azodicartoxylate (DIAD) l6niL, 5.58mmol) at 0 0 C. The resulting mixture was heated under reflux for 46 hr. After cooling, the mixture was evaporated off in vacua. The residue was dissolved in CH 2
CI
2 (10mL) and added TFA (IOmL). After the solution was stirred for 0.5 hr at room temp, the solution was evaporated in vacua. Water was added to the residue and washed with EtOAc. The aqueous layer was neutralized by the addition of sat.
NaHCO, and extracted with EtOAc. The extract was dried over Na 2 SO, and evaporated in vacuo to afford 0.698g methyl 3-nitro-4-(2-pyrrolidinylmethoxy) benzoate as a gum.
A mixture of methyl 3-nitro-4-(2-pyrrolidinylmethoxy)benzoate (0.668g, 2.38mmol), 3methoxy-4-[N'-(2-methylphenyl)ureidoJphenylacetic acid 12g, 3.57mmol), 1-hydroxybenzo triazole (HOBt) (0.482g, 3.S7nunol), 4-dimethylaminopyridine(DMAP) (43.6mg, 0.357mmo1), and 1-ethyl-3-(3-dimethylamiinopropyl)carbocliimide (EDC) (0.684g, 3.57mmol) in DMF (l1niL) was stiffed for 15 hr at room temp. EtOAc was added to the mixture and washed successively with I N HCI, sat. NaHCO 3 and brine. The organic layer was dried over Na 2
SO
4 and evaporated in vacua. The residue was chromatographed on silica-gel with EtOH-CHC 3 (1:20, v/v) as eluent to afford 0.927g methyl 4-[Il-[3-methoxy-4-[N' -(2-methylphenyl)ureidolphenylacethylj-2pyrrolidinylmethoxy]-3-nitrobenzoate as a yellow crystalline material.
A mixture of methyl 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido phenylacethyl]-2pyrrolidinylmethoxyJ-3-nitrobenzoate (0.917g, 1.S9mmol) in THF (lOmL) and 1 N NaOH (2.38mnL, 2.3 8mmol) 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
SQ
4 and evaporated in vacua to afford 0.826g 31 as a yellow crystalline solid. IH-NMR (400M~iz,
CDCI
3 5 1.91, 2.09 (1H, 3H, each in), 2.28 (3H, 3.54-3.62 (41I in), 3.64 (3H, 4.15, 4.59 (each IIH, each d, J=7.8Hz), 4.46 (lH, in), 6.66, 7.22 (each 1H, each 6.72 (IH, d, J=8.3Hz), 7.11-7.28 (4H, mn), 7.46 (lH, d, J=7.8Hz), 7.74 (111, d, J=7.8Hz), 7.85 (IH, 8.17 (lH, dd, 8.8Hz), 8.48 (1WK d, J=2.4Hz); MIS (FAB) m/zk 563 Example 28 3-aino-4-[ 1-[3-ffethoxy-4-[N' -(2-methylphenyl)ureido]phenylacetyl]-2-pyffolidinylmethoxy] WO 01/00206 WO 0100206PCT/USOOI 18079 benzoic acid H HH 2 COOH 32 A stirred mixture of 4-[l -[3-methoxy-4-[N'-(2-methylphenyl)ureidoI phenylacethyl]-2pyrrolidinylmethoxy]-3-nitrobenzoic acid (101mg, 0.l19OnuoI) and 5% Pd-C (0.247g) in methanol was hydrogenated at 1 atmn for 48 hr. Insoluble catalyst was removed, and the filtrate was evaporated in vacuo. T'he residue was chromatographed on silica-gel with EtOH-CC 3 1, vlv) as eluent to afford 61.0mg 32 as a crystalline material. 'H-NMR (4.00MHz, DMSO-d 6 8 1.95 (4H, in), 2.23 (3H, 3.60, 3.91, 4.10, 4.34 (5H, each in), 3.81 (3H, 4.88 (2H, in), 6.74 (LH, d, J=8.3Hz), 6.86-7.28 (5H, in), 7.78 (IH, d, J=7.8Hz), 7.99 (1H, d, J=83Hz), 8.30 (IH, s), 8.45, 8.55 (each IH, each MIS (FAB) m/z 533 1).
Example 29 4-[2-[1-[4-[N'-(2-fluorophenyl)ureido)-3-methoxyphenylacetyl]-2-pyrrolidinyllethynyl] benzoic acid H H AOOH 33 To a stirred solution of benzyl 4-amino-3-methoxyphenylacetate (1.36 g, 5 mmol) in THFf mL) was added 2-fluorophenyl isocyanate (561 ul, 5 inmol) and a catalytic amount of Et 3
N.
The resulting mixture was stirred for 3 hr. The mixture was quenched by the addition of H 2 0 miL) and extracted with EtOAc. The extract was washed with brine, dried over MgSO 4 and evaporated. The residue was chromatographed on silica-gel with CHC1 3 as eluent to give 2.06 g benzyl 4-[N'-(2-fluorophenyl)ureido-3-nethoxyphenylacetate as a green oil. 'H-NVR (CDC1) 3 8 3.63 (2H, 3.82 (3H, 5.14 (2H, 6.79-7.37 (12H, in), 8.01 (IH, d, J=7.8 Hz), 8.09-8.14 (1H, in).
To a stirred solution of benzyl 4-[N'-(2-fluorophenyl)ureido]-3-methoxyphenylacetate (2.04 g, 5 inmol) in THE (40 mL) was added 0.25 N NaOH (40 inL). The resulting mixture was 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 CHC1 3 -EtOH to give 1.04 g 4- WO 01/00206 WO 0100206PCT/USOO/18079 [N'-(2-fluorophenyl)ureidol-3-methoxyphenylacetic acid as a white crystalline powder. mp 185- 188 'H-NMR (DMSO-d 6 8 3.50 (2H, 3.82 (3H, 6.78 (1H, dd, J= 1.4 and 8.3 Hz), 6.92 (1H, d, J=1.4 Hz), 6.95-7.01 (1W in), 7.10-7.14 (1H, in), 7.19-7.24 (1H, in), 8.01 (1H, d, J=8.3 Hz), 8.14-8.18 (1 H, in), 8.72 (1 H, 9.17 (1 H, MS (FA.B) m/z 319 Anal. Calcd for
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~pyrrwlidine (195 mg, 0.8 innol), EDC (230 mg, 1.2 mmol), DMAP (98 mg, 0.8 mmol) in DMF (20 inL) was stirred overnight. The reaction mixture was poured into 1 N HCl and the resulting precipitate was collected with suction and dissolve in CHCl 3 The solution was dried over MgSO, and evaporated. The residue was chromatographed on silica gel with CHCI 3 -MeOH (100: 1, vfv) as eluent to give the desired compound, which was dissolved in THF (8 mnL. 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 CHC1 3 The extract was washed with brine, dried over MgSO 4 and evaporated. The residue was recrystallized from CHC1 3 -n-hexane to give 144 mg (3 33 as a pale yellow crystalline powder.
mp 152-155 'H-NMR (DMSO-d6) 5 1.92-2.27 (4 H, mn), 2.50 (2 H, 3.33-3.78 (2 HK in), 3.80 and 3.82 (total 3 H, s, each), 4.88-5.12 (1 H, in), 6.77-7.24 and 7.99-8.20 (total 7 H, in), 7.48 and 7.52 (2 H, d, J'-8.3 H-z, each), 7.91 (2K, d, J=8.3 Hz), 8.72 (1H, 9.18 (lH, 13.11 (1H, br-s); MS (FAB) m/z 516 Anal. Calcd for C29H2N 3 OF-2H 2 O: C, 63.15; H, 5.48; N, 7.62. Found: C, 63.58; H, 5.15; N, 7.22.
Example 4-f[ 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido~phenylacetyll-2-pyrrolidinyllmethoxy-3 methylbenzoic acid N-T 6(C
OOH
H H34 To a stirred solution of 4-iodo-2-methylphenol (465 ing, 1 .987inmo1), N-tertbutoxycarbonylprolinol (400 mng, 1.987 nunol), and Ph 3 P (625 ing, 2.384 iniol) in TI-I (7 inL) was added dropwise DIAD (0.5 inL, 2.404 imol) at room temp, and the mixture was stirred for 13 hr at 70 0 C. The reaction mixture was concentrated in vacuo and the residue was chromatographed on silica-gel with n-hexane EtOAc 1, v/v) as eluent to give 645.3 mng 4-[l1-(tertbutoxycarbonyl)-2-pyrrolidinyllmethoxy-l1-iodo-3-inethylbenzene as a pale yellow oil. IH-NMR WO 01/00206 WO 0100206PCT/USOO/1 8079 (400 MHz, CDC 3 6 1.47 9H), 1.83 1.89 (in, 1H), 1.96- 2.04 (in, 3H1), 2.16 31-1, 3.37 3.43 (br m, 3.81, 3.94 (br m each, 1H), 4.08 4.18 (in, 2H4), 6.62 (br s, 7.42 2H); MS (FAB) m/z 418 To a stirred solution of 1-QIert-butoxycarbonyl)-2-pyrrolidinyl~methoxy- 1-iodo-3methylbenzene (645.3 mng, 1.546 mmol) in DMSO (7 mL) and MeOH (6 mL) was added Et 3
N
(0.47 niL, 3.40 1 inmol), Pd(OAc) 2 (17.4 mng, 0.077 minol) and 1,3-bis(diphenylphosphino) propane (31.46 mg, 0.O77nunol). To the stirrede mixture was induced CO gas for 10 min. The mixture was stirred at 70*C for 2 days and concentrated. The residue was diluted with EtOAc, washed with brine, and dried over NaSO 4 The solvent was removed under a reduced pressure, and the residue was chromatographed on silica-gel with n-bexane EtOAc 1, v/v) as eluent to afford 301.6 mg (56 methyl4-[I -(tert-butoxycarbonyl)-2-pyrrolidinyl] methoxy-3-methylbenzoate as an oil. NMvR (400 MfHz, CDC1 3 8 1.47 9H), 1.86 2. 10 (br m, 4H), 2.33 3.32 3.50 (br m, 3.88 3H), 3.88, 4.04 (br m each, I 4.13- 4.20 2H), 6.89 (br m, 114), 7.82 (s, 1H), 7.85 (dd, J 2.0, 8.8 Hz, lH); MS (FAB) m/z 350 (MC+1).
TO a stirred solution of methyl 4-[1-(tert-butoxycarbonyl)-2-pyrrolidinyl]methyloxy-3methylbenzoate (301.6 mg, 0.863 mmol) in CH 2 CI, (6 niL) was added TFA (1.2 mL) at 0 0 C, 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
CHCI
3 The extract was washed with brine, dried over Na 2
SO
4 and concentrated under a reduced pressure to afford 192.5 mg methyl 3-methyl-4-(2-pyrrolidinyl)methoxybenzoate as an oil.
'H-NMR (400 MI-Iz, CDCI 3 5 1.58 1.65 (in, HI4), 1.78- 2.00 (in, 3H), 2.24 Cs, 3H), 2.97 (dt, J= 6.8, 10.2 Hz, 1H), 3.05 (dt, J 5.9, 6.8 Hz, 11-1), 3.54 3.58 (in, LH), 3.87 3.92 (dd, J 6.3, 9.3 Hz, 3.99 (dd, J 9.3 Hz, 111), 6.81l(d, J =8.3 Hz, 11-1), 7.83 1141), 7.85 (dd, J 8.3 Hz, I1H); MS (FAB) m/z 250 1).
A mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetate (211.3 mng, 0.44 nunol), methyl 3-methyl-4-(2-pyrrolidinyl)methoxybenzoate (109.7 mg, 0.44 mmol), Et 3 N (73.6 ul, 0.528 inmol) in DMF (2 niL) 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 on silica-gel with n-hexane EtOAc v/v) as eluent to afford methyl 4-[[l-13-methoxy-4-[N'- (2-methylphenyl)ureido] phenylacetyl]-2-pyrrolidinyl] methoxy]-3-methyl benzoate (241.6 mg, WO 01/00206 PTU0/87 PCT/USOO/18079 as an oil. To a stirred solution of this compound in THFf (4.4 mL) and H20(1.1 niL) was added LIOH (32mg, 1.32 mmol), and the reaction mixture was stirred at room temp overnight. The mixture was diluted with CHC 3 and acidified by the addition of IN HCI. The solution was washed with brine and dried over NaSO 4 The solvent was removed under a reduced pressure, and the obtained crude solid was recrystallized from n-hexane-EtOAc-CHC1 3 -MeOH to afford 126.3 mg 34 as a white crystalline powder. IR (KBr) 1685, 1606, 1454, 1257, 752cm-1; 'H-NMvR (400 DMSO-d) 8 1.87 2. 10 (in, 4H1), 2.12 3H1), 2.25 311), 3.51 3.71 (in, 411), 3.76 (s, 3 4.08 4.18 (in, 211), 4.34 (in, 111), 6.74 (dd, J 1. 5, 9.8 Hz, 111), 6.84 J 1. 5 Hz, 111), 6.94 (t,J 6.8 Hz, 111), 7.06 J= 8.8 Hz, 111), 7.12 J =7.8 Hz, 11H), 7.16 J =7.8 Hz, 11H), 7.72 IH), 7.76 (dd, J 2.0, 8.3 Hz, 11H), 7.79 J 7.8 Hz, 11H), 7.99 J =8.3 Hz, 11H), 8.46 114), 8.54 lH); MS (FAB) m/z 532 Anal. Calcd for C30HN 3 0 6 1/2H 2 0: C, 66.65; H, 6.34; N, 7.77. Found: C, 66.16; HK 6.37; N, 7.50.
Example 31 1-[3-methoxy-4-[N'-(2-nethylphenyl)ureido~phenylacetyl]-2-pyrrolidinylmethoxy] isophthalic acid
OOH
H HOH To a solution of diinethyl 4-acetoxyisophthalate (1.52 g, 6.03 inmol) 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 HCl and extracted with EtOAc. The extract was washed with sat. NaHCO 3 brine, and dried over MgSO 4 The solvent was evaporated to give 1.27 g (qy.) dimethyl 4-hydroxy isophithalate as a white crystalline powder. 'H-NMR (CDC 3 8 3.91 3 H), 3.99 3 7.01 1 H, J=8.8 Hz), 8. 11 (dd, 1 H, J=2.4, 8.8 Hz), 8.55 1 H, J=2.4 Hz) To a stirred solution of dimethyl 4-hydroxyisophthalate (1.27 g, 6.04 mmol), (S)-N-Boc- Prolmnol (1.22 g, 6.06 iniol), PPh 3 (l.90 g, 7.24 mmol) in TI-F (30 miL) was added DIAD (1.43 mL, 7.26 minol)at room temp. The resulting stirred mixture was then heated under reflux for 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 1, v/v) as eluent to give 2. 10 g dimethyl 1-QIert-butoxycarbonyl)-2-pyrrolidinylmethoxylisophthalate as a yellow oil.
'H-NMR (CDCI,) 8 1.26 9 1.85-2.16 (in, 3 3.36-3.46 2 3.90 6 4.11-4.31 (in, 2 4.95-5.02 (in, 2 7.09 (dd, 1 H, J=9.3, 24.9 Hz), 8.11-8.14 (in, 1 8.46 1 H, WO 01/00206 WO 0100206PCT/USOO/18079 J=9.3 Hz) A mixture of dimethyl 1-Qtert-butoxycarbonyl)-2-pyrrmlidinylmethoxy] isophthalate (2.01 g, 5.11 mmnol), TFA (20 mL), and CH 2 C1(25 mL) was stirred for 1.5 hr at room temp. The resulting mixture was concentrated in vacuo and made basic with sat. NaHCQ 3 The mixture was 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 CHCI 3 -MeOH 1, v/v) as eluent to give 0.80 g (531/) dimethyl (S)-4-(2-pyrrolidinylmethoxy)isophthalate as a yellow oil. 'H-NMR (CDCI,) 5 1.71 (in, 1 1.89 (in, 2 2.00 (in, 1 3.05-3.13 (in, 2 3.67 (in, 1 3.90 (s, 3 3.91 3 4.05-4.18 (in, 2 7.00 1 H, J=8.8 Hz), 8.14 (dd, 1 H, J=2.4, 8.8 Hz), 8.50 (d,1HKJ=2.4 Hz).
To a stirred solution of diinethyl (S)-4-(2-pyrrolidinylmethoxy)isophthalate (616 mg, 2. inmol) in DMF (13 inL) was added pentafluoro ester of 3-methoxy-4-[N'-(2-methylphenyl)ureidoJ phenylacetic acid (1.00 g, 2.08 minol) and Et 3 N (425 jA, 3.12 inmol), and the resulting mixture was stirred for 3.5 hr at room temp. The resulting mixture was diluted with EtOAc, washed with 1 N HCI, sat. NaHCO 3 brine, and dried over Na 2 504. After removal of the solvent, the residue was purified by column chromatography on silica-gel with CHC1 3 -MeOH (50: 1, v/v) as eluent to give 1.41 dimethyl 1-[3-inethoxy-4-[N'-(2-methylphenyl)ureido~phenylacetyl1-2pyrrolidinylmethoxy] isophthalate as a yellow oil. 'H-NMvR (CDCI,) 8 1.86-2.29 (mn, 4 2.30 (s, 3 3.47-3.57 (in, 2 3.58 3 3.59 2 3.83 3 3.91 3 4.22-4.37 (in, 2 4.42-4.47 (mn, 1 6.44-8.46 (series of m, 12 H).
To a stirred solution of diinethyl (S)-4-[1-[3-methoxy-4-[N'-(2inethylphenyl)ureido]phenyl acetyl] -2-pyrrolidinylmethoxy]isophthalate (1.41 g, 2.39 minol) in THfF (20 mL) was added 0.25 N NaOH (20 inL), 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 the solid was collected. The crude solid was recrystallized from CHC1 3 -MeOH to give 140 mng 35 as a white crystalline powder. 'H-NMvR (CDC1 3 8 1.83-2.18 (mn, 4 2.24 3H), 3.44-3.55 (in, 4H), 3.59 2 3.80 3 4.054.24 (in, 2 4.28-4.32 (in, 1 6.73-8.55 (series of in, total 12 MIS (FAB) m/lz 562 Anal. Calcd. for C,)l 3
'N
3 0 8 4H 2 O: C, 56.87; H, 6.20; N, 6.63. Found: C, 56.73; H, 5.56: N, 6.52.
Example 32 3-inethoxy-4-[2-[ 1-[3-inethoxy-4-[N'-(2-inethylphenyl)ureido]phenylacetylI-2- pyrrolidinyl] WO 01/00206 WO 0100206PCT/U 500/18079 ethynyl] benzoic acid 9H H 0O0OH 36 A stirred mixture of methyl 3-methoxy-4-nitrobenzoate (1.20 g, 5.7 minol) and 5% Pd-C in EtOH (30 mL) and THF (20 mL) was hydrogenated overnight at 1 atm. The midxture was filtered and the filtrate was evaporated. The residue was chromatographed on silica-gel wAith CHC1 3 as eluent, and the solid obtained was further purified by recrystallization from CHCI,-n-hexane to give 805 mg methyl 4-aniino-3-methoxybenzoate as white plates. mp 126-128; IR (KIBr) 3475, 1700 cm- 1 1H-NMR (CDC 3 8 3.86 (3-1 3.89 (3HK 4.21 (2H, br 6.66 (lH, d, J=8.3 Hz), 7.45 (1H, d, J=1.9 Hz), 7.54 (LH, dd, J=1.9 and 8.3 Hz); MS (FAB) m/z 182 Anal Calcd for C 9
H
11 N0 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 mniol) in EtOH mL) was added to dil.H 2 S0 4 (prepared from H 2 S0 4 0.5 mL and H 2 0 10 mL) at 0 0 C. A solution of NaNO 2 (331 mg, 4.8 mmol) in H 2 0 (10 roL) was added to the mixture. After stirring for 0.5 hr at the same temp, the mixture was poured into a cooled (0 0 stirred suspended solution of KI (1.83 &gI 1 nol) and cat. Cu in H 2 0 (100 mL). The mixture was vigorously stiffed for 1 hr at room temp and extracted with CHCI,. 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-iedo-3-methoxybenzoate and methyl 3-methoxybenzoate (748 mg) as a colorless oil.
To this oil was added Pd(PPh 3 mg, 0. 13 mmol), Cul (57 mg, 0.3 mmol) and i- Pr 2 NH (10 niL). The mixture was stirred for 1 hr under N, and a solution of 1-(tertbutoxycarbonyl)-2-ethynylpyrrolidine (488 mg, 2.5 mmol) in i-Pr 2 NI- (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 chromatographed on silica-gel with n-hexane-EtOAc 1, v/v) as eluent to give 431 mg 1 (tert-butoxycarbonyl)-2-[2-(2-methoxy-4-methoxycarbonylphenyl) ethynylipyrrolidine as a yellow oil. IH-Nl'vR (CDCl 3 5 1.49 (9 H, 1.77-2.14 (4 H, in), 3.36-3.51 (2 in), 3.90 (3 H, 3.91 (3 H, 4.604.81 (1 H, in), 7.36-7.39 (1 H, mn), 7.51 (1I H, 7.55-7.57 (1 H, mn).
WO 01/00206 WO 0100206PCT/USOO/18079 To a stirred solution of 1 -Qert-butoxycarbonyl)-2-[2-(2-methoxy-4-methoxycarbonyl phenyl)ethynyl~pyrrolidine (3 95 mg, 1. 1 mmol) in CH 2
C
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 CHC 3 The extract was washed with H 2 0, dried over MgSO 4 and evaporated to give 238 mg 2-[2-(2-methoxy-4-methoxycarbonylphenyl) ethynylipyrrolidine as ayellow oil. IH-NIR (CDC1 3 5 1.81-2.16 (4 H, in), 2.97-3.17 (2 H, in), 3.91 (6 H, 4.13-4.15 (1 H, in), 7.41 (1 H, d, J=8.3 Hz), 7.51 (1 H, 7.56 (1 H, d, J=8.3 Hz).
A mixture of 2-[2-(2-methoxy-4-methoxycarbonylphenyl)ethynylJpyrrolidine (233 ing, 0.9 minol), 3-methoxy-4-[N'-(2-methylphenyl)ureidolphentylacetic acid (314 mg, 1 inmol), EDC (268 mg, 1.4 inmol), DMAP (110 mg, 0.9 innol) in DNM (10 mL) was stirred overnight. The mixture was poured into 1 N HCI and the resulting solid was collected with suction. The solid obtaned 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 inL) and 0.25 N NaQH was added to this solution with stirring. The solution was poured into ice-i N HCI to give a solid. The solid was collected, washed with water, and air-dried. The crude solid was recrystallized from CHCI 3 -n-hexane to give 215 mg 36 as a white crystalline powder. mp 141-145; 1k (KBr) 3338, 2956, 2935, 2875, 2593, 1711 'H- NMvR (DMSO-d 6 8 1.91-2.14 (4 H, 2.24 (3 H, 3.38-3.68 (4 H, 4.88-5.08 (1 H, in), 6.76-8.56 (12 H, in); MS (FAB) m/lz 542 (W+41).
Example 33 3-N, N-dimethylanuino-4-[ 1-[3 -methoxy-4-[N'-(2-methylphenyl)ureidoj phenylacetylJ-2pyrrolidinylmethoxyjbenzoic acid H I~OOH 37 A stirred mixture of methyl 4-hydroxy-3-nitrobenzoate (3.22g, 0.0 163ino1) and 5% Pd-C (1 2. 9g) in MeOH (3OmL) was hydrogenated under an atmospheric pressure for 70 hr at room temp. Insoluble catalyst was removed, and the filtrate was evaporated in vacua. The residue was chroinatographed on silica-gel with EtOH-CHC 3 (1:20 v/v) as eluent to afford 1 .89g methyl 3-amino-4-hydroxybenzoate as a pale brown syrup.
A stirred mixture of methyl 3-amiino-4-hydroxybenzoate (1.07g, 6.4Ominol) and 5% Pd-C 14g) in MeOH (2OmrL) and 37% aq. formaldehyde (l.O8m-L, 0.0 122mo1) and I N HCI 1lML) 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 chromatogr aphed on silica-gel with EtOAc-n-hexane 10, v/v) as eluent to afford 0.817g methyl NVdimethylamino)-hydroxyfxnzoate as a syrup.
To a stirred mixture of methyl 3 N-dlmethylamino)-4-hydroxyvbenzoate 81 7g, 4l18mmol), N-tert-butoxycarbonylprolinol (0.926g, 4.60mmol), Ph 3 P (1.21g, 4 .6Ommol) in THF (2OmL) was added dropwise DIAD (0.953mL, 4.6Ommol) at 0 0 C. The resulting miuxture was heated under reflux for 41 hr. After cooling, the mixture was evaporated off in vacuo. The residue was chromatographed on silica-gel with EtOAc -n-hexane 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 CH 2
C
2 (lOm.L) and added TFA (10m.L). After the solution was stirred for 5 hr at room temp, thc solution was evaporated in vactio. Water was added to the residue and washed with
CHCI
3 The aqueous layer was neutralized by the addition of sat. NaJ-C0 1 and extracted with :15 CHCI3. The extract was dried over NaSO, and evaporated in vacuo to afford 1 .03g methyl :3 N-dimethylamnino)-4-2-pyrrolidinylmethoxy)bezoate as a gum.
UA m-ixtur of methyl N-dimethylamino)-4-(2-pyrolidinylmethox)benzoate (0.529g, I 1.9Ommol), 3-methoxy-4-[N' -(2-methylphenyl)ureido] phenylacetic acid 597g, 1 .9Ommol), HOBt (0.308g, 2.28mmol), 4-dimethylan-minopyridine(D)MAP) (23.2mg, 0. 190mmol), and 1 -ethyl- 3 3 -,dimethylaminopropyl)carbodiim-ide (EDC) (0.437g, 2.28mxnol) in DMIF (10m.L) was stirred for 15 hr at room temp. The mixture was neutralized by the carefully addition of I N HCI and extracted with EtOAc. The extract was dried over Na 2
SO
4 and evaporated in vacuo to afford 0.607g methyl 3-NN-dimethylamino-4-[ 1-[3 -methoxy-4-[N' -(2-methylphenyl)urcido] phenylacetylj-2-pyrrolidinyl methoxyjbenzoate as a white crystalline material.
A mixture of methyl 3-NN-dimethylamino-4-[ 1-[3 -methoxy-4-[N' -(2-methylphenyl) ureido]-phenylacetylJ-2-pyrrolidinylmethoybezote (0 6 00g, I .O4rtuol) in TI-F (l0m.L) and 0.25 N NaOH (5m.L, I .25mmol) was stirred for 21 hr at room 1=mp. CHC1 3 was added to the mixture and extracted with a mixture of water(lO0mL)- IN NaOH (4mL). The extract was neutralized with sat. NHCI and extracted withCHC 3 The extract was dried over NaSO, and evaporated in vacuo to afford 4 28mg 37 as a white crystalline solid. 'H-NMLR (400M1-z, 1.88, 1.99 and 2.11 (4H, each in), 2.24 (3H, 2.67 (6H, 3.33 (2H, in), 3.58 (2H, WO 01/00206 WO 0100206PCT/USOO/18079 in), 4.05-4.32 (3H, in), 6.75 (11, d, J=7.3Hz), 6.92-6.95 (1H, in), 7.05 (1H, d, J=8.3Hz), 7.11-7.17 (2H, in), 7.42 (111, 7.52 (1H, d, J=7.8H4z), 7.79 (111, d, J=7.8Hz), 8.00 (LH, d, J=7.8Hz), 8.31 (111, 8.46, 8.55 (each 111, each MIS (FAB) m/z 533 Examp~le 34 3-fluoro-4-[[I -[3-methoxy-4-[N'-(2-methylphenyl)ureidojphenylacetyl]-2-pyrrolidiny~methoxyI benzoic acid
OOH
38 To a stirred solution of 4-bromo-2-fluoinphenol (217 iii, 2.002 inmol), N-tertbutoxycarbonyl prolinol (403 mg, 2.002 mmol), and Ph3P (630 mg, 2.403 inmol) in THF (7 mL) was added DIAD (477 ul, 2.423 iniol) at room temp. The resulting mixture was stirred for 6 hr at roam temp and then overnight at 70 0 C. The mixture was concentrated in vacuo and the residue was chromatographed on silica-gel with n-hexane EtOAc 1, v/v) as eluent to give 549.4 mg 1-bromo-4-[ 1-Qert-butoxycarbonyl)-2-pyrrolidinyl~methoxy-3-fluorobenzene as an oil. 'H- NNvIR (400 Mz, CDC 3 8 1.46 9H), 1.85 (brmi, 1H), 1.90 -2.10 (brs,311), 3.30-3.47(in, 211), 3.85, 4.04 (hr s each, 111), 4.11 4.20 (in, 2H1), 6.82 6.98 (in, 111), 7.13 7.26 (in, 2H1); MS (FAB)m/nz 374 To a stirred solution of 1-bromo-4-[ 1-(tert-butoxycarbonyl)-2-pyrrolidinylI methoxy-3fluorobenzene (549.4 mg, 1.468 minol) in DMS0 (6 mL) and MeOH (5 mL) was added Et 3 N (448 ul, 3.229 mrnol), Pd(OAc) 2 (36.2 mg, 0. 161 mmol), and 1,3-bis(diphenylphosphino)propane (66.4 mng, 0.161Iimol). To the mixture was induced CO gas for 10 mmd. 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 chroinatogmaphed on silica-gel eluting with n-hexane:EtOAc 1, v/v) as eluent to afford 323.0 mg methyl 1-(terfbutoxycarbonyl)-2-pyrrolidinylj inethoxy-3-fluorobenzoate as a pale yellow oil. 'H-NIMR (400 MHz, CDC1 3 8 1.47 911), 1.87(br s, 1H1), 1.95 2. 10 (in, 314i), 3.34 3.44 (br m, 211), 3.89 (s, 311), 3.94 and 4.11 4.26 (br in each, 311), 7.03 7. 11 (in, 111), 7.75 7.80 (in, 211); MS (FAB) n/z 354 To a stirred solution of methyl 4-[l-(tert-butoxycarbonyl)-2-pyrrolidinyl Jinethoxy-3fluorobenzoate (323.0 mg, 0.9 14 minol) in CH 2
CI
2 (6.5 mL) was added TEA (1.3 mL) at 0 0 C, and WO 01/00206 WO 0100206PCT/USOO/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
CHCI
3 The extract was washed with brine, dried over Na 2
SO
4 and concentrated under a reduced pressure to afford 174.8 mg methyl 3-fluoro-4-(2-pyrrolidinyl)methoxybenzoate as a brownish oil. 1 H-NNMR(400 MHz, CDCI 3 8 1.54 1.63 (in, 111), 1.76 -2.02 (in, 311), 2.93 3.07 (in, 2H), 3.57 (ddd, J= 4.9, 6.9, 14.3 Hz, 111), 3.89 3H), 3.97 (dd, J= 6.8, 9.3 Hz, IH), 4.04 (dd, J 5.0, 8.8 Hz, 111), 6.98 J 17.6 Hz,1IH), 7.73 (dd, J 11.7 Hz, 7.78 (dt, J 8.8 Hz,1IH); MS (FAB) m/z 253 1).
A mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-inethylphenyl)ureidolphenylacetate (324.5 mng, 0.676 mmol), methyl 3-fluoro-4-(2-pyrrolidinyl)inethoxybenzoate (171.1 mg, 0.676 inmol), Et 3 N (113 ul, 0.811 mmol) in DMI (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 v/v) as eluent to afford methyl 3-fluoro-4-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-2-pyrrolidinyllmethoxyI benzoate (365.1 mng, 0.664 mmol) as an oil. To a stirred solution of this compound in THF (4.4 mL) and H20(1.1 inL) was added LiOH (46.3 ing, 1.932 inmol), and the reaction midxture was stirred at room temp overnight. The mixture was diluted with CHC1 3 and acidified by the addition of IN HCI. 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 crude solid was recrystallized from n-hexane-EtOAc-CHC 3 to afford 102 mng 38 as a white crystalline powder. mp 123 126; IR (KBr) 1616, 1537, 1282, 756cnr'; 'H-NMR (400 MlHz, DMSO-d 6 5 1.87 2.09 (in, 411), 2.25 3H), 3.48 3.57 (in, 211), 3.60 2H1), 3.83 311), 4.11 4.16 (in, lH), 4.24 (dd, J 2.9, 9.8 Hz, 111), 4.28 4.34 (br s, 111), 6.74 (dd, J 1.5, 8.3 Hz, IH), 6.87 (s,1IH), 6.94 J =7.3Hz, IH),7.12 J =7.8 Hz, IH), 7.15 J =8.3 Hz,IH), 7.34 J 8.8 Hz, 111), 7.66 (dd, J 2.0, 12.2 Hz, 111), 7.73 J 9.3 Hz, 111), 7.79 J 8.3 Hz, 111), 7.99 J= 7.8 Hz, 111), 8.46 lH), 8.55 111); MS (FAB) m/z 536 AnaL. Calcd for C,,H3FN 3 0 6 -0.5H 2 0: Q, 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 4-11 -[4-[N'-(2-fluorophenyl)ureidoJ-3-nethoxyphenylacetyl-2-pyrrolidinyljinethoxy-3-nethoxy benzoic acid WO 01/00206 WO 0100206PCT/USOO/18079 SH H 39 A mixture of 4-[N'-(2-fluorophenyl)ureidoj-3-methoxyphenylacetic acid (318 mg, 1 mmol), 2-(2-methoxy-4-etboxycarlbonyl)phenoxymethylpyrrolidine (279 mg, 1 mmol), EDC (288 mg, 1. 5 mmol), and DMAP (122 mg, 1 mmol) in DMP (20 niL) was stirred overnight. The mixture was poured into 1 N HC1 and the resulting solid was collected with suction. The solid was dissolved in CHC1 3 and dried over MgSO 4 After removal of the solvent, the residue was chromatographed on silica-gel with CHCI 3 MeOH (100: 1, v/v) as eluent to give an oil, which was dissolved in THiF:MeOH 1, v/v, 10 niL). 0.25 N NaOH (8 mL) was added to the solution and the resulting stirred mixture was heated under reflux for 3 hr. The midxture was poured into I N HCI.
The resulting solid was collected with suction, dissolved in CHCI 3 dried over MgSO 4 and evaporated. The residue was recrystallized from CHCI 3 -n-hexane-ether to give 329 mg 39 as a white crystalline powder. mp 140-144; IR (KBr) 3338, 2956, 2875, 2607, 1709 IH-NNM (CDC1 3 8 1.95-2.25 (4 in), 3.45-4.50 (12 H, in), 6.66-8.15 (12 H, mn); MS (FAB) m/z 552 Example 36 1 -[3-inethoxy-4-[N'-(2-methylphenyl)ureidojphenylacetylj-2-pyrrolidinyl]methoxy] carboxylic acid 04 To a stirred solution of 6-hydroxynicotinic acid (500 mg, 3.594 mmol) in benzene (8 niL) and MeQH (2 niL) was added dropwise TMSCHN, (1.97 mL, 3.953 minol) 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 chroinatographed on silica-gel with n-hexane-EtOAc v/v) as eluent to give 269.8 mg (491/) methyl 2as a white crystalline powder. JR (KBr) 3062, 1657, 1654, 1612, 1435, 1300, 1113,775, 642cm 1 '11-NiVR (400 M41z, CDCl 3 8 3.87 3H1), 6.58 J =9.8 Hz, 111), 7.99 (dd, J1 2.4, 9.8 Hz, 111), 8.19 J 2.4 Hz, 111); MIS (FAB) m/z 154 Anal Calcd for C 1
H
7 N0 3 1/4H,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 ing, 1 .762mmo1), WO 01/00206 WO 0100206PCT/USOO/18079 N-tert-butoxycarbonylprolinol (354.6 ing, 1.762 mmnol), and Ph 3 P (554.6 mng, 2.114 minol) in THE inL) was slowly added DIAD (0.42 inL, 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 1, vfv) as eluent to give 262.5 mg methyl 2-[[I1-Qert-butoxycarbonyl)-2-pyrrolidinylJmethoxy]pyridine-5-carboxylate as an oil. 'H- NMR (400 MlHz, CDCI 3 8 1.47 911), 1.85 1.98 (in, 411), 3.37 (br s, 211), 3.92 311), 4.12 4.33 (br m, 2H),4.4 (brs, IH),6.75(m, 1H),8. 15 (in, 111), 8.79 (in, IH); MS (FAB) nihz 337 To a stirred solution of methyl 2-t[[1-Qtert-butoxycarbonyl)-2-pyrrolidinyllinethoxy] (262.5mg, 0.870 nunol) in CH 2
CI
2 (5.3 mL) was added TEA 1 iL) at 0 0 C, and the resulting midxture 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 CHC13. The extract was washed with brine, dried over Na 2
SO
4 and concentrated under a reduced pressure to afford 173.1 mng methyl 2-[(2-pyrrolidinyl)inethoxyjpyridine-5carboxylate as a pale yellowish oil. 'H-NMR (400 MHz, CDCI 3 8 1.49 1.58 (ddt, J 6.8, 8.8 Hz, 111), 1.72 1.87 (in, 2H), 1.90 1.99 (in, 111), 2.92 -3.05 (mn, 211), 3.50 -3.57 (dddJ=4.4,7.3, 15.1 Hz, 11H), 3.91 31H), 4.23 (dd, J 10.7 Hz, I1H), 4.3 8 (dd, J 10.7 Hz, I 6.78 J 8.8 Hz, 111), 8.15 (dd, J 8.8 Hz, 111), 8.80 J 2.4 Hz, 11H); MS (FAB) nh/z 23 7 A mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetate (351.7 mng, 0.732 inmol), methyl 2-[(2-pyrrolidinyl)inethoxylpyridine-5-carboxylate (173.0 mg, 0.732 inunol), Et 3 N (122.4 jul, 0.878 minol) 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 SO4. The solvent was removed under a reduced pressure and the residue was chromatographed on silica-gel with nhexane:EtOAc vlv) as eluent to afford methyl 2-[Il-[3-methoxy-4-[N'-(2-inethylphenyl) ureidolphenylacetyll-2-pyrrolidinyl~methoxyJ pyridine-5-carboxylate (338.4 mng, 87%) as an oil.
To a stirred solution of this compound in THE (5.6 inL) and 1120(1.4 mL) was added LiOH (45.7 ing, 1.91 minol), and the reaction mixture was stirred at room temp overnight. The mixture was diluted with CHCI 3 and treated with sat. NH 4 CI, washed with brine, died 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 O-CHC1 3 -MeOH to afford 193.8 mng 40 as a white crystalline powder. mp 125 128; IR (KBr) 1716, 1600, 1533, 1255cm"'; 'H-NMIR (400 MlHz, DMVSO-d) 8 1.67 2.03 (in, 411, 2.50 311), 3.33 3.42 (mi, 111), 3.52 (mn, 211), 3.58 J =4.4 Hz, 114), 3.83 311), 4.27 4.31 (mn, 211), 4.42 4.47 (in, 111), 6.73 J 7.8 Hz, 1141), 6.87 6.95 (in, 311), 7.11 7.17 (mn, WO 01/00206 WO 0100206PCT/USOO/18079 2H), 7.79 J= 8.3 Hz, 1H), 7.99 J= 8.3 Hz, 1H), 8.14 (dd, J= 2.0, 8.8 Hz, 1H), 8.46 1H), 8.56 IH), 8.69 J=2.0 Hz, I1H), 13.06 (br s, lH); MS (FAB) m/z 519 (M+1);AnaL. Calcd for CaH 30
N
4
Q
6 d1/2HO: C, 63.75; H, 5.92; N, 10.62. Found: C, 63.61; H, 5.94; N, 10.27.
Example 37 3 -methoxy-4-[2-[4-[Ar-(2-methylphenyl)ureidojbenzylJ-4-thiazolyl~methoxybenzoic acid OOH 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 g, 12.33 mmol) at room temp. The resulting mixture was stirred for 17 hr at room temp. The mixture was diluted with EtOAc and washed with 10% K 3 P0 4 The aqueous layer was extracted with CH 2
CI
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-NNvI (400 Miz, CDCI 3 564.15 2H), 7.51 J 8.3 Hz, 2H), 8.24 J 8.8 Hz, 2H); MS (FAB) m/lz 197 Anal. Calcd for C 3
H
8
N
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. To a stirred solution of 4-nitrobenzylcarbothioamide (502.0 mg, 2.558 mxnol) in EtOH niL) was added 1,3-dichloro-2-propanone (649.6 mig, 5.16 nimol) and the mixture was heated under reflux for I hr. The mixture was concentrated and the residue was diluted with CHC 3 The solution was washed with IN 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 1, vfv) as eluent to afford 495.2 mg (72 4-[2-(4-nitrobenzyl)thiazolyljmethyl chloride as a pale yellow oil. 'H-NMR (400 MHz, CDC 3 6 4.43 2H), 4.68 2H), 7.23 IN), 7.49 J= 8.8 Hz, 2H), 8.20 J= 8.8 Hz, 2H); MS (FAB) m/z 269 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-mtrobenzyl)thiazolyll 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 CHCI 3 The extract was washed,%with H,0, brine, and dried over NaSO 4 The solvent was removed under a reduced pressure and the residue was WO 01/00206 PTUO/87 PCT/USOO/18079 chromatographed on silica-gel with n-hexane:EtOAc 1, vA') as eluent to afford 201.7 mg (60 ethyl 3-methoxy--2-(4-nitrobenzyl)-4-thiazolyljmethoxybenzoate as a pale yellow oil. IH-NMR (400 NMz, CDCl 3 8 1.39 J 7.3 Hz, 3H), 3.93 3H), 4.36 (q,J 7.3 Hz, 2H), 4.44 2H), 5.31 2H), 6.97 J 8.3 Hz, I 7.28 IlH), 7.48 J 8.8 Hz, 2H), 7.57 J 2.0 Hz, 1H), 7.64 (dd, J 8.3 Hz, 1H), 8.20 J= 8.8 Hz, 2H); MS (FAB) ,n/z 429 (MW+l).
A stirred solution of ethyl 3-methoxy-4-[2-(4-aminobenzyl)-4-thiazoly methoxybenzoate (201.7 mg, 0.47 1 mmol) and 5% Pd/C (40 mg) in EtQH (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, v/v) as eluent to afford 87.8 mg ethyl 3-methoxy-4- [2-(4-amninobenzyl)-4-thiazolyljmethoxybenzoate as a yellowish crystalline powder. 'H-NMR (400 MHz, CDCI 3 8 1.38 J 7.3 Hz, 3H), 3.93 3H), 4.21 2H), 4.35 J 7.3 Hz, 2H), 5.30 2H), 6.66 (dd, J 2.0, 6.4 Hz, 2H), 6.97 J =8.3 Hz, I 7.09 J =8.3 Hz, 2H), 7.18 (s, I 7.56 J 2.0 Hz, I 7.64 (dd, J 8.3 Hz, IlH); MS (FAB) m/z 3 99 To a solution of ethyl 3-methox-[2-(4-aminobenzyl)-4-tiazolyllmehoxybenzoate (87.8 mg, 0.220 mmol) in TI-F (2.0 mL) was added triethylamine (30.5 ul, 0.220 mmol) and otolyl isocyanate 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 CHCI 3 washed with H,0, and brine. The extract was dried over Na 2
SO
4 The solvent was removed under a reduced pressure to afford 110.4 mng (94 ethyl 3-methoxy-4-12-[4- [N'-(2-methylphenyl) ureido]benzylj-4-thiazolylJ methoxybenzoate as a pale yellow crystalline powder. lH-NMR (400 MHz, CDCI 3 8 1.38 J 7.3 Hz, 311), 2.28 3H), 3.92 3H), 4.28 (s, 2H), 4.3 5 J 7.3 Hz, 2H), 5.29 2H), 6.20(s, IlH), 6.47 I1H), 6.97 J 8.8 Hz, I1H), 7.20 I1H), 7.24 2H), 7.27 2H), 7.3 3 J =8.3 Hz, 2H), 7.49 J= 7.3 Hz, IlH), 7.56 J Hz, I1H), 7.63 (dd, J= 2.0, 8.3 Hz, IlH); MS (FAB) m/z 53 2 To a stirred solution of ethyl 3-methoxy-4-[2-[4-[N'-(2-methylphenyl)ureido] benzyl]-4thiazolyllmethoxybenzoate in THE (1.6 mL) and H 2 0 (0.4 mL) was added LiOH (6.0 mg, 0.249 rumol), 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 CHC1 3 The solution was made basic by the addition of IN NaOH. The aqueous extract was acidified by the addition of IN HCI and extracted with CHC 3 The extract was washed with brine and dried over Na 2
SQ
4 The solvent was removed under a reduced pressure and the obtained crude solid was recrystallized from n-hexane-EtOAc-EtOH to WO 01/00206 WO 0100206PCT/USJO/18079 afford 59.6 mg 41 as a white crystalline powder. mp 243 245; JR (KBr) 3282, 1685, 1637, 1600, 1554, 1516, 1278, 1234, 763, 748cm-1; 'H-NMR (400 MHz, DMSO-d,) 8 2.27 3H4), 3.83 3H), 4.30 (s,214), 5.21 2H), 6.97 J= 7.3 Hz, 111), 7.15 7.24 (in, 3H), 7.29 J =8.8 Hz, 2H), 7.47 (d,iJ 8.3 Hz, 2H), 7.50 1H), 7.58 J= 8.3 Hz, 7.62 111), 7.86 J 7.8 Hz, 1H), 7.97 I1H), 9.09 I1H), 12.70 (br s, 114); MS (FAB) m/z 504 AnaI. Calcd for CQ 7 HNOSd /4H 2 O: C, 63.83; HK 5.06; N, 8.27. Found: C, 63.74; H, 4.99; N, 8. Example 38 4-[I-f 3-methoxy-4-[N' -(2-methylphenyl)ureidojphenylacetylj-2-methyl-2-pyrrolidinyl methoxy]- 3-nitrobenzoic acid H IH -OOH 1 02P42 To a stirred solution of the N-tert-butoxycarbonylproline (6.00g, 0.0279mo1) in MeOHbenene(1:4, v/v) was added dropwise 2.0 M-n-hexane solution of TMSCHN, (16.7inL, 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 Nt-tert-butoxycarbonylproline methyl ester as a yellow syrup. 'H-NMR (CDC 3 8 1.41 914), 1.85-1.98 (in, 4H), 2.21-2.28 (mn, 214), 3.72 314), 4.29 (in, 1H).
To a stirred solution of diisopropylamine (2.O2mL, 0.0 l44mol) in THF (3OmL) was added dropwise 1. 59 M n-hexane solution of n-BuLi (9.O6mL, 0.0 144mo1) at minus 78'C for over 5 min.
The resulting solution was stirred for 20 min at minus 78'C. To the solution was added dropwise N-fert-butoxycarbonylproline methyl ester (3.00g, 0.0 13 lmmol) in THE (3OmL) at minus 78*C for over 5 min. The resulting solution was stirred for 10 min at minus 78*C. To the solution was added dropwise Mel (0.900mL, 0.0 144ino1) at minus 78'C. The resulting solution was stirred for minat minus 78 0 C. The solution was quenched by the addition of sat. NH 4 CI. Thieresulting mixture was extracted with CHC1 3 The extract was washed with water, dried over Na 2 SO,, and evaporated in vacuo to afford 3.20g N-tert-butoxycarbonyl-2-methylproline methyl ester as a yellow syrup. 'H-NMR (CDC 3 8 1.33 9H), 1.38 3H), 1.72-2.20 (in, 4H) 3.27-3.59 (in, 2H) 3.63 ,fr-6.3H1z, 3H).
To a stirred solution of N-terl-butoxycarbonyl-2-methylproline methyl ester (3.20g, 0.Ol3linol) in THE (2OnmL) was added IN NaOH (15.7mL) at room temp. After the resulting WO 01/00206 WO 0100206PCT/USOO/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 1IN HCI, and extracted with EtOAc. The extract was dried over Na 2 SO, and evaporated in vacuo to afford 1.7 1 g(57/6) N-ferfbutoxycarbonyl-2-methylproline as a yellow syrup. 'H-NMR (CDC 3 8 1.42 9H), 1.48 3H), 1.88-2.3 1 (in, 4H), 3.34-3.57 (in, 2H), 9.35 (br s, 1H) To a stirred solution of N-tert-butoxycarbonyl-2-methylproline 10g, 4.80rnnol) in THF (2OimL) was added dropwise BH 3 -SMe- 2 546inL, 5.76inmol) at room temp. After the resulting mixture was stirred for 6 hr at 80'C, the mixture was evaporated in vacuo. The residue was diluted with MeOH, washed with n-hexane and evaporated in vacuo to afford 0.648g N-tert-butoxycarbonyl-2-hydroxymethyl-2-methylpyrrolidine as a yellow syrup. 'H-NMR (CDC 3 8 1.47 911), 1.76-2.05 (in, 4H), 3.28-3.48 (in, 2H), 3.66 (in, 2H, d).
To a stirred solution of N-tert-butoxycarbonyl-2-hydroxymethyl-2-inethylpyrrwlidine (0.648g, 3.0iminol), methyl 4-hydroxy-3-nitrobenzoate (0.593g, 3.0 immol), and Ph 3 P (0.868g, 3.3 immol) in TI-F (lOmL) was added dropwise DIAD (O.686inL, 3.3 immol) at 0 0 C. After the resulting mixture was stirred for 24hr at 80*C, the mixture was evaporated in vacuo. The residue was diluted with CH 2
CI
2 (5inL) and added TWA (5mL). After the resulting mixture was stirred for 2hr at room temp, the mixture was evaporated in vacuo. The residue was diluted with HCI and extracted with CHCI 3 The aqueous layer was neutralized with sat. NaHCO 3 and extracted with CHC 3 The extract was dried over Na 2
SO
4 and evaporated in vacuo to afford 0. 188g methyl 3-nitro.4-(2-methyl-2-pyrrolidinylinethoxy) benzoate as a yellow syrup.
A mixture of methyl 3-nitro-4-(2-inethyl-2-pyrrolidinylmethoxy)benzoate 188g, 0.92Ommol), 3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetic acid (0.289g, 0.92Ominol), HOBt 149g, 1. l0minol), DMAP (I11.2mg, 0.092Oinmol) and EDC (0.21 Ig, 1. l0mmol) in DMF was stirred for l4hr at room temp. EtOAC was added to the mixture and the solution was 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 methyl 4-[[1-[3-methoxy-4-[N'-(2methylphenyl)ureido] phenylacetyl]-2-iethyl-2-pyrrolidinyllmethoxyj-3-nitrobenzoate as a yellow crystalline material. 'H-NMR (CDC 3 6 1.26 .1=5.9Hz), 1.85-4.50 (in, 1011), 2.30 3H), 3.67 2H), 3.92 6.36 2H), 6.75-7.52 (in, 7H), 8.02 J7.8Hz, 1H), 8.15 J=8.8Hz, 1Hf), 8.47 IH).
WO 01/00206 WO 0100206PCTUS0OI18079 A stirred mixture of 4-[I-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetyl]-2methyl-2-pyrrolidinyllmethoxyj-3-nitrobenzoate (0.489g, 0.828mmo1) in MeOH (SiuL) and IN NaOH (1.24mL) was heated under reflux for 2hr After cooling, the mixture was diluted with water and extracted with CHC 3 The aqueous layer was acidified with IN HCI, and extracted with CHCI,. The extract was dried over Na 2
SO
4 and evaporated in vacua to afford 0.366g 42 as a yellow crystalline material.
Example 39 4-[4-hydroxy- 1 -[3-methoxy-4-[NV'-(2-methylphenyl)urrido~phenylacetyl]-2-pyrrolidinylmethoxyj- 3- methoxybenzoic acid 9H Mf qOOH 43 A stirred mixture of 4-[4-benzyloxy-l -[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetylJ-2-pyrrolidinylmethoxy]-3-methoxybenzoate (440 mg, 0.645 mmol) and 5% Pd/C (400 mg) in AcOH:EtOH 1, v/v, 100 mL) was hydrogenated at 1 atmn for 5 hr. The mixture was filtered to remove the catalyst and the filtrate was concentrated in vacua. The residue was chromatographed on silica-gel with CHCI,:EtOH (10: 1, vfv) as eluent to give 90 mg ethyl 4- [4-hydroxy-1 -[3-methoxy-4-[N' -(2-methylphenyl)ureido] phenylacetyl]-2-pyrrolidinylmethoxyj-3methoxybenzoate as a pale yellow oil. 'H-NMR (CDCI 3 5 1.39 (3 H, t, J 7.3 Hz), 2.04-2.37 (total 5 H, in), 3.44-4.70 (16 H, series of in), 6.63 (1 H, 6.70-6.80 (2 H, in), 6.84 (1 H, d, J 8.3 Hz), 7. 11 (1 H, t, J 7.8 Hz), 7.20-7.24 (3 H, in), 7.45 (1 H, d, J 2.0 Hz), 7.59 (2 H, dd, J =8.3, 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-methylpheniyl)ureidoj phenylacetylJ-2-pyrrwlidinylmethoxy]-3-methoxyfbenzoate (90 mg, 0. 152 nimol) in 0.25 N NaOH mL, 1.25 minol) and THlE (5 mL) was heated under reflux: overnight. The mixture was poured into ice- I N HCI (200 mL). The precipitate was collected with suction and recrystallized from
CHCI
3 -MeOH-n-hexane to give 40 mng 43 as a colorless amorphous solid. 'H-NMR (DMSOd 6 5 1.92-2.11 (2 H, in), 2.24 (3 H, 3.31-5.07 (14 H, series of in), 6.73 (1 H, d, J =8.3 Hz), 6.84 (1 H, 6.93 (1 H, t, J= 7.8 Hz), 7.01-7.17 (3 H, in), 7.44 (1 H, 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, 8.55 (1 H, 12.67 (1 H, br s); MS (FAB) m/z 564 1).
WO 01/00206 WO 0100206PCT/LJSOO/18079 Example (2S,4R)-3-amino-4-[4-hydroxy-l1-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetylj-2pyrrolidinyl~methoxybenzoic acid 9H
NH
2 IHH OOH 4 To a stirred solution of (2S,4R)-4-benzyloxy-l1-Qtert-butoxycarbonyl)-2-prolinol (891 mg, 2.9 mmol), methyl 4-hydroxy-3-nitrobenzoate (572 mg, 2.9 mmol),and PPli 3 (839 mg, 3.2 mmol) in TBY (6 mL) was added DIAl) (630 raL, 3.2 nunol) and the mixture was heated under reflux overnight. After removal of the solvent, the residue was chromatographed on silica-gel with nhexane:EtOAc 1) and toluene:EtOAc (10: 1, v/v) as eluent to give 700 mg methyl (2S,4R)-4-[4-benzyloxy-lI-(tert-butoxycarbonyl)-2-pyrrolidinyl~methoxy-3-nitrobenzoate as a pale yellow oil.
To a stirred solution of methyl (2S, 4R)-4-[4-benzyloxy-l1-(tert-butoxycarbonyl)-2pyrrolidinyl] methoxy-3-nitrobenzoate (681 mg, 1.4 mmol) in CH 2
CI
2 (2 niL) was added TFA (2 niL), and the resulting mixture was stirred for 2 hr. After the reaction mixture was concentrated, the residue was made basic by the addition of sat. NaHCO 3 and extracted with CHC 3 The extract was washed with H 2 0, dried over MgSO 4 and evaporated to give 511 mg methyl (2S,4R)-4- [4-benzyloxy-2-pyrrolidinyllmethoxy-3-nitrobenzoate as a yellow oil.
A mixture of 3-methoxy-4-IN' -(2-methylphenyl)ureidolphenylacetic acid (409 mg, 1.3 mmol), methyl (2S,4R)-4- (benzyloxy-2-pyrrolidinyl)methoxy-3-nitrobenzoate (502 mg, 1.3 mmol), EDC (3 83 mg, 2 mmol), and DMAP (15 9 mg 1. 3 mmol) in DMF (20 niL) was stirred for 3 days. The mixture was poured into 1 N HCI 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 CHCI 3 :MeOH (200: 1, v/v) as eluent to give 680 mg methyl (2S,4R)-4-[4-benzyloxy-lI-[3-methoxy-4-[N'-(2-methylphenyl)ureidoj phenylacetylj-2-pyrrolidinylJmethoxy-3-nitrobenzoate as a white amorphous solid.
A solution of methyl (2S,4R)-4- [4-benzyloxy-l -[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenylacetyl]-2-pyrrolidinyl~methoxy-3-nitrobenzoate (676 mg, 0.99 mmol) and 5% Pd-C WO 01/00206 WO 0100206PCT/USOO/18079 (1 g) in EtOH:AcOH 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 CHCI 3 -EtOH-n-hexane as eluent to give 120 mng 44 as a pale yellow crystalline powder. MS (FAB) m/z 549 (M'+lI) Example 41 4-[14-fluoro-l1-[3-methoxy-4-IIN'-(2-methylphenyl)ureidolphenylacetyll-2-pyrrolidinylI methoxy]- 3-methoxybenzoic acid
F
~OOH A stirred mixture of ethyl 4-[4-benzyloxy-1 -(tert-butoxycarbonyl)-2-pyrrolidinylj methoxy- 3-methoxybenzoate 189 g, 2.449 mmol) and 5% Pd-C (240 mg) in EtOH (10 miL) 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-[l-Qert-butoxycarbonyl)-4-hydroxy-2pyrrolidinyl]methoxy-3-methoxybenzoate (735.3 mg, 76%) as a pale yellow oil. To a stirred cold (minus 78 0 C) solution of DAST (0.491 mL, 3.718 mmol) in CH 2
C
2 (7.4 niL) was added dropwise a solution of this compound in CH 2 C1 2 (2rnL), and the resulting mixture was stirred overnight.
The mixture was quenched with water and extracted with CHCI, 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 1, v/v) as eluent to afford 418.7 ing (57046) ethyl 1-Qtert-butoxycarbonyl)-4-fluoro-2-pyrrolidinyllmethoxy-3-methoxybenzoate as an oil. 'H-NMR (400MNHz, CDCIO)8 1:.39 J= 7.3 Hz, 3H), 1.49 9H), 2.16 (brmi, 1H), 2.58 (dd, J 15.6, 19.0 Hz, 1H), 3.60 3.75 (in, 2H), 3.91 3H), 3.97 J 9.3 Hz, 1H), 4.35 J 7.3 Hz, 2H), 4.33 4.53 (in, 2H), 5.25 J =52.7 Hz, 1H), 7.04 (dd, J 56.2 Hz, 1H), 7.55 (s, IH), 7.65 (br s, 1H); MIS (FAB) m/z 398 To a stirred solution of ethyl 4-[1 -Qtert-butoxycarbonyl)-4-fluoro-2-pyrrolidinyllmethoxy- 3-methoxybenzoate (482.2 mg, 1.2 13 nunol) in CH 2
CI
2 (10.0 niL) was added TFA (1.9 niL) at 0 0 C, 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 CHC 3 The extract was washed with brine, dried over Na 2
SO
4 and concentrated under a reduced pressure WO 01/00206 WO 0100206PCT/USOU/18079 to afford 348.7 mg ethyl 4-(4-fluoro-2-pyrrolidinyl)methoxy-3-methoxybenzoate as a brownish oil. 'H-NMR (400MNHz, CDCI 3 5 1.39 J =6.8 Hz, 3H), 1.97 (ddt, J= 1.5, 5.4, 14.7 H-z,l111), 2.27 (dddd, J 5.9, 8.8, 14.7, 32.7 Hz,1IH), 3.02 (ddd, J 13.1, 35.2 Hz' 1H), 3.36 (dd, J =12.7, 21.5 Hz,l1-1), 3.65 (in, lH), 3.90 3H), 4.09 (mn, lH), 4.35 J =6.8 Hz, 2H), 5.17, 5.31 (br meach, lH), 6.90 J= 8.3 Hz, 111), 7.75 J= 2.0 Hz, IH), 7.65 (dd, J 8.3 Hz, 1H); MIS (FAB) m/z 298 1).
A mixture of pentafluorophenyl 3-inethoxy-4-[N'-(2-methylphenyl)ureido ]phenylacetate (404.0 mg, 0.840 nunol), ethyl 4-(4-fluoro-2-pyrrolidinyl)methoxy-3-methoxybenzoate (250.0 mg, 0.840 mmol), Et 3 N (14 Ipi, 1.009 mmol) in DMIF (4.0 mL) was stirred for 1 hr at room temp. The mixture was diluted with EtOAc, washed with water, brine, and dried over NaSO 4 The solvent was removed under a reduced pressure and the residue was chromatographed on silica-gel with nhexane:EtOAc v/v) to afford 502 mg of ethyl 4-[[4-fluoro-l-[3-methoxy-4-[N' methylphenyl)ureido] phenylactylj-2-pyrrolidinyllmethoxyJ-3-methoxybenzoate as a yellow oil.
To a stirred solution of this compound in THE (8.0 mL) and H 2 0 (2.0 inL) was added LIOH (60.4mg, 2.520 inmol), and the mixture was stirred at room temp. overnight, and 50*C for 1 day.
The mixture was diluted with CHCI 3 and extracted with IN NaOH. The aqueous layer was acidified by the addition of IN HCI 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 obtained crude solid was recrystallized from EtOAc-CHC1 3 -EtOH-n-hexane to afford 294.8 mng 45 as a white crystalline powder. IR (KBr) 2958, 2937, 1687, 1601, 1531, 1454, 1419, 1267, 1214, 1029cm-'; 'H-NMR (400 M1Hz, DMSO-d 6 5 1.86 -2.09 (in, 511), 2.06 3H), 2.25 3.47 3.67 (in, 6H), 3.76 3H), 4.05 4.12 (in, 2H), 4.30 4.31 (in, 1H), 6.51 IH), 6.55 111), 6.73 6.95 (in, 2H), 7.11 7.17 (in, 2H), 7.64 111), 7.79 J 7.8 Hz, lH), 7.99 J 7.8 Hz, 8.47 11H), 8.55 lH); MS (FAB) niz 566 Anal. Calcd for C3,H 32
FN
3
O
7 ]1/2H 2 O: C, 62.7 1; H, 5.79; F, 3.3 1; N, 7.3 1. Found: C, 63.13; H, 6.17; F, 3.12; N, 7.04.
Example 42 3-acetylamiino- 4 -Il-13-methoxy-4-[N'-(2-methylphenyl)ureidojphenylacetyl]-2-pyrrolidinyI methoxyjbenzoic acid WO 01/00206 WO 0100206PCT/USOO/18079 U 46 A solution of 3-anlino-4-[ 1-[3 -methoxy-4-[N'-(2-methylphenyl)ureidoJ phenylacetylJ-2pyrrolidinylniethoxylbenzoic acid (130mg, 0.244mmo1) and DMAP (2.9mg, 0.0244nuno1) in pyridine (5mL) and acetic anhydride (5mL) was stirred for 2 hr at room temp. The mixture was evaporated off in vacuo (exess acetic anhydride was azeotropically removed with toluene). Water was added to the residue, and extracted with CHCI,. The extract was dried over Na 2
SO
4 and evaporated in vacuo. The residue was chromatographed on silica-gel with MeOH:CHC 3 (1:15 to 1:1, v/v) as eluent to afford 29mg 46 as a white crystalline material. 'H-NMR (DMSO-4) 1.80-2.30 (in, 4H), 2.04 3H1), 2.26 311), 3.33(s, 3H1), 3.40-4.80 (mn, 7H), 6.59 III), 6.74(d, .1=8.3Hz, I 6.79 J=8.8Hz, 11-1), 7.07-7.57 (in, 6H1), 7.75 J=8.8Hz, 1H1), 8.07 .1=8.3Hz, 111), 8.41 and 8.96 (each s, each IIH); MIS (FAB) m/z 575 Example 43 3-chloro-2-[[ 1-[3-methoxy-4-[N'-(2.methylphenyl)ureido~phenylacetylj-2-pyrrolidinyl methoxyJ acid H1H C OOH To a stirred solution of 3-chloro-2-hydroxypyridine-5-carboxylic acid (1 g, 5.762 nunol) in benzene (16 mL) and MeGH (4 mL) was added dropwise TMSCHN 2 (3.17 mL, 6.338 inmol) 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 suspended in water and precipitate was collected. The crude solid was washed with Et 2 O, and dried under a reduced pressure to give 728.1 mng methyl 3-chloro-2-hydroxypyridine-5carboxylate as a white crystalline powder. IR (KBr) 1655, 1282, 1245, 769cm-'; IH-NMR (400 M4Hz, DMSO-d) 8 3.79 3H1), 8.01 111), 8.06 III); MIS (FAB) "Vz 188 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; CI, 19.00; N, 7.34.
To a stirred solution of methyl 3-chloro-2-hydroxypyridine-5-carboxylate (300 mng, 1.599 minol), N-tert-butoxycalbonylprolinol (321.9 mng, 1.599 minol), and Ph 3 P (503 mg, 1.919 nunol) in WO 01/00206 WO 0100206PCT/USOO/18079 THE (3 mL) was slowly added DIAD (378 jud, 1.9 19 nunbI) at room temp, and the 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 1, v/v) as eluent to give 235.6 mg methyl 3-chloro-2- -Qert-butoxycarbonyl)-2-pyrrolidinyljmethoxylpyridine-5-carboxylate as a pale yellowish oil.
'H-NMR (400 MHz, CDCI 3 5 1.46 9H), 1.87 (in, 1H), 2.05 (br s, 311), 3.43 (br s, 2H), 3.92 (s, 311), 4.17, 4.26 (br s each, IH), 4.45 4.51 (in, 111), 4.50 114), 8.21 111), 8.67 J 2.0 Hz, I1H); MS (FAB) m/z 371 To a stirred solution of methyl 3-chloro-2-[[l-Qert-butoxycarboniyl)-2pyrrolidinyl~methoxyj pyridine-5-carboxylate (235.6mg, 0.635 mmol) in CH 2 C1 2 (5.0 niL) was added TEA (1.0 mL) at 0 0 C, and the reaction mixture was stiffed 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 CHCI 3 The extract was dried over Na 2 SO1, concentrated under a reduced pressure to afford 172.3 mg methyl 3-chloro-2-[(2-pyrrolidinyl)methoxylpyridine-5carboxylate as a pale yellowish oil. 'H-NMR (400 MHz, CDCl 3 8 1.55 1.63 (mn, 1H), 1.76 1.99 (in, 3H1), 2.93 2.99 (in, 11-1), 3.02 3.08 (mn, 114), 3.57 -3.62 (in, I1H), 3.92 3H4), 4.33 (dd, J 7.3, 10.7 Hz, 4.44 (dd, J1= 4.4, 10.7 Hz, 114), 8.21 J 2.0 Hz, 114), 8.67 J 2.0 Hz, 111); MS (FAB) ,n/z 271 1).
The mixture of pentafluorophenyl 3-methoxy-4-[N'-(2-inethylpheriyl)ureidoI phenylacetate (317.0 mng, 0.660 mmol), methyl 3-chloro-2-[(2-pyrrolidirnyl)methoxyJ carboxylate (172.0 mg, 0.635 mmol), Et 3 N (105 ul, 0.756 mmnol) 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-[fl-[3methoxy-4-[N'-(2-methylphenyl)ureidoI carboxylate as a brownish oil. To a stirred solution of this compound in THE (6.0 mL) and H 2 0 (2.0 niL) was added LiOH (45.3 mng, 1.89 inmol), and the reaction mixture was stirred for 5 hr at room temp. The mixture was diluted with n-hexane and extracted with I N-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 mng 47 as an orange crystalline powder. mp 122 125; IR (KBr) 3354, 1709, 1593, 1535, 1454, 1257cm-'; 'H-NMR (400 MI~z, DMSO-d) 5 1.67 2.03 (in, 411), 2.50 3H), 3.33 3.42 (mn, 111), 3.52 (in, 2H), 3.58 J =4.4 Hz, IlH), 3.83 314), 4.27 -4.31 (in, 2H), 4.42 4.47 (mn, 111), 6.73 J WO 01/00206 WO 0100206PCT1LJSO0118079 7.8 Hz, IlH), 6.87 6.95 (in, 3H), 7.11 7.17 (in, 2H), 7.79 J 8.3 Hz, IlH), 7.99 J 8.3 Hz, 1H), 8.14 (dd, J 8.8Hz, 1H), 8.46 11), 8.56 lH), 8.69 J=2.0 Hz, IlH), 13.06 (br s, 1H); MS (FAB) ink 553 (Wv+1);AnaL. Calcd for C7JH 29 CIN,0 6 C, 60.81; H, 5.29; N, 10.31.
Found: C, 60.98; H, 5.50; N, 9.46.
Example 44 1-[4-[N'-(2-fluorophenyl)ureidoJ-3-methoxyphenylacetylJ-2-pyrrolidiniy imethoxyj carboxylic acid H& OOH 48 To a stirred solution of 6-hydroxynicotinic acid (2 g, 14.38 mmol) in benzene (32 mnL and MeOH (8 mL) was added dropwise TMSCHN 2 (1.97 niL, 3.953 mmol) at 0 0 C, 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 Et 2 O, and dried in vacuo to give 1.566 g (7 methyl 2as a pale brown crystalline powder. IR (KBr) 1655, 1645, 16 1433, 1300, 1113, 777, 642cm-'; 'H-NMR (400 MHz, DMSO-d) 8 3.77 3H), 6.37 J 9.8 Hz, I 7.99 (dd, J 2.4, 9.8 Hz, I 8.03 J 2.4 Hz, IH); MS (FAB) miz. 154 Anal Calcd for C 7
H
7 N0 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 mmiol), Ntert-butoxycarbonylprolinol (1.31 g, 6.529 mmol), and Ph 3 P (2.06 g, 7.836 nunol) in TIIF (10 mL) was added DIAD (1.54 niL, 7.836 nunol) 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; 1, v/v) as eluent to give 712.3 mig methyl 2-[[1-(tertbutoxycarbonyl)-2-pyrrolidinylJ methoxyJpyridine-5-carboxylate as a pale yellow oil. 'H-NNMR (400 M1Hz, CDCI,) 5 1.47 9H), 1.85 1.98 (in, 4H), 3.37 (br s, 2H), 3.92 3H), 4.12 4.33 (br mi, 214), 4.48 (brs, lH), 6.75 (in, 1H), 8.15 (mn, lH), 8.79 (in, IH); MS (FAB) m/~z 337 To a stirred solution of methyl 2-[[1-(tert-butoxycarbonyl)-2pyrrolidinyllmethyloxylpyridine -5-carboxylate (232.3mg, 0.691 nirol) in CH 2
CI
2 (4.6 niL) was added TFA (0.9 niL) at 0 0 C, 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 IN NaOH. The aqueous solution was extracted with CHCI3, washed with brine, and the dried WO 01/00206 WO 0100206PCT/USOO/18079 over Na 2
SO
4 The solvent was evaporated under a reduced pressure to afford 146.2 mng methyl 2-(2-pyrrolidinyl)methoxypyridine-5-carboxylate as an oil. 'H-NMvR (400 MHz, CDCI 3 1.49 1.58 (in, 1H), 1.72 2.18 (in, 311), 2.92 3.05 (in, 2H1), 3.50 -3.57 (in, 111), 3.91 311), 4.23 (dd, J= 8.0, 10.7 Hz, 111), 4.38 (dd, J 4.4, 10.3 Hz, 1H), 6.78 J 8.8 Hz, 111), 8.15 (dd, J= 2.4, 8.8 Hz, lH), 8.80 J= 2.4 Hz, lH), MS (FAB) m/z 237 The mixture of pentafluorophenyl 4-[N'-(2-fluorophenyl)ureido]-3-methoxy-phenylacetate (314.8 mg, 0.650 mmol), methyl 2-[(2-pyrrmlidinyl)methoxy]pyridine-5-carboxylate (146.2 mg, 0.6 19 mmol), Et 3 N (103 ul, 0.743 mmol) in DMF (1.5 mL) was stirred for 1 hr at room temp. The m-ixture was diluted with Et 2 O, washed with brine, and dried over NaSO 4 The solvent was removed under a reduced pressure to afford methyl 2-If 1-[4-[N'-(2-fluorophenyl)ureidoj-3methoxyphenylacetylJ-2-pyrrolidinylj methoxyjpyridine-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 ing, 1.857 inmol), 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 layer was acidified by IN HCl and extracted with CHCI 3 The extract was washed with brine, and dried over NaSO 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 48 as a white crystalline powder. IR (KBr) 1602, 1537, 1456, 1265, 752cin'; H-NMR (400 MHz, DMSO-d 6 1.67 -2.03 (in, 411), 2.50 3H), 3.33 3.42 (in, 111), 3.52 (in, 211), 3.58 J =4.4 Hz, lH), 3.83 4.27 4.31 (in, 2H), 4.42 4.47 (mn, 111), 6.73 J 7.8 Hz, 1H), 6.87 -6.95 (mn, 3H), 7.11 7.17 (in, 2H), 7.79 J =8.3 Hz, 1H), 7.99 J =8.3 Hz, 11H), 8.14 (dd,J 8.8 Hz, lH), 8.46 11H), 8.56 1I-1), 8.69 J 2.0 Hz, IH), 13.06 (br s, 111); MIS (FAB) m/z 523 (M'+1);AnaL. Calcd for C 27
H
27 FN04/2H2O: C, 61.01; H, 5.3 1; N, 10.54. Found: C, 61.52; H, 5.39; N, 10.01.
Example 4-[11 ehx-4['(-ehypey~ridlhnlctyj2proidnlehl--piperazinyl acetic acid jOOH To a stirred suspension of 1-benzylpiperazine (5 g, 28.4 nunol) and K 2 C0 3 (5.89 g, 42.6 inmol) in DMIF (30 mL) was added ethyl broinoacetate (4.74 g, 28.4 minol) at room temp. The WO 01/00206 WO 01/1)206PCT/USOO/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 aver 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 ethyl 4benzyl-1-piperazinylacetate as a yellow oil. 'H-NMR (CDCI 3 8 1.27 (3 H, t, J 7.3 Hz), 2.88- 2.96 (8 H, in), 3.20 (2 H, 3.52 (2 H, 4.18 (2 H, q, J 7.3 Hz), 7.22-7.32 (5 H, in).
A stirred solution of ethyl 4-benzyl-1-piperazinylacetate (2.00 g, 7.62 nunol) and Pd/C (2 g) in AcOH:EtOH 1, 40 mL) was hydrogenated at I 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 rnL). The combined extracts were dried over KC0 3 and evaporated to give 1. 16g ethyl I1-piperazinylacetate as a yellow oil. 'H-NMvR
(CDCI
3 5 1.26-1.30 (3 H, in), 1.67(1 H, br 2.55(4 H, mn), 2.92-2.96(4 H, in), 3.19-3.20(2 H, in), 4.16-4.22(2 H, in).
to a stirred solution of N-Boc-L-prolinol (1.00 g, 5.02 mmol) and ethyl 1-piperazinyl acetate (864 mg, 5.02 minol) in MeOH: AcOH (10: 1, v/v, 11 InL) was added NaBH 3 CN (664 mg, 10.0 inmol) at room temp. After being stirred overnight, the mixture was poured into ice water (100 inL) and made basic by the addition of NaHCO 3 The mixture was extracted with CHC1 3 (2 x 200 inL). The combined extracts were dried over Na 2
CO
3 and evaporated. The residue was chromatographed on silica-gel with CHCI 3 :EtOH (10: 1, v/v) as eluent to give 1.20 g ethyl 4- [(l-Qert-butoxy carbonyl)-2-pyrrolidinylmethylJ-1-piperazinylacetate as a colorless oil. 'H-NMR
(CDCI
3 8 1.27 (3 H, t, J 7.3 Hz), 1.46-1.47 (9 H, in), 1.79-3.96 (total 19 H, series of in), 4.19 (2 H, q, J= 7.3 Hz).
A mixture of ethyl 1-Qtert-butoxycarbonyl)-2-pyrrolidinylinethylJ-l -piperazinylacetate (1.20 g, 3.38 inmol) in TFA (5 inL) and CH 2
CI
2 (5 rnL) was stirred overnight. After removal of the solvent, the residue was made basic by the addition of sat. NaHCO 3 The mixture was extracted with CHC1 3 (2 x 200 mL). The combined extracts ware dried over Na 2
CO
3 and evaporated to give 386 mng ethyl 4-(2-pyrrolidinylmethyl)-1-piperazinylacetate as a yellow oil. MS (FAB) 256 To a stirred solution of ethyl 4-(2-pyrrolidinylmethyl)-l-piperaziniylacetate (380 mg, 1.49 minol) and 3-methoxy-4-[N'-(2-methylpheryl)ureido]phenylacetic acid (468 ing, 1.49 inmol) in DMF (10 iii) was added EDC-HCI (428 ing, 2.24 minol), HOBt, and DMAP After being stirred overnight, the mixture was diluted with EtOAc (300 inL), washed with brine (2 x 200 niL), WO 01/00206 WO 0100206PCT/USOO/18079 and dried over MgSO 4 After removal of the solvent, the residue was chromatographed on silicagel with CHCI 3 :EtOH 1, v/v) as eluent to give 257 mg (3 ethyl -methoxy-4-[N'-(2methyiphenyl) ureidoJ phenylacetylj-2-pyrrolidinylmethyl]-l-piperazinylacetate as a yellow foam.
'H-NMR (CDCI,) 8 1.24-1.29 (3 H, in), 1.69-4.24 (total 29 H, series of in), 6.41 (1 H, in), 6.81 (2 H, in), 7.13-7.26 (4 H, in), 7.52 (1 HK d, J 7.3 Hz), 8.04 (1 H, d, J 8.3 Hz).
To a stirred solution of ethyl 4-f 1-[3-methoxy-4-[N'-(2-methylphenyl)ureidoJ phenylacetyll-2-pyrrolidiniylmethylj-1-piperazinylacetate (250 m&g 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 CHCI,:MeOH 3 x 100 mL).
The combined extracts were dried over MgSO 4 and evaporated. The crude solid was recrystallized from CHCI 3 -MeOH-n-hexane to give 40 mg 49 as a colorless crystalline powder. mp 160- 170 'H-NMR (DMSO-d 6 8 1.74-4.08 (total 27 H, series of in), 6.73 (1 H, d, J 7.8 Hz), 6.87 (I H, 6.93 (1 H, t,J =7.8 Hz), 7.11-7.17 (2 H, in), 7.79 (1 H, d, J =7.8 Hz), 8.00 (1 H, dd, J 7.8, 2.4 Hz), 8.47 (1 H, 8.56 (1 H, MS (FAB) 524 Anal Calcd for
C
28
H
3 7
N
5 0 5 HC1-H 2 0: C, 58.17; H, 6.97; N, 12.11. Found: C, 58.26; H, 7.26; N, 11.53.
Example 46 H H To a suspension of 4-aminophenylacetic acid (10 g, 66 minol) in 1: 1 CH 2
CI
2 :acetone (100 niL) was added o-tolyisocyanate (8.8 g, 66 minol). 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
CI
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 Example 47 H H 0 OH To a suspension of 2-ainino-4-thiazoleacetic acid (4 g, 25 inmol) in 1: 1 CH 2 CI:acetone (100 niL) was added o-tolylisocyanate (3.5 g, 26 inmol). 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
CI
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.
WO 01/00206 PCT/US00/18079 Example 48 4-[[1-[3-methoxy-4 2methylphenyl)ureidophenylacetyl-2-pyrrolidinylmethylamino]benzoic acid .52 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 mL) 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 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 H 2 0, 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 v/v) as eluent to afford 2.17g 4-[l-(tert-butoxycarbonyl)-2pyrrolidinylmethylamino] benzoate as a pale yellow oil. 'H-NMR (400 MHz, CDCI 3 5 1.48 (s, 9H), 1.51 2.09 4H), 3.05 3.07 and 3.43 3.48 (brm, IH), 3.18 (br s, 1H), 3.36 (br s, 2H), 3.84 1H),4.06 4.08, 4.20 4.24 (br m each, 1H), 6.49 6.65 2H), 7.84 J= 8.3 Hz, 2H); MS (FAB) m/z 335 To a stirred solution of methyl 4-[1-(tert-butoxycarbonyl)-2-pyrrolidinylmethylamino] benzoate (2.17 g, 6.490 mmol) in CH 2 CIl (44 mL) was added TFA (8.7 mL) at 0°C, and the 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
CI
2 The extract was washed with brine, dried over NaSO 4 and the solvent was concentrated under a reduced pressure to afford 1.34 g methyl 4-(2-pyrrolidinylmethylamino)benzoate as a brown oil, which is used to the subsequent reaction without further purification.
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 (HCI) (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 was chromatographed on silica-gel with CHCI 3 MeOH (50:1, v/v) as eluent to afford 882mg WO 01/00206 WO 0100206PCT/USOOI 18079 methyl 4-11 -methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetylJ-2-pyrrolidiny methylaininojbenzoate as a brown amorphous solid, which is used to the subsequent reaction without further purification.
To a stirred solution of the above methyl 4-[l-[3-methoxy-4-[N'-(2-methylphenyl)ureido phenylacetylJ-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 extractedwith CH 2
CI
2 The extract was washed with brine, dried over NaSO, 4 and evaporated in vacuo. The solid was recrystallized from n-hexane diisopropyl ether CHC1 3 MeOH to afford 180.5 mg (21o) 52as apale yellow amorphous solid. IR (KBr) 1604, 1535, 1511, 1454, 1255, 1224, 1174cm-'; 'H-NMvR (400 MEHz, DMSO-dA) 5 1.79 1.99 (br ma, 4H), 2.25 3H), 2.90 -2.94 (in, 1H), 3.35 3.62 (in, 6H), 3.87 3H), 4.12 4.15 (br s, 1H), 6.63 6.78 (in, 4H), 6.89 6.95 and 7.11 7.17 (m each, 314), 7.65 J= 8.3 Hz, 2H), 7.80 J= 8.3 Hz, I 8.02 J= 8.3 Hz, 1H), 8.47 1H), 8.57 lH), 12.0 (br s, 1H); MS (FAR) m/z 517 Anal Calcd for C2,H 3 2
N
4
O
5 d H 2 0: C, 65.15; H, 6.4 1; N, 10.48. Found: C, 65.45; H, 6.33; N, 10.02.
Example 49 1-[3-methoxy-4-[N'-(2-methylphenyl)ureidophenylacetyl] -2-pyrrolidinylmethyl]-Nmethylamninolbenzoic acid H H Ot~e COH53 To a mixture of methyl 4-[N-(2-pyrrolidinyl)methylainino]benzoate (600 mg, 1.794 mmol), 37%-formaldehyde (1.79 inL, 23.32 inmol), 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 NaSO,.
The solvent was removed under a reduced pressure, and the residue was chromatographed on silica-gel with n-hexane EtOAc 1, v/v) as eluent to afford 645 mg methyl 4-IN-[I- (iert-butoxycarbonyl)-2-pyrrolidinylnethyl-N-methylaminolbenzoate as a colorless oil. 'H-NMIR (400 MHz, CDC1 3 5 1.50 9H), 1.76 1.91 (in, 4H), 3.07 3H), 3.15 3.43 (in, 3H), 3.67 3.71 (mn, IH), 3.85 3H), 4.11 4.17 (mn, 1H), 4.37 lIi), 6.75 J 8.3 Hz, 2H), 7.89 J= 8.8 Hz, 2H); MS (FAR) m/~z 349 (W 4 WO 01/00206 WO 0100206PCT/1JSOO/18079 To a stirred solution of methyl 4.{N-[1-(terl-butoxycarbonyl)-2-pyrrolidinylmethyl]-Nmethylarninolbenzoate (645 mg, 1.794 mmol) in CH 2 C1 (6.5 mL) was added TFA (1.3 mL) at 0 0
C,
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
CH
2 CI,. 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 of methyl 4-[N..(2-pyrrolidinyl)methyl.Nmethyllaminobenzoate as a yellowish oil, which is used to the subsequent reaction without further purification.
The mixture of methyl 4-[N-(2-pyrrolidiniyl)methyl-N-methyl~aminobenzoate (191.8 mg, 0.772 mmol), 3-methoxy-4-(N'-2-metthylphenylureido)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 niL) was stirred for 3 hr at room temp. The reaction mixture was diluted with EtO, washed with brine, and dried over NaSO 4 The solvent was removed under a reduced pressure to afford 482.5 mg methyl 4-[N-[1-[3-methoxy-4-[N'-(2-methylphenyl)ureidoI phenylacetylJ-2-pyrrolidinylmiethylj-N-methylantino~bnzoate as a white amorphous powder, which is used to the subsequent reaction without further purification.
To a stirred solution of methyl 4-[N-[l-[3-miethoxy-4-[N'"-(2-methylpheryl)ureidolphenyl acetylj-2-pyrrolidinylmethyl-N-methylaminoJbenzoate in THF (5.0 mL) was added IN NaOH (6.2 niL, 6.2 nunol), and the mixture was heated under reflux for 3 days. The reaction mixture was concentrated in vacuo. The residue was neutralized with IN HCl, and extracted with CH 2 C1 2 The extract was washed with sat. NH 4 CI, brine, dried over Na 2
SO
4 and evaporated in vacuo. The crude solid was recrystallized from n-hexane-CHCI,-MeOH-isopropylether to afford 102.8 mg 2steps) 53 as a pale yellow amorphous solid. mp 144 146; IR (KBr) 3325, 1600, 1529, 1454, 1284, 1257, 1184cm'; '11-NMvR (400 MI-Iz, DMSO-d) 5 1.73 1.91 (in, 3H), 2.03 2.11 (in, IM1, 3.03 311), 3.16 (dd, J= 9.3, 14.2 Hz, 3.37 -3.60 (in, 411), 3.76 -3.80 (mn, 11-1), 3.86 4.25 (br s, 111), 6.75 (dd, J= 1.5, 8.3 Hz, 111), 6.86 J 1.5 Hz, 111), 6.90 J =8.8 Hz, 21M1, 6.95 -7.01 (mn, 11-1), 7.12 J= 7.8 Hz, 114), 7.20 -7.25 (in, 111), 7.73 (d,J 8.8 Hz, 211), 8.01 J 7.8 Hz, 111), 8.16 8.20 (in, 111, 8.73 1H), 9.19 J= 2.0 Hz, 1IM, 12.0 (br s, 111); MS (FAB) m/z 535 Anal. Calcd for C,,H,,FN 4
O
5 -1f2H 2 O: C, 64.08; H, 5.93; N, 10.3 1; F, 3.49. Found: C, 64.17, H, 5.84; N, 10.06; F, 3.26.
Example 1 -[-4-[N'-(2-fluorophenyl)ureido]-3-methoxyphentylacetylj-2-pyrrolidinylmethyl-N- WO 01/00206 WO 0100206PCT/USOO/18079 methylaniinoj-3-nitrobenzoic acid F Ce o 2 Na cooH 54 To a mixture of methyl 4-fluoro-3-nitrobenzoate (1.58 g, 4.666 mmnol) and [1-(tertbutoxycarbonyl)-2-pyrrolidinyljmethylamine (500 mg, 2.333 nol) in DMF (8.0 mL) was added
K
2 C0 3 (967 mg, 6.999 nunol), 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,. The solvent was removed under a reduced pressure, and the residue was chromatographed on silica-gel with nhexane EtOAc 1, v/v) as eluent to afford 834.9 mg (9 1%O) of methyl 4-[N-[lI-QIertbutoxycarbonyl)-2-pyrrolidinylmethyl]-N-methyllamino-3-nitrobenzoate as a pale yellow oil, which is used to the subsequent reaction without further purification.
To a ice-cooling solution of the above oil in CHC1 2 (8.3 ruL) 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 lN-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 553.6 mg methyl 4.IN-(2-pyrrolidinylmethyl)-N-methyljamino-3-nitrobenzoate as a pale yellow oil. 'H-NNvIR (400 MOHz, CDCI,) 5 1.31 1.40 (in, 111), 1.74 2.05 (in, 4H), 2.73 2.79 (in, I1H), 2.81 2.99 (mn, 11H), 2.94 3H), 3.29 -3.55 (mn, 2H), 3.89 7.14 J 9.3 Hz, IlH), 7.98 (dd, J 2.0, 8.8 Hz, 1H), 8.42 J 2.0 Hz, INH); MS (FAR) m/~z 294 (M 4 1).
A mixture of 3-inethoxy-4-[N'-(2-fluorophenyl)ureidoJphenylacetic acid (630.0 mng, 1.979 minol), methyl 3-nitr-o-4-[N-(2-pyrrolidinyl)inethyl-N-inethylaniino~benzoate (553.0 mg, 1.885 inmol), EDC(Hydrochloride) (542.0 ing, 2.827 inmol), HO~t (25.5 mg, 0. 189 nunol), and DMAP (23.1 mg, 0..189 inmol) in DMF (5.0 mL) was stirred at room temp. for 2 hr. The mixture was diluted with Et 2 O, 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 CHC1 3 MeOH (30: 1, v/v) as eluent to afford 1. 18 g (100%) methyl 4-[N-[1-[3-inethoxy-4-[N'-(2-fluorophenyl)ureidoJ phenylacetylJ-2-pyrrolidinyl incthylj-N-inthylainiinoJ-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-IN-[ 1-[3-methoxy-4-[N'-(2-fluorophcnyl) ureidoJphenylacetylJ-2-pyrrolidinylmethylJ-N-methylamino-3 -nitrobenzoate (2.50 mng, 0.421 WO 01/00206 WO 0100206PCT/USOO/18079 mmol) in THE (3.0 mL) was added IN NaOH (1.5 mL, 1.500 mmol), and the mixture was heated under refiux overnight. After coaling, the midxture was concentrated to a small volume. The residue was treated with IN HCI, and extracted with CHCI 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 diethyl ether CHC1 3 MeOH to afford 194.9 mg of 54 as a yellow amorphous solid. IR (KBr) 1685, 1610, 1529, 1454, 1284, 1259, 1228 'H-NMvR (400 MHz, DMSO-d) 5 1.63 1.91 (in, 3H), 2.04 2.07 (br s, lH), 2.60 (hr s, 1H), 2.80 lH), 2.99 2H), 3.05 3. 10 (in, 1H), 3.32 3.58 (in, 3H), 3.76 3.81 (in, 1H), 3.81 3H), 4.25 (hr s, 1H), 6.68 J 3.9 Hz, 1H), 6.71 J= 8.8 Hz, I1H), 6.81 6.96 (in, I1H), 7.07 J= 7.3 Hz, I1H), 7.17 (dd, J 9.8 Hz, IH), 7.48 J 7.8 Hz, 1H), 7.85 J 8.8 Hz, 1H), 7.97 J 8.8 Hz, 1H), 8.11 8.20 (in, 2H), 8.68 1H), 9.14 IH), 12.8 (hr s, IH); MS (FAR) m/z 580 Anal. Calcd for C2,H 3
,FN
5
O
7 l/4H 2 0: C, 59.63; H, 5.26; N, 11.99; F, 3.25.Found:C,59.68;H,5.34;N, 11.80; F, 3.2 1.
Example 51 3-amino-4-[N-methyl-[ 1-[4-[N'-(2-fluorophenyl)ureido]-3 -iethoxy-phenylacetylJ-2-pyrrolidinyl methyl] -N-inethylaniinolbenzoic acid A stirred solution of methyl 1-[4-IN'-(2-fluorophenyl)ureidol-3 -iethoxyphenylacetyl]-2-pyrrolidinylmethylJ-N-methylaminoJ-3-nitrobenzoate (901.0 mg, 1.518 nunol) in MeOH (18.0 mL) was hydrogenated over 5%Pd-C (1.35 g) at 45psi overnight. The mixture was 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,, and the solvent was removed under a reduced pressure. The residue was chromatographed on silica-gel with CHC1 3 MeOH (24: 1, v/v) as eluent to afford 283.7 mg methyl 3-anuno-4-[N-[l1-[4- [N'-(2-fluorophenyl)ureido] -3-methoxy-phenylacetylJ-2-pyrrolidinylmethylJ-N-methylamidnoJ 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 THE (3.0 mL) was added IN NaOH solution (1.5 mL, 1. 500 mnxol), and the mixture was refluxed overnight. The mixture was concentrated, treated with IN HCI, and extracted with CHC 3 The extract was washed with brine, dried over Na 2
SO
4 and evaporated in vacuo. The solid was recrystallized from n-hexane diethyl ether CHC1 3 MeOH to afford 179.8 mg 55 as a white amorphous solid. IR (KBr) 1614, WO 01/00206 WO 0100206PCT/USOO/18079 1601, 15 37, 14 54, 1228, 1219, 1184 'H-NMR (400 MHz, DMSO-Q~ 5 1.60 2.20 (in, 4H), 2.61 2.68 (in, 1H), 2.89 3H), 3.13 3.18 (in, IN), 3.40 3.61 (in, 4H), 3.85 3H), 4.01 (br m, IN), 4.93 (br s, 2H), 6.50 7.31 8N), 8.01 (dd, J 2.9, 8.3 Hz, INH), 8.18 J 8.3 Hz, 11N), 8.71 1H), 9.17(d, J= 1.5 Hz, IN), 12.3 (br s, 1N); MS (FAB) nVz 550(M'+1); Ana. Calcd for C 2 9
H
3 FN0 3 1l/4H 2 0: C, 62.86; H, 5.9 1; N, 12.64; F, 3.43. Found: C, 62.7 1; H, 6.00; N, 12.3 9; F, 3.16.
Example 52 1-[4-[A'-(2-methylpheniyl)ureido]phenylacetylJ-2-pyrrolidinyl]methylamino]benzoic acid e H H56 To a stirred mixture of methyl 4-[(2-pyrrolidinyl)methylamino]benzoate (220 mg, 0.94 nunol), 4- -methylphenyl)ureido~pheniylacetic acid (285 mg, 0. 94 inmol), 4-DMAP (140 mg, 1. 13 mmol) and catalytic amount of HOBT in DMF (7 ml) was added EDC-HCI (220 mg, 1. 13 minol) at room temperature. The resulting mixture was stirred at room temperature for 20 hr. The mixture was pored into iee-water. The solid was collected, washed 'with water and air-dried. The crude solid was purified by silica-gel (20 ml) column chromatography with CHCI,- EtOAc 1, v/v) to CNCI 3 EtON 1, v/v) as eluent to give methyl 4-(l -[4-[M-(2-methylphenyl)ureidoJ phenyl acetylJ-2-pyrrolidinyljmethylaminolbenzoate (400 ing, 85%) as a gum. 'H-NMR (CDCI 3 )8 1.75- 2.05 (series of m, 4 2.24 3 3.18 and 3.27 (each m, each 1 3.51 (in, 2 3.60 2 3.83 3 4.52 (in, 1 6.52 (in, 311), 6.81 1 7.11-7.25 (series of m, 7 7.53 (in, 1 7.81 J= 8.8 Hz, 2N).
A mixture of methyl 1-[4-[N'-(2-methylphernyl)ureido]phernylacetylJ-2-pyrrolidinylJ methylaminolbenzoate (280 mng, 0.56 mmol) in TI-F (3 ml) and 0.25N NaON (6.8 ml, 1.75 inmol) was stirred for 3 hr at 60-70 After cooling, the mixture was poured into ice-IN NCI (3 ml).
The solid was collected, washed with water and air-dried. The crude crystalline material was recrystallized from CHCl 3 -EtOH-IPE to give 56 (180 mg, 66%) as fine needles. MW 486.56 IR (KBr) n 3367, 3294, 1712, 1606, 1539 'H-NMR (CDC 3 -DMSO-d6) 8 1.80-2.05 (series of m, 4 2.26 3 2.94(m, 1 3.38 and 3.56 (series of in, 3 3.57 2 4.23 (mn,2 N), 6.48 (br s, 1 6.69 J =8.8 Hz, 2N), 6.91 J =7 Hz, 1 6.91 (in, 4 7.39 J =8.3 Hz, 2N), 7.66 J 8.8 Hz, 2N), 7.80 (in, 2 8.88 1 11.76 1 MS(FAB) m/lz 487 Anal. Calcd for CHN 4
O
4 -0.75xH 2 O: C, 67.24; HK 6.45; N, 11.20. Found: C, 67.13; N, 6.3 2; N, 11.0 1.
WO 01/00206 WO 01/11206PCT/USOOI 18079 Example 53 methyl 4-[l 44-[N'-(2-chlorophenyl)ureido]-3-methoxylphenylacetyl]-(2S)-pyrrolidinyl] methoxybenzoate 441 -[4-[N'-(2-chlorophenyl)ureido]-3-methxylphenylacetylJ-(2S)-pyrrolidinyllmethoxybenzoic acid I H 6Me58 To a stirred solution o2Syrlidinemetbanol (15.1 g, 149.5 mmol) in dioxane (100 ml) was added a solution of (Boc),0 (32.6 g, 164.4 mmol) in dioxane (100 ml). The reaction mixture was 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 as eluent to give (1-tertbutoxycarbonyl-(2S)-pyrrolidinyl)methanoI (31.6 g, quant.) as a colorless oil. 'H-NMR (CDCI 3 8 1.47 9H), 1.60-2.00 (in, 3.25-3.70 (4H, in), 3.92-4.00 (in, 111).
To a stirred solution of (1-tert-butoxycarbonyl-(2S)-pyrrolidinyl)methano (4-02 g, 20.0 mmol), methyl 4-hydroxybenzoate (3.04 g, 20.0 nunol) and Ph 3 P (6.28 g, 24 .0 mmol) in THF ml) was added DIAl) (4.85 g, 24.0 mmol) at 0 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, v/v) as eluent to give methyl 4- (1 -ert-butoxycarbonyl-(2S)-pyrrolidinyl)methoxybenzoate (5.4 g, 81%) as a pale yellow oil. 'H- NMvR (CDCl 3 8 1.47 9H1), 1.88-2.04 4H1), 3.41 (in, 211), 3.91 3H1), 3.90-3.92 (mn, 1H1), 4.11-4.16 (in, 211), 6.94 J 8.6 Hz, 7.94 J =8.3 Hz, 2H1).
To a stirred solution of methyl 1-tert-butoxycarbonyl-2S-pyrrolidinyl)inethoxybenzoate (2.1 g, 6.27 inmol) in CH 2
CI
2 (9.0 ml) was added TEA (6.0 ml) at 0 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 CHCl 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 ing, 2.0 inmol), 4-IN'-(2chlorophenyl) uredio]-3-methoxyphenylacetic acid (669 mng, 2.0 nunol), HOBt (405 ing, nunol), and triethylamine (554 ml, 4.0 mmol) in THF (10.0 ml) and MeCN (10.0 nil) was added WO 01/00206 WO 0100206PCT/USOO/18079 EDC-HCI (576 mg, 3.0 nunol) at 0 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 v/v) as eluent to give 57 (900 mg, as a colorless oil. MW 552.02 'H-NMvR (CDC1 3 8 2.04-2. 10 (in, 4H), 3.51-3.70 (in, 611), 3.87 3H1), 4.11-4.18 (in, 6.77-6.88 (in, 4M1, 7.23-7.34 (in, 4H1), 7.91-7.96 (in, 2M1, 8. 17-8.19 (in, 114).
The mixture was stirred at 70 'C for 24 hr. The mixture was concentrated in vacua, water was added thereto, and neutralized with IN HCI. The resulting solid was collected, washed with water, and dried in vacua to 58 (640 mg, as a white crystalline solid. MW 537.99 mp 126-130 IR (KBr) 3324, 2938, 2877, 1604, 1533, 1249, 1166, 750 1 H-NMR (DMSO-dA 8 1.93- 2.05 (in, 4H), 3.52-3.61 (in, 5H), 3.82 3H), 3.99-4.01 (in, 2H), 4.18-4.20 (in, lH), 4.29 (in, 11-1), 6.74-6.76 lH, J= 8.3 Hz), 6.87 111), 6.99-7.04 (mn, 3H), 7.25-7.29 (in, 111), 7.41-7.43 (d, 1H, J= 8.1 Hz), 7.86-7.91 (in, 211), 7.95-7.97 (in, 111), 8.09-8.11 1HJ= 8.3 Hz), 8.87-8.92 IH); MS (FAB) m/z 538 Anal calcd for C~gHnN 3
O
6 -0.5H 2 O: C, 61.48; HK 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 A nal Calcd for
C
28
H
2 7
N
3
O
6 -Na-1.5HO: C, 57.29; H, 5.15; N, 7.16. Found: C, 57.48; H, 5.04; N, 6.99.
Example 54 I-[4-[N'-(2-bromophenyl)ureidoj-3-methoxylphenylacetylj-(2S)-pyrrolidiniyljmethoxybenzoic acid Br H H "e59 To a stirred solution of methyl 4-(2S-pyrrolidinyl)inethoxybenzoate (470 mg, 2.0 inmol), broinophenyl)uredio]-3-inethoxyphenylacetic acid (758 ing, 2.0 numol), HOBt (405 mg. 3.0 minol), and triethylamine (554 m.17 3.0 inmol) in THF (10.0 ml) and MCCN (10.0 ml) was added EDC-HCl (576 mg, 3.0 mmol) at 0 The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacua. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 then dried over NaSQ, and concentrated in vacua. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc v/v) as eluent to give methyl 4-[I-[4-[N'-(2-bromophenyl)ureidoj-3-nethoxylphenylacetylJ -(2S)-pyrrolidinylj methoxybenzoate (1.0 g, 84%) as a colorless oil. 'H-NMR (CDCI 3 82.04-2.10 (in, 3.52-3.54 (mn, 111), 3.62 211), 3.70 3H1), 3.88 311), 4.13-4.19 (in, 211), 6.79-6.94 (in, 411), 7.20-7.31 WO 01/00206 WO 0100206PCT/LJSOO/18079 (in, 111), 7.91-8. 12 (in, 2H), 8.13-8.15 (in, H).
To a stirred solution of methyl 4-[I1-[4-[N'-(2-bromophenyl)ureido]-3-methoxylphenylacetyI]-(2S)pyrrolidinyljmethoxybenzoate (780 ing, 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 *C 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 vacua to give 59 (730 ing, 96%) as a white crystalline solid. NM 582.44 mp 120-125 JR (KBr) 3318, 2938, 1604, 1529, 1166, 1025 cm- I ;'H-NN4R (DMSO-d)8 1.92-1.96 (in, 4H), 3.52-3.60 (mn, 5H1), 3.82 311), 3.98-4.02 (m,lI H), 4.16-4.19 (in, 111), 4.29 (in, 111), 6.75 J 8.3 Hz, 111), 6.87 (mn,1H), 6.94-7.04 (in, 311), 7.29- 7.33 (in, 111), 7.57-7.59 (in, 111), 7.85-7.96 (in, 411), 8.72 111), 8.91 111); MS (FAB) m/lz 582 Anal. calcd for C28H~N 3
O
6 Br- L.H 2 O: C, 56.0 1; H, 5.04; N, 7.00; Br, 13.3 1. Found: C, 56.12; H, 4.98; N, 6.96;, Br, 13.57..
Example 3-amino-4-[ 1-[4-[AN'-(2-hydroxyphenyl)ureido]-3-inethoxyphenylacetamidol-2-pyrrolidiny methoxy]benzoic acid H HOMe
XNH
O)OH To a stirred solution of 2-nitrophenol (10.0 g, 72.0 inmol) and K 2 C0 3 (9.96 g, 72.0 nunol) in DMF (150 mL) was added dropwise benzyl bromide (9.40 inL, 79.2 nunol) at 0 After being stirred at room temperature for 3 hr, the reaction mixture was diluted with water, which was extracted with EtO. The extracts were washed with brine, dried over Na 2
SO
4 and concentrated to dryness.
Chromatography of the residue with hexane EtOAc 1, v/v) as eluent gave 2-benzyloxy nitrobenzene (14.7 g, as a yellow oil. 'H-NMvR (CDCl 3 )585.24 (s,211), 7.04 J =7.8 Hz, 111), 7.12 7.8 Hz,111, 7.31 -7.50(in, SH), 7.51 1.5 Hz, 11), 7.86 (dd,J=7.8, Hz, 111).
To a stirred solution of 2-benzyloxynitrobenzene (9.92 g, 43.3 iniol) and NiC1 2 (20.3 g, lS7inioI) in MeOH (350 rnL) was added portionwise NaBH 4 (8.09 g, 214 minol) at 0 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 WO 01/00206 WO 0100206PCT/USOO/18079 dryness. Chromatography of the residue with CHCI 3 as eluent gave 2-benzyloxy aniline (8.60 g, 100 as a reddish oil. 'H-NMvR (CDCI 3 8 3.71 (broad s 5.06 2H), 6.68 6.86 (in, 4H), 7.3 2 7.44 (mn, 5H); FAIB-MS m/z 200 1).
To a solution of 2-benzyloxyaniline 15 g, 5.77 mniol) in benzene (60 mL) was added triphosgene (1.27 g, 6.35 mmol) and Et 3 N (2.60 mL, 17.3 mmol) at 0 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 1, v/v) as eluent to afford tert-butyl 4-[N'-(2-benzyloxyphenyl)ureidoJ-3-methoxy phenylacetate (2.38 g, 89 as a yellow oil. 'H-NMR (CDCI 3 5 1.44 3.44 2H), 3.78 3H), 5.07 28), 6.73 (dd, J 8.0, 1.7 Hz, lH), 6.78 J =1.7 Hz, lH), 6.90 6.98 (in, 3H), 7.07 18), 7.29 lH), 7.33- 7.38 (in, 5H), 7.91 J 8.0 Hz, 111), 8.14 (in, 18).
To a solution of tert-butyl 4-[N'-(2-benzyloxyphenyl)ureido]-3-methoxyphenylacetate (2.35 g, 5.08 minol) in CH 2
CI
2 (25 mL) was added TFA (25 mL) at 0 After being stirred at room temperature. for 3 hr, the mixture was concentrated. The residue was dissolved in IN NaOH and washed with Et 2 O. The basic water layer was poured into ice 1IN HCI and the resulting mixture was extracted with CHCl 3 MeOH (4 vlv). 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)ureidoJ-3-methoxyphenylacetic acid (1.59 g, 77 as a brownish solid.
'H-NMR (DMSO-d 6 8 3.50 2H), 3.85 38), 5.26 28), 6.76 J 8.3 Hz, 18), 6.83 6.89 (in, 281), 6.91 18), 7.01 (dd, J= 8.3, 2.3 Hz, 18), 7.31 J= 7.3 Hz, 18), 7.39 J =7.3 Hz, 28), 7.49 J 7.3 Hz, 2H), 7.97 J =8.3 Hz, IH), 8.04 J =8.3 Hz, 18), 8.80 18), 8.86 184), 12.24 (broad s, 18); FAB-MS m.'z 407 To a solution of 4-[N'{(2-benzyloxyphenyl)ureido]-3-methoxyphenylacetic acid 12 g, 2.76 mmnol), methyl 4-(2-pyrrolidinylmethoxy)-3-nitrobenzoate (890 mng, 2.76 nunol), HOBt (74.0 mg, 0.55 mmol), DMAP (67.0 mg, 0.55 nunol), and Et 3 N (0.58 mL, 4.13 minol) in THF (15 mL) was added EDC-HCl (792 mng, 4.13 nunol). 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,, and concentrated to dryness. Chromatography of the residue with EtOAc as eluent gave methyl 4-[1-[4-[N'-(2-benzyloxyphenyl)ureidoj-3-methoxyphenylacetamido J-2-pyrrolidinylinethoxyJ-3-nitrobenzoate (1.52 g, 82 as a yellow amorphous solid. 'H-NMR WO 01/00206 WO 0100206PCT/USOO/18079
(CDCI
3 8 1.91 (in, 1H), 1.95 2.17 (in, 3H), 3.47 -3.53 (in, 2H), 3.56 2H), 3.60 (in, IH), 3.68 3H), 3.90 3H), 4.11 J =7.3 Hz, 111), 4.45 IH), 5.08 2H), 6. 70 (dd, J 1.9 Hz, I1H), 6.75 J= 1. 9 Hz, I1H), 6.91 6.99 (in, 3H1), 7.16 I 7.18 J= 8.3 Hz, I1H), 7.3 3 7.40 (in, 6H), 7.91 J 8.3 Hz, I1H), 8.11 8.16 (in, 2H), 8.46 I1H).
A solution of methyl 1-[4-[N'-(2-benzyloxyphenyl)ureidol-3-methoxyphenylactanido-2pyrrolidinylinethoxy]-3-nitrobenzoate (1.52 g, 2.27 inmol) in MeOH (20 mL) and THF (5 rnL) 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 3ainino-4-[ 1 .4-[N'-(2-hydroxyphenyl)ureido]-3-inethoxyphenylacetamido-2-pyrrolidinylmethoxy benzoate (1.12 g, 90 as a brownish amorphous solid. 'H-NMR (CDCI,) 8 1.97 2. 10 (in, 4H), 3.44 3H), 3.52 3.63 (in, 214), 3.85 3H), 4.10 -4.18 (in, 4.53 (in, 1IH), 6.65 -6.67 (in, 4H), 6.93 7.02 (in, 3H), 7.33 J 2.2 Hz, 7.36 (dd, J 8.3, 2.2 Hz, 7.60 7.69 J= 8.3 Hz, 8.08 1H), 9.47 (broad s, FAB-MS m/z 549 To a solution of methyl 3-axnino-4-(1-[4-[N'-(2-hydroxyphenyl)ureido]-3-inethoxyphenyI acetamiddo]-2-pyrrolidinylinethoxy]benzoate (1.12 g, 2.04 minol) in THF MeOH 1, v/v; mL) was added IN NaOH (4.20 mL, 4.20 nunol). After being stirred at room temperature for 24 hr, the reaction mixture was concentrated. The residue was diluted with water and neutralized with IN HCI at 0 The mixture was extracted with CHC 3 MeOH (4 vfv) ,which was washed with brine, dried over Na 2 SO,, and concentrated to dryness. Chromatography of the residue with CHC1 3 MeOH (5 1, v/v) as eluent gave 60 (352 mng, 32 as a pale yellow amorphous solid. MW 534.56 IR (KBr) 3282, 3062, 3025, 2952, 2865, 1629, 1546, 1509, 1454, 1419 'H-NMR (DMSO-d 6 8 1.87 -2.04 (in, 4H), 3.48 3.57 (in, 3.60 3.79 (s, 3.94 (dd, J 7.6 Hz, 4.12 (dd, J 3.9 Hz), 4.35 (in, 1$H, 4.87 (broad s, 6.70 -6.91 (in, 7.16 (dd, 1H, J 8.3, 2.0 Hz, 7.26 J 2.0 Hz, 7.96 J 8.3 Hz, 1$1, 7.97 J 8.3 Hz, 8.80 8.82 FAR-MS m/~z 535 Anal.
Calcd for C 28 H30NIO 7 -4.5H 2 C, 55-63; HL 6.39; N, 9. 10. Found: C, 55.08; HK 5.06; N, 8.69.
Example 56 1-[4-[AN'-(2-chlorophenyl)ureidoJ-3-nethoxyphenylacetylj-2-pyrolidinylinethylamino pyridine-2-carboxylic acid WO 01/00206 WO 0100206PCT/USOO/18079 1?i HMe H 6 5-1111-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetylj-2-pyrrolidinyllmethylaminoI pyridine-2-carboxylic acid To a stirred solution of 5-(methoxycarbonyl)pyridine-2-carboxylic acid (2.5 g, 13.8 nunol) and 4- DMAP (340 mg, 2.8 mmol) in tert-BuOH (15 ml) was added Boc 2 0 (6 g, 27.6 nunol) 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 NaSO,, and evaporated. The residue was chromatographed on silica gel (5fmI) with CH 2
C
2 as eluent to give methyl 6-tert-butoxycarbonylnicotinate (2.92 g, as needles. IR (K.Br) 2729, 1736, 1720, 1590, 1570 MS (FAB) nu/z 238 Anal. Calcd for C 1 2
H
15 N0 4 C, 60.75; H, 6.37; N, 5.90. Found: C, 60.72; HK 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 into ice-IN HCl (10 ml). The solid was collected, washed with water and air-dried. The crude solid was recrystallized from CHCI 3 -EtOH-IPE to afford 6-tert-butoxycarbonylnicotinic acid (850 mg, 76%) as needles. IR (KBr) n3095, 1728, 1705 cm'; 'H-NMR (DMIAO-d) 8 1.63 9 H), 8.09 (in, I 8.17 (in, 1 8.42 (dt, J 2-4 and 8.3 Hz, 1 9.21 J 2.4 and 8.8H1z, 1 H); MS (FAB) In/z 224 Anal. Calcd for C2J1 33
N
3 C, 36.18; H,3. 18; N, 3.84. Found: C, 36.85; HK 3.35; N, 3.79.
To a stirred mixture of 6-tert-butoxycarbonylnicotinic acid (1.9 g, 8.51 mmol) and triethylamine 17 g, 11.49 mmol) in tert-BuOH (30 and toluene (30 ml) was added a solution of diphenyl phosphoryl azide (2.93 g, 10.64 nunol) in toluene (3 ml) at room temperature. The resulting mixture was then heated at reflux for 5 hr. After cooling, ice and IN HCI (5n-d) was added to the mixture and extracted with toluene. The extracts were washed with brine, dried over Na 2 SO,, and evaporated. The residue was chromatographed on silica gel (50ml) with toluene- EtOAc 1, v/v) as eluent to give tert-butyl 5-tert-butoxycarbonyamino-2-pyridinecarboxylate (1.9 g, as a gum. 'H-NMR (CDCI 3 8 1.53 9 1.63 9 6.82 (br s, 1 8.01 J= 8.8 Hz, 1 8.17 (in, 1 8.46 J 2.4 Hz, 1 M).
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 CHCIl (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 *C for 10 min. The resulting stirred mixture was then heated at reflux for 10 hr. After cooling, N,-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
CI
2 The extracts were washed with brine, dried over Na 2
SO
4 and evaporated. The residue was chromatographed on silica gel (30ml) with CHCI 3 -EtOH (98:2, v/v) as eluent to give ethyl amino-2-pyridine carboxylate (700 mg, 65%) as a crystalline material. IR (KBr) n 3423, 3190, 1708, 1657, 15873338, 3296, 1691, 1641 cm'; 'H-NMR (CDCI,)8 1.42 J= 7.0 Hz, 3 4.11 (br s, 1 4.43(q, J 7.0 Hz, 2 6.99 (dd, J 2.7 and 8.5 Hz, 1 7.95 J 8.5 Hz, 1 H), 8.16 J 2.7 Hz, 1 MS (FAB) m/z 167 Anal. Calcd for CH,2CIN40 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 1tert-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 v/v, To the stirred solution was added NaBH 3 CN (500 mg, 7.90 mmol) at 0-5 The resulting mixture was stirred for a further 12 hr at room temperature. The mixture was piured into ice-sat NaHCO 3 (50 ml), and extracted with CH 2
CI
2 The extracts were washed with brine, dried over Na 2 SO,, and evaporated. The residue was chromatographed on silica gel (50ml) with CHCI 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 (CDC 3 8 1.38 9 1.42 6 H), 3.93 3 4.29 2 4.67 (br s, 1 7.15 J= 8.8 Hz, 1 8.18 (dd, J= 1.7 and 8.8Hz, 1 8.52 J= 1.7 Hz, 1H).
A mixture of ethyl 5-[[2-(l-tert-butoxycarbony)pyrrolidinyl]methylamino]pyridine-2carboxylate (800 mg, 2.29 mmol) in CH 2
CI
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
CI
2 The extracts were washed with brine, dried over Na 2 SO,-Na 2
CO
3 and evaporated to give ethyl 5-[(2-pyrrolidinyl)methylamino]pyridine-2carboxylate (460 mg, 81%) as a gum. 'H-NMR (CDCI 3 8 1.32 J= 7 Hz, 3 1.58-2.10 (series of m, 4 3.12-3.28 (series of m, 3 3.65 1 4.30 (be q, J 7 Hz, 2 6.27 (br, WO 01/00206 WO 0100206PCT/USOOI 18079 1 6.59 (dd, J= 2.4 and 8.5 Hz, 1 7.65 J= 8.5 Hz, 1 7.94 J =2.4 Hz, 1H).
To a stirred mixture of ethyl 5-[(2-pyrrolidinyl)methylaminojpyridinie-2-carboxylate (220 mg, 0.88 nunol), 4-[AN-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (300 mg, 0.88 nunol), 4- DMAP (135 mg, 1. 10 mmol) in DMIF (7 ml) was added EDC-HCl (215 mg, 1. 10 mniol) 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 (30m1) column chromatography with CHCl 3 EtOH (98:2, v/v) as eluent and crystallized with Et 2 O to give 5-[I-[4-[A-(2-chlorophenyl)ureidoJ-3-methoxyphenylacetyl]-2pyrrolidinyl]methylamino~pyridine-2-carboxylate (420 mg, 84%) as fine needles. IR (KBr) 3319, 1703, 1628, 1585, 1529 ciii'; 'H-NMR (CDCI 3 )8 1.38 J= 7 Hz, 3 1.73 17 (series of mn, 4 3.19 and 3.54 (each mn, each 1 3.63 2 3.70 3 4.39 (be q, J= 7 Hz, 2 4.55 (in, 1 6.02 (br s, 1 6.78-6.84 (series of s and mn, 3 6.98 (dt, J 2.4 and 8.0 Hz, 1 H), 7.16 1 7.21-7.26 (series of mn, 3 7.34 (dd, J =2.4 and 8.0 Hz, 1 7.90 J =8.3 Hz, IlH), 7.98 (in, 2H), 8.16 (dd, J 1. 2 and 8.8 Hz, I1H); MS (FA.B) m/z 566 Anal. Calcd for C191 32
CIN
5 0 5
-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-[I -[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyll-2pyrrolidinyl]methylanmunolpyridine-2-carboxylate (300 mg, 0.53 mmol) in THF:MeOH 1, vfv, 16 ml) and 0.25N NaOH (I11 ml, 2.75 mmnol) was stirred for 3 hr at room temperature. The mixture was poured into ice-IN HCI (3 The solid was collected, washed with water and airdried. The crude crystalline material was collected with CH 2
CI
2 -Et 2 O to give 61 (180 mg, as an amorphous solid MW 537.99 JR (KBr) 3319, 1701, 1620, 1585, 1533 cmn 1 'H-NMR (CDC 3 8 1H-NMR (DMSO-d) 8 1.80-2.05 (series of mn, 4 2.99 (in, 1 3.50- 3.59 (series of mn, 3 H), 3.60 2 3.86 3 4.11 (in, 1 6.78 J 8.5 Hz, 1 6.91 1 6.94 (mn, 1 H), 7.02 (in, 1 7.14 (dd, J= 2.5 and 8.5 Hz, 1 7.28 J= 7.0 Hz, 1 7.45 J= 8.0 Hz, 1 7.78 (d,J =8.8 Hz, IlH), 7.97 J =8.3 Hz, I1H), 8.08 (br s,1IH), 8.11 I1H), 8.89 1 8.94 1 MS (FAB) m/z 538 Anal. Calcd for C2 7 H28CN 5 1. 5XH 2 0: C, 57.39; H, 5.53; N, 12.39. Found: C, 57.37; HK 5.54; N, 11.74.
To a stirred mixture of ethyl 5-[(2-pyrrolidinyl)methylamuinolpyridine-2-carboxylate (230 mng, 0.923 inmol), 3-inethoxy-4-[Ml-(2-inethylphenyl)ureidoJphenylacetic acid (290 mg, 0.923 nunol), 4-DMAP (145 mng, 1. 15 mmnol) 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 WO 01/00206 WO 0100206PCT/USOO/18079 pored into ice-water. The solid was collected, washed with water and air-dried. The crude solid was purified by silica gel (30m1) column chromatography with CHCl 3 EtOH (98:2, vlv) as eluent to give ethyl -[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetylJ-2-pyrrolidinyl] methylaminojpyridine-2-carboxylate (400 mg, 80%) as fine needles. ER (KBr) n 3325, 1709, 1618, 1585, 1531 IH-NMR (CDCI 3 8 1.39 J= 7 Hz, 3 1.73-2.07 (series of m, 4 H), 2.28 3 3.12 and 3.49 (each m, each 1 3.60 2 4.39 (br q, J 7 Hz, 2 4.53 (in, 1 6.07 (br s, I 6.23 (br s, 1 6.75-6.77 (series of s and m, 2 6.82 (dd, J 3.0 and Hz, I 7.09-7.22 (series of m, 3 7.49 J 8.0 Hz, 1 7.90 J 8.5 Hz, 1H), 7.98 (d, J 2.6 Hz, lH), 8.06 J 8.8 Hz, 1H); MS (FAB) m/lz 546 Anal. Calcd for
CH
3 .5xH 2 O: C, 52.92; H, 6.69; N, 12.23. Found: C, 63. 11; H, 6.48; N, 11.96.
A mixture of ethyl 5-[[It-13 -methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetylj-2pyrrolidinyl~methylaminojpyridine-2-carboxylate (D9 14596) (290 mg, 0.53 mmol) in THF:MeOH 1, v/v, 16 ml) and 0. 25N NaOH (I I 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-dried. The crude crystalline material was collected with CH 2
CI
2 -Et 2 O to give 62 (170 mg, 62%) asan amorphous solid. MW 517.58 IR (KBr) 3283, 1701, 1618, 1529 'H-NMR
(CDCI
3 8 1H-NMR (DMSO-d) 5 1.78-2.04 (series of mn, 4 2.25 3 2.95-3.55 (series of m, 4 3.59 2 3.87 3 4.11 (in, I 6.75-7.24 (series of in, 7 7.83-7.97 (series ofim, 3 8.01 J= 8.3 Hz, 1 8.13 J= 2.6 Hz, I 8.11 (in, I 8.47 I 8.57 1 MS (FAR) n/z 518 Anal. Calcd for CnH 3
,N
5 5 2.5xH 2 O: C, 60.50; H, 6.39; N, 12.60. Found: C, 60.3 1; HK 6.28; N, 12. Example 57 I-[[4-[IN'-(2-chlorophenyl)ureidol-3-methoxyphenylacetylj carboxylic acid NR~y
COOH
1H H6e63 To a stirred solution of methyl 2-hydroxy-5-pyridinecarboxylate (2.0 g, 13.06 nunol), PPh 3 (4.2 g, 15.93 inrol) and 1-tert-butoxycarbony-(L)-prolinol (2.63 g, 13.06 minol) in THE (25 ml) was added a solution of DIAD (3.3 g, 15.67 minol) in TH-F (5 mld) at 0-10 OC. The resulting stirred mixture was then heated at reflux for 3 hr. After cooling, the mixture wsa evaporated in vacuo.
The residue was chromatographed on silica-gel (I 20in1) with n-hexane-EtOAc 1, v/v) as eluent to give methyl 1-tert-butoxycarbony)pyrrolidinyl]methoxypyridine-5-carboxylate (3.0 g, WO 01/00206 WO 0100206PCTUSOOII8079 68%) as a gumi. 'H-NMR (CDC 3 8 1.46 9 1.82-2.04 (series of m, 4 3.45 (in, 2 3.91 3 4.10-4.32 (series of m, 2 4.48 (mn, 1 6.32 (br, 1 6.75 J= 8.8 Hz, I 8.15 (dd, J:=2 and 8.8Hz, I 8.79 J= 2 Hz, I H).
A mixture of methyl 2-[2-(l-tert-butoxycarbony)pyrrolidinyl]methoxypyridine-5carboxylate (2.9 g, 8.62 mmol) in CH 2 CI, (80 ml) and TFA (20 mld) 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,-Na 2
CO
3 and evaporated to give methyl 2-(2-pyrrolidinyl)inethoxypyridine-5carboxylate (1.2 g, 59%) as a gum. 'H-NlvR (CDCI 3 )8 1.58-2.050 (series of m, 4 2.90-3.02 (series of m, 2 3.87 and 3.90 (each s, 3 4.23 (in, 1 4.37 (in, I 6.33 (br, 1 6.78 J =8.5 Hz, 1 8.15 (dd,J =2.2 and 8.8 Hz, 1 8.79 J =2.2 Hz, IlH).
To a stirred mixture of methyl 2-(2-pyrrolidinyl)methoxypyridine-5-carboxylate (370 mg, 1.57 mmol), 4-[N'-(2-chlorophenyl)ureidoj-3-methoxyphenylacetic acid (525 mg, 1.57 nunol), 4- DMAP (230 mg, 1.88 mmol) in DMF (10 ml) was added EDC-HCI (360 mg, 1.88 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 (30m1d) column chromatography with CHC1 3 EtOH (98:2, v/v) as eluent and crystallized with Et 2 O to give methyl 2-[1-[4-[N'-(2-chlorophenyl)ureidoJ-3-nethoxypheny (600 mg, 69%) as an amorphous solid.
'H-NMR (CDCI 3 8 .21 and 2.01 (each in, 4 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 aniide-isomers) A mixture of methyl I-[4-[M-(2-chlorophenyl)ureidoJ-3-inethoxyphenylacetylJ-2- (230 mg, 0.4 15 nmmol) in THF (1 ml) and 0.25N NaOH (4 ml, 1 minol) was stirred for 14 hr at room temperature and for 3 hr at 60 Mirter 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 CHC1 3 -EtOH 1, v/v) as eluent and crystallized with Et 2 O to give 63 (150 ing, as an amorphous solid. MW 538.98 IR (KBr) 3329, 1709, 1601, 1533 'H-NNM (CDCI 3 8 1.85- 2.05 (series of m, 4 3.50-3.60 (series of in, 2 3.82 3 3.86 2 4.29 (in, 1 H), 4.42 (in, 1 6.72-7.05 (series of in, 4 7.28 (in, 1 7.43 J =8 Hz, 2 7.95 J =8.3 Hz, 1 8.09 J 8.3 Hz, 2 8.64 (in, I 8.89 1 8.93 1 MS (FAB) m/lz 539 WO 01/00206 WO 0100206PCTLJSOOII8079 (W Anal. Calcd for C 7
H
28
CN
6 1.3xH 2 O: C, 57.55; H, 5.47; N, 9.94. Found: C, 57.94; H 5.00; N, 9.45.
Example 58 -(2-bromophenyl)ureidol -3-methoxyphenylacetyl] -2-pyrrolidinylmethoxyjpyridine-2carboxylic acid 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>HCI (360 mg, 1.88 nunol) 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 (30m1) column chromatography with CHCI 3 EtOH (98:2, v/v) as eluent and crystallized with Et 2 O to give methyl 5-fl -[4-[N'-(2-bromophenyl)ureidoJ-3-methoxypheny lacety1J-2pyrrolidinylmethoxy~pyridine-2-carboxylate (650 mg, 69%h) as an amorphous solid. IR (KBr) n 3323, 1720, 1624, 1601, 1527 'H-NMR (CDCI 3 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 (FAD) ni/z 597 (Mr and 599 Anal. Calcd for
C
28 HBrNO,-d.0xH 2 O: C, 54.55; H, 5.23; N, 9.09. Found: C, 54.13; H, 5.03; N, 9.33.
A mixture of methyl 5-fl -[4-[N'"-(2-bromophenyl)ureidoj-3-methoxyphenylacetylJ-2pyrrolidinylmethoxylpyridine-2-carboxylate (300 mg, 0.5 mmol) in THF:MeOH 1, v/v, 2 ml) and 0.25N NaOH (4 ml, 1 mmol) was stiffed for 3 hr at room temperature and for 5 hr at 60 'C.
AMrter 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 CHC1 3 -EtOH 1, v/v) as eluent and crystallized with Et 2 O to give 64 (180 mg, 62%) as an amorphous solid. MW 583.43 IR (KBr) n3319, 1705, 1685, 1601, 1529 cm-';'H-NMR (DMSOd 6 8 1.82-2.05 (series of m, 4 3.48-3.58 (series of m, 2 3.82 3 3.86 2 4.42- 4.55 (series of m, 3 6.72-6.98 (series of m, 4 7.32 J =8 Hz, 1 7.60 J =8 Hz, 1 7.95 (in, 2 8.08 (in, 1 8.63 (in, I 8.64 (in, 1 8.89 1 8.93 1 MS (FAD) m/z 5 83 Anal. Calcd for CH 2 sBrN 4
O
6 2.0xH 2 O: C, 52.26; H, 5.20; N, 9.03. Found: C, 52.72; H, 4.63; N, 8.50.
WO 01/00206 WO 0100206PCT/USOOI 18079 Example 59 441 -[3-[N'-(2-bromophenyl)ureido]-2-methoxy-6-pyridylacety]-(4S-luoro-(2S)-pyfrolidiny methoxyjbenzoic acid To a stirred solution of ethyl 3-aniino-2-methoxy-6-pyridylacetate (1.61 g, 7.66 mmol) in THF ml) were added 2-bromophenylisocyanate (948 mil, 7.66 nunol) and Et 3 N (107 ml, 0.776 niol).
After stirring overnight, the mixture was poured into H 2 0 (100 ml) and extracted with CHCI 3 MeOH 2 x 200 ml). The combined extracts were dried over MgSO 4 and evaporated. The residue was recrystallized from CHCI 3 -MeOH-hexane to give ethyl 3-[N'-(2-bromophenyl)ureidol- 2-methoxy-6-pyridylacetate (2.91 g, as a colorless crystalline powder. mp 160-163 'H- NMR (DMSO-d) 8 1.19 (dt, J= 7.1, 0.7 Hz, 3 3.69 2 3.95 3 4.07-4.13 (in, 2 H), 6.90 J= 7.8 Hz, 1 6.99 J= 7.8 Hz, 1 7.33 J= 7.8 Hz, 1 7.61 J= 7.8 Hz, 1 7.96 (dd, J=7.8, 1.5 Hz, 1 8.31 J= 7.8 Hz, I 8.82 1 9.12 1 MS (FAB) m/z 408 4 10 Anal. Calcd for C, 7 H,BrN 3
O
4 ,-0.25 H,0: C, 49.47; H, 4.52; 9.96.
Found: C, 49.34; H, 4.48; N, 9.6 A mixture of ethyl 3 -[N'-(2-bromophenyl)ureido]-2-methoxy-6-pyridylacetate (2.90 g, 7. 10 remol), 0.25 N NaOH (56.8 ml, 14.2 mmol), and THE (50 ml) was stirred for 5 h. The mixture was neutralized with I N HCI and the resulting precipitate was collected by filtration. The residue was recrystallized from CHCI 3 -MeOH-hexane to give 3-[N'-(2-bromophenyl)ureido]-2methoxy-6-pyridylacetic acid (2.40 g, 89%) as a colorless crystalline powder. mp 195-197 IH- NMvR (DMSO-d) 8 3.59 2 3.95 3 6.88 J 8.1 Hz, 1 6.97-7.01 (in, 1 7.33 J =7.3 Hz, 1 7.61 J =8.1 Hz, 1 7.95-7.97 (in, 1 8.29 J =8.1 Hz, 1 8.81 1 9. 10 1 12.35 (br s, I Anal. Calcd for C1 5 H,,BrN 3
O
4 C, 47.39; H, 3.71; N, 11.05. Found: C, 47.27; H, 3.59; N, 10.86.
To a stirred solution of 3-[N'-(2-bromophenyl)ureidoj-2-methoxy-6-pyridylacetic acid (751 mg, 1.97 minol) and methyl (4.S)-fluoro-(2S)-pyrrolidinylmethoxybenzoate (500 mag, 1.97 inmol) in DMIF (10 W) were added EDC-HCI (566 mg, 2.96 inmol), DMAP and HOBt 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 ml), dried over MgSO, and evaporated. The resulting residue was cliromatographed on silica gel with WO 01/00206 WO 0100206PCTIUSOO/18079
CHCI
3 -MeOH (20: 1) as eluent to give methyl 4-[1-[3-[N'-(2-bromophenyl)ureido]-2-methoxy-6pyridylacetylj-(4S)-fluoro-(2S)-pyrrolidinylmethoxybenzoate 16 g, as a yellow viscous solid.
A mixture of methyl 4-11-13 -[N'-(2-bromophenyl)ureidoj -2-methoxy-6-pyridylacetylj- (4S)-fluoro-(2S)-pyrrolidinylmethoxylbenzoate 16 g, 1.88 mmol), 0.25 N NaOH (15 ad, 3.75 mmol), and THE (15 mld) was stirred overnight. The mixture was neutralized with 1 N HCl and extracted with CHCI 3 -MeOH 2 x 200 ml). The combined extracts were dried over MgSO 4 and evaporated. The resulting residue was chromatographed on silica gel with CHC1 3 -MeOH (40:1 to 10: 1) as eluent to give 65 as a pale yellow amorphous solid. MW 601.42 'H-NMR (DMSO-d,) 8 2.27-2.39 (in, 2 3.33-4.84 (series of m, 10 5.33-5.53 (in, I 6.87-6.90 (mn, 1 6.99 1 H, J= 7.6 Hz), 7.08 2 H, J= 9.0 Hz), 7.34 1 H, J= 7.6 Hz), 7.61 1 H, J 7.8 Hz), 7.88 (dI, 2 H, J =9.0 Hz), 7.97 I H, J =8.3 Hz), 8.28-8.32 (in, 1 8.81-8.82 (in, 1 9.10-9.12 (mn, 1 12.66 (br s, 1 MS (FAB) mlz 601 603 Anal. Calcd for
C
27
H
26 BrFN 4
O
6 C, 53.92; H4,4.36; N, 9.32. Found: C, 52.37; H, 4.62; N, 8.38.
Example 4-[4.S)-fluoro- 1 -[4-[N'-(2-methylphenyl)ureidojphenylacetylj-(2S)-pyrrolidinylmethoxy~benzoic acid e HH 66 To a stirred solution of 4-[N'"-(2-methylphenyl)ureidojphenylacetic acid (337 mg, 1. 18 minol) and methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylnethoxy]benzoate (300 mg, 1. 18 inmol) in DMF (10 mld) were added EDC-HCl (339 mng, 1.77 nunol), HOBt (cat.) and DMAP The reaction mixture was stirred overnight. The mixture was partitioned between EtOAc (200) and H,0 (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 chroinatographed on silica gel with CHC1 3 -MeOH (50: 1) to give methyl 4-[(4S)-fluoro. 1-[4-[N'-(2-methylphenyl)ureidojphenylacetylJ -(2S)-pyrrolidinyl inethoxy]benzoate (613 mg, quant) as a yellow viscous oil. 'H-NMR (CDC1 3 )8 2.03-2.55 (series of m, total 5 3.47-4.21 (series of in, total 7 4.44-4.60 (in, 3 5.21 and 5.34 (in, each, total 1 6.87-7. 16 (in, 8 7.52-7.55 (mn, 3 7.93 J 8.8 Hz, 2 7.99 J 8.8 Hz, 1 H).
To a stirred solution of methyl 4-[(4.S)-fluoro-1-[4-[N'-(2-inethylphenyl)ureido] phenylacetyll -(2S)-pyrrolidinylnethoxyJbenzoate (613 mg, 1. 18 iniol) in THE (10 ml) was added WO 01/00206 WO 01111206PCTIUSOOII8O79 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 HCl (50 ml) and extracted with CHCl 3 -MeOH 2 x 200 ml). The combined extracts were dried over MgSO, and evporated. The residue was chromatographed on silica gel with CHCI 3 -MeOH (10: 1) to give 66 (378 mg, 63%) as a colorless amorphous solid. MW 505.54 'H-NMR (DMNSO-d 6 8 .08-2.30 (in, total 5 3.47-4.63 (series of m, 7 5.30-5.50 (in, 1 6.94 J =7.3 Hz, 1 7.02-7. 17 (in, 6 7.37-7.41 (in, 2 7.82-7.96 (in, 4 9.05 (s, 1 MIS (FAR) m/z 506 Anal. Calcd for CHFN 33 1.75 H 2 0: C, 62.62; HK 5.9 1; N, 7.82. Found: C, 62.23; H, 5.63; N, 7.18.
Example 61 4-[(4.S)-fluoro-1-13 -methoxy-4-[Nf'-(2-methylphenyl)ureido]phenylacetyl] -(2S)-pyrrolidinyl methoxylbenzoic acid To a stirred solution of l-QIert-butoxycarbonyl)-(4S)-fluoro-(2S)-proline (1.85 g, 7.93 inmol) in THF (15 ml) was added BH 3 DMS (0.75 ml, 7.93 minol) at room temperature. After being heated 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 CHC1 3 (2 x 200 ml). The combined extracts were washed with brine (100 ml), dried over MgSO 4 and evaporated. The residue was chroinatographed on silica gel with CHCl 3 -EtOAc 1) as eluent to give 1 -Qert-butoxycarbonyl)-(4S)-fluoro-(2S)-pyrrwlidinylmethano (1.76 g, quant) as a colorless oil. 'H-NMR (CDCI 3 8 1.48 9 1.64 (in, 1 1.97-2.28 (mn, 2 3.53-3.87 (series of in, 4 4.094.25 (in, 1 5.09 and 5.22 (mn, each, total 1 H).
To a stirred mixture of I-Qtert-butoxycarbonyl)-(4.S)-fluoro-(2S)-pyrrolidinylinethano (500 mg, 2.28 mmol), methyl 4-hydroxybenzoate (416 mg, 2.74 inmol), Ph3P (719 mg, 2.74 mmol) in THF (10 mld) was added DIAD (0.54 nil, 2.74 minol) at moom temperature. The mixture was 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 CHCI,-EtOAc as eluent (10: 1 to 4: 1) to give methyl 4-[l-Qtert-butoxycarbonyl)-(4S)-fluoro-(2S)-pyrrolidiny inethoxy]benzoate (597 mg, as a colorless oil. 'H-NMR (CDCI 3 8 1.49-1.59 (in, 9 2.05- 2.21 (in, 1 3.56-4.43 (series of in, 8 5.19 and 5.32 (in, each, total 1 6.97 (in, 2 7.98 J =8.5Hz, 2H).
WO 01/00206 WO 0100206PCT/USO0I 18079 A mixture of methyl 1 -Qtert-butoxycarbonyl)-(4S)-fluoro-(2S)-pyrrolidinylmethoxy] benzoate (590 mg, 1.67 mmol) and TVA (5 mld) in CHCI 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 CHCI 3 (2 x 200 ml). The combined extracts were dried over K 2 CO3 and evaporated to give methyl 4-f (4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (414 mg, 98%) as a yellow solid. IH- NMR (CDCI,) 5 1.89-2.02 (in, I 2. 16-2.31 (in, I 2.98 (mn, 1 3.35 (mn, 1 3.46-3.68 (in, 1 3.86 3 4.00-4.08 (in, 2 5.16 and 5.29 each, J =4.7 Hz, total 1 6.91 J =8.8 Hz, 2 7.96 J 8.8 Hz, 2 H).
A mixture of methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (205 mng, 0.8 minol), 3-methoxy-4-[N'-(2-methylphenyl)ureidolpheniylacefic acid (254 mg, 0.8 10 minol), EDC-HCI (233 mng, 1.22 minol), HOBt and DMAP (cat.) in DMF (10 ml) was stiffed 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 chroinatographed on silica gel with-CHC1 3 -MeOH 1) as eluent to give methyl 4-[(4S)-fluoro-1-[3-methoxy-4-[Ar-(2-inethylphenyl)ureido] phenylacetyl]-(2S)-pyrrolidirnylmethoxylbenzoate (445 mng, quant) as a light brown viscous. 'H- NMR (CDCI 3 )6 2.05-2.55 (in, total 6 3.554.13 (in, 11I 4.48-4.60 (in, 2 5.20 and 5.33 (each m, total 1 6.29 1 6.79 (in 2 6.96 (di, J= 8.8 Hz, 2 7.11-7.25 (in, 3 7.48 (di, J =7.6 Hz, 1 7.93 -8.09 (in, 4 H).
A mixture of methyl 4-[(4S)-fluoro- 1-f3-methoxy-4-[N'-(2-inethylphenyl)ureido] phenylacetylJ-(2S)-pyrrolidinylinethoxy]benzoate (445 ing, 1.62 inmol) and 0.25 N NaOH (15 ml, 3.75 inmol) in THF (15 ml) was stirred overnight at room temperature then for 2 h at reflux. The mixture was acidified with 1 N HCl and extracted with CHC1 3 -MeOH 2 x 200 mld). The combined extracts were dried over MgSO, and evaporated. The residue was chrointographed on silica gel with CHCl 3 -MeOH (10: 1) as eluent to give 67 (260 mg, 30%) as a pale yellow amorphous solid. MW 535.56 'H-NNM (DMSO-d) 862.25-2.51 (in, 5 3.33-4.41 (series of in, 5.30-5.50 (in, 1 6.75-7.17 (in, 7 7.79 (di, J 8.1 Hz, 1 7.87-8.04 (in, 3 8.48 (in, 1 8.58 (in, 1 MIS (FAB) m/z 536 AnaI.Calcd for C29H 3
DFN
3
O
6 -HO: C, 62.92; H, 5.83; N, 7.59. Found: C, 62.40; H, 5.82; N, 6.93.
Example 62 4-[1 -[4-[N--(2-Broinophenyl)ureidoj-3-nethoxyphenylacetyll-(4S)-fluoro-(2S)-pyfrolidinyI inethoxy]benzoic acid WO 01/00206 PCT/US00/18079 N 68 r H H 6Me, 68 A mixture of methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (501 mg, 1.98 mmnol), 4- [N'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (750 mg, 1.98 mmol), EDC-HCI (569 mg, 2.97 mmol), HOBt (cat.) and DMAP (cat.) in DMF (10 mi) was stirred overnight. The mixture was diluted with EtOAc (300 ml), washed with brine (100 mi), dried over MgSO, and evaporated. The residue was chromatographed on silica gel with CHCl 3 -EtOAc to CHC13- MeOH (10:1) as eluent to give methyl 4-f[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl acetyl]j-(4S)-fluoro-(2S)-pyrrolidinylmethoxylbenzoate (1.29 g, quant) as a brown viscous oil. 'H- NMR (CDCI 3 8 2.05-2.58 2 3.49-4.17 (series of m, 12 4.52-4.65 2 6.82-7.33 (series of m, 8 7.53 (dd, J= 8.1, 1.5 Hz,1H), 7.95-8.02 4H), 8.14 (dd, J 8.3, 1.7 Hz, 1H).
A mixture of methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-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 2 x 200 ml). The combined extracts were dried over MgSO, and evaporated. The residue was chromatographed on silica gel with CHC1 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 2 3.21-4.63 (series of m, 10 5.31-5.51 1 6.74- 7.10 5 7.32 J= 7.8 Hz, 1 7.60 J= 7.8 Hz, 2 7.87-7.99 4 8.74-8.75 1 8.92-8.94 (nm, 1 MS (FAB) m/z 601 Anal. Calcd for C 2 ,H,,BrFN306O2 H, 2 0: 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 OC; Anal. Calcd for CHBrFN 3 NaO 6 -0.75 HO 2 0: C, 52.88; H, 4.36; N, 6.61. Found: C, 52.97; H, 4.36; N, 6.61.
Example 63 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinyl methoxy]benzoic acid N y N" COOH 69 A mixture of methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (205 mg, 0.810 mmol), 3methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (254 mg, 0.810 mmol), EDC-HCI (233 WO 01/00206 WO 0100206PCT/USOO/18079 mg, 1.22 mmol), HQBt (cat.) and DMAP (cat.) in DMF (10 nml) was stirred overnight. The mixture was diluted with EtOAc (200 nil), washed with brine (2 x 100 mil), dried over MgSO 4 and evaporated. The residue was chromatographed on silica gel with CHCI 3 -MeOH (20: 1) as eluent to give methyl 1-[4-IN' -(2-chlorophenyl)ureidol-3-methoxyphenylacetyl]-(4S)-fluoro-(2S)pyrrolidinylmethoxylbenzoate (376 mg, 81%) as a yellow foam. 'H-NMR (CDCI 3 )82.07-2.56(in, 2 3.57-4.14 (series of mn, 11I 4.50-4.61 (in, 2 5.22 and 5.35 (series of in, total I H), 6.80-7.33 (series of mn, 9 7.93-8.00 (in, 3 8.16 J 8.1 Hz, 1 H).
A mixture of methyl methyl 4-fl -[4-[A'-(2-chlorophenyl)ureidol-3-methoxyphenylacetyl (4.S)-fluoro-(2S)-pyrrolidinylmethoxyjbenzoate (376 mg, 0.660 minol) and 0.25 N NaOH (15 ml, 3.75 mmol) in THF (15 mil) was stirred overnight at room temperature then for 2 h at reflux. The mixture was acidified with 1 N HCI and extracted with CHCI 3 -MeOH 2 x 200 ml). The combined extracts were dried over MgSO, and evaporated. The residue was chrointographed 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 IH-NMR (DMSO-d)8 2.24-2.50 1 2 3.48-4.65 (series of in, 10 5.30-5.50 (in, 1 6.75-7.08 (in, 5 7.29 J= 7.3 Hz, 1 7.43-7.45 (in, 1H), 7.89- 7.98 (in, 2 7.99 J =8.3 Hz, 1 8.09 J =7.1 Hz, 1 8.90-8.96 (in, MS (FAB) m/z 556 Anal. Calcd for C8HvCIFN 3
O
6 1/4H 2 0: C, 60.00; H, 4.95; N, 7.50. Found: C, 59.67; H, 5.08; N, 7. Example 64 1-f4-[N'-(2-bronophenyl)ureidophenylacetyl-(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoic acid N Nn NO-O-C OOH r H Methyl 4-fl -(4-benzyloxycarbonylaminophenylacetyl)-(4S)-fluoro-(2S)-pyrrolidinylnethoxy benzoate (300 mg, 0.576 minol) was tsdded EtOH-THF 30 mil) and the solution was 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 chroinatographed on silica gel with CHCI 3 -MeOH (20: 1) as eluent to give methyl am-inophenylacetyl)-(4S)-fluoro-(2S)-pyrrolidinylmethoxy~benzoate (200 mg, 90%) as a yellow oil.
IH-NMR (CDCI 3 8 2.0 1-2.56 (series of in, 2 3.50-4.14 (series of in, 5 4.454.62 (in, 2 H), 5.21 and 5.34 (each mn, total 1 6.60-6.65 (in, 2 6.88 J 8.8 Hz, 0.5 6.99-7.05 (in, 7.95-8.00 (in, 2 H).
WO 01/00206 WO 0100206PCT/USJO/18079 methyl 441 -(4-aminophenylacetyl)-(45)-fluoro-(2S)-pyrrolidinylmethoxylbenzoate (200 mg, 0.518 mmol) was dissolved TI-F (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 Miuted with EtOAc (200 ml). The solution was washed with brine (100 mld), dried over MgSO 4 and the solvent was removed under a reduced pressur. The residue was chromatographed on silica gel with CHCI,-EtOAc 1) to CHCl 3 -MeOH (10: 1) as eluent to give methyl 4-[1-[4-[N'-(2-bromophenyl) ureido]phenylacetyll-(45)-fluoro-(2S)-pyrrolidinylmethoxyjbenzoate (303 mg, quant) as a yellow oil. 'H-NMR (CDCI 3 8 2.08.2.60 (series of m, 2 3.56-4.69 (series of mn, 10 5.28 and 5.40 (in, each, total 1 6.84-6.92 (mn, 3 7.03.7. 10 (in, 3 14(d, J =8.1 Hz, 1 7.23 J 8.1 Hz, 1 7.39-7.44 (in, 2 7.89 J= 8.1 Hz, 1 7.98-8.03 (in, 2H), 8.09(d, J= 8.1 Hz, 1H).
Methyl 1 [4-[N'-(2-bromophenyl)ureido~phenylacetyl] -(4S)-fluoro-(2S)-pyrrolidinyl-methoxy] benzoate (300 mg, 0.5 13 mmol) was dissolved THF (5 ml), and 0.25 N NaOH (4.0 ml, 1.00 mimol) was added to this solution. After being stirred for 3 days, the mixture was poured into 1 N HCI (100 mld), and extracted with CHCl 3 -MeOH 2 x 200 mld). The combined extracts were dried over MgSO, and the solvent was removed under a reduced pressure. The residue was chromatographed on silica gel with CHCI 3 -MeOH (10:1) to give 70 (209 mng, as a colorless amorphous solid. MW 570.41 'H-NMR(DMSO-dj)8 2.24-2.51 (mn, 2 3.36-4.64 (series of mn, 7 5.31-5.50 1 6.97 (t,J =7.8 Hz, 1 7.04 J= 8.5 Hz, 1 7.09 J =8.8 Hz, 1 7. 14-7.20 (in, 2 7.34 J 7.8 Hz, 1 7.38-7.43 (in, 2 7.61 J 8.1 Hz, 1 H), 7.87-7.92 (mn, 2 8.08 J= 8.1 Hz, 1 8.15 1 9.45-9.47 (in, 1 12.66 (br s, 1 H); MS (FAB) m/z 572 570 Anal. Calcd for C 27
H
2 BrFN 3
O
3 -1.5 H 2 0: C, 54.28; H, 4.72;1 N, 7.03. Found: C, 54.67; HK 4.51; N, 6.61.
Example 4-[I-[4-[N'-(2-iodophenyl)ureido-3-methoxypheniylacetyl-(4S)-fluoro-<2S)-pyrrolidinylnethoxy benzoic acid H H 6Me71 To a stirred solution of tert-butyl 4-ainino-3-methoxyphenylacetate (1.94 g, 8.16 inmol) in TH-F ml) was added 2-iodophenylisocyanate (2.0 g, 8.16 inmol) and Et 3 N (114 ul, 0.8 16 inmol).
After stirring overnight, the mixture was poured into I N HCI (200 ml). The resulting precipitate was collected by filtlation and dissolved in CHC1 3 (200 ml). The solution was dried over MgSO, and evaporated to give tert-butyl 4-[N'-(2-iodophenyl)ureido]-3-methoxyphenylacetate (3.93 g, WO 01/00206 WO 0100206PCTIUS0OIISO79 quant) as a pale yellow amorphous solid. 'H-NMR (CDCI 3 )8 1.44 9 3.49 2 3.85 3 6.78-6.88 (in, 4 7.07 1 7.31-7.3 5 (in, 1 7.76 (dd, J= 7.8, 1.5 Hz, I 7.95 J =8.3 Hz, 1 7.99 (dd, J= 8.3, 1.5 Hz, IlH). MS nz 483 1).
A stirred mixture of tert-butyl 4-[N'-(2-iodophenyl)ureidoj-3-methoxyphenylacetate (3.93 g, 8.16 mmol) and TFA (5 ml) in CH 2 CI (5 ml) was refluxed for 3 h. After cooling to rt, the midxture 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 colun chromatography on silica gel with
CHCI
3 -MeOH 1) as eluent to give 4-[N'-(2-iodophenyl)ureido]-3-methoxyphenylacetic acid (2.89 g, 83%) as a pale yellow crystalline powder. IH-NMR (DMSO-d)8 3.62 2 3.88 3 6.78 J 8.3 Hz, 1 6.83-6.87 (in, 1 6.94 1 7.32-7.36 (mn, 1 7.69 (dd, J 8.3, 1.5 Hz, 1 7.84 (dd, J 1.5 Hz, 1 7.97-8.00 (im, 1 8.55 (in, 1 8.82 (in, 1 H), 12.26 (br s, I1H).
A mixture of 4-[N'-(2-iodophenyl)ureido]-3-methoxyphenylacetic acid (505 mg, 1. 18 minol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxylbenzoate (300 mg, 1. 18 inmol), EDC&HC1 (339 mg, 1.77 minol), DMAP (catatlytic amount) and HOBt (catalytic amount) in DMF (10 ml]) was stirred overnight. The mixture was diluted with EtOAc (300 mld) and washed with brine (2 x 200 ml). The solution was dried over MgSO, and evaporated. The resulting residue was chroinatographed on silica gel with CHCl 3 -EtOAc 1) as eluent to give methyl iodophenyl)ureido]-3-methoxyphenylacetyl] -(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (500 ing, 64%) as a colorless viscous oil. 'H-NMR (CDC 3 8 07-2.58 (in, 2 3.59-4.20 (in, 11I H), 4.51-4.64 (in, 2 5.24 and 5.37 (in, each, total 1 6.80-6.91 (in, 5 6.98 J =8.8 Hz, 2 7.34 J 7.8 H-z, 1 7.78 (dd, J 1.2 Hz, 1 7.95-8.02 (in, 4 H).
To a stiffed solution of methyl 4-[1-[4-[N'-(2-iodophenyl)ureidoJ-3-methoxyphenyI acetylj-(4S)-fluoro-(2.S)-pyrrolidinylinethoxyjbenzoate (500 mg, 0.756 minol) in TIHF (6 ml) was 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 CHCI,-MeOH 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 ing, as a colorless amorphous solid. MW 647.43 'H-NMR (DMSO-dj)6 2. 2.09-2.31 (in, 2 3.33-4.41 (series of in, 10 5.30-5.50 (in, 1 6.77-6.92 (mn, 3 7.03-7.09 (in, 2 7.34 J 8.1 Hz, 1 7.69 (dd, J 8.3, 1.5 Hz, 1I-H), 7.83-7.99 (in, 4 8.54 (in, 1 H), WO 01/00206 WO 0100206PCTUSOOII8079 8.82 (in, 1 MIS (FAR) m/z 648 Anal. Calcd for C 28
,H
2 1F1N 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)-fiuoro- 1 '-phenylureido)phenylacetylJ-(2S)-pyrrolidinylmethoxylbenzoic acid
F
(:)NYJO''XqO-Q-COOH To a stirred solution of ethyl 4-amninophenylacetate (6.43 g, 35.9 mniol) and Et 3 N (5.50 ml, 39.5 mmol inTBF(70 -d)wasadded phenyl isocyanate (3.90 ml, 35.9 inmol), 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) phenylacetate (9.64 g, as a white crystalline powder. mp 153-1 55'C; 'H-NMR (CDCl 3 )8 1.26 J =7.1 Hz, 3 3.52 2 4.15 J =7.1 Hz, 2 6.98-7.04 1 7.07-7. 11 (in, 4 7.18-7.2 5 (in, 5 7.42 1 MS (FAR) m/1z 299 (Mr+ Anal. Calcd for
C
17
H
28
N
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 inmol) in THE (80 ml) was added 0.5 N NaOH (80 ml) and the reaction mixture was heated under refiux for hr. After cooled to room temperature, the mixture was poured into ice-lI N HCL The resulting precipitate was collected under a reduced pressure and the crude solid was recrystallized from MeOH-CHCI 3 to give 4-(N'-phenylureido)phenylacetic acid (8.14 g, 93%) as a white crystalline powder. MS (FAR) m/z 271 (M 4 Anal. Calcd for C 15
H
14
N
2 0 3 C, 66.66; H, 5.22; N, 10.36.
Found: C, 66.45; H, 5.22; N, 10.30.
A mixture of 4-(N -phenylureido)phenylacetic acid (3 10 mg, 1. 15 minol), methyl 4-1(4S)fluoro-(2S)-pyrrolidinylinethoxy~benzoate (287 mg, 1. 13 minol), EDC-HCI (260 mg, 1.36 inmol), HQBt (185 mg, 1.37 inmol), and EtN (190 ml, 1.36 mmol) in DMF (5 mld) was stirred at room temperature overnight. The mixture was diluted with H,0, and extracted with EtOAc. The extract was washed with brine, dried over NaSO,, and evaporated. The residue was purified by column chromatography on silica-gel with CHCI,-MeOH (100: 1 to 50: 1, v/v) as eluent to give methyl 4- [(4S)-fluoro- 1 '-phenylureido)phenylacetylj-2S)-pyrrolidinylmethoxybelzoate (570 mng, 99%) as a pale yellow foam. 'H-NMR (CDCI,) 8 2.07-2.58 (in, 2 3.55-3.56 (in, 1 3.69- 3.98 (series of s and in, total 6 4.01-4.08 and 4.21-4.25 (each m, 1 4.46-4.65 (mn, 2 H), 5.23-5.25 and 5.38 (each m, 1 6.88-7.07 (in, 7 7.15-7.20 (in, 2 7.28-7.30 (in, 2 7.34 WO 01/00206 WO 0100206PCT/USOO/18079 and 7.40 (each s, 1 7.71 and 7.81 (each s, 1 7.9 1-7.95 and 7.99-8.0 1 (each mn, 2 MS (ESI) m/z 506 1).
To a stirred solution of methyl 4-[(4S)-fluoro-l '-phenylureido)phenylacetyl]-(2S')pyrrolidinylmethoxyjbenzoate (570 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 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-CHC 3 -LPE to give 72 (348 mng, 63%) as a white crystalline powder. NM 491.51 mp 169-1711C; 'H-NMR (DMSO-dE) 8 2.24-2.36 (in, 2 3.47-4.08 5 4.204.64 (mn, 2 5.3 1-5.50 (mn, 1 6.94-7.46 (series of m, total I11 H), 7.87-7.92 (in, 2 8.64-8.67 (mn, 2 12.63 (br s, 1 MS (FAB) m/lz 492 Anal Calcd for C 27
H
2 FN0 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-l1-[3 -iethyl-4-(N '-phenylureido)phenylacetyl]-(2S)-pyrrolidiniylmethoxylbenzoic acid To a stirred solution of tert-butyl 4-amino-3-methylphenylacetate (1.20 g, 5.42 minol) and Et 3
N
(830 ml, 5.95 minol) in THE (10 ml) was added phenyl isocyanate (650 ml, 5.98 minol) 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 3-methyl-4- (N'-phenylureido)phenylacetate 12 g, 61%) as a white crystalline powder. mp 143-145'C; 'H- NMR (CDCI 3 8 1.47 9 2.09 3 3.47 2 6.44 I 7.01-7.07 (in, 4 7.16- 7.27 (in, 2 7.30-7.33 (in, 2 7.45-7.47 1 H).
To a stiffed solution of tert-butyl 3-methyl-4-(N'-phenylureido)phenylacetate 12 g, 3.29 inmol) in CH 2
CI
2 (10 ml) was added TEA (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, to give 3-methyl-4-(N'-phenylureido)phenylacetic acid (680 mng, as white needles. 'H-NMR (DMSO-d6) 5 2.22 3 3.46 2 6.93-7.05 (mn, 3 H), WO 01/00206 WO 0100206PCT/USOO/18079 7.25-7.29 (mn, 2 7.43-7.46 (in, 2 7.72-7.74 (in, 1 7.90 1 8.98 1 12.26 (br s, I AnaL Calcd for C 26
H
16
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 minol), methyl 4- [(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (268 mg, 1.06 inmol), EDC-HCI (243 mng, 1.27 innol), HOBt (172 ing, 1. 27 mmol), and Et 3 N (180 ml, 1.29 iniol) in DMIF (5 ml) was stirred at room temperature overnight. The mixture was diluted with H,0, and extracted with EtOAc. The extract was washed with brine, dried over Na 2 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-I(4S)-ftuoro-1 -f3-methyl-4-(N '-phenylureido)phenylacetylJ-(2S)-pyrrolidinylmethoxy] benzoate (550 mng, as a white foam. 'H-NMR (CDCI 3 8 1.79 and 1.87 (each s, 3 2.04- 2.61 (mn, 2 3.52-3.54 (in, 1 3.73-4.27 (series of s and m, total 7 4.474.67 (mn, 2 H), 5.26-5.27 and 5.40 (each in, 1 6.79-6.99 (in, 6 7. 14-7.18 (in, 2 7.27-7.31 (mn, 2 H), 7.40-7.44 (in, 1 7.89-8.0 1 (in, 3 MS (ESI) m/z 520 1).
To a stirred solution of methyl 4-[(4S)-fluoro-1-[3-inethyl-4-(N -phenylureido) phenylacetylJ-(2.S)-pyrrolidinylinethoxylbenzoate (550 ing, 1.06 innol) in THF (5 ml) was added 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-i 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, as a white crystalline powder. NM 505.54 mp 130-135 0 C; 'H-NMvR (DMSO-d 6 862.18-2.30 (series of s and m, total 5 3.47-3.92 (series of mn, total 5 4.034.63 (mn, 2 5.31-5.50 (mn, I 6.94-7. 10 (in, 5 7.26-7.30 (rn, 2 7.45-7.47 (in, 2 7.70- 7.75 (nm, 1 7.87-7.92 (in, 3 8.96-8.98 (mn, 1 12.63 (br s, 1 MS (ESI) ink 506 (Ivr+ Anal Calcd for C,, 2 gJN30 5 112H 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.
Example 68 4-I(4S)-fluoro- l-[4-[N'-(2-fluorophenyl)ureido-3-nethylphenylacetyl-(2S)-pyrrolidinylnethoxyI benzoic acid To a stirred solution of tert-butyl 4-amino-3-inethylphenylacetate (1.09 g, 4.93 minol) and Et 3
N
(755 ml, 5.42 inxol) in THF (10 ml) was added 2-fluorophenyl isocyanate (610 1 5.44 nunol) WO 01/00206 WO 0100206PCTUSOOI18079 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 fiuorophenyl)ureidoj-3-methylphenylacetate (1.31 g, 74%) as a white crystalline powder. mp 89- 91'C; IH-NMR (CDCI 3 8 1.47 9 2.06 3 3.49 2 6.62 1 6.92-7.09 (in, 7.21 (br s, 1 7.49-7.51 (in, I 8.10-8.15 I AnaI. Calcd for CIH 23
FN
2
O
3
C,
67.02; H, 6.47; N, .7.82; F, 5.30. Found: C, 66.74; H, 6.3 5; N, 7.85; F, 5.69.
To a stirred solution of tert-butyl 4-[N '-(2-fluorophenyl)ureidoj-3-inethylphenylacetate (1.25 g, 3.49 minol) in CHCI 2 (10 ml) was added TEA (10 ml) and the reaction mixture was stirred at rom temperature overnight. The reaction mixture was concentrated to a small volume and poured into ice-H 2 0. The resuilting precipitate was collected under a reduced pressure and the crude solid was recrystallized from MeOH-CHCI,-IPE to give 4-[N'-(2-fluorophenyl)ureidoj-3methyiphenylacetic acid (830 mng, 79%) as white needles. 'H-NMR (DMSO-d 6 8 2.23 3 H), 3.47 2 6.96-7.30 (in, 5 7.74-7.76 (in, 1 8.17-8.20 (in, 1 8.33 1 8.94 1 12.27 (br s, 1 Anal. Calcd for C 1 6
H,
5
FN
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-inethylphenylacetic acid (321 mg, 1.06 mxnol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxylbenzoate (269 mg, 1.06 mmol), EDC-HCI (244 mng, 1.27 minol), HOBt (172 mg, 1.27 inmol), and Et 3 N (177 ml, 1.27 mmol) in DMPT (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 CHC1 3 -MeOH (100: 1, v/v) as eluent to give methyl 4-II(4S)-fluoro-l1-14-[N '-(2-fluorophenyl)ureido]-3-inethylphenylacetylJ-(2S)-pyrrolidiny methoxy]benzoate (560 mg, 98%) as a white foam. 'H-NMR (GDCI 3 )8 1.78 and 1.86 (each s, 3 2.16-2.65 (mn, 2 3.58-3.61 (in, 1 3.74-4.15 (series of sand mn, total 7 4.294.34 and 4.46-4.49 (each mn, 1 4.64-4.73 (mn, 1 5.29-5.34 and 5.43-5.47 (each in, 1 6.84-6.97 (in, 6 7.04-7.07 (mn, I 7.21 (br s, 1 7.55-7.59 (in, 1 7.85-8.02 (in, 3 8. 18-8.22 (in, 1 MS (ESI) m/z 538 (Mr+l).
To a stirred solution of methyl 4-[(4S)-fluoro-1 -[4-IN -(2-fluorophenyl)ureidoJ-3inethylphenylacetylJ-(2.S)-pyrrwlidinylmethoxy]benzoate (560 mg, 1.04 minol) in THE (5 nil) was added 0.5 N NaOH (5 ml), and the reaction mixture was heated under reflux for 5 hr. Afler cooled WO 01/00206 WO 0100206PCTIUSOO/18079 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-CHCI,-IPE to give 74 (297 mg, as a white crystalline powder. MW 523.53 mp 137-143'C; 'H-NMR (DMSO-<L) 2.20-2.31 (series of s and m, total 5 3.56-3.92 (series of m, total 5 4.03-4.63 (in, 2 5.3 1-5.50 (in, 1 6.96-7.26 (series of m, total 7 7.72-7.77 (in, 1 7.87-7.92 (in, 2 8. 1748.22 (in, 1 8.32-8.36 (in, 1 8.94-8.95 (in, 1 12.66 (br s, 1 MS (ESI) m/z 524 Anal. Calcd for Q 8 gH 2
.IF
2
N
3 05: 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 4-I(4S)-fluoro- -(2-trifluoromethylphenyl)ureidolphenylacetylJ-(2S)-pyrrolidinylmethoxy] benzoic acid
CF
3 H H To a stirred solution of ethyl 4-aminophenylacetate 13 g, 6.31 mmiol) and Et 3 N (965 ml, 6.92 inmol) in THE (10 ml) was added 2-trifluoroinethylphenyl isocyanate (953 ml, 6.31 inmol) and the 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 trifluoroinethylphenyl)ufeidolphenylacetate (1.93 g, 84%) as white needles. mp 137-1 39'C; 'H- NiAR (CDCl 3 5 1.25-1.29 (in, 3 3.59 2 4.154.20 (in, 2 7.05 (br s, 1 7. 13-7.23 (in, 6 7.47-7.5 1 (in, 1 7.54-7.56 (in, 1 8.0 1-8.03 (in, 1 H).
To a stirred solution of ethyl 4-[N-(2-trfluoromethylphenyl)ureidolphenylacetate (1.93 g, 5.27 iniol) in THE (10 ml) was added 1 N NaOH (10 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.
The resulting precipitate was collected under a reduced pressure and the crude solid was recrystallized from MeOH-CHCI 3 -IPE to give 4-[N'-(2-trifluoromethylphenyl)ureidolphenylacetic acid (910 mg, as a white crystalline powder. mp 224-225 0 C; H-NMR (DMSO-d) 83.50 (s, 2 7.18 J 8.3 Hz, 2 7.25-7.29 (in, 1 7.40 J 8.3 Hz, 2 7.62-7.69 (in, 2 H), 7.95-7.97 (in, 1 8.06 I 9.37 1 12.27 (br s, I Anal. Calcd for C,4-11 3 17 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 nmixture of 4-[N'-(2-trifluoroinethylphenyl)ureidolphenylacetic acid (302 mg, 0.89 nunol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxylbenzoate (226 mng, 0.89 nunol), EDC-HCl (205 WO 01/00206 WO 0100206PCT/USOO/18079 mg, 1.07 rmol), HOBt (145 mg, 1.07 nunol), and Et 3 N (150 ml, 1.08 minol) in DMF (5 nil) 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 CHCI 3 -MeOH (100: 1 to 60: 1, vA') as eluent to give methyl 4-[(4S)-fluoro-l1-[4-[N'-(24rifluoromethylphenyl)ureidolphenylacetyl-2S)pyrrolidinylinethoxyjbenzoate (463 mg, as a pale yellow foam. 'H-NMR (CDCl 3 8 2.09- 2.60 (in, 2 3.56-4. 12 (series of s and m, total 8 4.26-4.65 (in, 2 5.26-5.29 and 5.39-5.42 (each m, total 1 6.87-6.93 (in, 2 6.99-7.13 5 7.30-7.33 (in, 1 7.44-7.53 (in, 2 7.88-7.92 (mn, 1 7.99-8.04 (in, 2 8.09-8.15 (mn, 1 MIS (ESI) niz 574 To a stirred solution of methyl 4-I(4.S)-fluoro-1-[4-[N'-(2-trifluoromethylphenyl)ureidoJ phenylacetyl]-(2.S)-pyrrolidinylinethoxyjbenzoate (460 ing, 0.80 mxnol) in THF (5 ml) was added 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-lI N HCl, and the resulting precipitate was collected under a reduced pressure. The crude solid was recrystallized from MeOH-CHCI,-IPE to give 75 (169 mg, as a white crystalline powder. MW 559.51 mp 130-135'C; 'H-NMR (DMSO-d 6 8 2.24-2.30 (in, 2 3.51-4.24 (series of m, total 5 4.38-4.40 and 4.61 (each mn, total 2 5.31-5.50 (in, 1 7.03-7.42 (series ofmr, total 7 7.62-7.69 (in, 2 7.87-8.07 (in, 4 9.36-9.37 (mn, 1 12.64 (br s, 1 MIS (ESI) m/z 560 AnaI. Calcd fbr
C,HF
4
N
3 C, 60. 11; H, 4.50; N, 7.5 1; F, 13.58. Found: C, 60. 10; H, 4.85; N, 7.0 1; F, 12.90.
Example 4-[(4.S)-fluoro-1-[3-inethoxy-4-IN '-(2-trifluoroinethylphenyl)ureidolphenylacetylJ-(2S)-pyrrwlidinyI inethoxyjbenzoic acid
F
N N Me9 O-Q--OOH F3H H Me76 To a stirred solution of tert-butyl 4-amino-3-inethoxyphenylacetate (1.11 g, 4.68 nmnol) and Et 3
N
(720 ml, 5.17 nunol) in THF (10 mil) was added 2-trifluoroinethylphenyl isocyanate (707 ml, 4.68 nunol) 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'-(2trifiuoromethylphenyl)ureidolphenylacetate (1.11 g, 56%) as a white crystalline powder. mnp 13 1- 133 0 C; 'H-NMR (CDCI 3 8 1.44 9 3.49 2 3.85 3 6.83-6.88 (mn, 3 6.98 (hr s, 1 7.17-7.21 (in, 1 7.52-7.59 (in, 2 7.89-7.91 (mn, 1 8.04-8.06 (in, 1 H).
WO 01/00206 WO 0100206PCT/USOO/ 18079 To a stirred solution of tert-butyl 3-methoxy-4-[N '-(2-trifluoromethylpheniyl)ureido] phenylacetate (1.11 g, 2.62 mmol) in CH 2
CI
2 (10 ml) was added TEA (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 -IPE to give 3 -methoxy-4-[N (2-trifluoromethylphenyl)ureido~phenylacetic acid (839 ing, 87%) as a white crystalline powder.
mp 218-220*C; 'H-NMR (DMSO-A 8 3.51 2 3.87 3 6.76-6.79 (in, 1 6.93-6.94 (in, 1 7.27-7.30 (in, 1 7.6 1-7.69 (in, 2 7.82-7.84 (in, I 7.97-7.99 (mn, 1 8.71 (s, I 8.89 I 12.30 (br s, 1 AnaL Calcd for C 17 1
F
3
N
2 0 4 C, 55.44; H, 4. 11; N, 7.61; F, 15.47. Found: C, 55.30; H, 4.08; N, 7.63; F, 15.13.
A mixture of 3-methoxyl-4-[N'-(2-trifluoromethylphenyl)ureidoJphenylacetic acid (353 mng, 0.96 inmol), methyl 4-[(4S)-fiuoro-(2S)-pyrrolidinylinethoxy]benzoate (243 mg, 0.96 inmol), EDC-HCI (221 mg, 1. 15 inmol), HOBt (156 mg, 1. 15 mmol), and Et 3 N (160 nil, 1. 15 inmol) in DMIF (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 CHC1 3 -MeOH (100: 1 to 60: 1, v/v) as eluent to give methyl 4-f (4S)-fiuoro-l1-[3-inethoxy-4-[N -(2-trifluoroinethylphenyl)ureidoI phenylacetyl]- (2.S)-pyrrolidinylmethoxyjbenzoate (570 mng, 98%) as a white foam. 'H-NMR (CDC 3 8 2.05-2.58 (in, 2 3.56-4.21 (series of m, total I11 4.05-4.64 (in, 2 5.23-5.25 and 5.36-5.3 7 (each mn, total 11H), 6.79-6.82 (mn, 2 6.89-7.00 (mn, 2 7. 16-7.20 (in, 2 7.39-7.43 (mn, 1 7.5 1 7.59 (in, 2 7.93-8.02 (in, 4 MIS (ESI) nl/z 604 (Mr+ 1).
To a stirred solution of methyl 4-[(4S)-fluoro-1-[3-inethoxy-4-[N'-(2-trifluoromethyl phenyl)ureidolphenylacetylJ-(2S)-pyrrolidinylmethoxylbenzoate (570 mg, 0.94 minol) in THF nil) 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- I N HCI, and the resulting precipitate was collected under a reduced pressure. The crude solid was recrystallized from MeOH-CHCI,-IPE to give 76 (234 ing, 42%) as a white crystalline powder. MW 589.54 mp 129- 132'C; 'H-NMR (DMSO-4)6 2.23-2.29 (mn, 2 3.54-4.38 (series of s and in, total 8 4.40- 4.61 (in, 2 5.30-5.36 and 5.43-5.49 (each in, total 1 6.72-6.91 (in, 2 7.02-7.08 (in, 2 H), 7.25-7.29 (in, 1 7.59-7.67 (mn, 2 7.8 1-7.99 (mn, 4 8.69-8.70 (in, 1 8.87-8.90 (in, 1 12.67 (br s, I MIS (ESI) ink 589 Anal. Calcd for C9H2 FN 3
O
6 C, 59.08; H, 4.62; N, 7.13. Found: C, 59.22; H, 5. 10; N, 6.58.
WO 01/00206 WO 0100206PCTIUS0OO/8079 Example 71 4+{4S)-fluoro- 1 -[3-metbyl-4-[N '-(2-trifluoromethylphenyl)ureidopbenylacetyl-(2S)pyrrolidinylmethoxyjbenzoic acid 77F3 H e 7 To a stirred solution of tert-butyl 4-amino-3-methylphenylacetate (927 mg, 4.19 inmol) and Et 3 N (645 Ll, 4.63 mmol) in THF (10 ml) was added 2-trilluoromethylphenyl isocyanate (633 j4l, 4.19 mxnol) 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 3methyl-4-IN '-(2-trifluoromethylphenyl)ureido]phenylacetate (1.06 g, 62%) as a white crystal line powder. mp l78-180o2; 'H-NMvR (CDCI 3 8 1.44 9 2.25 3 3.51 2 6.38 (br s, 1 7.12-7.18 (in, 3 7.36-7.37 (in, 1 7.49-7.53 (in, 2 8.13-8.16 (in, 1 H).
To a stirred solution of tert-butyl 3-methyl-4-[N '-(2-trifluoromethylphenyl)ureidoj phenylacetate (1.06 g, 2.60 mmol) in CH 2
CI
2 (10 mlJ) 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,0. The resulting precipitate was collected under a reduced pressure and the crude solid was recrystallized fr-om MeOH-CHCI,-EPE to give 3-inethyl-4-[N'-(2trifluoromethylphenyl)ureido]phenylacetic acid (702 mg, 77%) as a white crystalline powder. mp 262-263*C; 'H-NMR (DMSO-d6) 8 2.24 3 3.48 2 7.03 J 8.3 Hz, I 7.08 1 7.26-7.30 1 7.61-7.69 (mn, 3 7.88 J 8.3 Hz, 1 8.39 1 8.55 1 H), 12.28 (br s, 1 Anal Calcd for C 1 7 1
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-inethyl-4-[N -(2-trifluoromethylphenyl)ureido]phenylacetic acid (359 mg, 1.02 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (258 mg, 1.02 inmol), EDC-HCI (234 mg, 1.22 minol), HOBt (165 mng, 1.22 minol), and Et 3 N (170 pLl, 1.22 inmol) in DMF (5 nil) 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,, and evaporated. The residue was purified by column chromatography on silica-gel with CHCI,-MeOH (100: 1 to 60: 1, v/v) as eluent to give methyl 4-[(4S)-fluoro-l1-[3-methyl-4-[N '-(2-trifluoromethylphenyl)ureidojphenylacetylj- WO 01/00206 WO 0100206PCT/USOO/18079 (2.S)-pyrrolidinylmethoxylbenzoate (612 ing, as a white foam. IH-NMR (CDC 3 8 1.92 and 2.00 (each s, total 3 2.09-2.61 (in, 2 3.564.29 (series of m, total 8 4.45-4.48 and 4.59- 4.64 (each in, total 2 5.24-5.30 and 5.38-5.44 (each m, total 1 6.90-7.14 (in, 5 7.22- 7.53 (mn, 5 7.90-7.92 (mn, I 8.00-8.06 (in, 2 MIS (ESI) m/z 588 (M t To a stirred solution of methyl 4-[(4S)-fluoro-1-13-methyl-4-[N'-(2-trifluoromethylphenyl) ureidojphenylacetylJ-(2ST)-pyrrolidinylmethoxy~benzoate (610 mng, 1.04 mxnol) 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- I N HCl, and the resulting precipitate was collected under a reduced pressure. The crude solid was recrystallized from MeOH-CHCl 3 -IPE to give 77 (186 mg, 3 as a white crystalline powder. MW 573.54 mp 123-126'C; 'H-NMR (DMSO-d 6 8 2.19-2.29 (series of s and in, totalS5 3.64-4.21 (series of m, total 5 4.36-4.60 (in, 2 5.30-5.36 and 5.43-5.49 (each m, total 1 6.97-7.08 (in, 4 7.24-7.28 (mn, 1 H), 7.59-7.68 (in, 3 7. 85-7.90 (in, 3 8.37-8.39 (in, 1 8.54-8.55 (in, 1 12.67 (br s, I H); MIS (ESI) m/z 573 (WI).
Example 72 4-[(4.S)-fluoro-l -[3-methyl-4-[N '-(2-inethylphenyl)ureido]phenylacetylJ-(2S)-pyrrolidinyl] methoxybenzoic acid me H r~e78 A mixture of 3-inethyl-4-[N'-(2-nethylphenyl)ureidojphenylacetic acid (250 mng, 0.84 minol), methyl 4-[(4.S)-fluoro-(2S)-pyrrolidinyllmethoxybenzoate (400 mg, 1.06 mrnol), EDC-HCl (242 mg, 1.26 inmnol) and DMIAP (154 mng, 1.26 minol) in DMF (5 ml) was stiffed 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., the extracts were concentrated in vacua. The residue was chroinatographed on silica gel [50 g, CHCI 3 /MeOH and then was purified on TLC ICHCl3/acetone to give methyl 4+[4S)-fluoro-1-[3methyl-4-[N'-(2-inethylphenyl)ureidojphenylacetylJ-(25)-pyrrolidinyljinethoxybenzoate (342 ing, 76%) as a colorless amorphous solid. IR (KBr) 3356, 2951, 1716, 1651, 1604, 1537, 1252 cmn-'; 'H-NMR (CDCI 3 8 2.07 J 6.6 Hz, 2H), 2.12 311), 2.27 (mn, 2.24 3H), 2.30-2.59 (in, 3.60 (d,J 5.3 Hz, 1H), 3.65-4.23 (in, 3H), 3.87 314), 4.50-4.62 (mn, 1H), 5.31 J= 52.4 Hz, lH), 6.23 (d,J 11.2 Hz, 111), 6.26 11.9 Hz, 111), 6.87-7.27 (in, 8H), 7.54-7.65 (mn, 311), 7.94-8.01 (mn, 2H); MS (FAB) m/z 534 (M4+ Anal. Calcd for C 3 0 11 32
FN
3
O
3 -0.7H4 2 O: C, WO 01/00206 WO 0100206PCT/USOO/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-[(4.S)-fluoro-1-(3 -methyl-4-IN '-(2-methylphenyl)ureido] phenylacetyl]-(2S)-pyrrolidinyljmethoxybenzoate (227 mg, 0.425 nunol) in THE (3.4 ml) was added 0.25 N NaOH (3.4 ml). After stirring at room temperature for 4 days, the mixture was acidified with IN HCI and extracted with CHCI 3 -MeOH The combined extracts were dried over Na 2 SO, and concentrated in vacua. The residue was purified on preparative-TLC [CHC13/MCOH to give 78 (190 mg, 86%) as a colorless amorphous solid. MW 519.56 IR (KBr) 3356, 2974, 1604, 1537, 1454, 1252 'H-NMR (DMSO-d,) 8 2.24 3H), 2.26 3H), 3.60 J 3.7 Hz, 2H), 3.65-4.65 (in, 8H), 5.3 1-5.50 (in, I1H), 6.92-7.18 (in, 711), 7.67-7.92 (in, 411), 8.22-8.32 (mn, 2H); MS (FAB) m/lz 520 Anal. Calcd for C 29 H3 FN 3
O
7 -1.lH- 2 Q: 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 '-(2-chlorophenyl)ureidoJ-3-methylphenylacetylJ-(4S)-fluoro-(2S)-pyrrolidinylI methoxybenzoic acid 1H H Me 79)-ao To a stirred mixture of tert-butyl 4-amino-3-methylphenylacetate (1.00 g, 4.52 mniol), 2chiorophenyl isocyanate (0.55 nil, 4.52 mxnol) in THE (10 ml) was added Et 3 N 13 ml, 0.90 mxnol) 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) ureido]-3-methylphenylacetate (1.57 g, 93%) as a pale yellow powder. mp 104-106 TC 'H- NMR (CDCI,) 8 1.45 9H), 2.28 314), 3.51 211), 6.33 (br, 1H), 6.96 J 7.6 Hz, 111), 7.08 (br, 114), 7. 16-7.30 (in, 4H4), 7.42 (in, 114), 8.2 8.1 Hz, 114).
To a stirred solution of tert-butyl 4-[N'-(2-chlorophenyl)ureidol-3-methylphenylacetate (1.57 g, 4.19 inmol) in CHC1 2 (10 ml]) was added TFA (6 mld) at room temperature. After 4 h stirring, the mixture was concentrated in vacua. The residue was triturated by the addition of water to give 4-[N'-(2-chlorophenyl)ureido]-3-methylphenylacetic acid (1.33 g, 100%/1) as a yellow powder. mp 243-245 TC 'H-NMR (CDC1 3 8 2.24 3H1), 3.47 2H), 6.99-7.08 (in, 3H), 7.28 J 7.6 Hz, lH), 7.44 (dt, J 2.4 Hz, 1H), 7.66 (dd, J 1.9 Hz, 111), 8.13 (dd, J 6.1, 1.7 Hz, IH), 8.61 J= 6.3 Hz, 2H); MS (ESI), m/z 319 (Mr+l1), 321 Calcd for WO 01/00206 WO 0100206PCT/USOOI 18079
C
16 H5C1N 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-f N' -(2-chlorophenyl)ureido]-3-methylphenylacetic acid (252 mng, 0.79 minol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinyllmethoxybenzoate (200 mg, 0.79 mmol), EDC-HCI (227 mg, 1.20 mmol) and DMAP (147 mng, 1.20 minol) in DMIF (5 nil) 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 vacua. The residue was purified on TLC ICHCIacetone to give methyl 4- -(2-chlorophenyl)ureidol-3-methylphenylacetyl] -(4.S)-fluoro-(2S)-pyrrolidiniyl] methoxybenzoate (390 mg, as a colorless amorphous solid. IR (KBr) 3340, 2951, 1712, 1624, 1604, 1533, 1438 'H-NMvR (CDCI,) 8 1.92-2.05 (in, 3H), 2.07-2.63 (in, 2H), 3.61 (d, 2K J 8.8 Hz), 3.70-4.15 (in, 5H1), 4.25-4.67 (in, 2H), 5.26-5.44 (in, I 6.84-8.19 (in, 13H); MS (FAB) m/~z 554 (Mr+ 5 56 To a stirred solution of methyl 4-fl '-(2-chlorophenyl)ureido]-3-methylphenyl acetylJ-(4S)-fluoro-(2S)-pyrrolidinyl~methoxybenzoate (268 ing, 0.484 mmol) in THE (3.8 ml) was added 0.25 N NaQH (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 The combined extracts were dried over Na 2 SO, and concentrated in vacua. The residue was purified on TLC [CHCIVMCOH (10/1)] to give 79 (124 mng, 47%) as a colorless amorphous solid. MW 539.98 JR (KBr) 3346, 2976, 1709, 1685, 1604, 1533, 1439 'H-NMR (DMSO-dJ 8 2.20 3H, one of isomers), 2.24 3K, one of isomers), 2.30 (in, 11H), 3.60 2H), 3.71-4.62 (in, 6H), 5.30-5.50 (in, 1H), 7.0 1-7.09 (in, 7.28 J =7.8 Hz, 11H), 7.44 J =8.1 Hz, lH), 7.66 J= 8.1 Hz, 11H), 7.87 (d,J 7.1 Hz, 2H), 8.13 J1=7.9 Hz, I1H), 8.62 J1=6.1 Hz, 2H); MIS (FAB) m/z 540 542 (MX+3).
For Na salt: Anal. Calcd for C 28
H,
7
CFN
3 7 Na-0.5EtQH- 1.51- 2 0: C, 56.9 1; HL 5.27; Cl, 5.79; F, 3. 10; N, 6.87. Found: C, 56.60; H, 4.98; Cl, 5.88; F, 3.08; N, 6.52.
Example 74 4-[I1-[4-IN '-(2-broinophenyl)ureido]-3-methylphenylacetyIJ-(4.S)-fluoro-(2.S)-pyrrolidinylj methoxybenzoic acid Br r1 r" Me to a stirred mixture of tert-butyl 4-ainino-3-inethylphenylacetate (780 mg, 3.30 innol), 2-bromo WO 01/00206 WO 0100206PCT/USOO/18079 phenyl isocyanate (0.41 ml, 3.30 mnmol) in THE (7 ml) was added Et 3 N (0.092 ml, 0.66 minol) at room temperature. After 3 h stirring, the reaction midxture was concentrated in vacuo. The residue was triturated by the addition of n-hexane, to give tert-butyl 4-IN'-(2-bromophenyl) ureidol-3methylphenylacetate (1.57 g, as a pale yellow powder. mp 138-145 OC 'H-NMR
(CDCI
3 5 1.44 9H1), 2.33 3 3.51 2H), 6.90 (dt, J1=9.0, 1.4 Hz, 1H1), 6.98 (br, I1H), 7.18-7.3 1 (in, 411), 7.3 9 (dd, J1=8.1, 2.9 Hz, I 7.44 J 8.0 Hz, I1H), 8.22 J1=8.3 Hz, 2H1); Anal. Calcd for C2,H 2 ,BrNO 3 -0.2H 2 O: C,56.80; H,5.58; N,6.62. Found: C,56.85; H,5.42; N, 6.62.
To a stirred solution of tert-butyl 4-[N'-(2-bromophenyl)ureido]-3-methylphenylacetate (1.27 g, 3.03 minol) in CH 2 C1 2 (10 ml) was added TFA (5 ml) at room temperature. After 1 h stirring, the mixture was concentrated in vacua. The residue was triturated by the addition of water, to give 4-[(N'-(2-bromophenyl)ureidoj-3-methylphenylacetic acid (1.05 g, 95%) as a pale yellow powder. mp 245-248 'C 'H-NMR (CDCI 3 5 2.24 3H), 3.48 2H), 6.96 (dt, J 7.3, 1.5 Hz, 111), 7.02 J1=8.3 Hz, 1H), 7.07 7.32 J= 8.1 Hz, 7.59-7.66 (in, 2H1), 8.44 I1H), 8.62 I1H); MIS (ESI), ni/z 3 63 (Mr+ 365 Anal. Calcd for
C
16
H
1 5 BrN 2 QgO0.7H 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)ureidoj-3-inethylphenylacetic acid (287 mng, 0.79 mmol), methyl 4-[(4S)-fluoro-(2.S)-pyrrolidinyllinethoxybenzoate (200 ing, 0.79 minol), EDC-HCl (228 mng, 1.20 iniol), HOBT (160 mg, 1. 19 mniol) and Et 3 N 55 ml, 3.95 inmol) 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 vacua. The residue was purified on TLC [CHCl,/acetone (10/ to give methyl 1-[4-[N'-(2-chlorophenyl)ureido]-3-inethylphenylacetyl]-4-fluoro-2pyrrolidinyllinethoxybenzoate (440 ing, 93%) as a colorless amorphous solid. 'H-NMR (CDC1 3 8 1.90 andl.97 (eachs, 3H, ainide isomers), 2.05-2.62 (in, 2H), 3.58 J= 8.1 Hz, 111), 3.77 (in, lH), 3.86 and 3.89 (eachs, 3H, amide isomers), 3.92-4.64 (mn, 5H), 5.24-5.42 (in, 1H), 6.83-7.23 (in, 611), 7.4 1-7.62 (in, 4H), 7.86-8.09 (in, 3H); MIS (ESI), m/z 598 600 To a stirred solution of methyl 4-[1-[4-[N'-(2-broinophenyl)ureidoJ-3-methylphenyl acetyl]-(45)-fluoro-(2S)-pyrrolidinyl]inethoxybenzoate (440 mng, 0.74 minol) in THE (6.0 ml), 0.25 WO 01/00206 WO (1100206PCTUSOOI18079 N NaQH (6.0 ml) was added. After stirring at room temperature for 1 days, the mixture was acidified with 1 N HCl and extracted with CHCI 3 -MeOH The combined extracts were dried over Na 2
SO
4 and concentrated in vacua. The residue was purified on TLC [CHCI)IMeOH (10/1)] to give 80 (229 mg, as a colorless amorphous solid. MW 584.44 IR (KBr) 3325, 2972, 1709, 1604, 1529, 1252 'H-NMvR (DMSO-d 6 8 2.25 3H), 2.31 (in, 1H), 3.17 3.60 J 4.7 Hz, 2H), 3.83-4.67 (in, 5H), 5.3 1-5.51 (in, 6.97 J= 7.3 Hz, 1H), 7.02-7.09 (in, 7.33 J= 8.0 Hz, 111), 7.61 J= 7.8 Hz, 1H), 7.64 J= 8.3 Hz, 111), 7.87 J =8.3 Hz, 2H), 7.90 J 8.8 Hz, 111), 8.44-8.65 (in, 2H4); MIS (ESI), m/z 584 (NC+ 586 Anal.
Caled for CuHl- 27 BrFN 3
O
7 70.4H 2 O: C, 56.84; H, 4.74; Br, 13.5 1; F, 3.2 1; N, 7. 10. Found: C, 56.9 1; H, 4.93; Br, 13.23; F, 3.15; N, 6.88. For Na salt of 80 Anal. Calcd for C~gH 2 ,BrFN 3 O-Na- 1.81- 2 0: C, 52.64; H, 4.67; Br, 12.5 1; F, 2.97; N, 6.58. Found: C, 53.04; H, 4.67; Br, 12.95; F, 3.28; N, 6.11.
Example 1-[4-[N'-(2-chlorophenyl)ureido] -3-methylphenylacetyl] -(2.S)-pyrrolidinyl]methoxybenzoic acid N N C~n o-Q(--CaaH H e81 To a stirred solution of methyl 4-(1-tert-butoxycarbonyl-(2.S)-pyrrolidinyl)methoxybenzoate (2.0 g, 5.9 mmol) in EtOH (10.0 mil) was added coned HCl (3.0 nil) at 0 The reaction mixture was stirred at room temperature for 4.0 hr. The mixture was concentrated in vacua. The resulting solid was collected and washed with EtOH-Et 2 O to give methyl 4-(2S-pyfrolidinyl)metioxybenzoate HCI salt (1.4 g, 87%) as a white crystalline solid. 'H-NMR (CDCI 3 8 1.90-2.25 (in, 4H), 3.25-3.45 (in, 2H), 3.88 3H), 3.90-4.00 (in, 1H), 4.25-4.45 (in, 2H), 6.96 J 8.5 Hz, 2H), 7.95 J Hz, 2H).
To a stirred solution of methyl 4-[(2S)-pyrrolidinyl]methoxybenzoate HC1 salt (135 mg, 0.5 iniol), 4-[AP-(2-chlorophenyl)uredioj-3-inethylphenylacetic acid (159 mg, 0.5 inmol), HOBt (68 mng, minol), and triethylamine (278 ml, 2.0 mxnol) in THE (5.0 nml) and MeCN (5.0 ml) was added EDC-HCl (144 ing, 0.75 minol) at 0 0 C. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacua. 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 vacua. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc( 1:2 ,vtv) as eluent to give methyl 441 -14-fN'-(2-chlorophenyl)ureidoJ -3-methylphenylacetylJ-2-pyfrolidinyl] inethoxybenzoate (220 mg, as a colorless oil. 'H-NMR (CDCI,) 8 1.91 and 1.97 (each s, WO 01/00206 WO 0100206PCT/USOO/18079 total 311), 2.00-2.20 (in, 4H), 3.55-3.65 (in, 4H1), 3.87 and 3.89(each s, total 3H),4.10-4.20 (m,21-1), 4.51 (in, 11H), 6.86-7.04 (im, 6H), 7.20-7.53 (mn, 411), 7.89-8.0 1 (in, 211), 8.22 J 8.3 Hz, 111).
To a stirred solution of methyl 4-[Il-[4-[N'-(2-chlorophenyl)ureidoJ-3-methylphenylacetylJ-2Spyrrolidinyl]methoxybenzoate (220 mg, 0.41 inmol) in THF (8.0 ml) and MeOH (4.0 ml) was added IN NaOH (0.8m1, 0.8 minol). The mixture was stirred at 70 'C for 24 hr. The mixture was concentrated in vacua, water was added thereto, and neutralized with IN HCI. The resulting solid was collected, washed with water, and dried in vacua to give 81 (220 mg, quant) as a white crystalline solid. MW 521.99 mp 122-124*C IiR (KBr) 3340, 1710, 1685, 1604, 1533, 1511, 1438 'H-NMR (DMSO-d 6 6 1.81-2. 11 (in, 4H), 2.18 and 2.20 (each s, total 311), 3.45-3.80 (in, 411), 3.95-4.05 (in, 111), 4.124.20 (in, 1H), 4.21-4.31 (in, 111), 6.99-7.06 (in, 511), 7.26-7.30 (in, IH), 7.44 J 7.8 Hz, 111), 7.62-7.64 (in, 11H), 7.85 -7.90 (in, 211), 8.13 J 6.8 Hz, 111), 8.60-8.62 (in, MIS (FAB) m/~z 522 Anal. caled for CUHUsN 3 OCl0.2H- 2 O: C, 63.99; H, 5.45; N, 7.99. Found: C, 63.90; H, 5.40; N, 7.72.
Example 76 1-[4-[N'-(2-bromophenyl)ureidoJ-3-methylphenylacetyl]-(2S)-pyrrolidinyInmethoxybenzoic acid N N4: 1 O-O-C OOH r H 1 e82 To a stirred solution of methyl [(2S)-pyrrolidinyl~methoxybenzoate HCI salt (135 ing, 0.5 minol), 4-[Iv-(2-bromophenyl)uredioJ-3-methylphenylacetic acid (181 mng, 0.5 mmol), HOBt (68 mg, inmol), and tniethylamine (278 ml, 2.0 inmol) in THE (5.0 ml) and MeCN (5.0 ml) was added EDC-HCl (144 mng, 0.75 minol) at 0 The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacua. 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 vacua. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc v/v) as eluent to give methyl 441 -[4-[N'..(2-bromophenyl)ureidoJ-3-methylphenylacetyll-(2S)pyrrolidinyljinethoxybenzoate (290 ing, quant) as a colorless oil. 1 1-NMR (CDC 3 8 1.95 and 2.01 (each s, total 311), 2.00-2.20 (in, 414), 3.50-3.65 411), 3.87 and 3.89 (each s, total 311), 4.10-4.20 (in, 211), 4.50 (in, 111), 6.85-7.06 (mn, 611), 7.24-7.28 (in, 111), 7.40-7.44 (in, 311), 7.89- 8.16 (in, 211), 8.17-8.18 (in, 111).
To a stirred solution of methyl 4-[l.14-[N-(2-bromophenyl)ureidoj-3-inethylphenylacetyll-2Spyrrolidinyllmethoxybenzoate (290 ing, 0.5 mmuol) in THE (8.0 mil) and MeOH (4.0 ml) was added WO 01/00206 WO 0100206PCT/USUO/18079 IN NaOH (1.0 ml, 1 .0 mxnol). The mixture was stirred at 70 'C for 24 hr. The mixture was concentrated in vacua, water was added thereto, and neutralized with IN HCL The resulting solid was collected, washed with water, and dried in vacua to give 82 (240 mg, 85%) as a white crystalline solid. MW 566.44 mp 125-130 0 C; IR(KBr) 3340, 1604, 1529, 1434 'H-NMR (DMSQ-d) 8 1.80-2. 10 (in, 414), 2.18 and 2.20 (each s, total 3H), 3.45-3.80 (in, 4H), 3.95-4.05 (in, 111), 4.15-4.20 (in, IH), 4.25-4-30 (mn, lH), 6.94-7.06 (in, 5H), 7.30-7.34 (in, 7.59-7.62 (mn, 2H), 7.85-7.90 (in, 2H), 8.01 J= 8.1 Hz, 1H), 8.44 1H), 8.62 IlH); MS (FAB) m/z 566 (M 4 Anal. calcd for C~sH2N 3
O
5 Br-0.5H 2 O: C, 58.44; H, 5.08; N, 7.30. Found: C, 58.57; H, 4.99; N, 7.18.
Example 77 1-[3-methyl-4-[N '-2-methylphenyl)ureidolphenylacetylJ-(2.S)-pyrrolidinylmethoxylbenzoic acid A mixture of 3-methyl-4-[N'-(2-methylphenyl)ureidojphenylacetic acid (438 mng, 1.47 mmol), methyl 4-[(2S)-pyrrolidinylmethoxylbenzoate (420 mng, 1.79 minol), EDCHCI (410 mg, 2.14 inmol), HOBt (228 mng, 1.69 minol) 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 NaSO 4 and evaporated. The residue was purified by column chromatography on silica-gel with CHC1 3 -MeOH (50:1 to 25: 1, v/v) as eluent to give methyl 4-11 -13-methyl-4-[N '-(2-methylphenyl)ureidojphenylacetylJ-(2S)-pyrrolidinylmethoxy benzoate (760 mg, quant.) as a white foam. 'H-NMR (CDCI 3 5 1.89 3 1.94-2. 14 (in, 4 H), 2.16 3 3.50-3.69 (in, 4 3.87 3 4.09-4.17 (in, 2 4.42-4.45 (in, 1 6.85-7.02 (in, 6 7.10-7. 16 (in, 3 7.51-7.53 (in, 1 7.62-7.64 (in, 1 7.91-7.94 (in, 2 MS (FAB) niz 516 (M t To a stirred solution of methyl 4-[1-[3-methyl-4-[N'-(2-inethylphenyl)ureidoJ phenylacetylj-(2S)-pyrrolidinylmethoxy]benzoate (420 mg, 0.71 mmol) in THF (7 ml) was added N NaOH (7 ml) and the reaction mixture was heated under refiux for 2 hr. After cooled to room temperature, the mixture was poured into ice-lI N HCl and the resulting precipitate was collected under a reduced pressure. The crude solid was purified by recrystallization from CHCI 3 [PE to give 83 (526 mng, as a white crystalline powder. MW 501.57 mp 191-193 0 C; 'H- NMR (DMSO-d) 8 1.87-2. 10 (mn, 4 2.20 3 2.26 3 3.44-3.79 (series of mn, total 4 3.99-4.45 (series of mn, total 3 6.91-7.17 (series of m, total 7 7.66-7.68 (in, 1 7.80- WO 01/00206 WO 0100206PCT/USOO/18079 7.90 (in, 3 8.19-8.21 (mn, 2 12.62 (br s, 1 MS (FAB) nI/z 502 Ana. Calcd for
C
2
,H
31
N
3 0 3 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-[(4.S)-fluoro-1 -[4-[N'-(2-methoxyphenyl)ureidol-3-methylphenylacetyl]-(2S)-pyrrolidiny methoxy]benzoic acid H Ae 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 THE (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 small volume in vacuo, and hexane was added to the residue to give precipitate, which was collected by filtration to give tert-butyl -(2-methoxyphenyl)ureido]-3-methylphenylacetate (1.74 g, 76%) as a white crystalline material, nip 157-158 'H-NMvR (CDCI 3 8 1.46 9 H), 2.30 3 3.50 2 3.76 3 6.43 (s,1I 6.83 (br d, J =8.4 Hz, 1 6.95 (br d, J Hz, 1 6.98-6.99 (in, 2 7.13 (brs, 1 7.23 (br s, 1 7.48 J =8.8 Hz, 1 8.14 J 8.4 Hz, 1 MS (ESI) m/z 371 (MW+H).
To a stirred solution of tert-butyl -(2-methoxyphenyl)ureidoj-3-methylphenylacetate (1.32 g, 3.56 mmol) in CH 2
CI
2 (15 ml) was added trifluoroacetic acid (10 ml), and the resulting midxture 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 Et0Hfhexane to give -(2-methoxyphenyl)ureido]-3-methylphenylacetic acid (932 mg, 83%/) as a white powder. mp 260-264 'H-NMvR (CD 3 OD) 8 2.30 3 3.55 2 4.87 3 6.87-6.92 (in, 2 6.97-6.99 (in, 2 7.10-7.24 (in, 2 7.53-7.58 (in, 1 8.04 J' 7.2 Hz, I1H);MNS (EST) m/z314 To a stirred solution of 4-[N'-(2-methoxyphenyl)ureidoJ-3-methylphenylacetic acid (336 mng, 1.07 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (271 mg, 1.07 mniol) and NNdimethylaminopyridine (130 mg, 1.07 nunol) in DMF (10 nil) was added EDC-HCI (226 mng, 1. 18 mxnol) at Mt and the resulting mixture was stirred for 20 h. The mixture was poured into IN-HCI aq. and extracted with EtOAc. The organic layer was washed with brine, drying over anhiydrous NaSO 4 then concentrated in vacuo. The residue was chroniatographed on silica gel with CHCI,- WO 01/00206 WO 0100206PCT/IJSOO/18079 MeOH (10 as eluent to give methyl 4-[(4.S)-fluoro-1-[4-[N'-(2-methoxyphenyl)ureidoj-3methylphenylacetyl]-(2S)-pyrrolidinylmethoxyjbenzoate (583 mg, 99%b) as a colorless amorphous solid. 'H-NMR (CDCI 3 mixture of rotamars, 8 2.05 and 2.12 total 3 2.05-2.61 (in, 2 H), 3.55-4.73 (series of mn, 13 4.51-4.66 (in, 2 5.26-5.40 (mn, 1 6.72-7.01 (series of m, 8 H), 7.38-8.13 (series of mn, 3 MS (ESI) m/z 550 (Mr+H).
To a stirred solution of methyl 4-[(4S)-fluoro--[4-N'-(2-methoxyphenyl)ureido]-3inethylphenylacetylJ -(2S)-pyrrolidinylinethoxylbnzoate (557 mg, 1.0 1 inmol) in MeOH-THF (1: 1, 10 ml) was added L.ON-NaOH aq. (4.05 ml, 4.05 mmol) at rt, and the resulting mixture was heated at 60'C for 2 h. The mixture was poured into lN-HCl aq. and extracted with EtOAc. The organic layer was washed with brine, drying over anhydrous Na 2 SOI, then concentrated in vacuo.
The residue was chromatographed on silica gel with CHCl 3 -MeOH (10 as eluent to give 84 (492 mg, 9 as a colorless amorphous solid. MW 535.56 'H-NMR (CD 3 OD), mixture of rotamars, 8 2.96 3 2.11-2.45 (in, 2 3.64-4.15 (series of m, 5 3.91 3 4.41-4.45 (in, 1 4.52-4.61 (in, 1 5.25-5.38 (in, 1 6.84-7. 10 (series of in, 7 7.54-7.58 (mn, 1 H), 7.93 J 8.8 Hz, 2 8.02 J 8.8 Hz, 2 MIS (ESI) m/z 536 (M 4 538 (MW+Na*).
Example 79 4-II(4S)-fluoro-lI-[4-[N' -(2-methoxyphenyl)ureido~phenylacetylj-(2.S)-pyrro-lidinylmethoxy] benzoic acid M Oe To a stirred solution of ethyl 4-amino-3-inethylphenylacetate (1.32 g, 7.37 nunol) and triethylamine (205 ml, 1.47 iniol) in THE (20 ml) was added 2-methoxyphenyl isocyanate (980 ml, 7.37 inmol), and the resulting mixture was stirred for 23 h. The mixture was concentrated to a small volume in vacua and hexane was added to the residue to give precipitate, which was collected to give ethyl 4-[N'-(2-methoxyphenyl)ureidojphenylacetate (2.44 g, quant.) as a white crystalline material. mp 107-109 0 C; 'H-NMR (CDCI 3 8 1.26 J 7.1 Hz, 3 3.56 3 H), 3.79 3 4.15 J 7.1 Hz, 2 6.82-6.85 (in, I 6.91-7.00 (in, 2 7.08 1 7.17 J 8.5 Hz, 2 7.27 J 8.6 Hz, 2 7.33 1 8.07-8. 10 (in, 1 MIS (ESI) m/z 329
(M
4 To a stirred solution of ethyl 4-[N'-(2-inethoxypheniyl)ureidojphenylacetate (2.22 g, 6.78 WO 01/00206 WO 0100206PCT/USOO/18079 mmol) in MeOH (30 ml) was added 1.0 M-NaOH aq. (10.2 ml, 10.2 rniol), and the resulting mixture was stirred overnight. 1N-HCI 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)ureidoJ phenylacetic acid as a white powder (1.87 g, mp 165-168 'H-NMR (CD 3 OD) 8 2.30 (s, 3 3.55 2 4.87 3 6.87-6.92 (in, 2 6.97-6.99 (in, 2 7. 10-7.24 (in, 2 7.53- 7.58 (in, 1 8.04 J 7.2 Hz, 1 MS (ESI) m/~z 300 To a stirred solution of 4-[N'-(2-methoxyphenyl)ureidolphenylacetic acid (353 mg, 1. 18 inmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinylmethoxy]benzoate (298 mg, 1. 18 nunol) and NNdimethylaminopyridine (144 mg, 1. 18 minol) in DMF (10 ml) was added EDC-HCI (226 mg, 1. 18 minol) at rt, and the resulting mixture was stirred for 22 h. The mixture was poured into IlN-HCl aq. and extracted with EtOAc. The organic layer was washed with brine, drying over anhydrous NaSO,, then concentrated in vacuo. The residue was chromatographed on silica gel with CHC1 3 MeOH (10 to give methyl 4-I(4.S)-fluoro l-[4-[N'-(2-methoxyphenyl)ureidojphenylacetylJ-(2S)pyrrolidinytmethoxyjbenzoate (594 mg, 94%) as a colorless amorphous solid. 'H-NMR (CDC1 3 mixture of rotamnars, 8 2.05-2.58 (series of in, 2 3.55-4.25 (series of mn, 5 3.77 3 H), 3.87-3.90 (in, 3 4.50-4.63 (in, 2 5.23-5.37 (in, 1 6.81-6.84 (in, 1 6.91-6.99 (in, 4 7.09-7.12 (in, 2 7.18-7.26 (in, 2 7.45-7.53 (in, 2 7.91-8.03 (in, 2 8.10-8.12 (mn, 1 MS (ESI) na/z 536 (MX+H).
To a stirred solution of methyl 4-[(4S)-fluoro-1-[4-[N'-(2-methoxyphenyl)ureido] phenylacetyl]-(2.S)-pyrrolidinylinethoxy]benzoate (568 mg, 1.06 mnnol) in MeOH-THF (1 mld) was added L.ON-NaOH aq. (4.24 ml, 4.24 minol) at Mi and the resulting mixture was heated at for I h. The mixture was poured into IN-HCI aq. and extracted with EtOAc. The organic layer was washed with brine, drying over anhydrous NaSO 4 then concentrated in vacuo. The residue was chroinatographed on silica gel with CHCI,-MeOH as eluent to give 85 (516 mg, 93%) as a colorless amorphous solid. MW 521.54 'H-NMR (CD 3 OD), mixture of rotamars, 8 2.12-2.46 (in, 2 3.65-4.19 (series of m, 5 3.88 3 4.42-4.45 (in, 1 4.52-4.62 (in, 1 5.24-5.39 (in, 1 6.85-6.91 (in, I 6.94-7.03 (series of m, 4 7.14-7.19 (in, 2 H), 7.3 5-7.40 (in, 2 7.92-7.96 (in, 2 8.02-8.04 (in, 1 MS (ESI) m/z 521 544 (M*+Na 1 Example 4-[(4S)-fluoro- l-[4-[N'-(2-methoxyphenyl)ureido]-3-nethoxyphenylacetyl]-(2S)-pyrrolidinyI WO 01/00206 WO 0100206PCT/USOO/1 8079 methoxy~benzoic acid CI 86 To a stirred solution of tert-butyl 4-amino-3-methoxylphenylacetate (1.41 g, 5.94 mmol) and triethylamine (165 ml, 1. 19 numol) in THIF (20 nil) was added 2-methoxyphenyl isocyanate (790 rid, 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)ureidoj-3-methoxylphenylacetate (2.06 g, as a white crystalline material. mp 132-134 'H-NMR (CDC1 3 8 1.46 9 3.50 2 3.87 3 3.88 3 6.84 I 6.87-6.90 (in, 2 6.98-7.03 (in, 2 7.12 1 H), 7.16 1 8.06 J =8.4 Hz, 1 8.13 (dd, J 2.0 Hz, I MIS (ESI) m/z 387 (Mr+H).
To a stirred solution of tert-butyl 4-[N'-(2-inethoxyphenyl)ureido]-3-methoxylphenylacetate (2.01 g, 5.20 inmol) in CHIC1 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 vacu.
Water was added to the residue to give precipitate, which was collected by filtration. The crude solid was recrystallized from EtOHlhexane to give 4-[N'-(2-inethoxyphenyl)ureidoJ-3-methoxy phenylacetic acid as white powder (1.40 g, mp 182-185 IH-NMR (CD 3 OD) 8 3.55 2 3.88 3 3.89 3 6.80-6.99 (in, 5 7.94 J =8.4 Hz, 1 8.00 J 7.2 Hz, 1 MS (ESI) m/~z 330 To a stirred solution of 4-[N'-(2-methoxyphenyl)ureidoJ-3-methoxyphenylacetic acid (353 ing, 1.07 nunol), methyl 4-[(4S)-fluoro-(2ST)-pyrrolidinylmethoxy]benzoate (271 mg, 1.07 minol) and N,N-dimethylaininopyridine (131 mg, 1.07 minol) in DMF (10 ml]) was added EDCHC1 (224 mug, 1. 18 mniol) at rt, and the resulting mixture was stiffed for 14 h. The mixture was poured into IN- HCI aq. and extracted with EtOAc. The organic layer was washed with brine, drying over anhydrous Na 2 SOI, then concentrated in vacua. The residue was chromatographed on silica gel with CHCI 3 -MeOH (10: 1) as eluent to give methyl 4-[(4S)-fluoro-1-[4-fN'-(2-methoxyphenyl) ureido]-3-methoxyphenylacetylj-(2S)-pyrrolidinylmethoxylbenzoate (372 mug, 6 as a colorless amorphous solid. 'H-NMR (CDC 3 mixture of rotamnars, 8 2.04-2.57 (series of in, 2 3.58-4.18 (series of in, 5 3.79 and 3.83 total 3 3.86 3 3.87 3 4.51-4.63 (in, 2 H), 5.22-5.36 (in, 1 6.80-6.89 (in, 3 6.94-7.03 (in, 4 7.15-7.25 (in, 2 7.94-8.01 (in, 2 WO 01/00206 WO 0100206PCT/USOO/18079 8.04-8.11 (in, 2 MS (ESI) m/z 566 (M 4 To a stirred solution of methyl 4-[(4S)-fluoro-1-[4-[N'-(2-methoxyphenyl)ureido]-3-methoxy phenyl-aceyl]-(2S)-pyrrolidinylmethoxylbenzoate (356 mg, 0.63 mmol) in MeOH-THF (1 1, ml) was added L.ON-NaOH aq. (1.88 ml, 1.88 mmol) at rt, and the resulting mixture was heated at 60 0 C for 2 h. The mixture was poured into 1N-HCI aq. and extractedhwith EtOAc. The organic layer was washed with brine, drying over anhydrous NaSO,, then concentrated in vacuo. The residue was chromatographed on silica gel with CHCI 3 -MeOH (10: 1) as eluent to give 86 (335 mg, as a colorless amorphous solid. MW 551.56 'H-NMvR (CD 3 OD), mixture of rotamars, 8 2. 14-2.48 (in, 2 3.69-4.20 (series of m, 5 3.88 3 3.89 3 4.464.57 (in, 2 H), 5.27-5.41 (in, 1 6.79-7.04 (in, 7 7.90-8.02 (in, 4 MS (ESI) m/z 552 (Mr+H).
Example 81 441 -[4-IN '-(2,6-dichloropheny1)ureidolphenylacety1J-(4S)-fluoro-(2.S)-pyrrolidinyllmethoxy benzoic acid
F
N-Cn o-Cj-COOH 1 87 To a mixture of ethyl 4-aminophenylacetate (1.62 g, 9.04 nunol) and 2,6-dichlorophenyl isocyanate (1.70 g, 9.04 inmol) in THF (40 ml) was added Et 3 N (0.25 ml, 1.81 minol) 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)ureidoJ phenyl acetate (3.19 g, as a colorless powder. mp 168-170 'C 'H-NMR (CDC 3 8 1.25 J= 7.1 Hz, 314), 3.56 2H), 4.14 J= 7.1 Hz, 2H), 6.50 (br, 1H), 6.67 (br, 1H4), 7. 12-7.52 (in, 7H).
To a stirred solution of ethyl 4-[N'-(2,6-dichlorophenyl)ureidojphenylacetate (3.19 g, 8.69 mmol) in TI-F (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-dichloropheniyl)ureidolphenylacetic acid (2.44 g, as colorless powder. mp 262-263 TC 'H-NMR (DMS0-l 6 8 3.48 2H), 7.14 J 8.3 Hz, 2H), 7.31 J 8.3 Hz, lH), 7.37 J 8.3 Hz, 2H), 7.52 J 8.0 Hz, 214), 8.18 IM, 8.90 IM), 12.22 (br, 1H), MIS (ESI) m/z 339 341 343 A mixture of 4-[N '-(2,6-dichlorophenyl)ureidolphenylacetic acid (268 ing, 0.79 mmol), methyl 4- WO 01/00206 WO 0100206PCTIJSOOI18079 [(2S,4.S)-4-fluoro-2-pyrrolidinyllmethoxybenzoate (200 mg, 0.79 nunol), EDCQHC1 (227 mg, 1. 19 mmol), H-OBT (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 extracts were concentrated in vacuo. The residue was purified on TLC [CHCI]MeOH to give tmethyl 44 f-(2,6-dichlorophenyl)ureidoJphenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinyl] methoxybenzoate (465 mg, 100%) as a colorless amorphous solid. 'H-NMR (CDCI 3 8 2.05-2. 57 (in, 2M), 3.60 2H, J 3.4 Hz), 3.64-3.84 (mn, 2H), 3.88 and 3.89 (each s, 3H, amide isomers), 3.92-4.63 (in, 3H), 5.22-5.38 (mn, 1H), 6.87 and 6.89 (each d, each J 7.9 Hz, 2H, amnide isomers), 7.01-7. 17 (in, 6H), 7.28 (in, 2H), 7.36 (br, lH), 7.92 J= 8.8 Hz, lH), 7.79 J= 8.8 Hz, IM); MS (ESI) in/z 574 576 578 To a solution of methyl 4-t(2S,4S)-1-14-[N -(2,6-diclorophenyl)ureidojphenylacetyl]-4-fluoro-2pyrrolidinyllmethoxybenzoate (465 mg, 0.809 inmol) in THF (40 ml), 0.25 N NaOH (40 ml) was added. After stirring at room temperature for 11I h, the mixture was acidified with 1 N HC1 and extracted with CHCI,-MeOH The combined extracts were dried over NaSO 4 and concentrated in vacuo. The residue was purified on TLC ICHCI 3 /MeOH to give 87 (340 mg, 75%) as a colorless powder. MW 560.40 mp 168-172 *C IR (KBr) 3340, 1711, 1685, 1604, 1240, 773 'H-NMvR (DMS0-dL) 8 2.22-2.30 (in, 2H), 3.61 J =7.4 Hz, 2H), 3.70- 4.75 (mn, 6H), 5.30-5.49 (in, IJH), 7.02-7. 18 (in, 511), 7.28-7.41 (in, 4H), 7.52 (dd, J= 8.0, 2.9 Hz, 2H), 7.86 (in, 2H), 8.29 (br, lH), 9.01 (br, lH), 12.66 (br, IN); MS (ESI) nik 560 562 564 (M t Anal. Calcd for C.
2 7H 24
C
2
FN
3 0,0.5H 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 -14-[N '(2,6-dichlorophenyl)uridoJ-3-nethoxyphenylacetyl-(4S)-fluoro-(2S)-pyrolidinyl inethoxybenzoic acid N' N -CJ-OOH 1 88 To a mixture of tert-butyl 4-amino-3-inethoxyphenylacetate (2.15 g, 9.04 iniol), 2,6dichlorophenyl isocyanate (1.70 g, 9.04 minol) in THFf (40 ml) was added Et 3 N (0.25 ml, 9.04 inmol) at room temperature. After 18 h stirring, the reaction mixture was concentrated in vacuo.
The residue was triturated by the addition of n-hexane, to give terI-butyl 4-[N WO 01/00206 WO 41100206PCTUSOOI18079 dichlorophenyl)ureidoj-3-methoxyphenylacetate (2.27 g, as a colorless powder. mp 177-181 0 C 'H-NMR (CDCI 3 8 1.43 9H), 3.74 2H1), 3.83 3H), 6.34 6.81 1H), 6.84 J= 8.3 Hz, 114), 7.06 (br, 11H), 7.27 J= 8.1 Hz, 11H), 7.39 8.1 Hz, 2H), 8.04 J 8.3 Hz, I M.
To a stirred solution of tert-butyl 4-IN '-(2,6-dichlorophenyl)ureido]-3-methoxyphenylacetate (2.27 g, 5.34 mmol) in CH 2
CI
2 (50 ml) was added TEA (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, as a colorless powder. mp, 246-249 0 C 'H-NN4R (DMSO-dA 8 3.49 211), 3.88 3H), 6.75 J 8.3 Hz, I1H), 6.93 IHM, 7.30 J =7.8 Hz, 7.52 J =8.0 Hz, 2H), 7.97 J =8.0 Hz, 111), 8.40 1H), 8.86 1H), 12.23 (br, lH); MIS (ESI) in/z 369 371 373 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 mmol), EDC-HC1 (227 mg, 1. 19 mmol), HOBT (161 mg, 1. 19 inol) and Et 3 N 55 ml, 3.95 mmol) in DMIF (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 NaSO,, the extracts were concentrated in vacua. The residue was chromatographed on silica gel [50 g, CHCl 3 IMeOH(40/ to give methyl '-(2,6-dichloropheryl)ureido]-3-miethoxyphenyI acetylj-(4S)-fluoro-(2S)-pyrrolidinyl~methoxybenizoate (530 mg, 100%) as a colorless amorphous solid. 'H-NMR (CDC1 3 8 2.03-2.62 (in, 2H), 3.61 2H, J 4.7 Hz), 3.62-3.66 (in, 211), 3.73 and 3.77 (each s, 3K, amide isomers), 3.78-3.85 (in, 1H), 3.87 and 3.88 (each s, 3H, arnide isomers), 3.95-4.63 (in, 4H), 5.22-5.36 (in, 1H), 6.82 6.88 J 8.8 Hz, 1H), 6.95 J 8.8 Hz, 211), 7.14-7.25 (in, 1H), 7.38 J= 8.1 Hz, 2H), 7.94-8. 10 (in, 3H); MIS (ESI) "nk 604 606 608 To a solution of methyl 1-[4-[N'-(2,6-dichlorophenyl)ureidol-3-methoxyphenylacetyl]-(4Sj)fluoro-(2S)-pyrrolidinyl~methoxybenzoate (530 mg, 0.78 mmol) in THF (40 ml), 0.25 N NaOH ml) was added. After stirring at room temperature for 11I h, the mixture was acidified with I N HCl and extracted with CHCl 3 -MeOH The combined extracts were dried over Na 2 SO, and concentrated in vacua. The residue was purified on TLC [CHClVMeOH to give 88 (420 mg, 75%) as a colorless amorphous solid. MW 590.43 mp 162-168 *C IR 3346, 2974, 1709, 1604, 1533, 1254 'H-NMR (DMSO-d 6 8 1.98-2.36 (in, 2H), 3.58 2H), 3.78- WO 01/00206 WO 0100206PCT/USOO/18079 3.95 (in, 6H1), 4.024.68 (in, 2H1), 5.3 1-5.50 1H1), 6.71-7.09 (in, 7.31 J =7.8 Hz, 111), 7.53 (dJ= 8.1 Hz, 2H), 7.87 8.1 Hz, 2H), 7.88-8.00(in, 111), 8.30-8.40(in, 1II), 8.89 (s, I1H); MS (ESI) m/z 590 592 594 Anal. Calcd for C 2 gH 26 Cl 2
FN
3 0 6 1l5H 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.
Example 83 4-I(2S,4S)-1 -14-[N '-(2,6-Dichlorophenyl)ureidol-3-methylphenylacetylj-4-fluoro-2-pyrrolidinylj methoxybenzoic acid o-CJ-CooH H H I~e89 To a mnixture of tert-butyl 4-amiino-3-methylphenylacetate (1.88 g, 8.51 mmol), 2,6-dichiorophenyl isocyanate (1.60 g, 8.51 mmol) in TI-F (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 vacua. The residue was triturated by the addition of n-hexane, to give terl-butyl 4-[N'-(2,6-dichlorophenyl)ureidol -3methylphenylacetate (2.58 g, 74%) as a colorless powder. mp 243-244 0 C 'H-NMR (CDCI,) 8 1.45 Cs, 9H), 2.30 3H), 3.49 Cs, 2H), 6.24 211), 7.12-7.16 (in, 3H), 7.35 J 8.3 Hz, 2H), 7.51 J= 7.8 Hz, I H).
To a stirred solution of tert-butyl 4-[N -(2,6-dichlorophenyl)ureido]-3-methylphenylacetate (2.58 g, 6.30 mmol) in CH 2
CI
2 (50 mil) was added TFA (20 ml) at room temperature. After 2 h stirring, the mixture was concentrated in vacua. The residue was triturated by the addition of water, to give 4- [N'-(2,6-dichlorophenyl)ureido]-3-methylphenylaoetic acid (2.12 g, as a colorless powder.
mp 274-283 'C 'H-NMR (DMSO-d6) 8 2.24 3M1, 3.46 2H4), 7.00 J 8.6 Hz, 111), 7.06 111), 7.30 J 7.8 Hz, 111, 7.52 J 8.3 Hz, 211), 7.65 J 8.2 Hz, 111), 8.12 (s, 111), 8.50 Cs, 111), 12.22 (br, 111); MIS (ESI) ,n/z 353 355 357 A mixture of 4-[N'-(2,6-dichlorophenyl)ureidol-3-nethylphenylacetic acid (181 mg, 0.51 mmol), methyl 4-[(2S,4S)-4-fluoro-2-pyrrolidinyllmethoxybenzoate (130 mg, 0.51 mmol), EDC>HC1 (147 mg, 0.77 mmol), HOBT C104 mng, 0.77 mmnol) and Et 3 N (0.35 ml, 2.55 inmol) 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.,
the extracts were concentrated in vacua. The residue was purified on TLC ICHClV/MeOH (20/ to give methyl '-(2,6-dchlorophenyl)ureido]-3 -iethylphenylacetylJ-(4S)-fluoro-(2S)- WO 01/00206 WO 0100206PCTIIJS00II8079 pyrrolidinyllmethoxybenzoate (283 mg, 94%) as a colorless amorphous solid. 'H-NMR (CDC1 3 8 1.95-2.61 (in, 3H), 3.55 (br, 2H), 3.67-3.81 (in, 2H), 3.87 6H), 3.89-4.68 (in, 2H), 5.23-5.43 (in, 1H), 6.81-7. 10 (in, 61D, 7.13-7.43 (in, 3H), 7.56 (br, 1H, one of isomers), 7.73 (br, 1H, one of isomers), 7.89-8.00 (in, 2H); MS (ESI) ilk 588 590 592 To a solution of methyl '-(2,6-dichlorophenyl)ureidol-3-methylphenylacetylj-4S)fluoro-(2.S)-pyrrolidinyl]methoxybenzoate (283 mg, 0.48 mmol) in THF (20 ml), 0.25 N NaOH ml) was added. After stirring at room temperature for I1I 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 NaSO, and concentrated in vacuo. The residue was on TLC [CHCI,/MeOH to give 89 (450 mag, 67%) as a pale brown amorphous solid. MW 574.43 rap 174-180 'C IR (KBr) 3330, 3288, 1711, 1685, 1604, 1512, 1242 cm-1; IH-NMR (DMSO-d) 582.24 (in, 3H), 3.61 2H, J= 6.1 Hz), 3.72-4.68 (in, 7H), 5.30-5.50 (in, 1H), 6.97- 7.20 (in, 414), 7.29-7.68 (in, 5H), 7.87 (in, 2H), 8.10-8.95 (in, 11H), 12.65 (br, lH);l MS (ESI) mnk 574 576 578 Anal. Calcd for C~sH,,Cl 2
FN
3 O,-0.5H 2 O: C, 57.64; H, 4.66; Cl, 12.15, F, 3.26; N, 7.20. Found: C, 57.37; H, 4.44; CI, 12.64; F, 3.23; N, 7.25.
Example 84 4-[1-[3-chloro-4-[N '-(2-inethylphenyl)ureidolphenylacetylj-(4S)-fluoro-(2S)-pyrrolidinylj methoxybenzoic acid
F
Me To a stirred solution of 3-chiorophenylacetic acid (21.76 g, 127.6 mmol) in dichloroethane (100 ml) was added MeOH (15.6 ml, 383 minol) and H 2 SOA (1 ml) at room temperature. After minutes stirring, the mixture was heated at 80 'C for 2 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 extract was concentrated in vacua to give methyl 3chlorophenylacetate (25.4 g, 100%) as a colorless oil. 'H-NMR (CDCl 3 8 3.60 2H), 3.70 (s, 3H), 7.15-7.26 (in, 4H).
To a stirred mixture of methyl 3-chlorophenylacetate (25.4 g, 128 mmol) in H 2 S0 4 (44 ml) was added HN0 3 (5.5 ml, 138 mmnol) at 0 TC. The reaction mixture was gradually raised to room temperature for 4 h. The reaction mixture was poured into ice water and extracted with EtOAc.
WO 01/00206 WO 0100206PCT/USOO/18079 The combined extracts were washed with aq. NaHCO, and brine. After dried over Na 2
SO
4 the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [1 kg, nhexane/EtOAc to give methyl 3-chloro-4-nitrophenylacetate (11.4 g, as a yellow oil.
'H-NMR (GDCI 3 8 3.69 2H), 3.74 3H), 7.33 (dd, J 8.3, 1.5 Hz, 1H), 7.49 J 1.5 Hz, 111), 7.87 J =8.3 Hz, I1H).
A mixture of methyl 3-chloro-4-nitrophenylacetate (10.9 g, 47.5 mmnol), reduced iron powder (8.58 g, 153.6 mmol), AcONa-3H 2 O (6.05 g, 44.5 mmol) and AcOH (17.6 ml) in MeOH/Hi 2 0 (100/400 ml) was heated at 110 0 C for Ilh. After cooled to room temperature, the reaction mixture was filtered through Celite and the filtered cake was washed with MeOH. The combined filtrate were evaporated and extracted with EtOAc. The combined extracts were washed with brine, dried over Na 2 SO, and concentrated in vacuo. The residue was chromatographed on silica gel [150 g, CHC13/EtOAc to give methyl 4-axnino-3-chloropheniylacetate (4.58 g, 48%) as a red oil.
'H-NMR (CDCI)8 3.49 2H), 3.68 3H4), 4.01 (br, 2H), 6.70 J= 7.4 Hz, IM), 6.96 (dd, J 8.1, 2.0 Hz, 1H), 7.17 J =2.0Hz, 111).
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 THE (20 ml) was added Et 3 N 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) ureidojphenylacetate (1.23 g, as a colorless powder. 'H-NMR (CDC1 3 8 2.34 3H), 3.54 2H), 3.68 3H), 6.24 (br, 1H), 6.99 (br, 111), 7.15 (dd, J 8.3, 2.0 Hz, 1H), 7.21-7.31 (in, 7.44 J 7.6 Hz, 1H), 8.20 J 8.5 Hz, 111); MIS (ESI) m/z 333 335 (MC+3).
To a stirred solution of methyl 3-chloro-4-[N'-(2-methylphenyl)ureido]phenylacetate (1.23 g, 3.70 m-mol) 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 HC1 and dried over 60 *C for 2 days under a reduced pressure to give 3-chloro-4-[N'-(2-methylphenyl) ureidoiphenylacetic acid (1.22 g, 100%/) as colorless powder. 'H-NMR (DMS0cl) 8 .26 3H), 3.40 2H), 6.95 J =7.3 Hz, IH), 7. 11 J= 7.6 Hz, 2H), 7.16 J =7.3 Hz, 114), 7.32 (s, lH), 7.76 J 8.0 Hz, lHf), 7.94 (dd, J 9.3, 1.0 Hz, 1H), 8.72 2H); MS (ESI) mnk 319 321 341 (MW+Na).
WO 01/00206 WO 0100206PCT/USOO/18079 A nudxture of 3-chloro-4-[N'-(2-methylphenyl)ureidojphenylacetic acid (319 mg, 1.00 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (253 mg, 1.00 mmnol), EDC-HCI (288 mg, 1.50 mmol), HOBT (203 mg, 1.50 mmnol) and Et 3 N (0.70 ml, 5.00 rnmol) in DMF (4 ml) was stirred at roam 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 purified on TLC [CHCI,/acetone (5/1)1 to give methyl 441 -f3-chloro-4-[N'-(2-methylphenyl)ureidolphenylacetyl]-(4S)-fluor-(2S)pyrrolidinyllmethoxybenzoate (480 mg, 87%) as a colorless amorphous solid. 'H-NMR (CDCI 3 8 2.10-2.60 (in, 2H), 2.29 3H), 3.56 J 6.8 Hz, 1H), 3.71-3.84 (in, IH), 3.87 and 3.89 (each s, 3H, amide isomers), 3.91-4.20 (in, 3H), 4.49-4.60 (in, 2H), 5.32 (dt, J 53.0, 4.2 Hz, IH), 6.80 (br, 1H), 6.89 and 6.95 (each d, J 8.8 Hz, 2H, amide isomers), 7.09-7.26 (in, 6H), 7.50 (di, J 7.3 Hz, I1H), 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, amidde isomers); MS (FAB) m/z 554 556 (MC+3).
To a solution of methyl 4-[I-[3iloro-4-[N-(2-nethylphenyl)ureido]phenylacetyl]-(4S)-fluoro- (2S)-pyrrolidinyllmethoxybenzoate (480 mng, 0.866 minol) 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 HCI. The precipitates were collected, washed with water and dried under a reduced pressure to give 90 (374 mg, as a colorless powder. MW 539.98 IR (KBr) 3354, 3060, 2976, 1709, 1604, 1244 IH-NMR (DMSO-ds) 8 2.27 3H), 2.31 (s, 2H), 3.66 J= 7.2 Hz, 2H), 3.71-4.67 (mn, 5M), 5.32-5.53 (in, 1H), 6.97 J= 7.3 Hz, 1H), 7.04- 7.22 (in, 5H), 7.32 and 7.35 (each d, J= 1.7 Hz, 1H, amide isomers), 7.77 J 7.6 Hz, 1H), 7.87 and 7.90 (each d, J 9.0 Hz, 2HK amnide isomers), 8.01 and 8.03 (each di, J =8.5 Hz, IlH, amnide isomers), 8.57 and 8.59 (each s, 1H, amide isomers), 8.63 and 8.65 (each s, 1H, amide isomers), 12.63 IH); MS (ESI) ,n/z 540 542 Example 4-[1 -[3-chloro-4-IN -(2-chlorophenyl)ureidojphenylacetylj-(4S)-fluoro-(2S)-pyrrolidinylinethoxy benzoic acid N 0O-.Y OOH 9 To a mixture of methyl 4-amino-3-chlorophenylacetate (1.00 g, 5.01 inmol) and 2-chiorophenyl isocyanate (0.60 ml, 5.01 inmol) in TI-F (20 ml) was added Et 3 N 14 ml, 1.00 numol) at room WO 01/00206 WO 0100206PCT/USOOI 18079 temperature. After 1 day stirring, 2-chiorophenyl isocyanate (0.60 ml, 5.01 mmol) was added to the reaction mixture and stirred 17 h. The reaction mixture was concentrated in vacua. The residue was triturated by the addition of n-hexane to give methyl 3-chloro-4-IIN'-(2-chlorophenyl) ureidojphenylacetate (1.35 g, 76%) as a colorless powder. 'H-NMR (CDC1 3 63.58 3H), 3.71 2H), 7.04 (in, 3H), 7.18 (dd, J= 8.5, 2.0 Hz, IM), 7.27-7.39 (in, 3H), 8.07 (in, 2H); MS (ESI) ni/z 3 53 3 55 3 57 To a stirred solution of methyl 3-chloro-4-[N'-(2-chlorophenyl)ureidojphernylacetate (1.35 g, 3.82 inol) in TI-F (30 ml) was added 0.25 N NaOH (30 ml). After stirring at roam temperature for 14 h, the solvent was concentrated in vacua. The residue was triturated by the addition of 1 N HCI and dried at 60 'C for 2 days under a reduced pressure to give 3-chloro-4-[N'-(2-chlorophenyl) ureidoiphenylacetic acid 12 g, as colorless powder. 'H-NN4R (DMSO-d) 8 3.52 211), 7.05 (in, lH), 7.17 J 8.5 Hz, IH), 7.30 J 7.6 Hz, IH), 7.37 1H), 7.46 (dd, J Hz, IM, 7.95 (dd, J 8.3, 1.2 Hz, IM, 8.07 J= 8.3 Hz, lH), 9.00 J =8.0 Hz, 2H); MIS (FAB) mnk 339 341 343 A mixture of 3-chloro-4-[N'-(2-chlorophenyl)ureidoJphenylacetic acid (339 mg, 1.00 inmol), methyl 4-[(2S,4S)-4-fluoro-2-pyrrolidinyl]methoxybenzoate (253 mng, 1.00 mmol), EDC-HCI (288 mg, 1.50 inmol), HOBT (203 mg, 1.50 mniol) and Et 3 N (0.70 ml, 5.00 mmol) in DMF (4 ml) was stirred at roam 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,,
the extracts were concentrated in vacua. The residue was chromatographed on silica gel [50 g, CHCI,/acetone(1 to give methyl 4-[I-[3-chloro-4-[N'-(2-chlorophenyl)ureido]phenylacetyl]- (4S)-fluoro-(2.S)-pyrrolidinyl]inethoxybenzoate (550 mg, 96%) as a colorless amorphous solid.
IH-NMvR (CDCI 3 8 2.14-2.64 (in, 2H), 3.59 J= 11.2 Hz, 2H), 3.78-3.82 (in, IN), 3.86 and 3.89 (each s, 3H, amide isomers), 3.91-4.28 (in, 2H), 4.504.79 (in, 2H), 5.34 and 5.39 (each dt, J 52.5, 4.4 Hz, 1H, amnide isomers), 6.89-6.98 (in, 3H), 7.09-7.13 (in, 2H), 7.22 (dtlJ= 7.3, 2.2 Hz, 7.29 (dd, J 8.1, 2.0 Hz, lH), 7.79 and 7.86 (each s, lH, amide isomers), 7.86-8.03 (in, 4H), 8.11 (dd, J= 8.3, 1.0 Hz, 1H);MNS (FAB) nkz574 576 578 To a solution of methyl 4-[1-[3-chloro-4-[N-(2-chlorophenyl)ureido]phenylacetyl]-(4S)-fluoro- (2S')-pyrrolidinyllmethoxybenzoate (550 mg, 0.957 minol) 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 HCI. The precipitates were collected, washed with water WO 01/00206 WO 0100206PCT/USOO/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 'H-NMR (DMSO-d 6 8 2.25-2.42 (in, 2H), 3.67 J 8.3 Hz, 211), 3.81-4.68 (in, 5H), 5.39 and 5.46 (each d, J 54.4 Hz, 111, anide isomers), 7.04-7. 10 (mn, 3H), 7.18 J 8.3 Hz, 111), 7.31 J 8.3 Hz, 111), 7.33 and 7.37 (each s, IH, amide isomers), 7.47 J 8.1 Hz, 111), 7.88 (dd, J 9.0, 3.2 Hz, 2M1, 7.98 (dd, J Hz, 11I) m, 11H), 8.09 J 8.3 Hz, 111), 8.99 J 2.9 Hz, 1H1), 9.02 111), 12.64 111); MS (ESI) m/z 560 562 564 Anal. Calcd for C 27
H
2 1 2
FN
3 0'0.2H 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 l-[4-[N'-(2-bromophenyl)ureidoJ-3-chlorophenylacetylj-4S)-fluoro-(2S)-pyrrolidinyl inethoxybenzoic acid 92 To a mixture of methyl 4-amiino-3-chlorophenylactate (1.00 g, 5.01 nimol) and 2-bromophenyl isocyanate (0.62 ml, 5.01 inmol) in Ti-F (20 ml) was added Et 3 N 14 ml, 1.00 inmol) at room temperature. After 1 day stirring, 2-broinophenyl isocyanate (0.60 ml, 5.01 mxnol) was added to the reaction mixture and stirred 24 h. The reaction mixture was concentrated in vacua. The residue was triturated by the addition of n-hexane to give methyl 4-[N'-(2-broinophenyl)ureido]-3chlorophenylacetate (1.34 g, 67%) as a colorless powder. 'H-NMR (CDC 3 8 3.58 311), 3.70 (s, 2H), 6.98 3H), 7.19 (dd, J 1.9 Hz, IlH), 7.32 I1H), 7.51 (mn,2H), 8.05 111); MS (ESI) m/z 398 400 402 To a stirred solution of methyl 4-[N'-(2-broinophenyl)ureido]-3-chlorophenylacetate (1.34 g, 3.37 inmol) in TI-F (30 mld) 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 TC for 2 days under a reduced pressure to give 4-IN '-(2-broinophenyl)ureidoj-3chlorophenylacetic acid (1.03 g, 80%) as colorless powder. 'H-NMR (DMS0cl) 8 3.56 211), 7.00 (in, 111), 7.17 (dd, J1=9.0, 1.7 Hz, 111), 7.32-7.40 (in, 2H1), 7.62 (dd, J= 8.0, 1.2 Hz, 111), 7.95 (in, 211, 8.83 9.01 12.41 (br, 111); MS(FAB) m/z 385 (M'+2),386(M+4),388 (Nr+6).
A mixture of 4-[N'-(2-bromophenyl)ureido]-3-chlorophenylacetic acid (384 mng, 1.00 WO 01/00206 PTUO/87 PCT/USOO/18079 mmol), methyl 4-[(4S)-fluoro-(2S)-pyrrolidinyllmethoxybenzoate (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 DMIF (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 NaSO 4 the extracts were concentrated in vacuo. The residue was chromatographed on silica gel g, CHCl,/acetone(l0/l)1 to give methyl -(2-bromophenyl)ureidoJ-3chlorophenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinyljmethoxybenzoate (530 mg, as a colorless amorphous solid. 'H-NMR (CDCI 3 8 2.14-2.63 (in, 2H), 3.58 J= 10.0 Hz, IH), 3.73-3.83 (in, 1H), 3.86 and 3.89 (each s, 3H, amnide isomers), 3.90-4.29 (in, 3H), 4.50-4.69 (in, 2H), 5.33 and 5.37 (each mn, lH amide isomers), 6.88-6.93 3H), 7.11-7.14 (mn 2H), 7.26 (mn, lH), 7.46 J 8.1 Hz, 1H), 7.62-7.78 (mn, 2H), 7.89 and 7.93 (each m, 2H, amide isomers), 8.01 (dd, J 8.8, 1.7 Hz, 2H); MIS (FAB) ink 618 620 622 (M 4 To a solution of methyl l-[4-[N-(2-broinophenyl)ureido-3-chlorophenylacetyl-(4S)-fluoro- (2S)-pyrrolidinyllinethoxybenzoate (530 mg, 0.856 mmol) in TI-F (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 HCI. The mixture was extracted with CHCI 3 IMeOH The combined extracts wevre washed with ice water and brine. After dried over NaSO,, the extracts were concentrated in vacuo. The residue was chroinatographed on silica gel [20 g, CHCljacetone( 0/)-CHC 3 /MeOH( 0/ to give 92 (59 mng, I11%6) as a colorless amorphous solid. NM 604.85 IR (KBr) 3329, 3060, 2976, 1712, 1526, 1435 cm 2 'H-NMIR (DMSO-4) 8 2.3 1 (in, 3.48-4.68 (in, 7H), 5.32-5.53 (mn, 11H), 6.99-7.19 (in, 4H), 7.36 I1H), 7.63 (dd, J 6.7, 1.2 Hz, 1H), 7.86-8.18 (mn, 4H), 8.83 lH), 9.02 1H), 12.67 (br, 1H); MIS (ESI) m/z 604 606 608 Anal. Calcd for C, 7
H
24 BrCIFN 3 O,-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.
Example 87 1-[3-chloro-4-(N '-phenylureido)phenylacetylJ-(4S)-fluoro-(2S)-pyrrolidinylJinethoxybenzoic acid CNyj)oIqOOH To a mixture of methyl 4-amino-3-chlorophenylacetate (1.31 g, 6.56 inmol) and phenyl isocyanate (0.71 ml, 6.56 mmol) in TI-F (20 nil) was added Et 3 N 19 ml, 1.33 inmol) at room temperature.
After 15 h stirring, the reaction mixture was concentrated in vacuo. The residue was triturated by WO 01/00206 WO 0100206PCTIUSOO/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 (CDCI,) 8 3.56 2H), 3.70 3H), 6.70 (in, lH), 7.06 IH), 7.14-7.18 (in, 2H), 7.26 (dd, J 1.9 Hz, 11H), 7.33-7.38 (in, 4H), 8.14 (dd, J= 8.3, 3.0 Hz, 1H),I MS (ESI) ,niz 319 321 (MW+3).
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 HC1 and dried at 60 OC for 2 days under a reduced pressure to give 3-chloro-4-(N'-phenylureido) phenylacetic acid 58 g, as pale brown solid. 'H-NMR (DMS0-cl) 8 3.55 2H), 6.99 J 7.3 Hz, IlH), 7.17 J= 8.3 Hz, 1H), 7.29 J= 7.6 Hz, 2H), 7.36 I1H), 7.46 J= 8.0 Hz, 2H), 8.07 (d,J 8.3 Hz, 1H), 8.28 IHM, 9.37 114), 12.37 (br, 11-).
A mixture of 3-.chloro-4-(N'-phenylureido)phenylacetic acid (305 mg, 1.00 minol), methyl 4- [(2S,4S)-4-fluoro-2-pyrrolidinyl]methoxybenzoate (253 mg, 1.00 minol), EDC-HCI (288 mng, 1.50 mmol), HOBT (203 mng, 1.50 mmol) and Et 3 N (0.70 ml, 5.00 iniol) in DMF (4 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., the extracts mere concentrated in vacuo. The residue was chromatographed on silica gel 130 g, CHCl,/acetone(20/1)J to give methyl I-[3-chloro-4-(N'-phenylureido)phenylacetylJ-(4S)-fluoro- (2S)-pyrrolidinyl]methoxybenzoate (720 mg, 100/6) as a colorless amorphous solid. MS (FAD) ilz 540 542 To a solution of methyl 4-[l -chloro-4-(N'-phenylureido)phenylacetylJ -(4S)-fluoro-(2S)pyrrolidinyljmethoxybenzoate (720 mg, 1.00 minol) in THIF/MeOH (30/30 ml) was added 0.25 N NaGH (30 ml). After stirring at room temperature for 2 h, the reaction mixture was heated at 'C for 22 h. 'After removed the solvent, the resulting residue was acidified with 1 N HCI. The 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 cmn*'; 'H-NMR (DMSO-d) 8 2.25-2.31 (in, 2H), 3.66 J 7.8 Hz, 2H), 3.714.67 (in, 5.31-5.52 (in, lH), 6.99 J 7.3 Hz, 1H), 7.04 and 7.07 (each d, J 8.7 Hz, 2K, amnide isomers), 7.14-7.18 (in, 1H), 7.29 J 7.3 Hz, 2H), 7.35 J 1.7 Hz, 1H), 7.46 J 7.8 Hz, 2H), 7,87 and 7.90 (each d, J 9.0 Hz, 2H, amide isomers), 8.04 and 8.06 (each d, J 8.5 Hz, I H, amide isomers), 8.26 and 8.28 (each s, IH, amnide isomers), 9.36 1Hf), 12.63 IH); MIS (ESI) m/z 526 WO 01/00206 PCT/USOO/18079 528 (M Anal. Calcd for C 2
,H
2 1CIFN30O-0.5H 2 O: 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; CI, 6.66; F, 3.39.
Example 88 4-[1-[3-bromo-4-[N '-(2-methylphenyl)ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrolidinyl ]methoxybenzoic acid Me V O-Q--COOH Me H H rOOH 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
SO
4 (0.5 ml) at room temperature. After 20 minutes stirring, the mixture was heated at 80 °C for 7 h. The reaction mixture was poured into ice water and extracted with CHCI 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 (CDC 3 8 3.60 2H), 3.71 J 1.0 Hz, 3H), 7.18- 7.44 4H).
To a stirred mixture of methyl 3-chlorophenylacetate (10.8 g, 47.1 mmol) in H 2
SO
4 (15.1 ml) was added HNO3 (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 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. The residue was chromatographed on silica gel [500 g, nhexane/EtOAc to give methyl 3-bromo-4-nitrophenylacetate (3.69 g, 29%) as a yellow oil.
'H-NMR (CDCI 3 8 3.68 2H), 3.73 3H), 7.38 (dd, J 8.3, 1.2 Hz, 1H), 7.67 J= 1.3 Hz, 1H), 7.83 J= 8.3 Hz, 1H).
A mixture of methyl 3-bromo-4-nitrophenylacetate (14.8 g, 53.8 mmol), reduced iron powder (9.62 g, 172 mmol), AcONa'3H20 (7.32 g, 53.8 mmol) and AcOH (20.0 ml) in MeOH/H,0 (150/600 ml) was heated at 90 °C for Ih. After cooled to room temperature, the reaction mixture was filtered 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, CHCI 3 /EtOAc to give methyl 4-amino-3-bromophenylacetate (9.01 g, 69%) as a brown oil. 'H-NMR
(CDCI
3 8 3.48 2H), 3.68 3H), 4.05 (br, 2H), 6.69 J 8.3 Hz, 1H), 7.00 (dd, J= 8.1, Hz, 1H), 7.32 J= 2.0 Hz, 1H).
WO 01/00206 WO 0100206PCT/USOO/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 TI-F (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)ureidoI phenylacetate (650 mg, as a pale brown powder. 'H-NMIR (CDC 3 82.34 3M1, 3.53 (s, 2H4), 3.68 3H), 6.18 (br, 11H), 6.96 (br, Il-H), 7.18-7.33 (in, 411), 7.29 J= 4.4 Hz, 111), 7.30 J 7.3 Hz, 11-1), 8.19 J 8.3 Hz, 1H1); MS (ESI) m/z 377 379 (Mr+2).
To a stirred solution of methyl 3-bromo-4-[N '-(2-methylphenyl)ureidojphenylaoetate (650 mg, 1.72 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 0 C for 2 days under a reduced pressure to give 3-bromo-4-IN'-(2methylphenyl)ureido] pheniylacetic acid (1.22 g, 100%/) as colorless powder. 'H-NMR (DMSO-d 6 8 2.26 3H), 3.32 2H), 6.93 (in, 2H1), 7.10-7.17 (in, 4H), 7.76 J =8.1 Hz, 214), 8.52 (s, I MS (ESI) m/z 385 387 A mixture of 3-broino-4-[NV'-(2-methylphenyl)ureidojphenylacetic acid (80 mg, 0.22 mmol), methyl 4-[(4.S)-fluoro-(2S)-pyrrolidinyl~inethoxybenzoate (56 mg, 0.22 miiol), EDC-HCI (63 mg, 0.33 mmol), HOBT (45 mg, 0.33 mmol) and Et 3 N 15 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 extracts were concentrated in vacuo. The residue was purified on TLC ICHCl,/acetone to give methyl 4-[Il-[3-bromo-4-[N'-(2-methylphenyl)uridophenylacetylJ-(4S)-fluoro-(2S)pyrrolidinyl ]methoxybenzoate (140 mg, 100%) as a yellow oil. 'H-NMvR (CDC 3 82.30 (s,3H), 2.55 (in, IlH), 3.56 J= 6.4 Hz, 214), 3.70-3.84 (in, 311), 3.87 3H), 3.99-4.59 (in, 3H), 5.23- 5.38 (in, 6.83-6.94 (in, 2H), 6.95 J 8.8 Hz, 111), 7.07-7.26 (in, 511), 7.36-7.63 (in, 21H), 7.94-8.15 (in, 3H); MS (ESI) m/z 598 (Mr+ 600 To a solution of methyl 1-[3-bromo-4-[N'-(2-inethylpheniyl)ureido]phenylacetylJ-(4S)-fluoro- (2S)-pyffolidinyljinethoxybenzoate (140 mng, 0.22 inmol) in TI-F (10 ml) was added 0.25 N NaOH ml). After stirring at room temperature for 14 h, the mixture was concentrated under a reduced pressure and acidified with 1 N HCI. The precipitates were collected, washed with water and dried under a reduced pressure to give 94 (109 mg, 85%) as a colorless powder. MW 584.43 IR 3313, 3060, 2976, 1687, 1604, 1525, 1244 cmf'; 'H-NMR (DMSO-d 6 8 2.27 3H), WO 01/00206 WO 0100206PCT/USOO/18079 2.29 (in, 2M), 3.66 J 8.1 Hz, 2H), 3.72-4.68 (in, 5H), 5.3 1-5.53 (in, 1H), 6.92-6.99 (in, 11-), 7.04 and 7.07 (each d, J= 8.3 Hz, 2H, amide isomers), 7.11-7.21 (in, 3H), 7.48 and 7.51 I1H, amide isomers), 7.75 and 7.79 (each d, J =8.1 Hz, I1H, amide isomers), 7.86-7.92 (in, 3H), 8.45 and 8.47 (each s, 1H, amide isomers), 8.59 1H), 12.64 IH); MIS (FAB) pn/z 584 586 Anal. Calcd far C.,H, 7 BrFN 3 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 89 441 -[3-bromo-4-[N '-(2-chlorophenyl)ureidolphenylacetyl-(4S-fluoro-(2S-pyrrolidinyJ methoxybenzoic acid
F
Toa mixture of methyl 4-amino-3-bromophenylacetate (587 mg, 2.40 inmol) and 2-chiorophenyl isocyanate (0.29 ml, 2.40 mrnol) in THF (2 nil) was added Et 3 N (33 ml, 0.24 nunol) 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] phenylacetate (710 mg, 74%) as a pale brown powder. IH-NMR (CDCI 3 83.57 Cs, 2H), 3.70 (s, 3H), 7.02-7.28 2H), 7.36 J 6.8 Hz, lH), 7.48 1H), 8.00-8. 11 (in, 211); MS (ESI) mkz 3 97 3 99 401 (MC+4).
To a stirred solution of methyl 3-bromo-4-[N-(2-cilorophenyl)ureido~phenylacetate (710 mng, 1.79 inmol) in THF (10 ml) was added 0.25 N NaOH (10 nil). After stirring at room temperature for 14 h, the solvent was concentrated in vacua. The residue was triturated by the addition of I N HC1 and dried at 60 'C for 2 days under a reduced pressure to give 3-bromo-4-[N'-(2-chlorophenyl) ureidoiphenylacetic acid (643 mg, 94%) as colorless powder. 'H-NMR (DMSO-d) 8 3.56 2H), 7.05 (in, IHM, 7.21 (dd, J= 8.6, 1.7 Hz, 114), 7.29 J= 7.8 Hz, l1H), 7.46 J= 8.1 Hz, 11-), 7.46 J= 8.1 Hz, 1H), 7.53 J= 1.7 Hz, 1H), 7.83 J= 8.3 Hz, IN), 8.06 J= 7.6 Hz, IHM, 8.86 8.89 lH), 12.40 Ili); MS (ESI) m/z 382 384(W+3).
A mixture of 3-broino-4-[N'-(2-chlorophenyl)ureidojphenylacetic acid (384 mng, 1.00 inmol), methyl 4-[(4.S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (253 mg, 1.00 inmol), EDC-HCI (288 mg, 1.50 minol), HOBT (203 mg, 1.50 nunol) 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, WO 01/00206 WO 0100206PCT/USOO/18079 CHC1,/acetone( 10/1)1 to give methyl 1-13-bromo-4-[N '-(2-chlorophenyl)ureido]phenylacetylJ- (4S)-fluoro-(2S)-pyrrolidinyllmethoxybenzoate (640 mg, 100%) as a colorless amorphous solid.
'H-NMR (CDCI 3 8 2.07-2.46 (in, 2H), 2.59 J 18.4 Hz, 1H), 3.57 J 10.5 Hz, 2H), 3.63- 4.67 (in, 7M, 5.26-5.44 (in, 1H), 6.89-6.96 (in, 3H), 7.13 J 7.6 Hz, 1H4), 7.1 J =7.3 Hz, 1H), 7.26-7.29 (in, 2H), 7.52-7.94 (in, 4H), 8.01(d, J =8.5 Hz, 1H), 8.09 J= 8.5 Hz, 1H); MS (FAD) mnk 618 620 622 To a solution of methyl 1-13-bromo-4-[N '-(2-chlorophenyl)ureidolphenylacetyl]-(4S)-fluoro- (2S)-pyrrolidinyljinethoxybenzoate (640 mg, 1.00 mmol) in THF (40 mld) was added 0.25 N NaOH ml). After stirring at room temnperature for 14 h, the mixture was concentrated under a 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-NMiR (DMSO-d 6 8 2.24-2.50 (in, 2H), 3.67 J= 8.3 Hz, 2M), 3.73-4.68 (in, 5M, 5.31-5.52 (in, 1H), 7.03-7.09 (in, 311), 7.22 (dt, J 1.7 Hz, 111), 7.30 J 7.3 Hz,2 IlH), 7.46 (dd, J 1.4 Hz, 111), 7.49 and 7.52 (each d, J 2.0 Hz, 1H, amide isomers), 7.80-7.9 1 (in, 3M1, 8.07 (dd, J 8.3, 1.2 Hz, 114), 8.85 and 8.86 (each s, 1H, amide isomers), 8.96 and 8.97 (each s, 1H, amide isomers), 12.62 111); MS (FAD) m/z 605 607 609 626 Anal. Calcd for
C
2 7
H
24 BrClFNO,-0.8H 2 O: 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.
Example 4-[I1-13-bromo-4-[N '-(2-broinophenyl)ureido~phenylaoetylj-(4S)-fluoro-(2S)-pyrrolidinylJ inethoxybenzoic acid H H r96 To a mixture of methyl 4-amino-3-bromophenylacetate (587 ing, 2.40 inmol) and 2-bromophenyl isocyanate (0.30 m.L 2.40 iniol) in TI-F (2 ml) was added Et 3 N (33 nil, 0.24 minol) 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-IN -(2-bromophenyl)ureido] phenylacetate (770 mg, 73%) as a pale brown powder. 'H-NMR (CDCI 3 8 3.55 214), 3.70 (s, 311), 6.97 (dd, J= 7.3, 1.5 Hz, 1H), 7.22 (dd, J= 8.5, 2.2 Hz, I 7.29-7.33 (in, 2H), 7.48 J 1.0, 2.2 Hz, IlH), 7.54 (dd, J 8.0, 1.2 Hz, 1H), 8.01 (in, 2H); MS (ESI) in/z 443 445 447 WO 01/00206 WO 0100206PCT/USOO/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 'C Ibr 2 days under a reduced pressure to give 3-bromo-4-[N'-(2-bromophenyl) ureidolphenylacetic acid (702 mg, as colorless powder. 'H-NMR (DMSO-ds) 8 3.56 2HM, 6.99 (dt, J 1.5 Hz, IHM, 7.21 (dd, J 1.7 Hz, IHM, 7.33 (dt, J 1.5 Hz, IM), 7.53 J= 1.7 Hz, 1H), 7.62 (dd, J= 8.1, 1.5 Hz, IHM, 7.82 J= 8.3 Hz, IHM, 7.93 (dd, J Hz, 111), 8.82 1H4), 8.86 1H), 12.39 IH); MIS (ESI) mkz 428 430(Mx+3).
A mixture of 3-bromo-4-[N'-(2-chlorophenyl)ureidoJphenylacetic acid (428 mg, 1.00 mmol), methyl 4-[(2S,4S)-4-fluoro-2-pyrrolidinyllmethoxybenzoate (253 mg, 1.00 mmnol), EDC-HCI (288 mg, 1.50 mmnol), HOBT (203 mg, 1.50 minol) and EtN (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 SO4, the extracts were concentrated in vacuo. The residue was chromatographed on silica gel 130 g, CHCI/acetone( 10/1)1 to give methyl 4-1 1-[3-bromo-4-[N -(2-bromophenyl)ureido~phenylacetylj- (4S)-fluoro-(2S)-pyrrolidinyl~methoxybenzoate (720 mg, 100%) as a colorless amorphous solid.
2 H-NMvR (CDCI 3 8 2.07-2.45 (in, 2HM, 2.58 (in, lH), 3.58 J 9.0 Hz, 2H), 3.63-4.69 (mn, 9H), 5.26-5.43 (in, IH), 6.88-6.99 (in, 3H), 7.16 J =8.3 Hz, lH), 7.23-7.32 (in, 2H), 7.46 (dd, J= 8.1, 1.5 Hz, IM, 7.5 1-8.20 (in, 5HM; MS (FAB3) m/z 664 666 668 To a solution of methyl 1-[3-bromo-4-[N -(2-broinopheniyl)uireido]phenylacetylJ-(4S)-fluoro- (ZS)-pyrrolidinyllmethoxybenzoate (720 mg, 1.00 inmol) in THF (40 ml) was added 0.25 N NaOH ml). After stirring at room temperature for 14 h, the mixture was concentrated in vacuo and acidified with 1 N HCI. The mixture was extracted with CHCIJ/MeOH 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, CHC13/acetone (l0/1)-CHC 3 /MeOH(20/1)j and triturated by the addition of ether to give 96 (489 mg, 75%) as a colorless amorphous solid. MW 649.30 JR (KBr) 3450, 3313, 3070, 1709, 1684, 1525, 1435 cin'; 'H-NMR (DMSO-d 6 8 2.25-2.50 (in, 2H), 3.67 J 8.3 Hz, 2H), 3.73-4.68 (in, 5H), 5.3 1-5.53 (in, 6.98-7.08 3H), 7.21 J =8.2 Hz, 1H), 7.34 J 8.8 Hz, IHM, 7.50 and 7.53 (each s, 1H, amide isomers), 7.62 J 8.0 Hz, IN), 7.80-7.96 (mn, 4H), 8.82 lH), 8.85 and 8.86 (each s, IlH, ainide isomers), 12.63 (br, I MS (FAB) m/z 650 (NC+ 652 654 672 Anal. Calcd for C 27 H7,Br 2
FN
3 O,-0.9H 2 O: C, 48.73; H, 3.9 1; N, 6.3 1; F, WO 01/00206 WO 0100206PCTIUSOO/18079 2.85. Found: C, 48.96; H, 3.98; N, 5.92; F, 2.77.
Exam We 91 1-I4-IN'-(2-methylphenyI)ureido]-2,3-dMfuorophenylacetyl-(4S)-fluoro-(2S)-pyrrolidifly methoxyjbenzoic acid.
~MeH H9 To a stirred solution of tert-butyl ethyl malonate (5.3 5 ml, 28.2 mmol) in DMF (150 ml) was added NaH (60% in oil, 3.38 g, 84.7 mmnol) at After 20 min, 2,3-difluoronitrobenzene (5 g, 28.2 mmol) in DMF (50 niL 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
CI
(100 mL). The mixture was extracted with EtOAc and the combined organic layer was washed with IM HCl 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 vacua, coevaporated with toluene (20 mL x 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 mmn x 300 mm, 15 m~fmin) to give ethyl 2,3-difluoro-4-nitrophenylacetic acid (5.85 g, 85%) as a yellow oil. 'H-NMR (CDCI,) 8 1.30 (mn, 3 3.78 211), 4.22 (mn, 2 7.22 (mn, I M, 7.84 (in, I MIS (FAB) m/z 246 (Mr+1).
To a stirred solution of ethyl 2,3-difuoro-4-nitrophenylacetate (5.85 g, 23.9 mol) in EtOH (100 mL), was added SnCI 2 (16.1 g, 71.6 nimol) at rt. The stirring was continued for 18 hours at reflux. After removal of the solvent, the residue was dissolved in CHC1 3 (100 niL) and poured into ice water-4M NaOH (40 nit of 4M NaOH in 300 mL of ice-water), extracted with CHC1 3 (100 nit x 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 mmn x 500 mmn, 15 m/mn) to give ethyl 4-amnino-2,3-dffluorophenylacetic acid (1.94 g, as a colorless oil. 'H-NMR (CDCI 3 8 1.25 J= 7.3 Hz, 3 3.55 J= 1.0 Hz, 2 3.78 (brs, 2 4.15 (dd, J =7.2 Hz, 14.2 Hz, 2 6.49 (dt, J 8, 8.2 Hz, 1 6.78 (in, I MIS (FAB) m/lz 216 (W+l1).
To a stirred solution of ethyl 4-amino-2,3-diffluorophenylacetate (323 mng, 1.5 inmol) in DMF (8 mQL, were added triethylamine (0.209 ml, 1.5 mmol) and 2-niethylphenyl isocyanate (0.372 ml, WO 01/00206 WO 0100206PCT/USOO/18079 mmol) at rt. The stirring was continued for 48 hour at 80 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 TH-F-MeOH 1, v/v, 20 niL, and was added 4M NaOH mL) at rt. The stirring was continued for 18 hours at rt. The reaction was poured into IM HCI, and the resulting precipitate was collected through filtration. The solid was recrystallized with CHCI 3 -n-hexane to give 4-[(2-methylphenyl)ureidoJ-2,3-difuorophenylacetic acid (200 mig, 42%) as a white solid. 'H-NMvlR (CDCI 3 5 2.30 3 3.35 2 6.98 (in, 1 7.04 (in, 1 Mf, 7.18 J =7.3 Hz, 2H), 7.69 (d,J =8.l1Hz, 1 7.90 1 H);MNS (FAB) m/z3 21 1).
To a stirred solution of methyl 4-(4-S-4-fluoro-2-pyrrolidinyl)methoxy benzoate (63 mg, 0.25 mmol) and 4-[N'-(2-methylpheniyl)ureidol-2,3-difluorophenylacetic acid (82 mg, 0.25 mmol) in DMVF (5 mL), were added EDC-HCI (72 mng, 0.38 minol), HOBt (69 mg, 0.48 mmol), and DMAP and the stirring was continued overnight at rt. The mixture was diluted with EtOAc niL), 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-H 2 0 (21 niL, 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 CHC1 3 -MeOH 1, The combined organic phase was dried over anhydrous MgSO4, and concentrated under a reduced pressure. The residue was purified with TLC (Whatinan, PLK- SF, CHCI 3 /MeOH, 20:1, vlv) to give 97 (69 mg, 51%) as a white powder. MW 541.52 JR (KBr) 3340, 1604, 1540, 1251, 1168, 754 'H-NMR (DMSO-dQ 8 2.25 3 2.32 (in, 2 3.68- 4.40 (in, 7 5.32-5.55 (in, 1 6.98 (mn, 2 7.05 J 8.8 Hz, 2 7.83 J 8.8 Hz, 2 7.82-7.92 2 8.40 1 9.14 1 MS (ESI) m/z 564 (Mr+Na); Anal. Calcd for CnH 26
F
3
N
3 O,-2.0H 2 O: C, 58.23; H, 5.24; N, 7.28. Found: C, 58.07, H, 4.84; N, 7.03.
Example 92 4-[I-[4-[IN'-(2-methylphenyl)ureido-2,5-difuorophenylacetyl-(4S)-fluoro-(2S)-pyrrolidinyI methoxylbenzoic acid Me H Hoo 98 To a stirred solution of di-tert-butyl ethyl malonate (6.32 nil, 28.2 inmol) in DMF (150 ml), was added NaH (60% in oil, 3.38 g, 84.7 minol) at rt. After 20 min, 2,5-difluoronitrobenzene (S g, 28.2 nimol) in DMF (50 inL) 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 Cl WO 01/00206 PCT/US00/18079 (100 mL). The mixture was extracted with EtOAc and the combined organic layer was washed with 1M 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 The residue was 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, 50 mm x 300 mm, 15 mL/min) to give ethyl 2,5-difluoro-4nitrophenylacetic acid (6.53 g, 90%) as a yellow oil. 'H-NMR (CDCI 3 5 3.75 2 3.76 3 7.29 (dd, J= 5.8 Hz, 10.5 Hz, 1 7.81 (dd, J= 6.0 Hz, 8.4 Hz, 1 MS (ESI) m/ 232 To a stirred solution of ethyl 2,5-difluoro-4-nitrophenylacetate (5.88 g, 25.4 mol) in EtOH (100 mL), was added SnCI, (17.2 g, 76.3 mmol) at rt. The stirring was continued for 18 hours at reflux. After removal of the solvent, the residue was dissolved in CHCl3 (100 mL) and poured into ice water-4M NaOH (40 mL of 4M NaOH in 300 mL of ice-water), extracted with CHCI 3 (100 mL x dried over anhydrous MgSO,, 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, 50 mm x 500 mm, 15 ml/min) to give ethyl 4-amino-2,5-difluorophenylacetic acid (2.85 g, 52%) as a colorless oil. H-NMR (CDCI 3 8 1.28 J= 7.3 Hz, 3 3.51 2 3.78 (brs, 2 4.15 (dd, J= 7.2 Hz, 14.2 Hz, 2 6.47 (dd, J 7.5, 10.4 Hz, 1 6.88 (dd, J 6.7, 11.0 Hz, 1 MS (FAB) m/z 216 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, 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 v/v, 20 mL), and was added 4M NaOH mL) at rt. The stirring was continued for 18 hours at rt. The reaction was poured into IM HCI, and the resulting precipitate was collected through filtration. The solid was recrystallized with CHC1 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 3 3.35 2 7.02 2 7.18 J WO 01/00206 WO 0100206PCT/US00118079 7.3 Hz, 2 7.69 J =7.8 Hz, 1 8.03 (in, 1 MS (FAB) 1nk 3 21 To a stirred solution of methyl 4-[(4S)-fluoro-(2S)-pyrrolidinyl]methoxybenzoate (63 mg, 0.25 mmol) and 2,5-difuoro-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (82 mg, 0.25 minol) in DMF (5 mL) was added EDC-HCI (72 mg, 0.38 minol), HOBt (69 mg, 0.48 mmol), and DMAP and the stirring was continued overnight at rt. The mixture was diluted with EtOAc mL), washed with IM NaOH, IM HCI, and brine, dried over anhydrous MgSO4, and concentrated under a reduced pressure. The residue was dissolved in TH-F-MeOH-H 2 0 (21 mL, 1: 1: 1, v/v/v) and the stirring was continued for 6 h at rt. The miixture was poured into IM HCl and extracted with CHCI,-MeOH 1, The combined organic phase was dried over anhydrous MgSO4, and concentrated under a reduced pressure. The residue was purified with TLC (Whatman, PLK- CHCI3/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 'H-NMvR (DMS0) 8 2.25 3 2.32 (in, 2 3.68-4.70 (in, 7 5.32-5.55 (in, 1 6.97 J =7.6 Hz, 1 7.06 J =8.5 Hz, 2 7.20 (in, 3 H), 7.87 J 8.8 Hz, 2 7.83-8.04 (in, 2 8.45 1 9.18 1 MIS (ESI) m/z 564
(M
t Anal. Calcd for C.,HXFNO 5 1.75 H20: C, 58.69; H, 5.19; N, 7.33. Found: C, 58.54, H, 4.85; N, 6.98.
Examnle 93 1-14-IN '-(2-chlorophenyl)ureido]-3-methoxyphenylacetylJ-4-fluoro-2-pyrrolidiny~methylamino benzoic acid Na0-3.-COOH H H6e99 I-[4-[N'-(2-bromophenyl)ureidol-3-mfethoxyphenylacetyl]-4-fluoro-2-pyrrolidinyllmethylamino benzoic acid H H 6Me100 To a stirred solution of methyll1-tert-butoxycarbonyl-4-fluoropyrrolidine-2-carboxylate( 1.2 g, 4.85 inmol) in MeOH (5 mld) was added 1 N NaOH (5 ml) and the mixture was stirred at room temperature for 1 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with IN HCI. The mixture was extracted with EtOAc. The extract was washed with water, then dried over Na 2
SO
4 and concentrated in vacua to give I -tert-butoxycarbonyl-4- WO 01/00206 PCT/US00/18079 fluoropyrrolidine-2-carboxylic acid (1.1 g quant) as a colorless oil. 'H-NMR (CDCI 3 6 1.47 (br s, 9H), 2.78-2.83 (br s, 3H), 4.37 2H), 6.73-6.76 3H), 7.17 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 BH3-THF(I.0 M solution in THF, 10.0 ml, 10.0 mmol) at 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°C, and extracted with EtOAc. The extract was washed with water, then dried over NaSO 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 (CDCI 3 8 1.48 9H), 2.29- 2.39 1H), 3.38-3.59 2H), 3.74-3.88 2H), 4.09-4.14 2H), 4.85 1H), 5.03 (br s, 1H), 5.16 (brs, 1H).
To a stirred solution ofoxalyl chloride (0.28 ml, 2.3 mmol) in CH 2
CI
2 (20.0 ml) was added DMSO (0.39 ml) at -78 OC. After 5 minutes, to the mixture was added 1-tertbutoxycarbonyl-4-fluoro-2-pyrrolidinylmethanol (500 mg, 2.28 mmol) in CH 2 CI, (5.0 ml). The mixture was stirred for 30 minutes at -78 OC, and triethylamine (1.6 ml) 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 C12. 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, mmol), and AcOH (0.13 ml) in DCE (10 ml) was added NaBH(OAc) 3 (656 mg, 3.09 mmol) at 0 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 ClI. The extract was washed with brine, dried over Na 2 SOI, and concentrated in vacuo. The residue was purified by column chromatography on silica gel with EtOAc- n-hexane v/v) as eluent to give methyl 4-(1-tert-butoxycarbonyl-4-fluoro-2-pyrrolidinyl)methylaminobenzoate (541 mg, 77%) as a pale yellow oil. 'H-NMR (CDCI3) 8 1.55-1.59 1H), 2.16-2.27 1H), 2.89-3.03 2H), 3.19-3.28 2H), 3.69-3.73 1H), 3.84 3H), 5.15 and 5.29 (each s, total 1H), 6.55-6.58 (m, 2H), 7.84-7.86 2H).
To a stirred solution of methyl 4-(1-tert-butoxycarbonyl-4-fluoro-2-pyrrolidinyl)methylamino benzoate (541 mg, 1.53 mmol) in CHCI 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, was added to the residue, and extracted with CH,C12. The extract was washed with brine WO 01/00206 WO 0100206PCT/USOO/1 8079 ,dried over Na 2 SO, and concentrated in vacua. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product(15l mg, 0.6 mmol), 4-[N'-(2-cblorophenyl)uredio]-3-methoxyphenylacetic acid (201 mg, 0.6 mmol), HOBt (94 mg, 0.7 mmol), and triethylamine (167 0i, 1, 1.2 mniol) in THF (10.0 mlJ) and MeCN (10.0 ml) was added EDC-HCI (173 mg, 0.9 minol) at 0 The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacua. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 2-M citric acid, and sat. NaHCQ 3 then dried over Na 2
SQ
4 and concentrated in vacua. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc(l :2 ,vtv) as eluent to give methyl 4-11-14-IN'- (2-chlorophenyl)ureido]-3-methoxyphenylacetylj-4-fluoro-2-pyrrolidinyl]methylaminobenzoate (320 mg, as an amorphous solid. 'H-NMR (CDCI 3 8 1.80-1.95 (in, 2.42-2.58 (in, 1lH), 3.20-3.51 (im, 3H), 3.51-3.76 (in, 5H), 3.84 311), 3.85-3.98 (in, 4.67-4.70 (in, IH), 5. and 5.23 each, total 111), 5.50 (br s, 111), 6.49-6.52 (in, 2H), 6.78-6.81 (in, 2H), 6.97-7.01 (in, 111), 7. 14-7.18 (in, 2H), 7.24-7.36 (Cm, 214), 7.80-7.82 (in, 2H), 7.99-8.01 114), 8. 15-8.18 (in, 1H).
To a stirred solution of methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido]-3-inethoxyphenyl acetylj-4-fluoro-2-pyrrolidinyl]inethylamiinobenzoate (320 mng, 0.56 innol) in THF (5.0 ml) and MeOH (3.0 ml) was added IN NaOH (0.8 ml, 0.8 inmol). The mixture was stirred at 70 *C for 24 hr. The mixture was concentrated in vacua, water was added thereto, and neutralized with IN HCI. The resulting solid was collected, washed with water, and dried in vacua to give 99 (280 mg, as a-white crystalline solid. MW 555.00 mp 132-136 0 C; JR (KBr) 3332, 2937, 1602, 1531, 1174, 752 cin*'; 'H-NMR (DMS-4) 8 2.00-2.40 (in, 211), 3.50-3.90 (in, 411), 3.75-3.85 (in, 4.27 (in, 111), 5.23 and 5.37 (each s, total 111), 6.51-7.03 (in, 5H), 7.25-7.29 (in, 111), 7.41- 7.44 (Cm, I1H), 7.64-7.68 (in, 2H), 7.92-8. 10 (in, 2H), 8.87-8.94 (in, 211); Anal. calcd for CnH28N 4 OFCl*0.6H 2 O: 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 1-tert-butoxycarbonyl-4-fluoro-2-pyrrolidinyl)inethylainino benzoate (541 mng, 1.53 inmol) in CH 2 C1 2 (8.0 ml) was added TFA (4.0 ml) at 0 The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacua. Sat.
NaHCO, was added to the residue, and extracted with CHC1 2 The extract was washed with brine, dried over Na 2
SO
4 and concentrated in vacua. The crude product was used to the subsequent reaction without fuirther purification. To a stirred solution of the crude product (151 mg, 0.6 inmol), 4-[N'-(2-broinophenyl)uredioj-3-methoxyphenylacetic acid (227 mng, 0.6 iniol), HOBt (94 WO 01/00206 WO 0100206PCT/USOOII 8079 mg, 0.7 mmol), and triethylamine (167 d, 1.2 mmol) in T1-F (10.0 ml) and MeCN (10.0 ml) was added EDC-HCI (173 mg, 0.9 mmol) at 0 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-EtQAc v/v) as eluent to give methyl (2-bromophenyl)ureido]-3-methoxyphenylacetylJ-4-fluoro-2-pyrrolidinyllmethylanuino benzoate (280 mg, 76%) as a colorless oil. 'H-NMvR (CDCI 3 8 1.80-1.98 (in, 1.42-1.58 (in, 111), 3.20- 3.52 (in, 3H), 3.67-3.79 (mn, 5H), 3.84 3H1), 3.94-3.97 (in, lH), 4.68-4.71 (in, 114), 5. 10 and 5.23 (each s, total 1H), 5.51 (br s, IH), 6.50-6.52 (in, 2H), 6.79-7.07 5H), 7.25-7.33 (in, 111), 7.51-7.53 (in, lH), 7.80-7.83 (in, 2H), 7.98-8.00 (in, lH), 8.11-8.14 (in, 111).
To a stirred solution of methyl 4-[l-[4-[N-(2-bromophenyl)ureidoj-3-methoxyphenyl acetylj-4-fluoro-2-pyrrolidinyl]methylaminobenzoate (280 mg, 0.46 inmol) in THF (8.0 ml) and MeOH (8.0 ml) was added IN NaOH (2.8 ml, 2.8 inxol). The mixture was stirred at 70 *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 100 (260 mg, as a white crystalline solid. MW 599.45 mp 13 1-135 JR (KBr) 3332, 2935, 1602, 1529, 1174 cin'; 'H-NMvR (DMSO-d) 8 1.95-2.01 (in, IN), 2.20-2.35 (in, 111), 3. 10-3.20 (in, 111), 3.50-3.70 (in,3H), 3.80-3.85 (in, 5N), 4.27 (in, IN), 5.24 and 5.37 (each s, total iN), 6.54-6.99 (in, 5H), 7.30-7.33 (in, IH), 7.58-7.94 (in, 3H), 7.94-7.98 (in, 2H), 8.73 (in, 111), 8.93 (in, lH); Anal.
calcd for CgHN 4 OBrF-0.7H 2 O: C, 54.95; H, 4.84; N, 9.15. Found: C, 54.98; H, 4.81; N, 8.93.
Example 94 4-[I-[3-methoxy-4-[N'-(2-methylphenyl)ureido~phenylacetyl-4R)-fluoro-(2S)-pyrrolidinyI methoxylbenzoic acid e e101 A mixture of methyl 4-[(4R)-fiuoro-(2S)-pyrrolidinylmethoxy~benzoate (634 mg, 2.50 inmol), 3methoxy-4-[N'-(2-inethylphenyl)ureidolphenylacetic acid (787 ing, 2.50 mmol), EDC-HCl (718 mg, 3.75 minol), HOBt DMAP (cat.) and DMF (10 ml) was stirred overnight. The mixture was diluted with EtOAc (300 mrl). The solution was washed with brine (2 x 100 ml), dried over MgSO 4 and concentrated in vacua. The residue was chromatographed on silica gel with CHCI,- WO 01/00206 WO 0100206PCT/USOIJ/18079 EtOAc as eluent to give methyl l-[3 -methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetylj-(4R)-fluoro-(2S)-pyrrolidinylmethoxy~benzoate (1.37 g, quant) as a pale yellow viscous solid. 'H-NMR (CDCI 3 8 2.24 3 2.26-2.47 (in, 2 3.46 3 3.49-3.64 (in, 4 3.87 3 4.06 (dd, J 9.5, 2.0 Hz, 1 4.51-4.62 (in, 2 5.20 and 5.33 (br s, each, total I1H), 6.63 1 6.72 J =8.3 Hz, 1 6.77 (d,J =9.0Hz, 2 7.05 J =7.6 Hz, 1 7.16-7.20 (in, 3 7.53 1 7.63 J 7.8 Hz, 1 7.91 J 9.0 Hz, 2 8.07 (d, J= 8.1 Hz, 1 H).
A mixture of methyl 1-13-methoxy-4-[N' -(2-methylphenyl)ureidolphenylacetylJ-(4R)fluoro-(2S)-pyrrolidinylmethoxy~benzoate (1.37 g, 2.49 mmol), 0.25 N NaOH (20 ml, 5.00 minol), and THF (20 ml) was stirred for 3 days. The mixture was poured into 1 N HCI (100 mlJ) and extracted with CHCI 3 -MeOH 2 x 200 ml). The combined extracts were dried over MgSO 4 and concentrated in vacuo. The residue was chromatographed on silica gel with CHCI,-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 total 5 3.42-4.66 (series of m, 10 5.31 and 5.44 (br s, each, total 1 6.71-7. 16 (series of mn, 7 7.79 J= 8.1 Hz, 1 7.85-7.89 (in 2 7.98-8.00 (in, 1 8.47 1 8.55 1 MS(FAB) m/lz 536 (M 4 1).
Example 4-I(4S)-chloro-l1-[3-methoxy-4-[N'-(2-chlorophenyl)ureido~phenylacetyl]-(2S)-pyrrolidinyl] methoxybenzoic acid
CI
C1 H H 6Me 102 To a stirred solution of methyl 4-Qtrans-1-tert-butoxycarbonyl-4-hydroxy-(2S)-pyrrolidinyl) inethoxybenzoate (351 mng, 1.0 ifnol) and P113P (393 mng, 1.5 ifnol) in CHC1 3 (5.0 ml) was added CC14, (5.Oml) at room temperature. The reaction mixture was stirred at 50*C for 24 hr. The reaction mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane to n-hexane-EtOAc(4: 1, v/v) as eluent to give methyl 4-(cis-lI-tertbutoxycarbonyl-4-chloro-(2S)-pyrrolidinyl)methoxybenzoate (340 mng, as a pale yellow oil.
IH-NMvR (CDCI,) 8 1.48 9H), 2.38-2.65 (in, 2H), 3.50-3.60 (mn, 1H), 3.88 3H), 3.894.05 4.26-4.41 (mn, 4H), 6.95-6.97 (in, 2H), 7.98 J 8.5 Hz, 2H).
To a stirred solution of methyl 4-(l-Iert-butoxycarbonyl-(4S)-chloro-(2S)-pyrrolidinyl) WO 01/00206 WO 0100206PCTIIJS0OI18079 methoxybenzoate (369mg, 1.0 iniol) in CH 2 C1 2 (3.0 ml) was added TEA (3.0 ml) at 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 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 (185 mg, 0.5 mmol), 4-[N'-(2-chlorophenyl)uredio]-3-methoxyphenylacetic acid (167 mg, nunol), HOBt (68 ing, 0.5 minol), and triethylamnine (208in1, 1.5 inmol) in THE (8.0 ml) and MeCN (8.0 ml]) was added EDC-HCI (144 mg, 0.75 inmol) at 0 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. NaIICO 3 2-M citric acid, and 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 v/v) as eluent to give methyl 4f(4.S)-cloro- 1 -[3-inethoxy-[N'-(2-chlorophenyl)ureido]phenylacetyl -(2S)-pyrrolidinyl]methoxy benzoate (2 10 ing, 72%) as a colorless oil. 1H-NMR (CDCl 3 8 3.35-3.50 (mn, 3.55-3.65 (in, lH), 3.6 1-3.66 (in, 3H), 3.75 3H), 3.88 3H), 3.99-4.04 (in, 1H), 4.35-4.40 (in, 3H), 4.48- 4.53 (in, 1H), 6.77-7. 10 (in, 7H), 7.25-7.36 (in, 2H), 7.93-8.00 (in, 2H), 8.18 J 8.0 Hz, 1H).
To a stirred solution of methyl 4-[(4S)-chloro--[3-iethoxy-[N-(2-chlorophenyl) ureido~phenylacetylJ-(2S)-pyrrolidinyl~methoxybenzoate(210 ing, 0.35 inmol) in THE (6.0 ml) and MeOH (3.0 ml) was added IN NaOH (0.7 ml, 0.7 inmol). The mixture was stirred at 70 *C for 18 hr. The mixture was concentrate 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 102 (200 mg, 98%) as awhite crystalline solid. M 572.44 mp 126-131 0 C; IR (KBr) 3330, 1685, 1604, 1533, 1438 cin-'; 'H-NMvR (DMSO-d) 8 2.15-2.25 (in, 1H), 2.58-2.63 (in, 1H), 3.58-3.78 (in, 3H), 3.83 3H), 4.134.42 (in, 4.73 (in, 114), 6.75-7.45 (in, 7.86-8. 10 (in, 4H), 8.90 lH), 8.95 1H); MS (EAB) m/z 572 (M 4 Anal. calcd for C.aH 27 NOCl: C, 58.75; H, 4.75; N, 7.34.
Found: C, 58.93; H, 4.85; N, 7.15.
Example 96 4-[41 '(-rmpey~rio-3mtoyhnlctl-4)-hoo(S-yrldnl inethoxybenzoic acid rH H6e103 To a stirred solution of methyl 4-[1-tert-butoxycarbonyl-(4S)-chloro-(2S)-pyrrolidinyl] WO 01/00206 WO 0100206PCT/USOO/18079 methoxybenzoate (369mg, 1.0 mmol) in CH 2
CI
2 (3.0 ml) was added TEA (3.0 ml) at 0 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 crude product was used to the subsequent reaction without fturther purification. To a stirred solution of the crude product (185 mg, 0.5 mniol), 4-[N'-(2-bromophenyl)uredioJ-3-methoxyphenylacetic acid (190 mg, nunol), HOBt (68 mg, 0.5 minol), and triethylamine (208m1d, 1.5 mmol) in THE (8.0 ml) and MeCN (8.0 mld) was added EDC-HC1 (144 mg, 0.75 mmol) at 0 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.
NaHCO3, 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 as eluent to give methyl 4- [1 -[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]-(4S)-chloro-(2S)-pyrrolidinyl methoxybenzoate (260 mg, 83%) as a colorless oil. 1H-NMR (CDC1 3 8 2.32-2.50 (in, lH), 2.53- 2.65 (mn, 1H), 3.6 1-3.67 (in, 3H), 3.75 311), 3.88 3H), 3.994.03 (in, 1H), 4.35-4.40 (mn, 3H), 4.45-4.55 (in, 1H), 6.78-7. 10 (in, 711), 7.28-7.33 (in, 111), 7.52 J 8.0 Hz, lH), 7.94-7.99 (in, 3H), 8.14 J =8.3 Hz, I H).
To a stirred solution of methyl 4-[l-[4-[N-(2-bromophenyl)ureido-3-nethoxypheny acetylJ-(4S)-chloro-(2S)-pyrrolidinyljmethoxybenzoate (260 mg, 0.4 minol) in THE (6.0 ml) and MCOH (3.0 nil) was added IN NaOH (0.8 ml, 0.8 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 103 (210 ing, as a white crystalline solid. MW 616.89 mp 127-132 0 C; JR (KBr) 3330, 1685, 1604, 1529, 1434 cm-' 3 'H-NNM (DMSOQA 8 2.18-2.28 (in, 1H), 2.60-2.70 (in, I1H), 3.55-3.75 (in, 3H), 3.83 311), 4.124.42 (in, 4H), 4.604.75 (in, lH), 6.75-7.06 (in, 5H), 7.30-7.34 (in, lH), 7.60 J 7.3 Hz, 1H), 7.86-7.94 5H), 8.75 1H), 8.94 MIS (FAB) m/z 616 618 A nal calcd for CgH 27
N
3
O
6 C1Br: C, 54.52; H, 4.41; N, 6.81. Found: C, 54.98; H, 4.54; N, 6.66.
Exampnle 97 4-[(4R)-chloro-1 -[3-methoxy-4-[N -(2-methylphenyl)ureido~phenylacetyl] -(2S)-pyrrolidinyl methoxyjbnzoic acid WO 01/00206 PCT/US00/18079 SN {OOH 104 r H 6Me104 4-[(4R)-chloro- 1 -[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-(2S)-pyrrolidinyl methoxyjbenzoic acid N N OO105 To a stirred solution of methyl 1-(tert-butoxycarbonyl)-(4S)-hydroxy-(2S)-pyrrolidinylcarboxylate (1.81 g, 7.34 mmol) in CCI 4
-CH
2
CI
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- EtOAc 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 (CDCI,) 8 1.42 9 2.32- 2.39 1 2.49-2.54 1 3.66-3.92 (series of s and m, total 5 4.44-4.55 2 H); MS(FAB) m/z 264 To a stirred solution of methyl 1 -(tert-butoxycaibonyl)-(4R)-chloro-(2S)-pyrrolidinyl carboxylate (1.35 g, 5.12 mmol) in THF (10 mi) 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-i N HCI and the mixture was extracted with CHCI,-MeOH The extract was was washed with brine, dried over Na 2
SO
4 and evaporated to give I -(tert-butoxycarbonyl)- (4R)-chloro-(2S)-pyrrolidinylcarboxylic acid (1.28 g, quant.) as a colorless oil. 'H-NMR (CDCI 3 8 1.44 9 2.37-2.54 2 3.68-3.88 2 4.42-4.45 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
CI
2 The solution was washed with HO 2 0, brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHC1 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 (CDC 3 8 1.48 9 1.98 1 2.26-2.32 1 3.56- WO 01/00206 WO 0100206PCTIJSOOI18079 3.65 (in, 2 3.77 (in, 2 4.24 (in, I 4.41-4.46 (in, 2 MS(FAB) m/z 236 1).
To a cooled (0 0 stirred solution of methyl 4-hydroxybenzoate (560 mg, 3.68 nunol), 1- (tert-butoxycarbonyl)-(4R)-chloroprolinol (870 mg, 3.69 mmol), Ph 3 P 16 g, 4.42 mniol) in THE mlJ) was added DIAD (870 ml, 4.42 mniol) and the reaction mixture was heated under reflux for 10 hr. Alter cooled to room temperature, the mixture was evaporated. The residue was purified by column chromatography on silica-gel with n-hexane-EtOAc 1, v/v) as eluent to give methyl 1-(tert-butoxycarbonyl)-(4R)-chloro-(2S)-pyrrolidinylmethoxyjbenzoate (890 mg, as awhite solid. mp 116-120'C; 'H-NMR (CDC1 3 81.47 9H), 2.39-2.53 3.69-3.70 and 4.13-4.17 (in, total 3 3.88 3 4.30-4.41 (in, 2 H4), 4.50-4.55 (im, I 6.90-6.92 (in, 2 7.96-7.98 (in, 2 H4); MS(FAB) mih 370 Anal Calcd for C 28
H
2 ,C1N0,: 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.-[I-(tert-butoxycarbonyl)-(4R)-chloro-(2S)pyrrolidinylmethoxyjbenzoate (840 mg, 2.27 mmol) in CH 2 CI (10 ml) was added TEA (10 ml) and the reaction mixture was stirred at room temperature overnight. The mixture was concentrated in vacua and made basic by sat. NaHCO,. The mixture was extracted with CHC1 3 washed with brine, dried over Na 2
SQ
4 and evaporated to give methyl 4-[(4R)-chloro-(2S)pyrrolidinylmethoxyl benzoate (580 mg, 95%) as a white solid. mp 61-64"C; 'H-NMvR (CDC1 3 8 1.85 (br s, I 2.03-2. 10 (in, 1 2.29-2.35 (in, 1 3.19-3.3 1 (in, 2 3.88 3 3.92- 4.06 (in, 3 4.53-4.56 (in, 1 6.91 J =8.8 Hz, 2 7.98 J 8.8 Hz, 2 MS (FAB) ,n/z 270 A mixture of 3-inethoxy-4-[N -(2-inethylphenyl)ureidoJphenylacetic acid (385 ing, 1.22 inmol), methyl 4-[(4R)-chloro-(2S)-pyrrolidinylnethoxyJbenzoate (330 mg, 1.22 iniol), EDCHCI (281 mg, 1.47 inmol), HOBt (200 mg, 1.48 innol) and Et 3 N (205 ml, 1.47 mniol) in THE (10 mrl) 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
SQ
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHCI 3 -MeOH (100: 1 to 50: 1, v/v) as eluent to give methyl 4-[(4R)-chloro-1-[3-inethoxy-4-[N '-(2-methylphenyl)ureidojphenylacetyl]-(2S)pyrrolidinylmethoxyjbnzoate (670 mg, 97%) as a white foam. 'H-NMR (CDCI 3 8 2.28 3 H), 2.33-2.57 (in, 2 3.50 Cs, 3 3.59-3.60 (in, 2 3.75-3.82 (in, 2 3.88 3 4.06-4.09 (in, 1 4.51-4.63 (in, 3 6.65-6.80 (in, 5 7.09-7.13 (in, 1 7.20-7.27 (in, 3 7.56- 7.58 (in, 1 7.91-7.93 (in, 2 8.05-8.07 (in, 1 MS(FAB) m/z 566 WO 01/00206 WO 0100206PCTIJSOOII8079 To a stirred solution of methyl 4-[(4R)-chloro-1-[3-methoxy-4-[N'-(2-methylphenyl) ureidojphenylacetyl]-(2S)-pyrrolidinylmethoxybenzoate (480 mg, 0.85 mmol) in TIHF (5 ml) was added 0.5 N NaOH (5 nil) and the reaction mixture was heated under refiux for 2 hr. After cooled to room temperature, the mixture was poured into ice-i N HCl and the resulting precipitate was collected under a reduced pressure. The crude solid was dissolved in CHCI 3 -MeOH and evaporated. The residue was washed with Et 2 O to give 104 (355 mg, 76%D/) as a white amorphous solid. MW 552.02 mp 128-132 0 C; 'H-NMR (DMSO4) 82.25 3H), 2.29-2.46 (in,2H), 3.57- 3.73 (in, 2 3.78 3 3.81-3.99 (in, 2 4.11-4.31 (in, 2 4.43-4.45 and 4.64-4.67 (each in, total 1 4.83-4.85 (in, 1 6.71-7.17 (in, 7 7.78-7.80 (in, I 7.87-7.91 (in, 2 H), 7.99-8.01 (in, 1 8.47 1 8.56 1 12.66 (br s, 1 MS(FAB) m/z 552 Anal Calcd for C2I-I0ClN 3
O
6 -3/4H 2 O: C, 61.59; H, 5.6 1; Cl, 6.27; N, 7.43. Found: C, 61.56; HK 5.5 1; Cl, 6.68; N, 7.26.
A mixture of 4-[N'-(2-chlorophenyl)ureidoj-3-methoxyphenylacetic acid (400 ing, 1. 19 mmol), methyl 4-[(4R)-chloro-(2S)-pyrrolidinylmethoxy~benzoate (320 mg, 1. 19 minol), EDC-HCl (275 ing, 1.43 minol), HOBt (195 rag, 1.44 minol) and Et 3 N (200 ml], 1.43 minol) 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 evaporated. The residue was purified by column chromatography on silica-gel with CHCI 3 -MeOH (100: 1, v/v) as eluent to give methyl 4-[(4R)-chloro-1 '-(2-chlorophenyl)ureidol-3-methoxyphenylacetyll-(2S)pyrrolidinylinethoxyjbenzoate (690 mg, 98%) as a pale yellow foam. 'H-NMR (CDCI 3 8 2.35- 2.41 (in, 1 2.49-2.59 (in, 1 3.55 3 3.57-3.70 (in, 2 3.73-3.86 (in, 2 3.88 3 4.06-4.09 (in, 1 4.54-4.66 (in, 3 6.67-6.81 (in, 4 6.95-6.99 (in, 1 7.23-7.25 (in, 1 7.29-7.33 (in, 1 7.47-7.49 (in, 2 7.90-7.99 (in, 3 8.18-8.2 1 (in, 1 MS(FAB) m/z 586 1).
To a stirred solution of methyl 4-I(4R)-chloro-l-14-[N'-(2-chlorophenyl)ureidoJ-3methoxyphenylacetylJ-(2S)-pyrrolidinylmethoxylbenzoate (410 mag, 0.70 minol) 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- I N HCI and the resulting precipitate was collected under a reduced pressure. The crude solid was dissolved in CHCl 3 -MeOH and evaporated. The residue was washed with Et 2 O to give 105 (282 mg, as an amorphous solid.
MW 572.44 mp 131-136 0 C; 'H-NMR (DMSO-d) 8 2.29-2.35 (in, 1 2.44-2.47 (in, 1 3.58- 3.74 (in, 2 3.78 3 3.81-3.99 (in, 2 4.10-4.32 (in, 2 4.44-4.46 and 4.66 (each in, WO 01/00206 WO 0100206PCT/U SOO18079 total 1 4.84 (mn, 1 6.74-7.04 (in, 5 7.26-7.30 (in, 1 7.43-7.45 (mn, 1 7.87-7.9 1 (in, 2 7.96 J 8.3 Hz, I 8.09 J 8.3 Hz, I 8.90 1 8.94 1 Anal.
Calcd for C-AH 27 Cl 2
N
3
Q
6 i3/4H 2 O: C, 57.39; H, 4.90; CI, 11.66; N, 7.17. Found: C, 57.57; H, 4.94; Cl, 11.66; N, 6.89.
Example 98 4-1(4S)-hydroxy-1 '-(2-inethylphenyl)ureido~phenylacetyl]-(2.S)-pyrrolidinylinmethoxybenzoic acid Me H 6Me106 To a stirred solution of methyl 4-[(4S)-acetoxy-1-tert-butoxycarbonyl-(2S)-pyrrolidinyl]methoxy benzoate (2.31 g, 5.87 mmnol) in CH 2
CI
2 (46 ml) was added TFA (10 mld) at room temperature.
After 3.5 h stirring, the mixture was concentrated in vacuo. The residue was diluted by the addition of CH 2
CI
2 and 1 N NaOH, which were extracted with CH 2
CI
2 The combined extracts were washed with brine, dried over Na 2
SO
4 which was concentrated in vacu. The residue was chromatographed on silica gel 1100 g, CHCI 3 /MeOH(20/1)J to give tmethyl 4-[4S)-acetoxy-(2S)pyrrolidinyl~methoxybenzoate (1.89 mg, 100%) as a pale purple solid. 'H-NMR (CDCI,) 8 2. 314), 2.14 (mn, 1H), 2.65 (in, 114), 3.52-3.63 (mn, 3.89 3H), 4.18 (mn, lH), 4.28 J= 5.9 Hz, 214), 5.38 (mn, 1H), 6.93 J 8.8 Hz, 2H), 7.99 J 8.8 Hz, 2H).
A mixture of 3-methoxy-4-[N -(2-inethylphenyl)ureidolphenylacetic acid (343 ing, 1.09 inmol), methyl 4-[(4S)-acetoxy-(2S)-pyrrolidinyl]inethoxybenzoate (320 ing, 1.09 iniol), EDC-HCI (313 mg, 1.64 inmol), HOBT (222 ing, 1.64 inmol) and Et 3 N (0.76 ml, 5.45 minol) in DMF (7 nil) 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 NaSO 4 the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHClV/Aectone to give methyl 4-[(4S)-acetoxy-l1-[3-inethoxy4-[N -(2-inethylphenyl) ureido]phenylacetylJ-(2S)-pyrrolidinyl]inethoxybenzoate (520 ing, 81%) as a brown amorphous solid. 'H-NMR (CDCI 3 8 2.00 3H, one of isomers), 2.03 3H, one of isomers), 2.28 (mn, 3.54 lH), 3.58 2H), 3.64 lH), 3.67 and 3.69 (each s, 3H, ainide isomers), 3.85 J =5.4 Hz, lH), 3.88 314), 4.04 J= 9.3 Hz, 1H), 5.27-5.34 (in, 1H), 6.51 (mn, 1H), 6.76-6.89 (in, 2H), 6.94 J 8.1 Hz, lH), 7.14 (mn, 114), 7.25 (in, 414), 7.53 J= 8.3 Hz, 1H), 7.96 J 0 Hz, IlH), 8.00-8. 10 (mn, 2H); MS (ESI) m/z 590 (M4+1).
WO 01/00206 WO 0100206PCT/USOO/18079 To a solution of methyl 4-[(4S)-acetoxy- 1-14-IN -(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 midxture was extracted with EtOAc. The aqueous layer was acidified with 1 N HCI and extracted with CHCI,-MeOH The combined extracts were washed with brine. After dried over Na 2 SO,, the extracts were concentrated in vacuo. The residue was crystallized by the addition of CHCI~, EtOH and ether to give 106 (68 mg, 14%) as a colorless powder. MW 533.57 mp 148-152 *C IR (K-Br) 3356, 2939, 1687, 1604, 1533, 1454, 1255 cm';'H-NMR (DMSO-d) 8 1.95-2.09(i,2H), 2.25 3H), 3.59 (d,J= 5.9 Hz, 2H), 3.71 (in, 1H), 3.81 and 3.85 (each s, 3H, amide isomers), 4.13-4.47 4H), 5.19 (br, 1 6.70-7.21 (in, 7H), 7.79 J =7.9 Hz, I 7.86 J 8.8 Hz, 2H1), 8.01 J =8.3 Hz, 114), 8.47 114), 8.57 I MS (ESI) m/z 533 (M 4 Anal. Calcd for C 4-I 3
N
3
O
7 dIH 2 Q: C, 63.15; H, 6.03; N, 7.62. Found: C, 63.29; H, 5.76; N, 7.46.
Example 99 4-11-14-IN '-(2-chlorophenyl)ureidoj-3-methoxyphenylacetyl]-(4S)-hydroxy-(2S)-pyrrolidinyl methoxybenzoic acid Amixture of -(-hohnyl)ureido-3-methoxyphentylacetic acid (342 mg, 1.02 mniol), 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 mmnol) and Et 3 N (0.71 ml, 5. 10 mmol) in DMIF (6 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 NaSO,, the extracts were concentrated in vacuo. The residue was chromatographed on silica gel g, CHClV/Aectone (511 to give methyl 4-(4.S)-acetoxy- I -14-IN'-(2-chlorophenyl)uireidoJ -3 methoxyphenylacetylj-(2S)-pyrrolidinylmethoxybenzoate (510 mg, 82%) as a pale brown amorphous solid. 'H-NM~R (CDCI,) 8 2.0Oland 2.04 (each s, 3H4, amide isomers), 2.17 (in, 214), 3.56-3.66(m3H), 3.61 3H), 3.88 3H), 3.89(mn, 4.07 9.6Hz, 1H), 4.45 (dd,J= 9.2, 3.4 Hz, 1H), 4.56 (in, 1H), 5.3 1-5.39 (mn, 114), 6.80-7.01 (in, 4H), 7.23 J 8.1 Hz, 414), 7.34 J 8.1lHz, 1 7.95 J 8.5 Hz, 114), 8.00 (in, lH), 8.18 J 8.3 Hz, 1H); MS (ESI) m/lz 6 10 612 (MW+3).
To a solution of methyl 4-[(4.S)-acetoxy-l-[4-[N-(2-chlorophenyl)ureido]-3-methoxyphenylaceyl -(2.S)-pyrrolidinyllmethoxybenzoate (5 10 mng, 0.836 inmol) in THF (30 ml), 0.25 N NaOH (30 ml) WO 01/00206 WO 0100206PCT/USOOI 18079 was added. After stirrng at room temperature for 2 days, the mixture was extracted with EtOAc.
The aqueous layer was acidified with 1 N HCI and extracted with CHCI,-MeOH 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 (22 mg, as a colorless powder. MW 553.99 mp 138-142 0 C III (KBr) 3334, 2939, 1685, 1604, 1533, 1439, 1248cm-'; 2 H-NMR(DMSO-d)8 1.93-2.14(m,2H), 3.60(d,J=5.7 Hz, 2H), 3.71 (in, 1H), 3.81 and 3.85 (each s, 3H, amnide isomers), 4.14-4.50 (mn, 4H), 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 (in, 3H), 7.28 J 7.3 Hz, 1H), 7.43 (dd, J 8.1, 1.2 Hz, 1H), 7.86-7.95 (in, 2H), 7.97 J 8.1 Hz, 11H),8. 10(dd, J 1.5 Hz, 1H), 8.90 IH), 8.94 1H), 12.64 (br, 1H); MS (ESI) ni/z 554 556 (W 4 Anal. Caled for CgH~gClN 3 C, 60.7 1; H, 5.05; Cl, 6.40; N, 7.58.
Found: C, 60.47; H, 5.37; Cl, 6.3 1; N, 7.19.
Example 100 4-I(4S)-acetoxy-1 '-(2-bromophenyl)ureido]phenylacetylj-(2S)-pyrrolidinyl~methoxybenzoic acid r H 6Me108 A mixture of r(-oinpeyl)ureido]-3-methoxyphernylacetic acid (387 mg, 1.02 nunol), methyl 4-[(4S)-acetoxy-(2S)-pyrrolidintyl]inethoxybenzoate (300 mng, 1.02 mmol), EDC-HC1 (293 mg, 1.53 minol), HOBT (207 mg, 1.53 mmnol) and Et 3 N (0.71 ml, 5. 10 minol) in DMF (6 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 the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHClI/Aectone to give methyl 4-[(4S)-acetoxy-1-[4-[N'-(2-bromophenyl)ureidoj phenylacetyll-(2S)-pyrrolidinyl~methoxybenzoate (510 mg, as a yellow oil. 'H-NMR (CDCI,) 8 2.01 and 2.04 (each s, 3H, amiide isomers), 2.31 (in, 2H), 3.54-3.68 (in, 3H), 3.76 (s, 2H), 3.88 3H), 3.89-4.58 (in, 4H), 5.3 1-5.36 (mn, 1H), 6.81-6.96 (mn, 5H), 7.19-7.32 (in, 3H4), 7.51 J 8.0 Hz, 11H), 7.93-8.00 (in, 3H), 8.13 J 8.3 Hz, 1H); MS (ESI) m/z 654 656 To a solution of methyl 4-[4S)-acetoxy-I -(2-broinophenyl)ureido]phenylacetyl pyrrolidinyl]methoxybenzoate (5 10 mng, 0.779 inmol) in THE (30 ml), 0.25 N NaQH (30 ml) was added. After stirring at room temperature for 2 days, the mixture was extracted with EtOAc. The WO 01/00206 WO 0100206PCT/U 500/I8079 remaining aqueous layer was acidified with 1 N HC1 and extracted with CHCl 3 -MeOH The combined extracts were washed with brine. After dried over Na 2
SQ
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 'C IR (KBr) 3332, 2937, 1685, 1604, 1529, 1529, 1435 cm"; 'H-NDM (DMSO-d,) 8 1.92-2.14 (mn, 2H), 3.60 J =5.9 Hz, 214), 3.72 (im, 114), 3.81 and 3.85 (each s, 3H, amide isomers), 4.14-4.49 (in, 4H4), 5.19 (br, 1H), 6.72 and 6.75 (each in, 1H, anide isomers), 6.85 and 6.90 (each in, 1H, amide isomers), 6.97 J= 6.1 Hz, 1H), 7.06 J =8.8 Hz, 2H), 7.32 J =7.1 Hz, 1H), 7.60 (dd,J 7.8, 1.2 Hz, 111), 7.86 J 8.8 Hz, 211), 7.87-7.97 (in, 3H), 8.74 1H), 8.93 1H), 12.60 (br, 1H); MS (ESI) ni/z 559 561 Anal. Calcd for CuH112B 3
O
7 -0.1H 2 0: C, 56.03; HL 4.74; Br, 13.3 1; N, 7.00. Found: C, 55.80; H, 4.84; Br, 13.64; N, 6.66.
Example 101 1-14-[N '-(2-chlorophenyl)ureidol-3-inethoxyphenylacetyll-(4R).hydroxy-(2S)-pyrrolidiny methoxylbenzoic acid
_,OH
MI5ii r'e O~k~~Q109 To a stirred solution of methyl 4-[(4R)-acetoxy-l -Qert-butoxycarbonyl)-(2S)-pyrrolidinylinethoxy benzoate (835 mg, 2.12 minol) in CH 2
CI
2 (5 ml) was added TFA (5 ml) and the reaction midxture 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 CHCI 3 washed with brine, dried over K 2 C0 3 and evaporated to give methyl 4-[(4R)-acetoxy-(2S)-pyrrolidinylinethoxy]benzoate (580 mg, as a brown oil. 'H-NUR (CDCI 3 8 1.86-1.93 (in, I 2.00-2. 12 (series of s and in, totalS5 H), 3.03-3.29 (in, 1 3.73-3.80 (in, 1 3.88 3 3.93-4.01 (in, 2 5.27-5.30 (in, 1 6.91 J =9.0 Hz, 2 7.98 J 9.0 Hz, 2 MS (FAB) m/z 294 A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (365 mng, 1.09 inmol), methyl 4-[(4R)-acetoxy-(2S)-pyrrolidinylmethoxy~bnzoate (320 mng, 1.09 inmol), EDCI (250 mng, 1.30 inmol), HOBt (180 mg, 1.33 iniol) and Et 3 N (182 ml, 1.31 inmol) in THE (5 ml) was stiffed at room temperature for 2 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 chrmatography on silica-gel with CHC1 3 -MeOH (50:1, v/v) as eluent to give methyl 4-[(4R)-acetoxy-l-[4-[N'-(2-chlorophenyl)ureido]-3-methoxypheny acetylj-(2S)-pyrrolidinylmethoxy]benzoate (500 ing, 75%) as a white foam. 'H-NMR (CDC1 3 WO 01/00206 WO 0100206PCTUSOO/18079 2.01 3 2.03-2.05 (in, 1 2.20-2.26 (in, 1 2.37-2.43 (in, 1 3.59 2 3.62 3 3.66-3.87 (in, 2 3.89 3 4.074.09 (in, 1 4.48-4.51 (mn, 1 4.59 (in, 1 6.70- 6.82 (in, 4 6.97-7.01 (in, 1 7.24-7.35 (in, 4 7.92-7.98 (in, 3 8.12-8.21 (in, 1 MS (FAB) m/z 610 (M 4 1).
To a stirred solution of methyl 4-[(4R)-acetoxy-1-[4-[N -(2-chlorophenyl)ureidoj-3inethoxyphenylacetyl]-(2S)-pyrrolidinylmehoxyjbenzoate (500 ing, 0.82 mmol) in THE (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- I N HCl and the resulting precipitate was collected under a reduced pressure. The crude solid was purified by recrystallization from CHCl 3 -IPE to give 109 (223 mg, as a white crystalline powder. NM 553.99 mp 137- 142*C; 'H-NMR (DMSO-d 6 8 1.95-2.09 (in, 2 3.41-3.43 (in, 1 3.57 3 3.78 3 4.07-4.40 (series of m, total 4 5.07 (in, 1 6.72-6.74 (in, 1 6.85 (in, 1 6.99-7.03 (in, 3 7.25-7.29 (in, 1 7.42-7.43 (mn, 1 7.85-7.87 (in, 2 7.93-7.95 (in, 1 8.07- 8.09 (in, 1 8.88 1 8.92 1 12.65 (br s, 1 MS (FAB) m./z 554 (W 4 Anal Calcd for CnHuCIN 3 O7-1/2H 2 O: C, 59.73; H, 5.19; Cl, 6.30; N, 7.46. Found: C, 59.58; H, 5.32; Cl, 6.99; N, 7.2 1.
Example 102 4-[(4R)-hydroxy-l1-[3-methoxy-4-[N '-(2-methylphenyl)ureidojphenylacetyll-(2S)-pyrrolidiny inethoxy~benzoic acid e e110 A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureidojphenylacetic acid (320 mg, 1.02 nunol), methyl 4-[(4R)-acetoxy-(2S)-pyrrolidinylmethoxy~benzoate (300 ing, 1.02 inmol), EDC-HCl (235 mng, 1.23 inmol), HQBt (166 mg, 1.23 mmol) and Et 3 N (171 ml, 1.23 inmol) in THE (5 ml) was stirred at room temperature for 2 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 CHCI 3 -MeOH (50: 1, v/v) as eluent to give methyl 4-[(4R)-acetoxy- 1-[3-inethoxy-4-[N '-(2-methylphenyl)ureidolphenylacetyl]-(2S)pyrrolidinylinethoxy~benzoate (420 ing, as a white foam. 'H-NMR (CDC1 3 5 1.99 3 H), 2.02-2.05 (mn, 1 2.15-2.41 (series of s and m, total 5 3.55 3 3.57 2 3.63-3.73 (mn, 2 3.89 3 4.07-4. 10 (mn, 1 4.45-4.48 (mn, 1 4.57 (mn, 1 6.56 1 6.66 (in, 1 6.75-6.82 (in, 3 7.11-7.24 (mn, 4 7.54-7.56 (in, 1 7.92-7.94 (mn, 2 8.05- WO 01/00206 WO 0100206PCTLJSOOII8079 8.07 (in, 1 MIS (FAB) m/z 590 (Mr+1).
To a stirred solution of methyl 4-[(4R)-acetoxy-l-[3-methoxy-4-[N'-(2-methylphenyl) ureido]phenylacetylj-(2S)-pyrrolidinylmethoxybenzoate (420 mg, 0.71 innol) in THF (5 mld) was added 0.5 N NaOH (5 mld) and the reaction mixture was heated under reflux overnight. 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 purified by recrystallization from CHC1 3 -IPE to give 110 (182 mg, 48%) as a white crystalline powder. MW 53 3.57 mp 178-182 0
C;
IH-NMR (DMSO-d) 851.92-2. 10 (in, 2 2.23 3 3.40-3.44 (in, 1 3.56-3.67 (in, 3 H), 3.78 3 4.05-4.39 (series of mn, total 4 5.06 (in, 1 6.71-7.01 (in, 5 7. 10-7. 16 (in, 2 7.77-7.79 (in, 1 7.85-7.89 (in, 2 7.98-8.00 (in, I 8.45 1 8.54 1 12.59 (br s, 1 MS (FAB) m/lz 534 Anal. Calcd for C1H 313 1/27-H 2 O: 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)ureidolphenylacetylJ-2-pyrrolidinylmethyl- 1piperazinylacetic acid
N
0 Y j
COGH
H H OMe11 To a stirred solution of Nf-(tert-butoxycarbonyl) (4S)-fluoroprolinol (1.26 g, 5.75 minol), Et 3 N (4 ml, 28.5 iniol) and DMSO (4.1 ml, 57.5 inmol) in CH 2 C1 2 (20 ml) was added SO 3 -pyridine (2.74 g, 17.2 nunol). After 5 h stirring, the mixture was evaporated to remove CH 2
CI
2 and diluted with Et 2 O (200 nil). The solution was washed with 1 N HCI (200 mld) and brine (200 ml), dried over MgSO, and evaporated. The resulting residue was chroinatographed on silica gel with hexane- EtOAc 1) to give N-(tert-butoxycarbonyl) (4S)-fluoroprolinal (628 mng, 50%) as a yellow oil.
'H-NMR (CDC 3 51.41-1.47 (in, 9 2.02-2.48 (in, 2 3.47-3.94 (in, 2 4.16 and 4.29 (each d, each J 9.8 Hz, total 1 5.13 and 5.26 (each s, total 1 H).
To a stirred solution of N-Qtert-butoxycarbonyl) (4S)-fluoroprolinal (1.44 g, 6.63 inmol), ethyl 1-piperazinylacetate (1.71 g, 9.94 nunol) and AcOH (759 ul, 13.3 innol) in MeOH (20 ml) was added NaBH 3 CN (880 mng, 13.3 nunol). 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 MgSQ 4 and evaporated. The oily residue was WO 01/00206 WO 0100206PCT/USOO/18079 chromatographed on silica gel with CHCI 3 -MeOH (20:1) to give ethyl 4-[l-(tert-butoxycarbonyl)- (4S)-fluoro-2-pyrrolidinylmethyl]-l-piperazinylacetate (2.38 g, 95%) as a yellow oil.
A mixture of ethyl 4-fl -(tert-butoxycarbonyl)-(4S)-fluoro-2-pyrrolidinylmethyl]-1 piperazinylacetate (2.38 g, 6.37 mmol), TEA (5 ml) and CH 2
CI
2 (5 ml) was stirred for 3 h. The mixture was evaporated and the residue was made basic with sat. NaHCO 3 (100 ml). The mixture was extracted with CHCI 3 -MeOH 2 x 150 ml) and the combined extracts were dried over
K
2 C0 3 and evaporated to give ethyl 4-[(4S)-fluoro-2-pyrrolidinylmethylj-1 -piperazinylacetate (1.44 g, 83%) as a brown oil. IH-NMR (CDC 3 8 1.27 (dt, J 7.1, 2.0 Hz, 3 1.66-3.35 (series of m, 17 4.18 (dq, J= 7.1, 2.0 Hz, 2 5.09 and 5.22 (each m, total 1 H).
A mixture of ethyl 4-[(4S)-fluoro-2-pyrrolidinylmethyl]-1-piperazinylacetate (1.44 g, 5.27 mmol), 3 -methoxy-4-[N'-(2-methylphenyl)ureidojphenylacetic acid (1.66 g, 5.27 mxnol), 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 silica gel with CHCI 3 -MeOH (20: 1) as eluent to give ethyl 4-[(4S)-fluoro--3-methoxy--N'r-(2methyiphenyl) ureido~phenylacetylJ-2-pyrrolidinylmethyl]-1 -piperazinylacetate (2.47 g, 82%) as a yellow viscous solid. 'H-NMR (CDCI 3 5 1.24-1.29 (in, 3 1.92-4.36 (series of mn, 7 5.16 and 5.29 (each mn, total I 6.43 1 6.74-6.81 (mn, 2 7.12-7.2 9 (in, 4 7.50 J 7.8 Hz, 1 8.0 1-8.07 (mn, 1 H).
A mixture of ethyl 4-[(4S)-fluoro-l1-[3-inethoxy-4-[N'-(2-methylphenyl)ureidolphenylacetylJ-2pyrrolidinylmethylj-1-piperazinylacetate (1.0 g, 1.76 inmol) and 0.25 N NaOH (14 ml, 3.50 mrnol) in THE (15 ml) was stirred overnight. The mixture was neutralized with 1 N HCl and evaporated.
The residue was purified by ion exchage resin (DIAJON, HP2O) with H 2 0 to MeOH as eluent to give 111 MW 541.61 (400 mg, 40%) as a pale yellow amorphous solid. 'H-NMR (CD 3 OD) 2.00-3.95 (series of in, 24 4.34-4.40 (in, I 5.23 and 5.36 (in, each, total I 6.78-6.82 (in, 1 6.92 (in, 1 7.00-7.04 (in, 1 7.09-7.23 (in, 4 7.59 J 7.1 Hz, 1 7.99-8.02 (in, 1 MS(EAB) ni/z 542 (Mr+ 1).
Example 104 4-[I1-[4-[N'-(2-chlorophenyl)ureidoj-3-nethoxyphenylacetylJ-4,4-difluoro-2-pyrrolidinylnethyl- 1piperazinylacetic acid WO 01/00206 WO 0100206PCT/USOO/18079 I H 6Me112 To a stirred mixture of 1-(tert-butoxycarbonyl)-4,4-difluoro-2-pyrrolidinylmethano (2.11 g, 8.89 mmol), EtN (6.2 ml, 44.5 mmol), DMSO (6.3 ml, 88.9 mmol) in CH 2
CI
2 (20 mld) was added SO,-pyridine (4.25 g, 26.7 mmol). After 3 h stirring, the mixture was concentrated in vacua and diluted with Et 2 O (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 1) as eluent to give I1-(tert-butoxycarbonyl)-4,4-diluoro-2pyrrolidinecarbaldehyde (1.40 g, 67%) as a yellow oil. 'H-NMR (CDCI,) 8 1.45-1.52 (in, 9 H), 2.49 (in, 2 3.75-3.88 (in, 2 4.29-4.42 (in, I 9.54 and 9.60 each, total 1 H).
To a stirred solution of l-Qter:-butoxycarbonyl)-4,4-difluoro-2-pyrrolidinecarbadehyde (1.40 g, 5.95 mmol) and ethyl 1-piperazinylacetate (1.02 g, 5.95 minol) in MeOH-AcOH (12:1, 13 mld) 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 CHCl 3 (2 x 200 ml). The combined extracts were dried over MgSO, and concentrated in vacua. The residue was chromatographed on silica gel with CHCl 3 -MeOH (20: 1) as eluent to give ethyl 4-[l1-Qert-butoxycarbonyl)-4,4-difluoro-2pyrrolidinylmethylj-1.piperazinylacetate (822 mg, 35%) as a yellow oil. 'H-NMR (CDC 3 8 1.27 J= 7.1 Hz, 3 1.46 (in, 9 1.64 (in, 2 2.39-2.64 (in, 10 3.19 2 3.42-4.05 (series of m, 3 4.18 J 7. 1 H-z, 2 H).
A solution of ethyl 4-[1 -(tert-butoxycarbonyl)-4,4-difluoro-2-pyrrolidinylinethyl]-1piperazinylacetate (820 mg, 2.09 mmol) and TEA (5 ml) in CH 2
CI
2 (5 ml) was stirred for 1 h. The mixture was concentrated in vacua and the residue was made basic with sat. NaHCO 3 The resulting mixture was extracted with CHCI,-MeOH 2 x 200 ml). The combined extracts were dried over K 2 C0 3 and concentrated in vacua to give ethyl 4-(4,4-difluoro-2-pyrrolidinyl methyl)- 1 piperazinylacetate (493 mg, 81%) as a brown oil. 'H-NMR (CDCI,) 8 1.27 J 7.1 Hz, 3H), 1.91 (in, 2 2.27-2.60 (in, 10 3.09-3.34 (in, 4 3.46-3.53 (mn, 1 4.19 J= 7.1 Hz, 2
H).
A mixture of ethyl 4-(4,4-diluoro-2-pyrrolidinylmethyl)-1-piperazinylacetate (490 mg, 1.69 mniol), -(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (567 ing, 1.69 mmol), EDC-HCl (486 mng, 2.54 minol), HOBt (cat.) and DMAP (cat.) in DMF (10 ml) was stirred overnight. The mixture was diluted with EtOAc (250 ml), washed with brine (2 x 200 ml), dried over MgSQ 4 and concentrated in vacua. The residue was chromatographed on silica gel with
CHCI
3 -EtOAc 1) to CHCI 3 -MeOH (10: 1) as eluent to give ethyl 4-[l-[4-[N'-(2-chlorophenyl) WO 01/00206 WO 0100206PCT/USOO/18079 ureidoJ-3-methoxyphenylacetylJ-4,4-difluoro-2-pyrrolidinylmethyl] -1-piperazinylacetate (973 mg, as a yellow viscous oil. 'H4-NMR (CDC 3 8 1.25 J 7.1 Hz, 3 2.31-2.68 (in, 12 H), 3.17-3.20 (in, 2 3.52-3.91 (in, 4 4.10-4.48 (series of mn, 3 6.75-6.84 (in, 2 7.00 (dt, J 1.5 Hz, I1H), 7.16-7.29 3H), 7.35 (dd, J 1.5 Hz, 1 8.00 J =8.3 Hz, 1 H), 8.18 (dd, J 1.5 Hz, 1 H).
To a stirred solution of ethyl 4-[1 -[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-4,4difluoro-2-pyrrolidinylmethylJ- 1-piperazinylacetate (292 mg, 0.480 minol) in TI-F (4 ml) was added 0.25 N NaOH (3.8 ml, 0.96.0 inmol). After 2 days stirring, the mixture was neutralized with 1 N HCI and extracted with CHCl 3 -MeOH 2 x 200 ml). The combined extracts were dried over MgSO 4 and concentrated in vacua. The residue was purified by tin layer column chromatography on silica gel with CHCl 3 -MeOH 1) to give 112 MW 580.02 (81.7 mg, 29%) as a pale yellow amorphous solid. MW 580.02 'H-NMR (DMSO-4) 8 2.24-2.50 (series of m, 12 H), 3.404.47 (series of m, 10 6.76 J 8.1 Hz, 1 6.88 1 7.02 J 8.1 Hz, 1 H), 7.28 J= 8.1 Hz, 1 7.44 J=8.1 Hz, 1 7.97 J 8.1 Hz, 1 8.08 J =8.1 Hz, 1 8.96-8.99 (in, 2 MIS (FAB) m/z 580 (MX+1).
Example 105 4-[I-[3-methoxy-4-[N '-(2-methylphenyl)ureido]phenylacetyl]-(4S)-phenoxy-(2S)-pyrrolidinylj methyl-l1-piperazinylacetic acid Me H 6Me kN,-COOH 113 To a stirred mixture of methyl (2S,4R)-1-tert-butoxycarbonyl-4-hydroxy-2-pyrrolidinylcarboxylate (4.69 g, 19.1 mmol), phenol (1.98 g, 2 1.0 mmnol) and PPh 3 (5.51 g, 2 1.0 mmol) in THE (80 mld) was added DIAD (4.13 ml, 2 1.0 inmol) 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, CHCljEtOAc to give methyl (2S,4S)-1-tertbutoxycarbanyl-4-phenoxy-2-pyrrolidinylcarboxylate (5.31 g, 86%) as a colorless oil. 'H-NM~R (CDC1 3 8 1.43 (br, 9K, one of isomers), 1.48 (br, 9H, one of isomers), 2.48 (in, 1H), 3.75 (br, 3H), 4.42-4.96 (mn, 2H), 6.88-7.35 (mn, To a stiffed solution of methyl (2S,4S)-lI-Iert-butoxycarbonyl-4-phenoxy-2-pyrrolidiniylcarboxylate (5.31 g, 16.5 inmol) in THE (132 ml) was added 0.25 N NaOH (132 ml, 33.0 minol) 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 CHC1 3 The combined extracts were washed with brine, dried over Na 2
SO
4 and evaporated. The residue was WO 01/00206 WO 0100206PCT/USOO/18079 recrystallized from n-hexane-CHC1 3 to give (2S,4S)-1-tert-butoxycarbonyl-4-phenoxy-2pyrrolidinylcarboxylic acid (2.96 g, as a white powder. 'H-NMR (DMSOAd) 8 1.36 9H), 2.16 J 13.2 Hz, 111), 2.56 114), 3.46 (in, 111), 3.71 (dt, J =12.0, 5.4 Hz, 1H1), 4.26 (dt, J= 7.1 Hz, 111), 4.99 (mn, 6.85 (in, 211), 6.94 J 7.3 Hz, 114), 7.28 J 7.3 Hz, 111).
To a stirred solution of (2S,4S)-1-teri-butoxycaibonyl-4-phenoxy-2-pyrrolidinylcarboxylic acid (2.39 g, 7.76 iniol) in THE (50 ml) was added BH 3 -DMS (1.55 ml, 15.5 mmol) at 0 0 C. After min. stirring at the same temperature, the mixture was allowed to room temperature and then heated at 50 0 C for 2 h. After cooling to room temperature, the mixture was concentrated in vacuo and quenched by the addition of water at 0 The mixture was extracted with EtOAc. The combined extracts were washed with brine, dried over NaSO, and evaporated. The residue was chromatographed on silica gel [60 g ,CHCI 3 /MeOH to give (2S,4S)-1-tert-butoxycarbonyl- 4-phenoxy-2-pyrrolidinylmethaol (2.83 g, 100%/) as a colorless oil. '11-NMvR (CDCI3) 8 1.47 (s, 911), 1.95 (br, 111), 2.36 (in, 111), 3.56-3.74 31-I), 3.89-4.52 (in, 311), 4.85 (br, 1141), 6.84 (dd, J 8.8, 1.2 Hz, 211), 6.97 J 7.2 Hz, IH), 7.29 2H, J 7.8 Hz).
To a stirred mixture of (2S,4S)- I -tert-butoxycarbonyl-4-phenoxy-2-pyrrolidinylmethano (2.75 g, 9.37 inxol), Et 3 N (7.84 ml, 56.2 mniol), DMSO (6.66 ml, 9.37 mmol) in CH 2 C1 2 (30 ml) at 0 0
C
was added SO 3 pyridine (4.47 g, 28.1 inmol), then the resulting mixture was allowed to raise to room temperature. After 2.5 h stirring, the mixture was concentrated in vacua. To the resulting mixture was added water and extracted with Et 2 O. The combined extracts were washed with brine, dried over Na 2
SO
4 and concentrated in vacua. The residue was chroinatographed on silica gel [100 g, CHCl,/acetone to give (2S,4S)-1-tert-butoxycaibonyl-4-phenoxy-2-pyrrolidine carbaldehyde (2.54 g, 93%) as a yellow oil. 'H-NfM (CDC 3 5 1.45 9K1 one of isomers), 1.49 9H, one of isomners), 2.17 (br, 21), 3.65-4.31 3H), 4.91 (br, I1H), 6.79 J =7.8 Hz, I1-H), 6.77 (in, 11-1), 7.28 (in, 211), 9.66 (in, I1-H).
To a stirred mixture of 1-tert-butoxycarboniyl-(4S)-phenoxy-(2S)-pyrrolidinecarbaldehyde (1.36 g, 4.67 naiol), ethyl 1-piperazinylacetate (1.61 g, 9.37 mfnol) in TIHE (30 ml) was added NaBH(OAc) 3 (1.98 g, 9.34 inmol) 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, and concentrated in vacua.
The residue was chromatographed on silica gel [50 g, CHClV/MeOH to give ethyl 4-[1iert-butoxycarbonyl-(4S)-phenoxy-(2S)-pyrrolidinyl]methyl-l -piperazinylacetate (1.05 g, 50%) as a colorless oil. 'H-NMR (CDCI 3 5 1.27 J= 7.3 Hz, 31-4), 1.57 91-I), 2.18 (in, 111), 2.33-2.74 (in, 911), 3.17 211), 3.52-4. 10 (in, 4.17 J 7.3 Hz, 211), 4.89 (br, 111), 6.84 J 6.8 Hz, 2H1), 6.95 (in, 11-I), 7.26 (in, 311).
WO 01/00206 WO 0100206PCTI USOOI 18079 To a stiffed solution of ethyl 4-[l-tert-butoxycarbonyl-(4S)-phenoxy-(2S)-pyrrolidiniyl~methyl-lpiperazinylacetate (1.05 g, 2.35 mmol) in CHCl 2 (20 ml) was added TEA (20 ml) at room temperature. After 3 h stirring, the mixture was concentrated in vacuo, which was diluted with CHCI,-MeOH (10/1) and made basic by the addition of I N NaOH. The combined reaction mixture was extracted with CHCl 3 -MeOH The organic layer was washed with brine, dried over NaSO, and concentrated, to give ethyl 4-[(4S)-phenoxy-(2S)-pyrrolidinyljmethyl-lpiperazinylacetate 12 g, quant.) as a brown oil, which was used without further purification.
'H-NMR (CDCI,) 8 1.25 (tt, J= 7.1, 7. 1 Hz, 314), 1.91 J= 12.0 Hz, 11-1), 2.42-2.85 (in, 101-1), 3.22 2H), 3.50 214), 3.54-3.82 (in, 214), 4.15 J= 7.1 Hz, 214), 4.98 (br, I1H), 6.84 J 8.1 Hz, 214), 6.76 J 7.1 Hz, 11H), 7.26-7.31 (in, 314), 7.40 (br, 1H4).
A mixture of 4-[N'-(2-methylphenyl)ureido]-3-methoxyphenylacetic acid (337 mng, 1.07 mmol), ethyl 4-[(4S')-phenoxy-(2.S)-pyrrolidinyljmethyl- 1-piperazinylacetate (373 mg, 1.07 minol), EDC-HCI (308 mg, 1.61 mmnol) and DMAP (197 mg, 1.61 inmol) in DM (6 mld) 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 [50 g, CHC1 3 IMeOH (50/1)1, to give ethyl 4-[1 -[3-methoxy-4-[N'-(2-methylphentyl)ureido~phenylacetyl]- (4S)-phenoxy-(2S)-pyrrolidinyl~metbyl-l-piperazinylacetate (430 mg, 62%) as a colorless amorphous solid. 'H-NMR (CDC1 3 6 1.25 J= 7.8 Hz, 3H), 1.98-2.17 (mn, 214), 2.26 3H4), 2.36-2.82 (in, 1114), 3.13 114), 3.17 11-1), 3.55 J= 2.4 Hz, 11-1), 3.66 J= 0.9 Hz, 314), 3.67-3.84 (in, 214), 3.98-4.35 (in, 3H4), 4.854.95 (in, 114), 6.27-6.89 (mn, 614), 7.08 J= 7.3 Hz, 1141), 7.19 J= 6.8 Hz, 11H), 7.28 (mn, 214), 7.41 J= 4.9 Hz, 114), 7.55 J 7.8 Hz, 1141), 8.05 (dd, J 7.3, 2.2 Hz, 114). For HCI salt: a pale brown amorphous solid. IR (KBr) 3265, 3059, 1747, 1533, 1225 MIS (FAB) m/z 644 Anal. Calcd for C3A4 4
N
5
Q
6 -HCI-2. 1H 2 0: 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-[I-[3-inethoxy-4-[N'-(2-inethylphenyl)ureidolphenylacetyl]-(4S)-phenoxy- (2S)-pyrrolidinyllmethyl-1-piperazinylacetate (240 mg, 0.373 minol) in THE (3.0 ml), 0.25 N NaOH (3.0 mlJ) was added. After stirring at room temperature for 20 h, the mixture was neutralized with 1 N HCI and extracted with CHCI,-MeOH The combined extracts were dried over Na 2 SO, and concentrated in vacuo. The residue was triturated by the addition of ether, to give 113 MW 615.72 (143 mng, as a white powder. LR (KiBr) 3346, 2949, 1633, 1533, 1227 'H-NMR (DMSO-d 6 5 1. 76 (mn, 114), 2.18 (br, 214), 2.25 314), 2.42-2.83 (mn, 914), 3.17 114), 3.20 1141), 3.38 (in, IH), 3.70 214), 3.72-3.78 (in, 214), 3.85 314, one of isomers), 3.87 314 one of isoiners),.3.95 (in, 1141), 4.27 (br, 114), 5.08 (in, 114), 6.75 J =8.3 Hz, 114), 6.93 (in, 514), 7.14 (in, 214), 7.30 J 7.6 Hz, 114), 7.79 J 9.0 Hz, 114), 8.02 (in, 114), 8.50 114), 8.58 114); MIS (FAB) n/z 616(M'+ Anal. Calcd for WO 01/00206 WO 0100206PCT/USOO/18079
CH
2 N5O 6 1EtOH-2H 2 O: C, 62.58; H, 7.00; N, 10.67. Found: C, 62.73; H, 6.58-; N, 10.24.
Example 106 1-14-LA' -(2-chlorophenyl)urido-3-methoxyphenylacetyl]-(4S)-phenoxy-(2S)-pyrrolidinyl] methyl-I -piperazinylacetic acid mixture of hnyl)ureido]-3-methoxyphenylacetic acid (358 mg, 1.07 minol), ethyl 4-[(4S)-phenoxy-(2.S)-pyrrolidinylmethylJ-1-piperazinylacetate (373 mng, 1.07 mmol), EDC-HCI (308 mg, 1.61 mmol) and DMAP (197 mng, 1.61 inmol) in DMF (6 m-l) 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 vacua. The residue was chromatographed on silica gel [150 g, CHC13/MeOH to give ethyl 1-[4-[N'-(2-chlorophenyl)ureidoj-3-methoxyphenylacetylJ- (4S)-phenoxy-(2S)-pyrrolidinyl]methyl-l -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 (in, 214), 2.35-2.80 (in, I 3.12 and 3.18 (each s, 1I1, amide isomers), 3.57 J= 5.4 Hz, I1H), 3.66 and 3.3 8 (each s, 3K, amide isomers), 3.70-3.90 (in, 2H), 4.024.43 (in, 3H), 4.88-4.97 (in, 11H), 6.83-6.99 (mn, 61-f), 7.18-7.32 (in, 314), 7.68 J 8.3 Hz, 11H), 7.74 1H), 8.16 (dd, J 8.3, 1.4 Hz, 114).
For HCI salt: a pale brown amorphous solid. IR (KBr) 3300, 2978, 1745, 1533, 1225 MIS (FAB) mlz 664 666 Anal. Caled for C 35 HI4C1N 5
O
6 -1H1l-2.4H4 2 O: C, 56.51; H, 6.48; N, 9.41. Found: C, 56.51; H, 6.18; N,9.28.
To a solution of ethyl 4-[1-[4-[M,-(2-chlorophenyl)ureido]-3-inethoxyphenylacetyl]-(4S)-phenox(y- (2S)-pyrrolidinyl]methyl-l-piperazinylacetate (181 ing, 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 CHCI 3 -MeOH The combined extracts were dried over Na 2
SO
4 and concentrated in vacua. 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, 163 1, 1595, 1225 'H-M (DMSO-dj 5 2.13-3.05 (in, 1 1H), 3.22 and 3.36 (each s, 2H, amide isomer), 3.38 (mn, 3.60 214), 3.71 (in, 1ff), 3.85 3Hf), 3.95 (in, 1ff), 4.28 (br, 5.06 (in, 1ff1), 6.76 J= 8.3 Hz, 1ff1), 6.9 1-7.03 (in, 5ff), 7.29 (in, 3ff), 7.44 J 7.9 Hz, 1ff), 7.97 (dd, J= 8.1, 4.1 Hz, 1ff), 8.08 J= 7.0 Hz, 1ff), 8.91 1ff), 8.95 1ff1); MS (FAB) m/z 636(M'+1), 638(M'+3); Anal. Calcd for C3H3,CN,O 6 -0.2EtOH- 1 .3H 2 0: C, 59.98; H, 6.30; N, 10.47. Found: C, 60.25; H, 6.12; N, 10. 11.
WO 01/00206 WO 01/1)206PCT/USOO/18079 Example 107 441 '-(2-bromopheniyl)ureidoJ-3-methoxyphenylacetyl-4S)-phenoxy(2S)-pyrrwlidinyl methyl-1-piperazinylacetic acid ~r H Me K..N -COOH 115 A mixture of 4-[NV'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (406 mg, 1.07 rnmol), ethyl 4-[(4S)-phenoxy-(2S')-pyrrolidinyl]methyl-1-piperazinylacetate (373 mg, 1.07 mmol), EDC-HCI (308 mg, 1.61 mmol) and DMA" (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 extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHCI,/MeOH to give ethyl 4-[1-[44[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetyl]- (4S)-phenoxy-(2S)-pyrrolidinyl~methyl-1-piperazinylacetate (560 mg, as a colorless amorphous solid. 'H-NNM (CDCI 3 5 1.25 (tt, J= 7.1, 7.1 Hz, 2.04-2.84 (in, 13H-), 3.12 and 3.18 (each s, I1H, amide isomers), 3,57 J =4.4 Hz, 111), 3.66 and 3.68 (each s, 3H, anide isomers), 3.68-3.87 (in, 311), 4.054.41 (in, 211), 4.87-4.96 (in, 11-1), 6.76-6.98 (in, 611), 7.20-7.33 (mn, 311), 7.46 J= 8.1 Hz, 11-1), 7.67 and 7.71 (each s, I1H, amide isomers), 7.78 and 7.81 (each s, 1H, amide isomers), 7.95 J 8.3 Hz, 8.09 J 8.1 Hz, 111). For HCl salt: a pale brown amorphous solid. JR (KBr) 3 384, 2978, 1745, 1340, 1120 MIS (FAB) m/lz 708 1), 710 Anal. Calcd for CH51-4BrN 5
O
6 -H10-2.5H 2 O0: C, 53.20; H, 6.12; N, 8.86. Found: C, 52.98; H, 5.79; N, 8.66.
To a solution of ethyl 441 -[4-[M,-(2-broinophenyl)ureido]-3-methoxypheniylacetyl]-(4S)-phenoxy- (2S)-pyrrolidinyl~methyl-l-piperazinylacetate (330 mg, 0.466 mmol) in TI-F (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 The combined extracts were dried over Na 2 SO, and concentrated in vacuo. The residue was triturated by the addition of ether, to give 115 (217 mg, as a white powder. MW 680.59 IR (KBr) 3315, 3095, 2941, 163 1, 1529, 1435 cm'; 'H-NMikR (DMSO-d) 8 1.75 (in, 111), 2.18 and 2.23 (each s, 2K, amide isomers), 2.30-2.78 (in, 911), 3.15 211), 3.47 (mn, 111), 3.58 211), 3.60-3.82 (in, 311), 3.84 (in, 311), 3.93 (in, 111), 4.26 (br, 11-1), 5.08 (in, 111), 6.75 J= 7.8 Hz, 114), 6.98 (in, 51-1), 7.30 311), 7.60 (dd, J= 8.1, 2.2 Hz, 111), 7.94 (in, 211), 8.75 IH), 8.93 111); MS(FAB) n/z 680(MW+1), Anal. Calcd for C33H3BrN 5
O
6 -0.2EtOH-2HO: C, 55.27; H, 6.00; N, 9.65. Found: C, 55.32; H, 5.56; N, 9.25.
WO 01/00206 WO 0100206PCT/USOO/18079 Example 108 4-[(4S)-(4-carboxyphenoxy)-1 -[3-methoxy-4-[N '-(2-methylphenyl)uriedo]phenylacetyl]-(2S)pyrrolidinylmethoxylbenzoic acid 10-0-CO00H e HOOHM 116 To a stirred solution of methyl 1-QIert-butoxycarbonyl)-(4R)-hydroxy-(2.S)-pyrrolidinylcarboxylate (10.4 g, 0.04 mol) and imidazole (8.66 g, 0. 13 mol) in DMF (40 ml) was added mBs-cl (7.03 g, 0.05 mol) and the reaction mixture was stirred at 60'C for 3 hr. After cooled to room temperature, the mixture was diluted with brine, and extracted with Et 2 O. The extract was washed with brine, dried over Na 2 SO,, and evaporated. The residue was purified by column chromatography on silica-gel with n -hexane-EtOAc 1, v/v) as eluent to give methyl 1 -(terI-butoxycarbonyl)-(4R)- (ieri-butyldimethylsilyloxy)-(2S)-pyrrolidinylcarboxylate (15.0 g, as a colorless oil. 'H-NMR (CDC1 3 5 0.06 6 0.87 9 1.41 and 1.46 (each s, 9 1.99-2.03 (in, I 2.16-2.18 (mn, 1 3.3 1-3.42 (in, 1 3.56-3.63 (in, 1 3.73 and 3.74 (each s, 3 4.31-4.42 (in, 2 H); MS (ESI) ,n/z 360 (MW+1).
To a stirred solution of methyl 1 -Qert-butoxycarboniyl).<4R)-(tert-butyldimethylsilyloxy)-(2S)pyrrolidinylcarboxylate (15.0 g, 0.04 inol) in THE (60 mld) was added I N NaQH (60 mld) and the reaction mixture was stirred at 60*C for 2 hr. After cooled to room temperature, the mixture was concentrated to a small volume, 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 -Qert-butoxycarbonyl)-(4R)- (tert-butyldimethylsilyloxy)-(2S)-pyrrolidinylcarboxylic acid (12.8 g, 89%) as a colorless oil. IH- NMR (CDCI 3 850.07 and 0.08 (each s, 6 0.87 9 1.49 9 2.06-2.11 (in, I 2.4 1- 2.44 (in, I 3.40-3.59 (mn, 2 4.36-4.50 (in, 2 MS m/z 346 1).
To a cooled (0 0 C) stirred solution of 1 -Qert-butoxycarbonyl)-(4R)-Qtert-butyldimethylsilyloxy)- (2S)-pyrrolidiniylcarboxylic acid (12.8 g, 0.04 mol) in THF (150 mlJ) was added dropwise BH 3
DMS
(5.30 ml, 0.06 mol) and the reaction mixture was stirred at room temperature overnight. The reaction mixture was quenched by sat. NH4CI, 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 1, v/v) as eluent to give 1-(tert-butoxy carbonyl)-(4R)-(tert-butyldimethylsilyloxy)-(2S)-prolinoI (10.5 g, as a colorless oil. IH-NMR
(CDCI
3 5 0.06 6 0.87 9 1.47 9 1.58-1.63 (in, 1 1.93-1.98 (in, 1 3.32- WO 01/00206 WO 0100206PCT/US00118079 3.44 (in, 2 3.51-3.57 (in, 1 3.67-3.71 (in, 1 4.13-4.15 1 4.27 (in, 1 4.87- 4.89 (in, 1 H).
To a cooled (0 0 stirred solution of methyl 4-hydiroxybenzoate (4.81 g, 0.03 mol), 1-Qertbutoxycarbonyl)-(4R)-(tert-butyldimetbylsilyloxy)-(2S)-prolino (10.5 g, 0.03 niol), and Ph 3 P (9.96 g, 0.04 mol) in THE (160 ml) was added dropwise DIAD (7.48 nml, 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 1, v/v) to give methyl 4-[l1-Qert-butoxycarboniyl)-(4R)-(Qer-butyldiinethylsilyloxy)-(2S)pyrrolidinylmethoxy]benzoate (9.58 g, as a white solid. mp 86-88 0 C; IH-NMvR (CDCI,) 8 0.08 6 0.88 9 1.46 9 2.04-2.15 (in, 2 3.29-3.48 (mn, 2 3.88 3 H), 4.064.30 (in, 3 4.46-4.51 (in, 1 6.91-6.93 (in, 2 7.96-7.98 (in, 2 MS (ESI) m/z 466 To a cooled (0 0 stirred solution of methyl 4-[1-(tert-butoxycarbonyl)-(4R)-(tert-butyldiniethyI silyloxy)-(2.S)-pyrrolidinylinethoxylbenzoate (1.49 g, 3.20 nimol) in THE (15 ml) was added TBAF (6.40 ml, 6.40 inmol, 1 M solution in THEF) 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,, and evaporated. The residue was purified by column chromatography on silica-gel with toluene-acetone 1, v/v) as eluent to give methyl 4-[1 -QerI-butoxycarbonyl)-(4R)-hydroxy-(2S)pyrrolidinylmethoxyjbenzoate (1.05 g, 93%) as a white solid, nip 103-105 0 C; 'H-NMvR (CDCI 3 8 1.46 9 2.11-2.28 (in, 2 3.49-3.60 (in, 2 3.88 3 4.154.34 (in, 3 4.53-4.57 (in, 1 6.91 J 8.6 Hz, 2 7.97 J 8.6 Hz, 2 MIS (ESI) m/z 352 (W 4 To a cooled (0 0 stirred solution of methyl 4-hydroxybenzoate (0.56 g, 3.68 innol), methyl 4-[1- (tert-butoxycarbonyl)-(4R)-hydroxy-(2S)-pyrrolidinylinethoxyjbenzoate (1.30 g, 3.70 inuol), and Ph 3 P 16 g, 4.42 inmol) in THF (20 ml) was added dropwise DIAD (0.87 ml, 4.42 inmol) and 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 1, vfv) as eluent to give methyl 4-[1-QIert-butoxycarbonyl)-(4S)-(4-methoxycarbonylphenoxy)-(2S)pyrrolidinylmethoxylbenzoate (1.80 g, as a colorless oil. 'H-NMR (CDCI,) 8 1.49 9 H), 2.3 1-2.38 (in, 1 2.45-2.49 (in, 1 3.64-3.77 (in, 2 3.88 6 4.07-4.15 (in, 1 H), 4.33-4.44 (in, 2 4.95-5.01 (in, 1 6.85 J 8.8 Hz, 2 6.94 (br s, 2 7.97 J =8.8 Hz, 4 MIS (ESD) m/z 486 1).
WO 01/00206 WO 0100206PCT/USOO/18079 To a stirred solution of methyl 4-11 -Qeri-buioxycarbonyl)-(4S)-(4-methoxycarbonyl phenoxy)-(2S)-pyrrolidinylmethoxy~benzoate (1.80 g, 3.71 mmuol) in CH 2
CI
2 (15 mld) was added TFA (15 ml) and the reaction mixture was stirred at room temperature for 1.5 hr. The mixture was concentrated in vacua, made basic by sat. NaHCO 3 and extracted with CHC 3 The extract was washed with brine, dried over K 2 C0 3 and evaporated to give methyl methoxycarbonyl phenoxy)-(2S)-pyrrolidinylmethoxyjbenzoate (1.50 g, as a pale yellow oil.
'H-NMR (CDC1 3 6 1.87-1.92 (in, 1 2.41-2.48 (in, 1 3.18-3.23 (in, 11-1), 3.34-3.37 (in, 1 3.60-3.66 (in, 1 3.88 3 3.89 3 4.04-4.13 (in, 2 4.94-5.00 (in, 1 6.87- 6.93 (in, 4 7.96-8.00 (in, 4 MS (ESI) m/lz 386 1).
A mixture of 3-methoxy-4-[N-(2-methylphenyl)ureidolphenylacetic acid (400 mg, 1.27 inmol), methyl 4-[(4S)-(4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxy]benzoate (491 mg, 1.27 minol), EDCHC1 (293 mg, 1.53 inmol), HOBt (207 mng, 1.53 minol), and Et 3 N (215 1 dl, 1.54 mmol) in THF (10 ml) was stirred at room temperature overnight. The mixture was diluted with H,0, and extracted with EtOAc. The extract was washed with brine, dried over NaSO,, and evaporated. The residue was purified by column chromatography on silica-gel with CHC1 3 -MeOH 1 to 50: 1, v/v) as eluent to give methyl -methoxy-4-[N'-(2-methylphenyl)uriedoI phenylacetyll-4S)-(4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxybenzoate (532 mg, as a white foam. 'H-NMR (CDC1 3 8 2.26-2.49 (series of s and m, total 5 3.56-3.93 (series of sand mn, total 13 4.07-4.59 (series of in, total 3 5.01 (in, 1 6.69-6.94 (mn, 7 7.09-7.13 (in, I 7.20-7.3 1 (in, 3 7.52-7.57 (mn, 1 7.92-8.00 (in, 4 8.06-8.09 (in, 1 MS (ESI) m/z 682 To a stirred solution of methyl 44 1-[3-inethoxy-4-[N'-(2-inethylphenyl)uriedojphenylacetyl (4-inethoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxybnzoate (532 mg, 0.78 minol) in THF nil) 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-lI N HCI and the resulting precipitate was collected. The crude solid was recrystallized from MeOH-CHC 3 -Et 2 O to give 116 (125 mng, 25%) as a pale yellow crystalline powder. MW 653.68 mp 154-159*C; 'H-NMR (DMSO-d)2.24 3 2.38-2.49 (in, 2 3.63 2 3.67-3.88 (series of s and in, total 4 H), 4.01-4.06 and 4. 15-4.19 (each m, total 2 4.27-4.31 and 4.38-4.42 (each in, total 2 5.18- 5.25 (mn, 1 6.72-6.77 (in, 1 6.85-7. 16 (series of mn, total 8 7.78-7.89 (in, 5 7.99-8.02 (in, 1 8.46 1 8.57 1 12.65 (br s, 2 MIS (ESI) m/z 654 Anal Calcd for
C
36
H
35
N
3 0 9 11/2H 2 0: C, 65.25; H, 5.48; N, 6.34. Found: C, 65.29; H, 5.54; N, 6.20.
WO 01/00206 WO 0100206PCT/USO0I 18079 Example 109 4-[(4S)-(4-carboxyphenoxy)- '-(2-chlorophenyl)uriedo]-3-methoxyphenylacetyl]-(2S)pyrrolidinylmethoxyjbenzoic acid
__OOH
~AN-Kr&OOH I H 6Me117 A mixture of -(chroenyl)ureido]-3-methoxyphenylacetic acid (420 mg, 1.25 mmol), methyl 4-[(4S)-(4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxylbenzoate (483 mg, 1.25 mmol), EDCHCI (288 mg, 1. 50 mmol), HOBt (203 mg, 1. 50 mmol), and Et 3 N (2 10 pl, 1. 51 mmol) in THE (10 mld) was stirred at room temperature overnight. The mixture was diluted with H,0, and extracted with EtOAc. The extract was washed with brine, dried over Na 2 SO., and evaporated. The residue was purified by column chromatography on silica-gel with CHC1 3 -MeOH 1, v/v) as eluent to give methyl 4-[1-[4-[N'-(2zhldorophernyl)uriedoj-3-methoxypheniylacetyl]- (4S)-(4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxylbenzoate (488 mg, as a white foam. IH-NMR (CDCI 3 8 2.28-2.51 (in, 2 3.62-3.94 (series of s and m, total 13 4.07-4.62 (series of m, total 3 4.99-5.03 (in, 1 6.78-6.99 (in, 7 7.23-7.34 (in, 2 7.42-7.52 (mn, 2 7.92-8.01 (in, 5 8.17-8.20 (in, 1 MS (EST) m/z 702 To a stirred solution of methyl 4-f 1-f4-[N'-(2-chlorophenyl)uriedol-3-inethoxyphenylacetylJ-(4S)- (4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxylbenzoate (488 mg, 0.70 innol) in THF ml) was added 0.5 N NaOH (5 mlJ) and the reaction mixture was heated under reflux for 3 hr.
After cooled to room temperature, the mixture was poured into ice- I N HC1 and the resulting precipitate was collected. The crude solid was recrystallized from MeOH-CHC1 3 -Et 2 O to give 117 (137 mug, as a white crystalline powder. MW 674.10 np 150-153*C; 'H-NMR (DMSO-d) 8 and 4.17-4.21 (each in total 2 4.30-4.34 and 4.40-4.45 (each m, total 2 5.20-5.27 (mn, 1 H), 6.77-6.81 (in, 1 6.89-6.92 (in, 1 7.0 1-7.08 (mn, 5 7.27-7.3 1 (in, 1 7.43-7.46 (in, 1 7.86-7.92 (in, 4 7.97-8.00 (in, 1 8.10-8.12 (in, 1 8.91 1 8.96 1 12.65 (br s, 2 MIS (ESI) m./z 674 AnaL Calcd for C 33
H
32
CIN
3 0 9 1/4H 2 0: C, 61.95; 114.83; N,6.19; CI,5.22. Found: C,61.77; HK4.86; N,6.13; Cl, 5.49.
Example 110 4-[I1-14-IN '-(2-bromophenyl)uriedoj-3-methoxyphenylacetyl]-(4S)-(4-carboxyphenoxy)-(2S)pyrrolidinylmethoxy]benzoic acid WO 01/00206 WO 0100206PCT/USOOI 18079 H OOH 118 A mixture of 4-[N'-(2-bromophenyl)ureidoJ-3-methoxyphenylacetic acid (464 mg, 1.22 mmol), methyl 4-[(4S)-(4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxylbenzoate (472 mg, 1.22 mmol), EDC-HC1 (282 mg, 1.47 mmol), HOBt (200 mg, 1.48 inmol), and Et 3 N (205 jil, 1.47 mniol) in THE (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 NaSO 4 and evaporated. The residue was purified by column chromatography on silica-gel with CHCI 3 -MeOH (60:1 to 50: 1, v/v) as eluent to give methyl 4-[1-[4-IjN'-(2-bromophenyl)uriedoj-3-methoxyphenyl acetyl]-(4S)-(4-methoxycarbonylphenoxy)-(2S)-pyrrolidinylmethoxybelzoate (379 mg, 41%) as a white foam. 'H-NMR (CDCI 3 5 2.28-2.51 (in, 2 3.59-3.95 (series of s and m, total 13 H), 4.074.62 (series of m, total 3 4.99-5.03 (mn, I 6.79-6.95 (mn, 7 7.27-7.36 (in, 3 H), 7.49-7.51 (mn, 1 7.93-8.01 (mn, 5 8.11-8.14 (in, 1 MS (EST)m/Vz 747 To a stirred solution of methyl 4-[1-[4-[N'-(2-bromophenyl)uriedo]-3-methoxyphenylacetyl]-(4S)- (4-methoxycarbontylphenoxy)-(2S)-pyrrolidinylmethoxy~benzoate (379 mg, 0.51 mmol) in THF mld) 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-l N HCI and the resulting precipitate was collected. The crude solid was purified by preparative TLC to give 118 (51 mg, as a pale yellow amorphous solid. MW 718.55 'H-NMR (DMSO4),3 2.20-2.40 (in, 2 H), 3.65-3.89 (series ofim, total 6 4.024.63 (series of mn, total 4 5.19-5.26 (in, 1 6.74-7.06 (in, 7 7.30-7.34 (in, 1 7.59-7.6 1 (in, 1 7.83-7. 96 (in, 6 8.74 1 8.93 1 H); Anal Calcd for C 31 1-lBrN 3
O
9 -2H 2 O: C, 55.71; HK 4.81; N, 5.57. Found: C, 55.92; H, 4.80; N, 5.30.
Example I111 4-[(4S)-(4-carboxyphenoxy)- 1-14-IN '-(2-methylphenyl)uriedolphentylacetylJ-(2S)-pyrrolidiny methoxyjbenzoic acid N
OOH
H OH 119 A mixture of 4-[N -(2-methylpheryl)ureidolphenylacetic acid (328 ing, 1. 15 iniol), methyl 4- [(4S)-(4-methoxycarbonylphenoxy)-(2.S)-pyrrolidinylinethoxy~bnzoate (444 mg, 1. 15 nunol), WO 01/00206 WO 0100206PCT/USOO/1 8079 EDCHCI (265 mg, 1.38 mmol), HOBt (187 mg, 1.38 nunol), and Et~N (195 p.1, 1.40 nimol) in THE (10 ml) was stirred at roam 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 (60:1 to 50: 1, v/v) as eluent to give methyl 4-[(4S)-(4-methoxycarbonylphenoxy)-1-[4-[N '-(2-methyl phenyl)uriedo]phenylacetylJ-(2S)-pyrrolidinylmethoxy~benzoate (332 mg, 441%) as a white foam.
'H-NMR (CDC1 3 8 2.13 3 2.24-2.48 (in, 2 3.52-3.90 (series of s and m, total 10 H), 4.05-4.58 (series of mn, total 3 5.01 (in, 1 6.78-6.90 (in, 4 6.98-7.20 (in, 8 7.51-7.56 (in, 2 7.90-8.00 (in, 4 MIS (ESI) m/z 652 1).
To a stirred solution of methyl 4-[(4S)-(4-inethoxycarbonylphenoxy)-1-[4-[N'-(2-nethyI phenyl)uriedo]phenylacetyl]-(2S)-pyrrwlidinylnethoxy]benzoate (332 mg, 0.51 minol) in TJHE 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-i N HC1 and the resulting precipitate was collected. The crude solid was recrystallized from MeOH-CHC 3 -Et 2 O to give 119 (118 mg, 37%) as a white crystalline powder. MW 623.65 mp 157-160'C; 'H-NMR (DMSO-d) 8 2.20-2.25 (series of s and in, total 4 2.39-2.47 (in, 1 3.64 2 3.68-3.89 (in, 1 H), 4.024.08 and 4. 16-4.20 (each in, total 2 4.29-4.33 and 4.39-4.43 (each mi, total 2 5.20- 5.26 (in, 1 6.92-6.96 (in, 1 7.02-7.08 (mn, 4 7. 12-7.18 (in, 4 7.39-7.41 (in, 2 H), 7.84-7.92 (in, 6 9.01 1 12.65 (br s, 2 MS (ESI) m/~z 624 Anal. Calcd for C3,H 33
N
3
O
8 1lH 2 O: C, 65.5 1; 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]-2pyrrolidinylinethoxybenzoic acid (~J~J~f 4 NOOH 12 To a stirred solution of methyl 1-(tert-butoxycarbonyl)-4-hydroxyproline (4.0 g, 16.3 inmol), Ph 3
P
(5.14 g, 19.6 inmol), and 2,4-difuorophenol (2.55 g, 19.6 iniol) in THE (50 n-l) was added DIAD (3.9 ml, 19.6 minol), and the mixture was heated at reflux for 3 h. After cooling to room temperature, the mixture was concnitrated in vacua and the residue was chromatographed on silica gel with CHCI 3 -EtOAc 1) to give methyl 1 -Qtert-butoxycarbonyl)-4-(2,4-difluorophenoxy) pyrrolidine-2-carboxylate (5.82 g, quant) as yellow oil.
WO 01/00206 WO 0100206PCT/USOO/18079 To a stirred solution of methyl l-(tert-butoxycarbonyl)-4-(2,4-difuorophenoxy)pyrrolidine-2carboxylate (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 ml). The extracts were dried over MgSO, and evaporated. The residue was chromatographed on silica gel with CHCI 3 -EtOAc 1) as eluent to give 1-(tertbutoxycarbonyl)-4-<2,4-difluorophenoxy)pyrrolidine-2-carboxylic acid (2.55 g, as a colorless foam. 'H-NMvR (CDCI 3 8 1.42-1.47 (in, 9 2.29-2.74 (series of m, 2 3.66-3.71 (mn, 2 H), 4.46-4.51 (in, 1 4.83 (in, 1 6.73-6.95 (in, 3 H).
To a stirred solution of 1-Qter:-butoxycarbonryl)-4-(2,4-difluorophenoxy)pyrrolidine-2-carboxylic acid (2.55 g, 7.43 inmol) 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 concntrated in vacuo and quenced by the addition of H 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 1) as eluent to give 1-(teri-butoxycarbonyl)-4-(2,4-difluorophenoxy)-2pyrrolidinylmethanol (1.76 g, 72%) as a colorless oil. 'H-NMR (CDC 3 8 1.45 9 2.28-2.36 (in, 2 3.58-4.99 (series of m, 8 6.74-6.90 (in, 3 H).
To a stirred solution of 1-Qtert-butoxycarbonyl)-4-(2,4-difluorophenoxy)-2-pyrrolidinylmethano (500 mg, 1.52 mmol), methyl 4-hydroxybenzoate (277 mg, 1.82 mniol),-and Ph 3 P (477 mg, 1.82 nunol) in THE (10 mld) was added DIAD (358 ul, 1.82 inmol), and the mixture was heated at reflux for 5 h. After cooling to room temperature, the mixture was concentrated in vacuo and the residue was chromatographed on silica gel with CHCI 3 -EtOAc (20: 1) as eluent to give methyl 4-[lI -(tertbutoxycarbonyl)-4..(2,4-difluorophenoxy)-2-pyrrolidinylmethoxy]benzoate (529 mg, 75%) as a colorless oil. 'H-NM~R (CDCI,) 5 1.46 9 2.20-2.47 (in, 2 3.64 (in, 2 3.86 3 H), 4.07-4.43 (in, 3 4.86 (mn, 1 6.74-6.87 (in, 3 6.94 2 H, J 8.5 Hz), 7.95 2 H, J 8.5 Hz).
To a stirred solution of methyl 4-f 1-Qert-butoxycaxbonyl)-4-(2,4-difluorophenoxy)-2-pyrrolidiny methoxyJbenzoate (529 mg, 1. 15 inmol) in CH 2
CI
2 (5 mld) was added TEA (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 were dried over K2CO 3 and evaporated to give methyl 4-[4-(2,4-difuorophenoxy)-2-pyrrolidiny methoxy]benzoate (385 mg, 92%) as a yellow oil. IH-NMR (CDC 3 8 1.89-1.95 (in, 1 2.28- WO 01/00206 WO 01/02116PCT/UJSOO/18079 2.3 5 (in, 1 3.09 (dd, J =12.5, 4.9 Hz, 1 3.33 J =12.5 Hz, 1 3.60 (in, 1 3.86 3 4. 10 J= 5.6 Hz, 2 4.84 (in, 1 6.73-6.89 (in, 3 6.91 J= 8.5 Hz, 2 H4), 7.96 (d, J 8.5 Hz, 2 H).
A midxture of methyl 4-[4-(2,4-difluorophenoxy)-2-pyrrolidinylmethoxylbenzoate (380 mg, 1.05 mmol), 3-inethoxy-4-[N'-(2-methylphenyl)ureido~phenylacetic acid (329 ing, 1.05 inmol), EDC-HCI (302 mg, 1.58 mmnol), and catalytic amount of HQBt 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 CHCI 3 -EtOAc 1) to CHCI 3 -MeOH (10: 1) as eluent to give methyl 4-[4-(2,4-difluorophenoxy)-1I-[3-inethoxy-4- [N'-(2-methylphenyl)ureidojphenylacetyl]-2-pyrolidinylnethoxybenzoate (693 ing, quant). 'H- NMR (CDCl 3 82.16-2.53 (in, 5 3.61-4.93 (series of mn, 14 6.48-8.12 (series of in, 16 H).
To a stirred solution of methyl 4-[4-(2,4-duorophenoxy)--[3-methoxy-4-[N'-(2-methylphenyl) ureido~phenylacetylJ-2-pyrrolidinylinethoxybenzoate (693 mg, 1.05 mimol) in TH F (8 mld) was added 0.25 N NaOH (8.4 ml, 2. 10 mmol). The mixture was stirred overnight. The mixture was poured into 1 N HCI (200 ml) and the resulting precipitate was collected with suction. The solid was chroinatographed on silica gel with CHCI 3 -MeOH (50:1 to 10: 1) as eluent to give 120 (323 mg, as a colorless amorphous soid. MW 645.65 'H-NMvR (DMS04) 8 2.25 3 2.35 (in, 2 3.33-5.18 (series of in, I I 6.75 (dd, 1 H, J= 8.3, 1.7 Hz), 6.87-7.30 (series of mn, 8 7.79 1 HJ 8.3 Hz), 7.85-7.90 (in, 3 8.01 1 HJ 8.3 Hz), 8.49 1 8.57 1 MIS (FAB) nzz, 646 1).
Example 113 1-[4-fN'-(2-chlorophenyl)ureido]-3-methoxyphenylacetylJ-4-(6-quinolyloxy-2S-pyrrolidilyl inethoxybenzoic acid H OOH 121 To a stiffed solution of methyl (trans-lI-tert-butoxycarbonyl-4-hydroxy-2-pyrrolidinyl) inethoxy benzoate (1.0 g, 3.0 inxol), 6-hydroxyquinoline (435 ing, 3.0 iniol), and Ph 3 p (943 mg, 3.6 iniol) in TI-F (10 ml) was added DIAD (727 mg, 3.6 inmol) at 0 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 To a WO 01/00206 WO 01/11206PCTIJSOOII8079 stirred solution of the product in CH 2
CI
2 (6.0 ml) was added TFA (6.0 mI) at 0 The reaction midxture 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 CI1. The extract was washed with brine ,dried over Na2SS0 4 and concentrated in vacuo. The residue was purified by column chromatography on silica gel with MeOH-CH 2 Cl 2 1% to I as eluent to give methyl [(6-quinolyloxy-(2.S)-pyrrolidinyl)Jmethoxybenzoate (900 mg, as a pale yellow oil. 'H-NMR (CDCI,) 8 1.92-2. 10 (in, IH), 2.45-2.55 (in, 114), 3.20-3.30 (in, 114), 3.38-3.50 (in, 114), 3.60-3.70 (in, 111), 3.88 314), 4.05-4.18 (in, 2H), 5.03 (in, 114), 6.91 J= 8.5 Hz, 1H), 7.02 J= 2.7 Hz, 11-1), 7.35-7.38 (in, 2H4), 7.96 J= 8.5 Hz, 11-1), 8.00-8.05 (in, 2H), 8.76 J= 3.2 Hz, 114).
To a stirred solution of methyl 4-(4S-(6-quinolyloxcy-2S-pyrrolidinyl)methoxybenzoate (300 mg, 0.79minol), 4-[N'-(2-chlorophenyl)uredioj-3-methoxyphenylacetic acid (264 mg, 0.79 inmol), HOBt (107 mg, 0.79 mmol), and triethylaxnine (330 ml, 2.37 inxol) in THF (10.0 ml) and MeCN (10.0 ml) was added EDC-HCI (228 mg, 1.2 mniol) at 0 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 to EtOH-EtOAc v/v) as eluent to give methyl 4-[l-[4-[N'-(2-chlorophenyl)ureidoj-3inethoxyphenylacetylj-4-(6-quinolyloxy-(2S)-pyrrolidinylJmethoxybenzoate (520 mg, as a colorless oil. 'H-NMR (CDCI 3 8 2.30-2.60 (in, 314), 3.64 2H), 3.73 3H), 3.80-3.95 (in, 114), 3.87 314), 4.154.30 (in, HA4), 4.504.70 (in, 2H), 5. 11 (br s, lH), 6.81-7.01 (mn, 614), 7.26 -7.39 (in, 614), 7.93-8.03 (mn, 5H), 8.19 J 8.3 Hz, IH), 8.80 114).
To a stirred solution of methyl 1-[4-[N'-(2-chlorophenyl)ureidoj-3-inethoxypheny acetylj-4-(6-quinolyloxy-2S-pyrrolidinyl~inethoxybenzoate (520 mg, 0.75 ininol) in THF (10.0 nil) and MeOH (5.0 ml) was added IN NaOH (1.5 ml, 1.5 inmol). The mixture was stirred at 60 0
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 121 (450 mg, 88%) as awhite crystalline solid. 681.13 mp 129-133 IR (KBr) 3332, 1704, 1604, 1531, 1419, 1222, 1166 'H-NMIR (DMSO-d) 6 2.25-2.55 (in, 2H), 3.67 2H), 3.82 3H), 3.81-3.92 (mn, 114), 4.02-4.15 (mn, 2H), 4.40-4.50 (in, 214), 5.25-5.40 (in, IH), 5.33-7.07 (in, 7.26-7.49 (mn, 5H4), 7.83-8.23 (in, 6H), 8.73-8.74 (in, 1IH), 8.90 1H), 8.94 IH); MS (FAB) m/lz 681 AnaL calcd for C 37
H
33
N
4 0 7 CI-0.5H 2 0: C, 64.39; H, 4.97; N, 8.12. Found: C, 64.22; H, 4.90; N, 7.96.
WO 01/00206 WO 0100206PCT/USOO/1 8079 Example 114 4-[I-[4-[N'-(2-bromophenyl)ureidoJ-3-methoxyphenylacetyl]-(4S)-(6-quinolyloxy-(2S)pyrrolidinyljmethoxybenzoic acid OOH 12 To a stifred solution of methyl 4-(4S-(6-quinolyloxy-2S-pyrrolidinyl)methoxybenzoate (300 mg, 0.79mmol), 4-IMP-(2-bromophenyl)urediol-3-methoxyphenylacetic acid (299 mg, 0.79 mmnol), HOBt (107 mg, 0.79 mmol), and triethylanune (330 ad, 2.37 mniol) in THF (10.0 mil) and MeCN (10.0 ml) was added EDC-HCl (228 mg, 1.2 mmol) at 0 The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacua. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaiHCO, then dried over NaSO 4 and concentrated in vacua. The residue was purified by column chromatography on silica gel with EtOAc to EtOH-EtOAc v/v) as eluent to give methyl '-(2-bromophenyl)ureidoj -3methoxyphenylacetylj-(4S)-(6-quinolyloxy-(2S)-pyrrolidinyl]methoxybenzoate (530 mg, 91%) as a colorless oil. 'H-NMR (CDCI 3 5 2.30-2.62 (in, 3.65 211), 3.75 3.80-3.95 (mn, 11-), 3.93 4.10-4.30 (in, 111), 4.504.70 (mn, 2H), 5.11 (br s, 6.82-6.98 (in, 6H1), 7.15-7.39 (in, 5H), 7.52 J1=8.0 Hz, 11H), 7.93-8.03 (in, 8.14 J 8.3 Hz, 1I-H), 8.80 I H).
To a stirred solution of methyl 4-[1-[4-[N'-(2-broinophenyl)ureido]-3-inethoxyphenyl acetylJ-4-(6-quinolyloxy-2S-pyrrolidinyllmethoxybenzoate(530 mng, 0.72 inmol) in THF (10.0 nil) and MeOH (5.0 ml) was added IN NaOH (1.4 mld, 1.4 inmol). The mixture was stirred at 70 0
C
for 24 hr. The mixture was concentrated in vacua, water was added thereto, and neutralized with 1N HCI. The resulting solid was collected, washed with water, and dried in vacua to give 122 (460 mg, 88%) as a white crystalline solid. NM 725.59 mp 149-153 JR (KBr) 3332, 1704, 1604, 1527, 1222, 1164 'H-NMR (DMSOAd) 852.28-2.58 (in, 2H1), 3.67 211), 3.82 31-), 3.85-3.90 (in, 1H), 4.054.15 (in, 4.404.50 (in, 21H), 5.20-5.32 (mn, 111), 6.77-7.07 (in, 51-), 7.3 1-7.61 (in, 7.83-7.97 (in, 5141), 8.21-8.22 (in, 11-1), 8.73-8.74 (in, 8.92 IH); MS (FAB) m/z 725 727 Anal calcd for C 37
H,,N
4 0 6 Br-0.5H 2 O: C, 60.50; H, 4.67; N, 7.63; Br, 10.88. Found: C, 60.5 1; H, 4.60; N, 7.52; Br, 11.06.
Example 115 4-I(2S,4S)- 1-[3-methoxy-4-[N'-(2-inethylphenyl)ureido]phenylacetyl]-4-(2-naphthyloxy)-2- WO 01/00206 WO 0100206PCT/USOO/18079 pyrrolidinyl~methoxybenzoic acid M H Me 123 To a stirred mixture of methyl (2S,4R)-1-zert-butoxycarbonyl-4-hydroxy-2pyrrolidinylcarboxylate (4.22 g, 17.2 mmol), 2-naphthol (2.73 g, 18.9 mmol) and PPh 3 (4.96 g, 18.9 mmol) in THIF (80 mlJ) 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, CHCI 3 JEtOAc (10/ to give methyl (2S,4S)- 1 teri-butoxycarbonyl-4-(2-naphthyloxy)-2-pyrrolidinylcarboxylate (5.37 which was used without further purification.
To a stirred solution of methyl (2S,4S)-l-Iert-butoxycarbonyl-4-(2-napthhyloxy)-2pyrrolidinyl 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 mixture was acidified by the addition of 1 N HC1 and extracted with CHC1 3 The combined extracts were washed with brine, dried over Na 2 SO, and evaporated. The residue was recrystallized from n-hexane-CHC1 3 to give (2S,4S)-l-tert-butoxycarbonyl-4-{2-naphthyloxy)-2pyrrolidinylcarboxylic acid [4.44 g, 85%(2 steps)] as a white powder. 'H-NMR (DMSO-d 6 8 1.37 and 1.41 9H, amiide isomers), 2.26 J= 13.9 Hz, 114), 2.65 (in, 111), 3.47 J= 11.5 Hz, 3.81 (in, 111), 4.30 (in, lH), 5.14 (in, 1H1), 7.02-7.86 (in, 7H).
To a stirred solution of (28,4S)-i -tert-butoxycarbonyl-4-(2-naphthyloxy)-2pyrrolidinylcarboxylic acid 12 g, 3.13 iniol) in TI-F (30 ml) was added BH 3 -DMS (0.63 ml, 6.3 iniol) at 0 The mixture was raised to room temperature immediately and then heated at 50 'C for 1.5 h. After cooling to room temperature, the mixture was quenched by the addition of water at 0 *C and extracted with EtOAc. The combined extracts were washed with brine, dried over NaSO, and evaporated. The residue was chromatographed on silica gel [50 g, CHCI 3 /MeOH to give (2S,4S)-l-ter-butoxycarbonyl-4-(2-naphthyloxy)-2-pyrrolidinylmethano (1.10 g, 100%) as a pale yellow oil. IH-NMR (CDCI 3 5 1.48 9H), 2.45 (in, 11-1), 3.58-4.80 (in, 411), 5.01 (br, 111), 7.04-7.99 (in, 711).
To a stirred mixture of (28,45)- 1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2-pyrrolidinyI WO 01/00206 WO 0100206PCTUSOO/18079 methanol (640 mg, 1.86 mmol), methyl 4-hydroxybenzoate (283 mg, 1.86 mmol) and PPh 3 (488 mg, 1.86 mmol) in TI-F (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 1100 g, n-hexanelEtOAc(2/11)], to give methyl 4-[(2S,4S)-1 -tert-butoxycarbonyl-4-(2-naphthyloxy)-2-pyrrolidinyljmethoxybenzoate (830 mg, 93%) as a colorless oil. 'H-NMIR (CDC1 3 5 1.50 J 8.3 Hz, 911), 2.34 (in, 1T-i), 2.53 J 14.2 Hz, lH), 3.72-3.85 (in, 1H1), 3.86 and3.87 3H, amide isomers), 4.17 (in, 111), 4.26- 4.52 (in, 2H1), 5.06 (br, 111), 6.87 J 8.8 Hz, 111), 6.94 J 8.8 Hz, 21-0, 7.04 (br, 211), 7.33 J 7.3 Hz, 111), 7.42 J 7.3 Hz, 111), 7.64-8.02 (in, 511).
To a stirred solution of methyl 4-[(2S,4S)-1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2pyrrolidinyllmethoxybenzoate (870 mg, 1.74 inmol) in CH 2 Cl 2 (24 ml) was added TFA (6 ml) at room temperature. The mixture was stiffed over night, which was concentrated in vacua. 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
CI
2 The organic layer was washed with brine, dried over NaSO, and concentrated. The residue was chromatographed on silica gel [100 g, n-hexane/EtOAc(211)J, to give methyl 4-[(2S,4S)-4-(2-naphthyloxy)-2-pyrrolidinylJmethoxybenzoate (750 mng, 100%/) as a black oil 'H-NMR (CDCI,) 81.99 (dd, J= 14.2, 5.6Hz, 11), 2.48 3.22 (dd, J=12.2, 4.6 Hz, 3.43 J= 12.5 Hz, 1H1), 3.67 (in, I1-H), 3.86 and 3.87 3K1 amide isomers), 4.11 7.12 (dJ=9.0Hz 11), 7.33 (dtJ= 8.1,1.2Hz,l11-), 7.44 (dJ= 6.8, 1.2 Hz,11), 7.70 (d,J= 8.1 Hz, 11), 7.75 (dd, J 5.1 Hz, 2H), 7.90 J =8.5 Hz, 11), 7.96 (dd,J 2.0Hz,211).
A mixture of 3-inethoxy-4-[N'-(2-nethylphenyl)ureido]phenylacetic acid (333 ing, 0. 106 nunol), methyl 4-[(2S,4S)-4-(2-naphthyloxy)-2-pyrrolidinyljmethoxybenzoate (400 mg, 1.06 inmol), EDC-HCI (305 mg, 1.59 inmol) and DMAP (194 mg, 1.59 inmol) in DMF (10 ml) was 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 NaSO 4 the extracts were concentrated in vacua. The residue was chroinatographed on silica gel [100 g, nhexane/EtOAc(1I/l)CHC1/MeOH(50/ 1)1, to give methyl 1-[3-methoxy-4-[N-(2methylphenyl)ureidoJphenylacetyl]-4-(2-naphthyloxy)-2-pyrrolidinyljmethoxybenzoate (520 mng, as a pale brown amorphous. 'H-NMR (CDCl 3 8 2.28 314), 2.29 (in, 111), 2.55 J 14.2 Hz, I1H), 3.60 J =3.4 Hz, 211), 3.66 J =3.7 Hz, 3H), 3.68-4.00 (in, 5H1), 4.05-4.67 (in, 3H1), 5.09 (br, 111), 6.61 11H), 6.77 (in, 211), 6.87 J= 8.8 Hz, 111), 6.94-7.54 (in, 811), 7.68- WO 01/00206 WO 0100206PCT/US001 18079 8.09 (in, 8H); MS (ESI) m/~z 674 I).
To a solution of methyl 4-[(2S,4S)-1 -[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-4-(2-naphthyloxy)-2-pyrrolidinyllinethoxybenzoate (415 mg, 0.616 inmol) in THF (4.9 ml), 0.25 N NaOH (4.9 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 The combined extracts were dried over Na 2 S0 4 and concentrated in vacuo. The residue was purified on TLC [CHCI3/MeOH to giv e 123 (180 mg, 441/) as a colorless amorphous. IM 659.73 JR (KBr) 3354, 2937, 1685, 1601, 1533, 1255 'H-NMR (DMSO-d 6 5 2.24 3H), 2.25-2.43 (in, 3.65 211), 3.81 311), 3.83 (in, 111), 4.054.70 (mn, 5.21-5.33 (hr, 111), 6.76 J 7.3 Hz, 111), 6.86-7.35 (in, 9H1), 7.44 J= 7.3 Hz, 114), 7.76-7.89 (in, 611), 8.01 (d,J 8.3 Hz, 1H), 8.48 1H), 8.56 IH); MS (FAB) m/~z 660(M~+1).
Examp~le 116 4-[(2S,4S)-1-[4-[N'-(2-hlorophenyl)ureido-3-nethoxyphenylacetyl]-4-(2-naphtyloxy)-2pyrrolidinyl]methoxybenzoic acid %?LH H OMe JXO H124 A mixture of 4-[N'-{2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (3 10 mg, 0.93 mmol), methyl 4-[(2SA4S)-4-(2-naphthyloxy)-2-pyrrolidinyllinethoxybenzoate (350 mg, 0.93 inmol), EDC-HCI (267 mg, 1.40 inmol) and DMAP (171 ing, 1.40 minol) in DMIF (10 ml) was 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 chroinatographed on silica gel [100 g, nhexane/EtOAc( 1/ )CHC13]MeOH(50/1 to give methyl 4-I(2S,4S)- 1-[4-[N'-(2-chlorophenyl) ureidoJ-3-inethoxypheniylacetyl]-4-(2-naphthyloxy)-2-pyrrolidinyl~methoxybenzoate (450 mg, 68%) as a pale brown amorphous. 'H-NMR (CDC 3 8 2.32 (in, 111), 2.58 J 14.5 Hz, 111), 3.63 J= 2.7 Hz, 11-1), 3.70 311), 3.86 311), 3.84-3.95 (in, 4.15-4.64 411), 5.11 (br, 11-1), 6.79-7.06 (in, 711), 7.21-7.46 (in, 7H1), 7.66-7.77 (in, 7.92 J 8.8 Hz, 11-1), 7.97 (in, 11-1), 8.17 J 8.4 Hz, I1H); MS (ESI) m/z 694 696 (Mr+3).
To a solution of methyl 4-I(2S,4S)-l '-(2-chlorophenyl)ureido]-3-inethoxyphenyl acetylj-4-(2-naphthyloxy)-2-pyrrolidinyl]methoxybenzoate (381 mng, 0.535 mmol) in THE (4.3 ml), WO 01/00206 WO 0100206PCT/USOO/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 HC1 and extracted with CHCI,-MeOH (10/ The combined extracts were dried over Na 2
SO
4 and concentrated in vacua. The residue was purified on TLC [CHCIIMeOH (10/1)] to give 124 (140 mng, as a colorless amorphous. MW 680.15 IR (KBr) 3323, 2935, 1704, 1601, 1529, 1529, 1508 cm-1; 'H-NMvR (DMSO-d) 8 2.27-2.49 (in, 2H), 3.65 2H4), 3.81 314), 3.83 (in, I1H), 4.05-4.71 (mn, 4H4), 5.30 (br, I1H), 6.77 J= 7.8 Hz, 114), 6.87-7.16 (in, 4H), 7.14 (dd, J 8.8, 2.2 Hz, I1H), 7.27 J 7.3 Hz, I1H), 7.29-7.46 (mn, 414), 7.76-7.86 (in, 514), 7.96 J 8.3 Hz, 11H), 8.08 (dd, J 8.3, 1.2 Hz, 114), 8.90 1H), 8.93 1H); MS (FAB)m/z 680 682 Anal. Calcd for CmH3,CN 3
O
7
IH
2 0: C, 65.37; H, 5.20; N, 6.02. Found: C, 65.43; H, 5.11; N, 5.93.
Example 117 1-[3-methoxy-4-[N'-(2-inethylphenyl)ureido~phenylacetyl]-4-(2-naphthyloxy)-2acid M~e H H 6Me 125 To a stirred mixture of methyl (2S,4R)-1-tert-butoxycarbonyl-4-hydroxy-2pyrrolidinylcarboxylate (4.22 g, 17.2 inrol), 2-naphthol (2.73 g, 18.9 inmol) and PPh 3 (4.96 g, 18.9 mol) in THF (80 nil) was added DIAD (3.72 ml], 18.9 mmol) at room temperature under an atmosphere of itrogen. After stirring over night, the mixture was concentrated in vacua. The residue was chromatographed on silica gel 1600 g, CHCI 3 /EtOAc (10/ to give methyl (2S,4S)- 1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2-pyrrolidinylcarboxylate (5.37 which was used to the next reaction without further purification.
To a stirred solution of methyl (2S,4S-1-teri-butoxycarbonyl-4-(2-naphthyloxy)-2pyrrolidinyl carboxylate (5.37 g) in THF (116 ml) was added 0.25 N NaOH (116 ml, 29.0 mnimol) 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 HCl and extracted with CHC 3 The combined extracts were washed with brine, dried over Na 2
SO
4 and evaporated. The residue was recrystallized with n-hexane-CHCI 3 to give (2S,4S)-l1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2pyrrolidinylcarboxylic acid [4.44 g, 851/o(2 steps)) as a white powder. 'H-NMR (DMSO-A 8 1.37 and 1.41 9Hf, amide isomers), 2.26 J 13.9 Hz, 114), 2.65 (in, 1ff), 3.47 J 11.5 Hz, 1ff), 3.81 (in, 1ff), 4.30 (in, 1ff), 5.14 (in, 1ff), 7.02-7.86 (in, 7Hf).
WO 01/00206 WO 0100206PCT/USOO/1 8079 To a stirred solution of (2S,4S)-1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2pyrrolidinylcaiboxylic acid 12 g, 3.13 mmol) in THE (30 mlJ) was added BH, 3 DMS (0.63 ml, 6.3 mmol) at 0 TC. The mixture was raised to room temperature immediately and then heated at 50 TC for 1. 5 h. After cooling to room temperature, the mixture was quenched by the addition of water at 0 *C and extracted with EtOAc. The combined extracts were washed with brine, dried over Na 2 SO, and evaporated. The residue was chromatographed on silica gel [50 g, CHCI3/MeOH to give (2S,4S)- l-tert-butoxycarbonyl-4-(2-naphthyloxy)-2-pyrrolidinylmethanol 10 g, 100%) as a pale yellow oil. 'H-NMR (CDCI 3 8 1.48 911), 2.45 (in, 1H), 3.58-4.80 (in, 4H), 5.01 (br, 1H), 7.04-7.99 Cm, 7H4).
To a stirred mixture of (2S,4S)-l-ter-butoxycarbonyl-4-(2-naphthyloxy)-2pyrrolidinylmethanol (484 mg, 1.41 mmol), methyl 2-hydroxcy-5-pyridinecarboxylate (216 mng, 1.41 inmol) and PPh 3 (370 mng, 1.41 minol) in THE (15 ml) was added DIAD (0.28 Ml, 1.41 inmol) 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, nhexane/EtOAc(21 to give methyl-2-[(2S,4S)-1-tert-butoxycarbonyl-4-(2-naphthyloxy)-2- (170 mg, as a colorless oil. 'H-NMR (CDC 3 8 1.47 9H), 2.37 (mn, 114), 2.46 J 14.2 Hz, I 3.71-4.00 (in, 2H), 3.89 3H), 4.304.56 (in, 2H), 4.74 (dd, J 9.8, 4.6 Hz, 1H), 5.06 (br, 111), 6.70 J 8.8 Hz, 2H), 7.05-7.09 (in, 2H), 7.33 J 6.9 Hz, 1H), 7.42 J 6.9 Hz, 7.67-7.75 (in, 3H), 8.09 Cd, J 8.8 Hz, IN), 8.77 J =2.2 Hz,I To a stirred solution of 5-carboxyinethyl-2-[(2S,4S)-1-tert-butoxycarbonyl-4-(2napthyloxy)-2-pyrrolidinyl]methoxypyridine (170 mg, 0.36 minol) in CH 2
CI
2 (5 ml) was added TEA (2 ml) at room temperature. After 2 h stirring, the mixture was concentrated in vacuo, which was diluted with CHCl 2 and basified by the addition of 1 N NaQH. The combined reaction 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 ICHCIVMCOH to give methyl 2-[(2S,4.S)-4-(2-naphthyloxy)2-pyrrolidinyllmethoxypyridine-5-carboxylate (107 mng, as a colorless oil 'H-NMR (CDCI 3 5 1.95 (in, I 2.27 (br, 2.46C(m, 1H), 3.19 (dd, J= 12.2, 4.9 Hz, 111), 3.41 J 12.2 Hz, 1H), 3.65 IH), 3.89 3H), 4.58 (mn, 2H), 5.00 (br, IN), 6.77 J =8.8 Hz, 111), 7.06 (br, lH), 7.11 (dd, J 8.8, 2.7 Hz, IN), 7.3 1-7.45 2H), 7.69-7.76 3H), 8.13 (dd, J 8.8, 2.4 Hz, 1H), 8.78 J 2.2 Hz, 11-).
WO 01/00206 WO 0100206PCT/USOO/18079 A mixture of 3-methoxy-4-[N' -(2-methylphenyl)ureidojphenylacetic acid (89 mg, 0.283 mmol), methyl-2-[(2S,45)-4-(2-ntaphthyloxy)-2-pyrrolidinyl~methoxypyridine-5-caboxylate (107 mg, 0.78 mmnol), EDC-HCI (81 mg, 0.425 mmol) and DMAP (52 mg, 0.425 mmol) in DMF (3 mlJ) 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 NaSO 4 the extracts were concentrated in vacua. The residue was purified on TLC [CHC1V/MeOH to give 1-[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetylJ-4-(2acid methyl ester (193 mg, 100%) as a colorless amorphous. 'H-NMR (CDCI 3 8 2.27 J 3.2 Hz, 3H4), 2.30 (in, 1H), 2.49 (dd, J 14.2, 2.0 Hz, 3.60 J 3.9 Hz, 3.67 J 5.9 H{z, 3H1), 3.81 1H4), 3.85 1H), 3.88 and 3.91 3H, amide isomers), 3.95 (mn, 4.02-5.09 (in, 4H), 6.67 J 8.8 Hiz, I H), 6.73-7.13 (in, 314), 7.20-7.45 (in, 7H4), 7.53 J= 7.8 Hz, LH), 7.67-7.77 (in, 314), 8.02-8.84 (mn, 3H).
For HCI salt: a pale brown amorphous. IR (KBr) 3346, 2951, 1720, 1601, 1533, 1281 MS (FAB) ,nk 675 1).
To a solution of 2-[(2S,4S)-l -[3-methoxy-4-[N'-(2-inethylphenyl)ureidojphenylacetylJ-4- (2-naphthyloxy)-2-pyrrolidinylJmethoxcy-5-pyridinecarboxylic acid methyl ester (158 mng, 0. 23 mmol) in THF (1.8 ml), 0.25 N NaOH (1.8 mld) was added. After stirring at room temperature for 22 h, the mixture was neutralized with 1 N HCl and extracted with CHCl 3 -MeOH The combined extracts were dried over Na 2
SO
4 and concentrated in vacua. The residue was purified on TLC ICHCIVMeOH (5/1)1 to give 125 (51 mng, 34%) as a colorless amorphous solid. MW 660.72 JR (KBr) 3354, 2956, 1601, 1533, 1255, 1022 'H-NMR (DMSO-d 6 8 2.24 3H), 3.30-5.32 (in, 13HM, 6.72-8.82 (in, 19H); MS (FAB) m/lz 661(M+1); Anal. Calcd for C3,HN 4 O,-0.5EtOH-1H2O: C, 66.75; H, 5.89; N, 7.98. Found: C, 66.39; H, 5.55; N, 7.66.
Example 118 4-[5-(R)-benzyloxymethyl-1 -[4-[N'-(2-mehylphenyl)ureidoJphenylacetylJ-2-(S)-pyrrolidintyl inethoxy~benzoic acid Me~~r HOH 126 To a stirred solution of benzyl-(S)-glycidyl ether (5.0 g, 30.5 iniol) in TI-F (100 nil) was added allylinagnesiuin chloride (1.0 Mmi Et 2 O, 30.5 ml, 30.5 inmol) at -781C, and the resulting WO 01/00206 WO 0100206PCT/U 500/18079 mixture was gradually warmed up to rt with stirring. The mixture was poured into water and concentrated in vacuo, then extracted with CHCI,. The organic layer was dried over anhydrous Na 2 SO, and concentrated in vacuo. The residue was chroinatographed on silica gel with hexane- EtOAc (5 as eluent to give 1-benzyloxy-2-(R)-hydroxy-5-hexene (2.18 g, 35%) as a colorless oil: 'H-NMR (CDCI 3 5 1.52-1.60 (in, 2 2.11-2.25 (in, 2 2.34 J 3.2 Hz, 1 3.35 (dd, J 9.6, 8.0 Hz, 1 3.52 (dd, J 9.6, 3.2 Hz, 1 3.84-3.86 (in, 1 4.57 2 4.96- 5.07 (series of mn, 2 5.78-5.88 (in, 1 7.29-7.38 (in, 5 MS (EST) m/lz, 224 (Mr+NH4,).
To a stirred solution of 1-benizyloxy-2-(R)-hydroxy-5-hexene (2.18 g, 10.5 minol), triphenylphosphine (3.32 g, 12.7 iniol) and phthalimide (1.86 g, 12.7 mmol) was added diisopropyl azodicarboxylate (2.62 ml, 12.7 mniol) 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 as eluent to give I- (2.95 g, 83%) as a colorless oil: 'H-NMiR (CDCl 3 8 1.76- 1. 84 2 2.06 (dd, J =14.4, 6.8 Hz), 2 H, 2.12-2.22 I1H), 3.69 (dd, J= 10.0, 5.6 Hz, 1 4.00 J =9.6 Hz, 1 4.46 (d,J 12.0 Hz, 1 4.53 J= 12.0 Hz, 1 4.51-4.58 (mn, 1 4.91-4.99 (series of m, 2 5.72-5.79 (in, 1 H)7.21-7.26 (mn, 5 7.71-7.83 (series of m, 2 MIS (ESI) m/z, 336 To a stirred solution of 1-benzyloxy-2-(S)-phthalimido-5-hexene (2.95 g, 8.80 mmol) in EtQH (30 mlJ) was added hydrazine hydrate (80% in water, 460 ml, 11.4 mniol) 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, and extracted with CHC1 3 The organic layer was dried over anhydrous Na 2 SOI, then concentrated in vacuo to give 2- (1.90 mg, quant.) as a colorless oil. 'H-NMR (CDCl 3 8 1.37- 1.55 (series of mi, 4 2.08-2. 19 (in, 2 2.99-3.03 (in, 1 3.25 (dd, J 7.6 Hz, 1 H), 3.45 (dd, J 9.2, 4.0 Hz, 1 4.53 2 4.94-5.06 (series of in, 2 5.76-5.85 (in, 1 H), 7.27-7.37 (in, 5 MS (ESI) nVz, 206 247 (M'*+H+CH 3
CN).
To a stirred solution of 2-(S)-amino-l-benzyloxy-5-hexene (1.89 g, 9.21 inmol) and triethylamine (1.28 ml, 9.21 inmol) in CH 2
CI
2 (20 ml) was added benzoyl chloride (1.07 ml, 9.21 iniol) at it, and the resulting mixture was stirred for 23 h. The mixture was poured into water and extracted with CH 2
CI
2 The organic layer was washed with water, drying over anhydrous WO 01/00206 WO 0100206PCT/USOO/18079 Na 2
SO
4 then concentrated in vacuo. The residue was chromatographed on silica gel with hexane- EtOAc (5 as eluent to give N-[2-(S')-(1.benzyloxy)-5-hexenyl]benzamide (2.67 g, 94%) as a colorless needles. mp 78-79 'H-NMR (CDCI 3 5 1.76-1.82 (in, 2 2.11-2.17 (in, 2 3.59 (brs, 2 4.29-4.35 (mn, 1 4.54 (dd, J 19.2, 12.0 Hz, 2 4.96-5.05 (series of m, 2 5.78- 5.89 (mn, 1 6.39 J 8.0 Hz, 1 7.27-7.51 (in, 8 7.74 J 7.2 Hz, 2 MS (ESI) ni/z, 3 10 To a stirred solution of N-[2-(S)-(1-benzyloxy)-5-hexenyllbenzamide (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 rmol) in one portion, and the resulting mixture was stirred for 20 h. The mixture was poured into aq.Na 2
S
2
O
3 and concentrated in vacuo, 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 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 inmol) and N, IVdimethyl aminopyridine (180 mg, 1.47 nunol), and the resulting mixture was stirred overnight at rt. The mixture was poured into water and extracted with CH 2
C
2 The organic layer was washed with water, drying over Na 2
SO
4 and concentrated in vacuo. The residue was chromatographed on silica gel with hexane-EtOAc (5 to give methylpyrrolidine (1.27 g, as a colorless oil. 'H-NMR (CDCI 3 8 1.41 and 1.49 total 9 H), 1.85-2.00 (series of m, 4 3.33-4.59 (series of in, 8 7.26-7.32 (in, 5 7.4 1-7.46 (in, 2 H), 7.54-7.57 (in, 1 8.02 J 7.6 Hz, 2 MS (ESI) m/z, 426 448 (M+Nae).
To a stirred solution of (1.23 g, 2.89 mniol) in MeOH (30 ml) was added NaOH (1.0 M in water, 3.47 ml, 3.47 inmol) at rt, and the resulting mixture was stirred for 4 h. The mixture was neutralized with aq. lN-HCl and concentrated in vacuo, then extracted with CHC1 3 The organic layer was washed with brine, drying over anhydrous Na2S0 4 then concentrated in vacuo. The residue was chroinatographed on silica gel with hexane-EtOAc (3 as eluent to give N-Boc-5-(S)-hydroxyinethyl-2-(S)benzyloxymethyl pyrrolidine (847 ing, 91%) as a colorless oil. 'H-NMR (CDC1 3 8 1.41 9 H), 1.57 (brs, 1 1.95-1.97 (in, 2 2.05-2.18 (in, I 3.36 J 8.4 Hz, 1 3.56-3.62 (in, 2 3.67-3.72 (in, 2 3.95 (brs, 1 4.03 (brs, 1 4.51 1 7.28-7.37 (in, 5H); MIS (FAB) m/lz, 322 (M 4 To a stirred solution of (388 mng, 1.21 imol), triphenylphosphine (380 ing, 1.45 inmol) and methyl 4-hydroxybenzoate WO 01/00206 WO 0100206PCT/USOO/18079 (220 ing, 1.45 inmol) was added diisopropyl azodicarboxylate (200 ml, 1.45 minol) 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 vacuc. The residue was chromatographed on silica gel with hexane-EtOAc (3 :1) as eluent to give methyl 4-[2-(S)-(N-Boc-5-(S)-benzyloxymethyl)pyrrolidinylmethoxylbenzoate (462 mg, 84%) as a colorless oil: 'H-NMR (CDCI,) 8 1.40 and 1.48 total 9 1.98-2.13 (mn, 4 3.83 and 3.85 3 3.33-4.25 (series of m, 4 4.47-4.59 (mn, 2 6.91-6.96 (in, 2 H), 7.26-7.34 (in, 5 7.95-7.98 (in, 2 MIS (FAB) m/z, 456 478 To a stirred solution of methyl 4-[2-(S)-(N-Boc-5-(S)-benzyloxymethyl)pyrrolidinyl inethoxy] benzoate (446 mg, 0.98 minol) in CH 2
CI
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
SQ
4 then concentrated in vacuo to give methyl benzyloxymethyl) pyrrolidinylmethoxyjbenzoate (363 mng, quant.) as yellowish oil. The product was used for next reactions without further purification. 'H-NMR (CDC1 3 5 1.43-1.65 (mn, 2 H), 1.93-2.07 (mn, 3 3.36-3.68 (series of in, 4 3.89 3 3.86-3.93 (over lap, 2 4.55 2 6.90 J 8.4 Hz, 2 7.26-7.37 (mn, 5 7.97 J 8.4 Hz, 2 MIS (FAB) m/z, 356 To a stirred solution of methyl 4-[2-(S)-(5-(S)-benzyloxymethyl)pyrrolidinylnethoxy benzoate (115 mng, 0.32 inmol), 4-[N'-(2-inethylphenyl)ureidolphenylacetic acid (92.0 ing, 0.32 nunol) and NN-diinethylaminopyridine (52.0 mng, 0.42 mmnol) in DMF (10 mlJ) was added EDC HCI (81.0 mng, 0.42 minol) at rt, and the resulting mixture was stirred overnight. The reaction mixture was poured into water and extracted with CHCI,. The organic layer was washed with brine, drying over anhydrous Na 2 SOI, then concentrated in vacuo. The residue was chroinatographed on silica gel with CHC1 3 -MeOH (20 as eluent to give methyl oxyinethyl- 1 -N-2mtyphnlucdlhnlaeaio--S-yroiiymtoy benzoate (169 mng, as a colorless amorphous solid. 'H-NMR (CDCl 3 mixture of rotainars 8 1.92-2.18 (mn, 3 2.24 and 2.25 total 3 2.20-2.3 1 (overlap, 1 3.39-3.70 (series of mn, 4 3.87 and 3.89 total 3 4.17 and 4.18 total 2 4.304.45 (series of m, 2 4.53 2 6.43-7.13 (series of in, 9 7.20-7.36 (series of mn, 7 7.58-7.99 (series of in, 3 MS (FAB) 622 (Mr+H).
WO 01/00206 WO 0100206PCT/USO0/1 8079 To a stirred solution of methyl 4-[5-(R)-benzyloxymethyl- 1 -14-IN' -(2-methyiphenyl) ureidol phenyla eyl]-2-(S)-pyrrolidinylmethoxylbenzoate (156 mg, 0.24 mniol) in MeOH-THF (1 12 mld) was added 1.OM-NaOH (1.2 nml, 1.20 mmol) at rt, and the resulting mixture was heated at 80*C with stirring lbr 7 h. The mixture was poured into lN-HCI, then extracted with CHC1 3 The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 then concentrated in vacua. The residue was chromatographed on silica gel with CHCI 3 -MeOH (5 as eluent to give 126 (94.0 mig, as a colorless amorphous solid. MW 607.70 'H-NMR (CD 3 QD), mixture of rotamars 5 1.85-2.35 (series of m, 4 2.43-2.92 (series of m, 5 2.28 3 3.55-4.55 (series of m, 10 6.85-7.95 (series of m, 17 MS (ESI) m/z, 630 Example 119 -[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyJ-2-(S)pyrrolidinylmethoxylbenzoic acid I HOOHM 127 To a stirred solution of methyl 4-[2-(S)-(5-(S)-benzyloxymnethyl)pyrrolidinylmethoxyI benzoate (117 mig, 0.33 minol), 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (110 mig, 0.33 mmol) and NN-dimethylaminopyridine (50.0 mig, 0.40 mmol) in DMF (10 ml) was added EDC-HCI (76.0 mig, 0.40 mmol) at Mt and the resulting mixture was stirred overnight. The reaction mixture was poured into water and extracted with CHC 3 The organic layer was washed with brine, drying over anhydrous NaSO,, then concentrated in vacua. The residue was chromatographed on silica gel with CHC1 3 -MeOH (20: 1) as eluent to give methyl 4-15-(R)benzyloxyimethyl-l1-[4-[N'-(2-chlorophenyl)ureido-3-methoxypheylacetylJ -2-(S)-pyffolidinyl methoxy]benzoate (189 mig, 85%) as a colorless amorphous solid. 'H-NMR (CDC1 3 mixture of rotamars; 5 1.91-2.30 (series of m, 4 3.39-3.74 (series of m, 3 3.73 2 4.15-4.02 (in, 2 4.32-4.44 (mn, 1 4.54 2 6.71-7.02 (series of in, 5 7.06 I 7.17 I H), 7.24-7.40 (series of m, 7 7.89-8.22 (series of m, 4 MS (FAB) m/z, 672 (M 4 To a stirred solution of methyl 4-[5-(R)-benzyloxymnethyl-l-[4-[N'-(2-chlorophenyl) ureido]-3-methoxyphenylacetylJ-2-(S)-pyrrolidinylmethoxy]benzoate (169 mg, 0.25 nimol) in MeOH-THF (2 5, 7 ml) was added 1.OM-NaQH (750 ml, 0.75 mimol) at rt, and the resulting mixture was heated at 80 0 C with stirring for 2 h. The mixture was poured into 1N-HCI, then extracted with CHC1 3 The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 WO 01/00206 WO 0100206PCT/1JSOO/18079 then concentrated in vacuo. The residue was chromatographed on silica gel with CHCI 3 -MeOH as eluent to give 127 (114 mg, 69%) as a colorless amorphous solid. MW 658. 14 IH- NMR (CDOD), mixture of rotamars 5 1.88-2.37 (series of m, 4 3.51-4.49 (series of m, 8H) 3.64 and 3.73 total 3 4.86 2 6.85-7.95 (series of m, 16 MS (ESI) m/z, 658 680 Anal Calcd for CmH36CN 3 O7-H 2 O: C, 63.95; HL 5.66; N, 6.2 1. Found: C, 63.65; H, 5.40: N, 5.95.
Example 120 1.14-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetylj-2-(S) pyrrolidinylmethoxy]benzoic acid rH H6Me 128 To a stirred solution of methyl 4-[2-(S)-(5-(S)-benzyloxyrinethyl)pyrrolidinylmethoxyJ benzoate (119 mg, 0.34 mmol), 4-[N'-(2-bromophenyl)ureidoJ-3-methoxyphenylacetic acid (127 mg, 0.34 mmol) and NN-dimethylaminopyridine (50.0 mg, 0.40 mmol) in DMF (10 ml) was added EDC-HCI (77.0 mg, 0.40 mmol) at rt, and the resulting mixture was stiffed overnight. The reaction mixture was poured into water and extracted with CHCI,. The organic layer was washed with brine, drying over anhydrous Na 2
SO
4 then concentrated in vacua. The residue was chromatographed on silica gel with CHCI,-MeOH (20: 1) as eluent to give methyl benzyloxymethyl-l1-[4-[N'-(2-bromopheniyl)ureido]-3 -methoxyphenylacetylJ-2-(S)-pyrrolidinyl methoxylbenzoate (217 mg, 90%) as a colorless amorphous solid. 'H-NMR (CDC1 3 mixture of rotamars 5 1.92-2.31 (series of mo, 4 3.39-3.73 (series of m, 3 3.65 214), 4.15-4.02 (in, 2 4.32-4.44 (mn, 1 4.54 2 6.71-6.99 (series of m, 5 7.04 1 7. 11 1 H), 7.22-7.39 (series of mn, 7 7.51-8.17 (series of mn, 4 MS (FAB) ink, 716 718 To a stirred solution of methyl 4-[5-(R)-benzyloxymethyl-lI-[4-[N'-(2-bromophenyl) ureidoJ-3-inethoxyphenylacetylj-2-(S)-pyrrolidinylmethoxy~benzoate (178 mg, 0.25 minol) in MeOH-THF (2 5, 7 ml) was added 1.OM-NaOH (750 ml, 0.75 minol) at rt, and the resulting mixture was heated at 80 0 C with stirring for 1.5 h. The reaction mixture was poured into 1N-HCl, then extracted with CHC 3 The organic layer was washed with brine, drying over anhydrous Na 2 SO., then concentrated in vacuo. The residue was chromatographed on silica gel with CHCI,- MeGH (15 as eluent to give 128 (159 mng, as a colorless amorphous solid. MW 702.59 'H-NM4R (C13,01), mixture of rotamars 5 1.88-2.35 (series of mn, 4 3.51-4.49 (series of m, 8 WO 01/00206 WO 0100206PCTUSOOII8079 3.64 and 3.72 total 3 4.87 2 6.65-8.05 (series of m, 16 MS (ESI) nvz, 702 704 Anal Calcd for C36H 36 BrN 3 O7-H 2 O: C, 60.00; H, 5.32; N, 5.83. Found: C, 59.66; H, 5.04: N, 5.65.
Example 121 3-5()bnyoynty--4[-2mtypey~rio--ehxpeyaey]2() pyrrolidinylmethoxy~benzoic acid Me H COOH 129 To a stirred solution of (415 mg, 1.29 mmol), triphenylphosphine (410 mg, 1.55 mmol) and methyl 3-hydroxybenzoate (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 as eluent to give methyl (513 mg, 87%o) as a colorless oil: IH- NMR (CDCl 3 8 1.40 and 1.46 total 9 1.95-2.20 (series of m, 4 3.33-3.72 (series of m, 2 3.82-4.00 (in, 1 3.90 and 3.91 total 3 4.094.21 (in, 3 4.21-4.57 (in, 2 7.11- 7.15 (in, 1 7.26-7.37 (in, 6 7.53-7.65 (mn, 2 MIS (ESI) ni/z, 456 To a stirred solution of methyl 3-[2-(S)-(N-Boc-5-(.S)-benzyloxymethyl)pyrrolidinylinethoxyl benzoate (501 mg, 1. 10 mmol) in CH 2
CI
2 (10 ml) was added trifluoroacetic acid (10 ml) at it, and the resulting mixture was stirred for 1 h. The mixture was concentrated in vacuo and poured into aq.NaHCO 3 the extracted with CHCI 3 The organic layer was washed with water, diying over anhydrous NaSO 4 and concentrated in vacuo to give methyl benzyloxymethyl) pyrrolidinyl-methoxylbenzoate (387 ing, quant.) as yellowish oil. The product was used for next reactions without further purification. 'H-NMR (CDCI 3 5 1.26-1.65 (mn, 2 H), 1.94-2.04 (in, 3 3.37-3.52 (mn, 2 3.63-3.66 (mn, 1 3.85-3.93 1 3.91 3 4.55 2 7.09-7.11 (in, 1 7.27-7.54 (in, 6 7.54-7.55 (mn, IH), 7.61-7.63 (in, 2 MIS (ESI) m/z, 35 To a stirred solution of methyl 3-[2-(.S)-(5-(.S)-benzyloxymethyl)pyrrolidinylinethoxyI benzoate (140 ing, 0.39 inmol), 4-[N'-(2-inethylphenyl)ureidoJ-3-inethoxyphenylacetic acid (125 mng, 0.39 inmol) and NN-diinethylaminopyridine (58.0 mng, 0.47 inmol) in THF (15 ml) was added WO 01/00206 WO 0100206PCT/1JSOO/18079 EDC-HCI (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 CHO, 3 The organic layer was washed with brine, drying over anhydrous Na 2 SO,, then concentrated in vacua. The residue was chromatographed on silica gel with CHCl 3 .MeOH (10 as eluent to give methyl benzyloxymethyl- 1-[4-[NV'-(2-methylpheniyl)ureidoJ-3-methoxyphenylacetylJ-2-(S)-p'rrolidiny methoxylbenzoate (256 mg, quant.) as a colorless amorphous solid. IH-NMR (CDC1 3 mixture of rotamars 581.67 3 1.97-2.40 (series of m, 4H), 3.42-3.85 (series of m, 5 3.60 3 H), 3.95 and 3.97 total 3 4.154.26 (in, 2 4.36-4.49 (mn, 1 4.59 2 6.32 and 6.36 (s, total 1 6.75-6.87 (series of mn, 2 M, 7.20 (brs, 2 7.16-7.72 (series of mn, 10 8.03 -8.09 (in, 1 MS (ESI) m/z, 652 (MW+H).
To a stirred solution of methyl 3-[5-(R)-benzyloxymethyl-1I-[4-[N'-(2-methylphenyl) ureidoj-3-methoxypheniylacetylj -2-{S)-pyrrolidinylmethoxylbenzoate (185 mng, 0.28 inmol) in MeOH-THF (2 5, 7 ml) was added 1 OM-NaOH (860 ml, 0.86 mniol) at rt, and the resulting mixture was heated at 60'C with stirring for 1 hL The reaction mixture was poured into lN-HCI, then extracted with CHCI,. The organic layer was washed with brine, drying over anhydrous Na 2 SO4, then concentrated in vacuo. The residue was clurinatographed on silica gel with CHCI,- MeGH 1) as eluent to give][29 (171 mg, 94%) as a colorless amorphous solid. MW 637.72 IH-NMR (CDOD), mixture of rotamars 5 1.89-2.37 (series of in, 4 2,29 3 3.52-4.53 (series of m, 8 3.66 and 3.74 total 3 4.85 2 6.66-7.98 (series of in, 16 MIS (ESI) m/z, 638 660 Anal. Calcd for C 37 H9N 3
Q
7
-H
2 O: 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-l1-[4-[N'-(2-chloropheniyl)ureidol-3-nethoxyphenylacetylJ-2-(S)pyrrolidinylinethoxylbenzoic acid COHOH6M 130 To a stirred solution of methyl 3-[2-(S)-(5-(S)-benzyloxyinethyl)pyrrolidinylmethoxyI benzoate (118 ing, 0.33 minol), 4-[N'-(2-chlorophenyl)ureido]-3-inethoxyphenylacetic acid (112 ing, 0.33 inmol) and NN-diinethylaminopyridine (50.0 mg, 0.40 inmol) in THF (15 ml) was added EDC-HCI (80.0 ing, 0.40 iniol) at rt, and the resulting mixture was stirred overnight. The reaction mixture was poured into water and extracted with CHCI,. The organic layer was washed WO 01/00206 WO 0100206PCT/USOO/18079 with brine, drying over anhydrous Na 2 SOI, then concentrated in vacuo. The residue was chromatographed on silica gel with CHCI,-MeOH (10 as eluent to give methyl bcnzyloxymethyl-l1-[4-[N'-(2-chlorophenyl)ureidoJ-3-methoxyphenylacetamido]-2-(S)pyrrolidinylmethoxylbenzoate (226 mg, quant.) as a colorless amorphous solid. 'H-NMR (CDCI,), midxture of rotamars 5 1.99-2.40 (series of m, 4 3.43-3.92 (series of m, 5 3.67 3 3.98 and 4.02 total 3 4.18-4.29 (in, 2 4.36-4.51 (in, I 4.60 2 6.75-6.92 (series of m, 2 7.01-7.22 (series of m, 4 7.29-7.53 (series of m, 9 7.62-8.26 (series of M, 3 H); MIS (ESI) 672 To a stirred solution of methyl 3-[5-(R)-benzyloxymethyl-1-[4-[N'-(2-chlorophenyl) ureido]-3-methoxyphenylacetylj-2-(S)-pyrrolidinylmethoxyjbenzoate (169 mg, 0.25 mmol) in MeOH-THF (2 5, 7 mlJ) was added 1.OM-NaOH (760 ml, 0.76 mmol) at rt, and the resulting mixture was heated at 60'C with stirring for 1.5 h. The reaction mixture was poured into lN-HC1, then extracted with CHCI 3 The organic layer was washed with brine, dryed over anhydrous NaSO,, then concentrated in vacuo. The residue was chromatographed on silica gel with CHCI,- MCOH 1) as eluent to give 130 (155 mg, 94%) as a colorless amorphous solid. MW 658.14 'H-NMR (CD 3 OD), mixture of rotaniars 5 1.89-2.35 (series of m, 4 3.52-4.53 (series of m, 8 3.68 and 3.75 total 3 4.85 2 6.68-8.03 (series of M, 16 MIS (ESI) m/z, 658 680 Anal. Calcd for C36H3CIN 3 0QHO: C, 63.95; H, 5.66; N, 6.2 1. Found: C, 64.01; H, 5.38: N, 5.96.
Example 123 3-[5-(R)-benzyloxymethyl-l1-[4-[N'-(2-bromophenyl)ureidoJ-3-methoxyphenylacetylJ-2-(S)pyrrolidinylmethoxy~benzoic acid COHHOM 131 To a stirred solution of methyl 3-[2--(5-(S)-benzyloxymethy)pyrrolidinylmethoxyj benzoate (116 mg, 0.33 inmol), 4-[N'-(2-bromophenyl)ureidol-3-methoxyphenylacetic acid (124 mg, 0.33 inmol) and NN-dimethylan-inopyridine (48.0 mg, 0.39 inmol) in THF (15 ml) was added EDC-HCI (75.0 mg, 0.39 inmol) 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 NaSQ 4 then concentrated in vacuo. The residue was chromatographed on silica gel with CHC1 3 -MeOH (10 as eluent to give methyl WO 01/00206 WO 0100206PCT/USOO/18079 benzyloxymethyl-l1-[4-[N'-(2-bromophenyl)ureidoJ-3 -methoxyphenylacetyl]-2-(S)-pyrrolidinyl methoxyjbenzoate (209 mg, 89%) as a colorless amorphous solid. 'H-NMR (CDC3), mixture of rotamars 8 1.99-2.37 (series of m, 4 3.43-3-91 (series of m, 5 3.70 3 3.93 and 3.96 total 3 4.19-4.28 (in, 2 4.37-4.51 (in, 1 4.60 2 6.77-7.11 (series of m, 6 H), 7.28-7.74 (series of m, 10 7.91-7.95 (series of m, I 8.20-8.23 (series of mn, 1 MIS (ESI) m/z 716 718 To a stirred solution of methyl 3-[5-(R)-benzyloxymnethyl-1-[4-[N'-(2-broinophenyl) ureidoJ-3-inethoxyphenrylacetyl-2()-pyrrolidinylmethoxylbenzoate (176 mg, 0.25 mmol) in MeOH-THF 5, 7 ml) was added I OM-NaOH (760 ml, 0.76 inmol) at rt, and the resulting mixture was heated at 60'C with stirring for 1.5 h. The reaction mixture was poured into 1N-HCI, then extracted with CHCI 3 The organic layer was washed wAith brine, drying over anhydrous Na 2 SOI, then concentrated in vacua. The residue was chromatographed on silica gel with CHCI 3 MCOH 1) as eluent to give 131 (156 mg, as a colorless amorphous solid. MW 702.59 'H-NMR (CD 3 OD), mixture of rotainars 8 1.89-2.37 (series of in, 4 2,29 3 3.52-4.53 (series of in, 8 3.68 and 3.75 total 3 4.85 2 6.67-7.95 (series of mn, 16 MIS (ESI) m/z, 702 (WMH, 704 Anal Calcd for C36H 3 6 BrN 3
O,-H
2 O: 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-l1-[3-inethoxy-4-[N '-(2-methylphenyl)ureidojphenylacetylJ-2-pyrrolidinyl methoxybenzoic acid ~§vx~yCOOH eH H Me 132 To a stirred mixture of (2S4R)-1-:ert-butoxycarbony1-2-tert-butyldiphenylsilyloxymethy- 4-hydroxypyrrolidine (21.7 g, 47.6 mmuol), acetic acid (3.0 ml, 52.4 nunol) and PPh 3 (12.5 g, 52.4 inmol) in THF (330 ml) was added DIAD (9.4 mil, 47.6 nunol) at room temperature under an atmosphere of nitrogen. After 2 h stirring the same temperature, the mixture was heated at 50 'C for 2 h After cooling to room temperature, the reaction mixture was concentrated in vacua. The resulting residue was chromatographed on silica gel [1 Kg, n-hexane/EtOAc(511)J, to give (2S,4S)-4-acetoxy-l1-iert-butoxycarbonyl-2-tert-butyldiphenylsilyloxynethypyrolidine (23.3 g, 99%) as a colorless oil. 'H-NMR (CDCI,) 8 1.06 9H), 1.35 and 1.43 9H, amide isomers), 1.92 (br, 3H), 2.20-2.45 (mn, 2H), 3.31-4.07 (mn, 5H), 5. 17-5.30 (mn, 111), 7.36-7.44 (mn, 6H), 7.65- WO 01/00206 WO 0100206PCT/IJSOO/18079 7.71 (in, 4H).
To a stirred mixture of (2S,4S)-4-acetoxy- 1-tert-butoxycarbonyl-2-tert-butyldipheny silyloxy methypyrrolidine (23.3 g, 46.9 inmol) and acetic acid (6.0 ml, 104.8 inmol) in THE (470 ml) was added TBAF (93.8 ml, 93.8 nimol) at 0 0 C. After 24 h stirring, the mixture was 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, and concnitrated in vacuo. The residue was chromatographed on silica gel [700 g, CHC131 EtOAc to give (2S,4S)-4-acetoxy-l-tert-butoxycarbonyl-2-pyrrolidinemethanol (9.70 g, as a colorless oil. 'H-NMR (CDCI 3 8 1.47 9H), 1.63 (in, 1H), 1.81 (in, 1H), 2.07 3H), 2.34 (in, 114), 3.42 (dd, J 12.7, 0.9 Hz, 3.62-3.85 (in, 3H), 4.48 (br, 1H), 5.20 (br, 11-).
To a stirred mixture of (2S 4S)-4-acetoxy-l1-tert-butoxycarbonyl-2-pyrrolidinemethao (9.70 g, 37.4 mmol), p-hydroxybenzoic acid methyl ester (5.69 g, 37.4 inmol) and PPh 3 (10.8 g, 41.1 mmol) in THF (200 ml) was added DIAD 10 ml, 41.1 inmol) at roamn temperature. After h stirring, the mixture was concentrated in vacuo. The resulting residue was chromatographed on silica gel 1700 g, CHCI/EtOAc to give methyl 4-(2.S4.S)-4-acetoxy-l-tertbutoxycarbonyl-2-pyrrolidinyl~inethoxybenzoate (11.8 g, 81%) as a pale yellow oil. 'H-NMR
(CDCI
3 5 1.48 9H), 2.03 311), 2.27 (mn, 2H), 3.46 (in, IN), 3.72 (in, IN), 3.88 3H), 3.98 J= 9.0 Hz, 1H), 4.21-4.47 (in, 2H1), 5.31 (br, lH), 6.96 (br, 2H), 7.98 J =8.8 Hz, 2H).
To a stirred solution of methyl 4-[(2.S4S)-4-acetoxy-1-tert-butoxycarbonyl-4-hydroxy-2pyrrolidinylJinethoxybenzoate (7.43 g, 18.9 mmol) in MeOH (150 mld) was added cat. K 2 C0 3 at roam temperature. After 1 day stirring, the mixture was concentrated in vacuo. The resulting residue was recrystallized by the addition of CHCI 3 -n-hexane, to give methyl 4+II2S 4S)- 1 -tertbutoxycarbonyl-4-hydroxy-2-pyrrolidiny]nethoxybenzoate (5.76 g, 87%) as a colorless solid. 'H- NMR (CDCl 3 8 1.46 9H), 2. 11 (mn, 114), 2.35 (br, 3.27-3.65 (in, 2H), 3.89 3H), 4.07- 4.54 (in, 4H), 6.96 J 6.9 Hz, 2H), 7.99 J =6.9 Hz, 2H).
To a stirred solution of methyl 4-[(2SA4S)-1-tert-butoxycarbonyl-4-hydroxy-2-pyrrolidinylJ methoxybenzoate 10 g, 5.98 inmol) in THE (60 ml) was added 60% oil NaH (359 ing, 8.97 iniol) at 0 After 15 minutes stirring, Mel (1.20 ml, 8.97 inmol) was added to the midxture was added at same temperature, and the resulting mixture was allowed to raise to roam temperature for over I h. Then 60% oil NaH (3 59 mg, 8.97 inmol) and MeT 20 ml, 8.97 inmol) was added to WO 01/00206 WO 0100206PCT/USOO/18079 the reaction mixture at room temperature and stirred for 14 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 NaSO 4 the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, n-hexane/EtOAc(411)J, to give methyl 44(2S,4S)-1-tertbutoxycarbonyl-4methoy-2-pyrrolidinyllmethoxybenzoate (1.32 g, 60%) as a colorless oil. 'H- NMR (CDCl 3 8 1.48 9H), 2.05 1H), 2.29 J =14.2 Hz, 1H), 3.30 3H4), 3.36-4.38 (in, 4H), 6.76 (br, 2H), 7.97 J 8.8 Hz, 2H).
To a stirred solution of methyl 4-[(2S,4S)-1-tert-butoxycarbonyl-4-methoxy-2pyrrolidinyl] methoxybenzoate (2.38 g, 3.61 nunol) in CH 2
CI
2 (46 mlJ) was added TFA (23 ml) at room temperature. After 14 h stirring, the mixture was concentrated in vacuo. The residue was diluted by the addition of CH 2 C1 2 and 1 N NaOH, and extracted with CH 2
CI
2 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, CHC1 3 /MeOH (20/ to give methyl 4S)- 4-methaxy-2-pyrrolidinyl~methoxybenzoate (950 mg, as a yellow oil. 'H-NMR (CDCI,) 2.16 J= 5.3 Hz, lH), 2.72 iN), 2.95 J =6.8 Hz, lH), 3.11 J =11.0 Hz, 111), 3.26 J 1.9 Hz, 3H), 3.52 (br, 111), 3.84 J 1.7 Hz, 3H), 3.92 1H), 4.00 J 4.1 Hz, 2H), 6.88 (in, 2H), 7.94 (in, 214).
A mixture of 3-methoxy-4-[N '-(2-methylphenyl)ureido~pheriylacetic acid (375 mg, 1. 19 inmol), methyl 4-[(2S,4S)-4-methoxy-2-pyrrolidinyllmedioxybenzoate (317 mg, 1. 19 inmol), EDC-HCl (342 mg, 1.79 minol), HOBT (242 mg, 1.79 mmol) and Et 3 N (0.83 mld, 5.95 minol) 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., the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHC13VAectone (10/l)CHCI/MeOH to give methyl 4-[(2S,4S)-4-methoxy-l-[3methoxy-4-[N '-(2-methylphenyl)ureidolphenylacetylJ-2-pyrrolidinyl~methoxybenzoate (650 mg, 98%) as a pale brown amorphous solid 'H-NMR (CDCI 3 5 2.03 (in, 1H), 2.31 3H), 2.32 (in, INH), 3.29 J 1.0 Hz, 311), 3.57-3.68 (in, 5H), 3.88 J =1.0 Hz, 3H1), 3.99-4.06 (mn, 2H), 4.46 (in, IM1, 6.19 (mn, lH), 6.80 lH), 6.81 J= 9.0 Hz, IH), 6.96-7.19 (in, 4H), 7.29 (in, 211), 7.50 J 6.7 Hz, 7.95-8. 10 (in, 3H); MIS (ESI) m/z 562 1).
To a solution of methyl 4-[(2S,4S)-4-methoxy- 1 -inethoxy-4-[N -(2-methylphenyl) ureido] phenylacetyl]-2-pyrrolidinyl~methoxybenzoate (650 mg, 1. 16 inmol) in THE (18.5 ml), WO 01/00206 WO 0100206PCT/USOO/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 CHCI 3 -MeOH The combined extracts were dried over NaSO 4 and concentrated in vacuo. The residue was chromatographed on silica gel [50 g,
CHCI
3 IMeOH to give 132 (540 mg, as a colorless amorphous solid. MW 547.60 IR (KBr) 3354, 2937, 1709, 1685, 1604, 1533, 1454 'H-NMR (DMSQ-d 6 5 2. 11 (in, 2H), 2.25 3H), 3.22 3H1), 3.49-3.78 (in, 4H4), 3.82 and 3.86 3H, amide isomers), 3.87-4.52 (in, 4H4), 6.7 1-7.17 (in, 7H1), 7.79 J 8.1 Hz, 1H1), 7.86-8.03 (in, 3H), 8.45.8.57 (in, 2H1), 12.64 (br, 11H); MIS (ESI) m/z 548 Anal. Calcd for C30H- 33
N
3
O
7 dlNa l.5H- 2 0: C, 60.29; H,6.07; N, 7.03.
Found: C, 59.90; H, 5.59; N, 6.69.
Example 125 '-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-4-methoxy-2-pyrrolidinyl inethoxybenzoic acid 0COOH I0 133 A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (398 mg, 1. 19 mmol), methyl 4-[(2S,4.S)-4-inethoxy-2-pyrrolidinyl~methoxybenzoate (317 mng, 1. 19 mmol), EDC&HCI (342 mg, 1.79 inmol), HOBT (242 mg, 1.79 minol) and Et 3 N (0.83 mil, 5.95 inmol) in DMIF (5 mil) 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, CHClVAectone (10/1)CHCI 3 /MeOH to give methyl chlorophenyl)ureidoJ.3-inethoxyphenylacetylJ-4-methoxy-2-pyrrolidinyl~inethoxybenzate (600 mg, as a colorless amorphous solid 1 H-Nlv (CDCI 3 5 1.99-2.06 (in, 1H), 2.34 J 13.9 Hz, IH), 3.30 3.59 J= 7.4 Hz, 11H), 3.62 J=3.2 Hz, 2H), 3.83 3H), 3.88 3H), 4.00-4.18 (in, 3H), 4.42-4.51 (in, 2H), 6.82-7.07 (in, 7H), 7.28 J 8.3 Hz, 1H), 7.35 (dd, J 7.9, 1.5 Hz, 114), 7.94-8.00 (in, 3H),8. 18(d, J 8.3Hz, 1H); MIS (ESI) nI/z 582 584 To a solution of methyl 4-[(2S,4S)-1 -[4-[N-'(2-chlorophenyl)ureido]-3-inethoxyphenyl acetyl]-4.inethoxy-2-pyrrolidinyl]methoxybenzoate (600 mg, 1.03 inmol) in THE (16 mil), 0.25 N NaOH (16 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 The combined extracts were dried over NaSO 4 and concentrated in vacuo. The residue was purified on TLC [CHCI 3 MeOH to WO 01/00206 WO 0100206PCT/USOO/18079 give 133 (495 mg, as a colorless amorphous solid. MW 568.02 IR (KBr) 3330, 3070, 2937, 1709, 1685, 1604, 1533 cm" 1 'H-NMIR (DMSO-d) 8 2.11 (in, 2H), 3.22 3H), 3.56-3.78 (in, 4H1), 3.81 and 3.85 3H, amide isomers), 3.88-4.56 (in, 4H), 6.73 and 6.77 J 8.1 Hz, III, amide isomers), 6.85 and 6.91 1H, amide isomers), 7.01-7.07 (in, 3H), 7.28 J 8.1 Hz, I H), 7.43 J 8.1 Hz, 7.85-7.94 (in, 2H), 7.97 J 8.6 Hz, 111), 8.09 J 8.3 Hz, I1H), 8.90-8.95 (in, 2H); MIS (FAB) m/z 570 572 Example 126 4-I(2S,4S)- 1-14-IN '-(2-bromophenyl)ureidol-3-inethoxyphenylacetyl]-4-methoxy-2-pyrrolidinyl inethoxybenzoic acid (?NY4 0 BrH H 6e134 A mixture of 4-[N'-(2-bormophenyl)ureidoJ-3-inethoxyphenylacetic acid (451 mg, 1. 19 inmol), methyl 4-[(2S,4S)-4-methoxy-2-pyrrolidinyljmethoxybenzoate (317 mg, 1. 19 iniol), EDC-HCI (342 mg, 1.79 inmol), HOBT (242 mg, 1.79 imol) and Et 3 N (0.83 ml, 5.95 inmol) 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 NaSO 4 the extracts were concentrated in vacuo. The residue was chrotnatographed on silica gel [50 g, CHCI 3 /Aectone (10/1)1, to give methyl 4-[(2S,4S)-l -14-[N'-(2-bromophenyl)ureidoJ-3 methoxy phenylacetyl]-4-inethoxy-2-pyrrolidiny~methoxybenzoate (760 ing, 100%) as a yellow oil. 'H-NMvR (CDCI 3 5 1.99-2.32 (mn, 1H), 2.34 J 13.4 Hz, 111), 3.30 3H), 3.59 (mn, 111), 3.63 J =3.2 Hz, 2H), 3.68 (dd, J 12.2, 5.1 Hz, 1H), 3.81 (br, 3H), 3.88 311), 3.91-4. 16 (in, 214), 4.49-4.51 (in, 2H), 6.82-7.15 (in, 7H), 7.31 J= 8.1 Hz, 111), 7.52 J 8.1 Hz, IH), 7.93- 8.00 (in, 311), 8.14 J 8.3 Hz, IlH); MS (ESI) ink~ 626 628 (MW+3).
To a solution of methyl 4-[(2S,4S)-1-[4-[N'-(2-broinophenyl)ureido~phenylacetyl]-4methoxy-2-pyrrolidinyl~methoxybenzoate (760 ing, 1. 19 mmol) in THF (19 mld), 0.25 N NaOH (19 ml) was added. After stirring at room temperature for 12 h, the mixture was acidified with 1 N HCI and extracted with CHCI 3 -MCOH The combined extracts were died over Na 2
SO
4 and concentrated in vacuo. The residue was chromatographed on silica gel [50 g, CHCIVMeOH to give 134 (580 ing, 78%) as a colorless amorphous solid. NM 612.47 IR (KBr) 3330, 2935, 1709, 1685, 1604, 1529, 1434 'H-NNIR (DMSO-d) 8 2.11 (in, 2H), 3.22 3H), 3.58- 3.78 (mn, 4H), 3.81 and 3.86 3H, amide isomers), 3.92-4.52 (mn, 4H), 6.72 J =8.6 Hz, 11-), 6.77 J 8.3 Hz, 1H), 6.85 and 6.91 IH, amnide isomers), 6.97 J= 7.1 Hz, 1H), 7.02 and WO 01/00206 WO 0100206PCT/USOO/18079 7.06 J =8.6 Hz, 2H, amide isomers), 7.32 J =7.3 Hz, 1H), 7.59 (dd, J 1.0Hz, 1H), 7.94 (dd, J= 8.1, 1.2 Hz, 2H), 7.95-7.98 (in, 2H), 8.74 I1H), 8.94 I 12.63 (br, I MS (FAB) ni/z 612 (W t 614 Example 127 4-I(4R)-methoxy-1 -13-methoxy-4-IN '-(2-mci hylphenyl)ureidojphenylacetylJ-(2S)-pyrrolidinyl methoxyjbnzoic acid PMe YN N-Y Q O-D-COOH To a stirred solution of 1 -(tert-butoxycarbonyl)-(4R)-methoxy-(2S)-pyrrolidinylcarboxylic acid (2.87 g, 11.7 mmnol) in THF (25 ml) was added BH 3 -DMS (1.66 ml, 17.5 inmol) at roam temperature and the reaction mixture was stirred at room temperature overnight. The mixture was evaporated and the residue was dissolved with CH 2
CI
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 silicagel with CHCI 3 -MeOH (50: 1, v/v) as eluent to give 1-(tert-butoxycarbornyl)-(4R)-methoxy-(2S)prolinol (1.79 g, as a colorless oil. IH-NMVR (CDCI 3 5 1.47 9 1.69-1.73 (in, 1 H), 2.12-2. 17 (in, 1 3.31 3 3.37-3.40 (in, 1 3.53-3.62 (in, 2 3.68-3.73 (in, 1 3.83- 3.87 1 4.044.07 (in, I 4.90-4.92 (in, 1 MS (FAB) ,n/z 232 To a stirred solution of methyl 4-hydroxybenzoate 18 g, 7.76 inmol), I1-(tert-butoxy carbonyl)-(4R)-methoxy-(2S)-prolinol (1.79 g, 7.74 inmol) and Ph 3 P (2.44 g, 9.30 inmol) in THE ml) was added DIAD (1.83 ml, 9.29 inmol) and the reaction mixture was heated under reflux for 5 hr. After cooled to room temperature, the mixture was evaporated. The residue was filtered on silica-gel with toluene-acetone 1, v/v) as eluent to give the crude product. The crude product was dissolved in CH 2
CI
2 (20 nil). The solution was added TEA (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
K
2 C0 3 and evaporated. The residue was purified by column chromatography on silica-gel with
CHCI
3 -MeQH (30:1 to 30:2, v/v) as eluent to give methyl 4-[(4R)-methoxy-(2S)-pyrrolidinyl methoxyjbeuzoate (1.67 g, 81% for 2 steps) as a reddish brown oil. 'H-NDAR (CDCl 3 6 1.65-1.72 (in, 1 1.89 Cbs, 1 2.05-2.22 (in, 1 2.95-3.15 (in, 2 3.31 3 3.69-3.76 (Mn 1 H), 3.88 3 3.91-4.06 (in, 3 6.89-6.92 (in, 2 7.96-7.98 (mn, 2 MS (FAB) m/lz 266 1).
WO 01/00206 WO 0100206PCT/USOOI 18079 A mixture of 3 -methoxy-4-[N -(2-methylphenyl)ureidojphenylacetic acid (470 mg, 1.50 rumol), methyl 4-[(4R)-methoxy-(2S)-pyrrolidinYlmethoxyjbenzoate (396 mg, 1.49 minol), EDC-HC1 (343 mg, 1.79 mnxol), HOBt (242 mg, 1.79 mimol) and Et 3 N (250 ml, 1.79 mmol) in THF (10 mil) 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 NaSO,, and evaporated.
The residue was purified by column chromatography on silica-gel with CHCI 3 -MeOH (100: 1, v/v) as eluent to give methyl 4-[(4R)-methoxy-1-[3-methoxy-4-[N'-(2-methylphenyl)ureidojpheny acetylj-(2S)-pyrrolidinylmethoxyJbenzoate (822 mg, as a white foam. 'H-NMR (CDC 3 6 2.14-2.24 (im, 2 2.27 3 3.25 3 3.51 3 3.58-3.73 (in, 4 3.88 3 H), 3.98-4.09 (in, 2 4.40-4.53 (in, 2 6.67-7.29 (series of mn, total 9 7.57-7.59 (in, 1 H), 7.9 1-7.93 (in, 2 8.04-8.06 (in, 1 MS (FAR) m/lz 562 (Mr+l).
To a stiffed solution of methyl 4-[(4R)-methoxcy-1-[3-methoxy-4-[N'-(2-methylphenyl) ureidoJ phenylaceylJ-(2S)-pyrrolidinylmethoxy]benzoate (517 mg, 0.92 inmol) in THF (5 ml) was added 0.5 N NaOH (5 n-l) and the reaction mixture was heated under reflux for 3 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 purified by recrystallization from MeGH- CHC1 3 -IPE to give 135 (144 mg, 29%) as a white crystalline powder. MW 547.60 mp 112- 11 5*C; 'H-NMR (DMSO-d) 8 2.04-2.17 (in, 2 2.25 3 3.21 3 3.56-3.75 (in, 4 H), 3.79 3 4.04-4.35 (in, 4 6.73-7.17 (series of m, total 7 7.79-7.81 (in, 1 7.87-7.89 (mn, 2 7.99-8.0 1 (in, 1 8.47 1 8.55 1 12.63 Cbs, 1 MS (FAR) m/z 548 Anal. Calcd for C3OH 3 3
N
3
O
7 l/4H 2 0: C, 65.26; HL 6.12; N, 7.6 1. Found: C, 65.36; H, 6.45; N, 7.24.
Example 128 1-[4-[N'-(2-fluorophenyl)ureido]-3-nethoxyphenylacetyl]-(4R)-methoxy-(2S)-pyrolidinyI methoxy~benzoic acid PMe Y'HH Me 136 A mixture of 4-[N'-(2-fluorophenyl)ureidoJ-3-methoxyphenylacetic acid (476 mng, 1.50 inmol), methyl 4-[(4R)-methoxy-(2S)-pyrrolidinylmethoxy~bnzoate (3 97 mg, 1. 50 minol), EDC-HCI (344 ing, 1.79 inmol), HO~t (243 ing, 1.80 innol) and Et 3 N (250 nml, 1.79 inmol) 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 evaporated.
WO 01/00206 WO 0100206PCT/USOO/18079 The residue was purified by column chromatography on silica-gel with CHCI 3 -MeOH (100: 1, v/v) as eluent to give methyl 1-[4-f N'-(2-fluorophenyl)ureidol-3 -methoxyphenylacetyl]-(4R)methoxy-(2S)-pyrrolidinylmethoxy]benzoate (806 mg, as a pale yellow foam. 'H-NMR
(CDCI
3 5 2.14.2.37 (in, 2 3.28 3 3.44 3 3.48-3.74 (in, 4 3.88 3 4.02- 4.15 (mn, 2 4.43-4.58 (mn, 2 6.63-7. 10 (series of m, total 7 7.68-7.73 (in, 1 7.89-802 (in, 4 8.16-8.20 (mn, 1 MS (FAR) m/z 566 To a stirred solution of methyl 4-fl -[4-[N'-(2-fluorophenyl)ureidoJ-3-methoxyphenyl acetyl]-(4R)-methoxy-(2.S)-pyrrolidinylmethoxylbenzoate (491 mg, 0.87 mmiol) 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-i N HCl and the resulting precipitate was collected under a reduced pressure. The crude solid was purified by recrystallization from MeOH- CHC1 3 -l.PE to give 136 (173 ing, as a white crystalline powder. MW 551.56 mp 111-1 16 0
C;
'H-NMR (DMSO-dQ 582.08-2.17 (in, 2 3.21 3 3.56-3.73 (mn, 4 3.78 3 4.04- 4.33 (in, 4 6.74-7.22 (series of in, total 7 7.87-7.89 (mn, 2 7.99-8.01 1 8.16-8.20 (mn, 1 8.70 1 9.18 1 12.64 (br s, 1 MS (FAR) m/z 552 Anal. Calcd for C-2H30FN 3 O0.l,5H 2 O: 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-f l-f4-[N '-(2-chlorophenyl)ureido-3-methoxyphenylacetyl-(4R)-methoxy-(2S)-pyrrolidinyI methoxy]benzoic acid PMe ~I H Me 137 A mixture of 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (460 mg, 1.37 inmiol), methyl 4-f(4R)-inethoxy-(2S)-pyrrolidinylmethoxy~benzoate (365 mg, 1.38 nunol), EDC-HCI (316 mng, 1.65 inmol), HOBt (223 mng, 1.65 inmol) and Et 3 N (230 ml, 1.65 iniol) in TI-F (10 ml) was stiffed 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, v/v) as eluent to give methyl 4-[1-[4-[N'-(2-chlorophenyl)ureido-3-nethoxyphenylacetylJ-(4R)inethoxy-(2S)-pyrrolidinylmethoxylbenzoate (801 mng, q. as a white foam. 'H-NMR (CDCl 3 2. 13-2.36 (in, 2 3.27 3 3.58 3 3.61-3.73 (mn, 4 3.88 3 4-06-4.14 (in, 2 4.43-4.56 (in, 2 6.70-6.99 (series of mn, total 5 7.23-7.42 (mn, 4 7.90-8.00 (mn, 3 H), 8.17-8.20 (in, I MIS (FAR) m/z 582 1).
WO 01/00206 WO 0100206PCT/USOO/18079 To a stirred solution of methyl l-[4-[N'-(2-chlorophenyl)ureido]-3-methoxYPhenyl acetylJ-(4R)-methoxy-(2S)-pyrrolidinylmethoxy~benzoate (541 mg, 0.93 mmol) in THFf (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 m-ixture was poured into ice-i N HCI and the resulting precipitate was collected under a reduced pressure. The crude solid was purified by recrystallization from MeOH-
CHC
3 -IIPE to give 137 (281 mg, 53%) as a white crystalline powder. MW 568.02 mp 116-1 19'C; 'H-NMR (DMSO-d) 5 2.08-2.17 (in, 2 3.21 3 3.56-3.73 (in, 4 3.79 3 4.04- 4.33 (in, 4 6.75 J 8.3 Hz, 1 6.87 1 7.02 J =8.3 Hz, 3 7.28 J =7.8H-14 1 7.44 J= 7.8 Hz, 1 7.87-7.89 (in, 2H), 7.96 J= 8.3 Hz, 1 8. 10 J= 8.3 Hz, 1 8.89 1 8.93 1 12.63 (br s, 1 MS (FAB) mih 568 A naL Calcd for
C
2 ,430CIN 3
O
7 -1/4H- 2 O: C,60.84;H,5.37;CI,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)ureidol-3 -iethoxyphenylacetyl]-(4R)-methoxy-(2S)-pyrrolidiny inethoxylbenzoic acid PMe Me138 A mixture of 4-[N'-(2-broinophenyl)ureidoj-3-methoxyphenylacetic acid (600 mg, 1.58 inruol), methyl 4-[(4R)-inethoxy-(2S)-pyrrolidinylmethoxylbenzoate (420 ing, 1.58 mmol), EDC-HCI (364 mg, 1.90 inmol), HOBt (214 mg, 1.58 minol) and Et 3 N (265 ml, 1.90 inrol) in THF (15 ml) was stiffed 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
SQ
4 and evaporated.
The residue was purified by column chromatography on silica-gel with CHC1 3 -MeOH (100: 1, v/v) as eluent to give methyl 1-14-IN '-(2-bromophenyl)ureidoJ-3-methoxyphenylacetyl]-(4R)methoxy-(2S)-pyrrolidinylmethoxy]benzoate (1.01 g, as a pale yellow foam. 'H-NMR
(CDCI
3 5 2.13-2.33 (in, 2 3.27 3 3.57 3 3.61-3.72 (in, 4 3.88 3 4.05- 4.14 (in, 2 4.43-4.57 (in, 2 6.70-7.00 (series of mn, total 5 7.29-7.52 (in, 4 7.92-8.01 (in, 3 8.12-8.15 (in, 1 MS (FAB) m/lz 626 To a stirred solution of methyl 1-[4-[N'-(2-bromophenyl)ureido]-3 -iethoxyphenyl acetylj-(4R)-methoxy-(2S)-pyrrolidinylnethoxyjbenzoate (697 mng, 1. 11 inmol) in THE (8 ml) was added 0.5 N NaOH (8 and the reaction mixture was heated under reflux for 2 hr. After cooled to room temperature, the mixture was poured into ice-l N HCI and the resulting precipitate was collected under a reduced pressure. The crude solid was purified by recrystallization from MeOH- WO 01/00206 WO 0100206PCT/USOO/18079 CHCI,-IPE to give 138 (252 mng, as a white crystalline powder. MW 612.47 mp 125-130 0
C;
1 H-NMR (DMS04) 5 2.08-2.17 (mn, 2 3.21 3 3.60-3.72 (in, 4 3.79 3 3.95- 4.33 (mn, 4 6.75-7.08 (series of Rn total 5 7.3 1-7.34 (in, 1 7. 59-7.61 (in, 1 7.87-7.89 (in, 2 7.93-7.96 (in, 2 M, 8.73 1 8.91 I 12.63 (br s, 1 MIS (FAB) In/z 612 Anal Calcd for C ,H13BrN 3
O
7 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-difuoro-lI-[3-inethoxy-4-[N'-(2-inethylphenyl)ureidojphenylacetyl]-2-pyrrolidinylmethoxy benzoic acid N Q--OKOWJCOOH Me H H 6Me 139 To a stirred solution of N-Boc proline methyl ester (2 .0 g, 8.15 inmol) in CH 2 Cl 2 were added 3 A molecular sieves (2 g) and PDC (4.60 g, 12.2 inmol). 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-(tertbutoxycarbonyl)4-oxopyrrolidine-2-carboxylate 13 g, 57%) as a colorless oil. 'H-NMvR (CDC 3 1.46-1.48 (in, 9 2.56-2.6 1 (in, 1 2.88-3.00 (in, 1 3.77 3 3.82-3.88 (in, 2 H), 4.71-4.83 (in, 1 H).
To a cold (-78 0 stirred solution of methyl 1 -Qert-butoxycarbonyl)-4-oxopyrrolidine-2carboxylate 13 g, 4.65 inmol) in C11 2 C1 2 (20 ml) was added inethylDAST (1.1 min, 11.6 minol).
The mixture was allowed to warm to room temperature. After 15 h stirring, the mixture was poured into H 2 0 (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 CHCI 3 -EtOAc (20: 1) as eluent to give methyl Il-(tert-butoxycarbonyl)-4,4-difluoropyffolidine -2-carboxylate (885 mng, 72%) as a yellow oil. 'H-NMR (CDCl 3 8 1.42 and 1.47 each, total 9 2.46 (ddd, d 26.9 13.7, 5.1 Hz,l1H), 2.62-2.78 (in, 111), 3.75-3.95 (in, 5H), 4.43-4.57 (in,111).
To a stirred solution of methyl 1 -Qert-butoxycarbonyl)-4,4-difuoropyrrolidine-2-carboxylate (885 mg, 3.34 iniol) in THE (25 ml) was added 0.25 N NaOH (26.7 ml, 6.67 inmol) and the stirring was continued for I h. The mixture was poured into I N HCI (100 ml) and'extracted with CHCI, (2 x 200 ml). The combined extracts were washed with brine (100 ml), dried over MgSQ 4 and evaporated to give 1-(tert-butoxycarbonyl)-4,4-difluoropyrrolidine-2-caboxyic acid (775 mng, WO 01/00206 WO 0100206PCT/USOO/ 18079 92%) as a yellow crystalline solid. mp 113-117 0 C; 3 H-NMR (CDC 3 8 1.44 and 1.49 each, total 9 2.53-2.80 (in, 2 3.71-3.90 (in, 2 4.20-4.61 (in, 1 MS (FAB) m/z, 252 (Wi+ Anal. Calcd for C, 0
H
13
F
2 C, 47.8 1; H, 6.02; N, 5.58. Found: C, 48.06; H, 6.05; N, 5.45.
To a stirred solution of N-Qtert-butoxycarbonyl) 4,4-difluoroproline (3.00 g, 11.9 minol) in TIHF ml) was added BH 3 D!MS -IMl, 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 CHC1 3 (2 x 200 mlJ).
The combined extracts were dried over MgSO, and evaporated. The residue was chromatographed on silica gel with CHCI 3 -EtOAc 1) as eluent to give 1-(tert-butoxycarbonyl)-4,4-difluoro-2pyrrolidinylmethanol (2.11 g, 75%) as a colorless oil. IH-NMR (CDC 3 8 1.48 9 2.04-2.55 (in, 2 3.59-4.17 (mn, 5 H).
To a stiffed mixture of l-Qtert-butoxycarbonyl)-4,4-difluoro-2-pyrrolidinylmethano (600 mng, 2.53 mmol), methyl 4-hydroxybenzoate (462 ing, 3.03 mmol), PhyP (795 mng, 3.03 iniol) in THF ml) was added DIAD (597 ul, 3.03 iniol) 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 1) as eluent to give methyl 4-Q(ert-butoxycarbonyl)-4,4-difluoro-2-pyrrolidinyhnethoxyjbenzoate (831 mng, 880/o) as a colorless oil. 'H-NMR (CDCI 3 5 1.48 9 2.53-2.61 (mn, 2 3.63-4.41 (series of in, total 8 6.94 J= 8.8 Hz, 2 7.99 J =8.8 Hz, 2 H).
A mixture of methyl 44 l-QIert-butoxycarbonyl)-4,4-difluoro-2-pyrrolidinylmethoxyJ benzoate (830 ing, 2.23 inmol) and TFA (5 ml) in CHC1 2 (5 ml) was stirred for 3 h and concntrated in vacuo. The residue was made basic with sat. NaH-C0 3 and extracted with CHC1 3 (2 x 200 ml). The combined extracts were dried over K 2 C0 3 and concntrated in vacua to give methyl 4-(4,4-dilluoro-2-pyrrolidinylnethoxy)benzoate (550 ing, 91%) as a pale yellow solid. 'H-NMiR
(CDCI
3 8 2.19 (in, 1 2.43 (in, 1 3.19-3.41 (in, 2 3.77 (mn, 1 3.89 3 4.00-4.09 (in, 2 6.92 J =9.0 Hz, 2 7.99 J 9.0 Hz, 2 MS (FAB) m/'z 272 Anal.
Calcd for C131H1F 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 mng, 1.99 minol), 3-inethoxy-4-[N'-(2-inethylphenyl)urcido]phenylacetic acid (626 mg, 1.99 mmol), WO 01/00206 WO 0100206PCT/USOEJ/18079 EDC-HCI (572 mg, 2.99 mmol), HQBt (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 nil), dried over MgSO,, and concntrated in vacuo. The residue was chromatographed on silica gel with
CHCI
3 -MeOH (20:1) as eluent to give methyl 4-[4,4-difluoro-l-[3-methoxy-4-[N'-(2methylphenyl)ureidoj phenylacetylJ-2-pyrrolldinylmethoxy~benzoate (1.00 g, 89%) as a colorless foam. 'H-NMR (CDCI 3 8 2.31 3 2.47-2.63 (in, 2 3.52-3.97 (series of s and m, total 4.07-4.30 (in, 2 4.67-4.69 (in, 1 6.45 1 6.65 J= 1.7 Hz, 1 6.74-6.76 (mn, 1 6.84 J 8.8 Hz, 2 7.14 (mn, 2 7.24 (in, 2 7.52-7.54 (in, 1 7.94 J 8.8 Hz, 2 8.09 J =8.1 Hz, I H).
A mixture of methyl 4-[4,4-difluoro-1-[3-inethoxy-4-[N'-(2-methylphenyl)ureidolphenylacetylJ-2-pyrrolidinylinethoxyjbenzoate (1.00 g, 1.76 inmol) and 0.25 N NaOH (14 ml, 3.50 minol) in THF (14 ml) was stirred overnight. The mixture was acidified with 1 N HCl 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 CHCI 3 -MeOH (20:1 to 10: 1) as eluent to give 139 (658 mg, as a colorless crystalline powder. NM 553.55 mp 135-140 OC; IH-NMR (DMSO-d) 5 2.23 3 2.49-2.73 (in, 21-H), 3.36-4.55 (series of in, 10 6.73 J= 8.3 Hz, 1 6.84 1 6.93 J =7.3 Hz, 1 7.00 J =8.3 Hz, 2 7.10-7. 16 (mn, 2 7.78 (d, J =8.3 Hz, 1 7.86 J= 8.3 Hz, 2 8.00 (d,J 8.3 Hz, I 8.47 I 8.56 1 H); MS (FAB) m/z, 554 Anal. Calcd for CH1 F 2
N
3
O
6 3/41- 2 O: C, 61.42;, H, 5.44; N, 7.06.
Found: C, 61.30; H, 5.44; N, 7.06.
Example 132 1-[4-[N'..(2-chlorophenyl)ureido]-3-inethoxyphenylacetyll-4,4-diluoro-2-pyrrolidinylnethoxy benzoic acid Me o-J-oo 140 A mixture of methyl 4-(4,4-difluoro-2-pyrrolidinylmethoxy)benzoate (229 ing, 0.845 inmol), 4- -(2-chlorophenyl)ureidol-3-nethoxyphernylacetic acid (283 mg, 0.845 inmol), EDC-HCI (243 mg, 1.27 minol), HO~t DM"P and DMIF (10 ml) was stirred overnight. The m-ixture 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 CHCl 3 EtOAc (20:1 to 4: 1) as eluent to give methyl 4-[l-[4-[N'-(2-chlorophenyl)ureidol-3-nethoxy WO 01/00206 WO 0100206PCTILJS00II8079 phenylacetyl]-4,4-difuoro-2-pyrrolidinylmethoxyjbenzoate (482 mg, 97%) as a colorless viscous solid. 'H-NMR (CDCI 3 8 2.50-2.67 (in, 2 3.54-4.71 (series of mn, 13 6.69 J= 1. 5 Hz, 1 6.76 J= 8.3 Hz, 1 6.84 J= 8.8 Hz, 2 6.98 (dt, J= 7.8, 1.5 Hz, 1 7.23-7.27 (in, 1 7.33 J= 8.3 Hz, 1 7.39 2 7.94 J =8.8 Hz, 2 8.00 J= 8.3 Hz, 1 8.19 (dd, J= 8.3, 1.5 Hz, 1 H).
A mixture of methyl methyl 4-li1-[4-[N' -(2-chlorophenyl)ureidoj-3-methoxyphenylacetyl]- 4,4-difuoro-2-pyrrolidinylmethoxyjbenzoate (480 mng, 0.816 inmol), 0.25 N NaOH (6.5 ml, 1.65 mmol), and THE (20 was stirred for 3 days. The mixture was poured into 1 N HCI (100 ml) and extracted with CHCI,-MeOH 2 x 200 ml). The combined extracts were dried over MgSO 4 and concnitrated in vacuo. The residue was chromatographed on silica gel with CHCI 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-Q 8 2.45-2.74 (mn, 2 3.63-4.83 (series of m, 10 6.76 J= 8.3 Hz, I H), 6.87 1 7.00-7.05 (in, 3 7.26-7.30 (mn, 1 7.44 (dd, J 1.2 Hz, 1 7.88-7.93 (in, 2 7.98 J =8.3 Hz, 1 8. 10 J= 8.3 Hz, 1 8.92 I 8.96 H)M; MIS (FAB) m/1z 574 Anal. Calcd for C H 26 C1 3
O
6 -HO: C, 56.81; H, 4.77; N, 7. 10. Found: C, 56.75; H, 4.69; N, 6.79.
Example 133 4-[(2R,3R,4.S)-3,4-isopropylidenedioxy-l1-[4-[N'-(2-inethylphenyl)ureido]-3-methoxyphenylacetylJ- 2-pyrrolidinyliinethoxybenzoic acid N Y -Q O-CJCOoH Me H 6e141 To a solution of methyl (2S,3R,4S)- 1 -benzyloxycarbonyl-3,4-isopropylidenedioxy-2-pyrrolidinyI carboxylate (10.7 g, 31.9 innol) in THE (250 ml), 0.25 N NaOH (255 mld) was added. After stirring at room temperature for 24 h, the mixture was acidified with I N HCI and extracted with EtOAc. The combined extracts were washed with brine, which were dried over NaSO, and concentrated in vacuo, to give (2S,3R,4.S)-lI-benzyloxycarbonyl-3,4-isopropylidenedioxy-2pyrrolidinylcarboxylic acid (9.87 g, as a colorless oil. 'H-NMR (CDCl 3 8 1.32 3H), 1.46 J 2.7 Hz, 3H), 3.61(in, 1H), 3.82 and 3.92 J 12.7 Hz, IH, aniLide isomers), 4.58 and 4.64 lH, anide isomers), 4.77 J 5.1 Hz, 114), 4.83 and 4.89 J 5.9 Hz, 1H,amude isomers), 5.15 and 5.19 (in, 2H, amideisoiners), 7.3 1-7.37 (in, WO 01/00206 WO 0100206PCT/USOOI 18079 To a stirred solution of (2S,3R,4 1 -benzyloxycarbonyl-3,4-isopropylidenedioxy-2-pyrrolidiny carboxylic acid (9.87 g, 30.7 mmol) in THF (200 ml) was added BH, 3 DMS (6.14 mld, 61.4 innol) at 0 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 of water at 0 The mixture was extracted with EtOAc. The combined extracts were washed with water and brine, which were dried over NaSO, and concentrated in vacuo. The residue was chromatographed on silica gel 1200 g, CHCl3fMeOH (20/ to give (2R, 3R,4S)-1I-benzyloxy carbonyl-3,4-isopropylidenedioxy-2-pyrrolidinylmethanol (10. 1 g, 100%/) as a colorless oil. IH- NMR (CDC1 3 8 1.31 3H), 1.45 311), 3.56-4.74 (mn, 711), 5.14 211), 7.34 (in, To a stirred mixture of (2R,3R,4S)-lI benzyloxycarbonyl-3,4-isopropylidenedioxy-2-pyrrolidinyI methanol (312 ing, 0.64 imnol), methyl p-hydroxybenzoate (67 ml, 0.70 inmol), PPh 3 (184 mg, 0.70 imnol) in THF (7 nil) was added DIAD (138 ml, 0.70 inmol) at 0 'C 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 chroinatographed on silica gel [10 g, n-hexane/EtQAc to give methyl 4-[(2R,3R,4,S)-benzyloxycarbonyl-3,4-isopropylidenedioxy-2-pyrrolidnyl methoxybenzoate (321 ing, as a colorless oil. 'H-NMvR (CDCI 3 5 1.01 6H), 1.03 3H), 2.23 (in, IlH), 2.63 (in, I 3.61 J= 12.5 Hz, 1H), 3.804.27 (in, 4H), 4.84 (br, 1H), 5.01 and 5.08 (ABq, J= 12.2 Hz, 1H, amide isomers), 6.75-6.87 (in, 3H), 7.19-7.63 (in, 151-).
A suspension of methyl 4-[(2R,3R,4S)-l1-benzyloxycarbonyl-3,4-isopropylidenedioxy-2pyrrolidinyllinethoxybenzoate (2.37 g, 5.76 inmol) and 10% Pd/C (240 ing) 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 mnl). The suspension was stiffed 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 chroinatographed on silica gel [100 g, CHCld/acetone to give methyl 4-[(2R,3R,4S)-3,4-isopropylidenedioxy-2pyrrolidinyljinethoxybenzoate (930 mag, as a brown oil. 'H-NMR (CDCI 3 8 1.35 3M), 1.50 3H), 3.02 (dd, J= 13.7, 4.1 Hz, 3.13(d, J =13.7Hz, 3.58 J =6.3 Hz, iH), 3.88 3H), 3.90 (dd, J= 9.3, 6.6 Hz, 1H), 4.02 (dd, J 9.5, 3.9 Hz, IN), 4.74 J =5.6 Hz, I 4.79 (in, INH), 6.90 J= 9.0 Hz, 2H), 7.98 J 9.0 Hz, 211).
A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (437 mg, 1.39 minol), methyl 4-[(2R,3R,4S)-3,4-isopropylidenedioxy-2-pyrrolidinyl]methoxybenzoate (428 ing, 1.39 WO 01/00206 WO 0100206PCT/USOO/18079 mmol), EDC-HCI (400 mg, 2.09 nunol) and DMAP (170 mg, 1.39 mmol) in DMF (12 mld) was stirred at roam 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,
the extracts were concentrated in vacua. The residue was chromatographed on silica gel [70 g, CHCld/acetone to give methyl 4-[(2R,3R,4S)-3 ,4-isopropylidenedioxy- methylphenyl)ureidoJ-3-methoxypherylacetylJ-2-pyrrolidinyllmethoxybenzoate (840 mg, 100%/) as acolorless amorphous solid IR (KBr) 3354, 2985, 2939, 1716, 1533, 1254 cnm-; 'H-NMR (CDC1 3 5 1.31 311), 1.42 311), 2.05 311), 3.50 311), 3.55-3.88 (in, 4H), 3.89 311), 4.13 (in, 111), 4.67 (br, 111), 4.78 J= 6.1 Hz, 111), 4.88 J= 5.6 Hz, 111), 6.46 111), 6.62 J =1.5 Hz, 111), 6.74 (in, 311), 7.05 111), 7.14 J= 7.3 Hz, 11-1), 7.23 (in, 2M1, 7.57 J= 7.8 Hz, 111), 7.91-8.08 (in, 311); MIS (ESI) m/lz 604 Anal. Calcd for C 33 1 37
N
3 s0.6H,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-l1-[4-[N'-(2-methylphenyl)ureido]-3methoxyphenylacetyl]-2-pyrrolidiniyllmethoxybenzoate (183 mg, 0.303 minol) and g.HCI-MeOH (6 ml) was stirred at roam temperature for 17 h. The mixture was concentrated in vacuo. The residue was purified on TLC [CHCI3/MeOH (10/1)1, to give methyl 4-(2R,3R,4S)-3,4-dihydroxy- 1 -[4-[N'-(2-methylphenyl)ureidoJ-3-methoxyphenylacetylj-2-pyrrolidinyl]methoxybenzoate (162 mg, 95%) as a colorless amorphous solid JR (KBr) 3342, 1716, 1604, 1535, 1255 cur'; IH-NMR (CDC1 3 562.25 (br, 311), 3.33-3.75 (in, 711), 3.87 311), 4. 10 J= 8.3 Hz, 111), 4.24 211), 4.37 (in, 214), 6.62-7.94 (in, 1311); MIS (ESI) m/z 564 To a solution of methyl 4-[(2R,3R,4S)-3,4-isopropylidenedioxy-1 -[4-[N'-(2-inethylphenyl)ureidoJ- 3-methoxyphenylacetylJ-2-pyrmolidinyl]inethoxybenzoate (490 mg, 0.8 12 inmol) in THF (9.8 mld), 0.25 N NaOH (9.8 mld) was added. After stirring at room temperature for 4 days, the mixture was acidified with 1 N HCI and extracted with CHC1 3 -MeOH The combined extracts were dried over Na 2 SO, and concentrated in vacua to give 141 (445 mg, 93%) as a colorless amorphous solid.
MW 689.64 JR (KBr) 3354, 2983, 2937, 1707, 1604, 1533 '1-NIVR (DMSO-d) 8 .24 and 1.26 311, amide isomers), 1.26 and 1.32 311, amnide isomers), 2.24 311), 3.40 (dd, J= 14.0, 5.1 Hz, 111), 3.60 (in, 211), 3.71 (mn, 111), 3.76 311), 3.82 311), 3.92-4.96 (mn, 511), 6.74 and 6.78 (in, 111, amide isomers), 6.83-7.16 (in, 611), 7.79 J 8.3 Hz, 111), 7.87 J 9.1 Hz, 211), 8.01(in, 111),8.49 (dJ= 3 4 .Hz, 111), 8.57 111); MS (FAB) m/z 590(M'+1); Anal. Calcd for C32H 3 5N 3 Os-2.3H 2 O: C, 60.90; H, 6.32; N, 6.66. Found: C, 61.00; H, 6.00; N, 6.27.
WO 01/00206 WO 0100206PCT/USOO/18079 Example 134 4-[(2R,3R,4S)-3,4-dihydroxy-l-[4-[N '-(2-methylphenyl)ureido]-3-methoxyphenylacetylj-2pyrrolidinyljmethoxybenzoic acid
,OH
H H Me 0 o-aC- 142 To a solution of methyl 4-[(2R,3R,45)-3,4-dihydroxy-l-[4-[N-(2-methylphenyl)ureidoJ-3methoxyphenylacetylJ-2-pyrrolidinyllmetioxybenzoate (63 mg, 0. 112 mmol) in THE (0.89 mld), 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 CHCl 3 -MeOH The combined extracts were dried over Na 2
SQ
4 and concentrated in vacuo to give 142 (54 mg, as a colorless amorphous solid.
MW 549.57 IR (KBr) 3356, 2958, 2927, 1685, 1604, 1535, 1255 cm- 1 'H-NNM(DMSO-d) 8 2.24 3H), 3.40 (in, 1H), 3.58 2H), 3.66 (dd, J= 9.8, 6.6 Hz, 1141), 3.80 3H), 3.99-4.30 (in, SF1), 5.10(br, IH), 6.72 J=8.1 Hz, 111), 6.85 1H), 6.93 (tJ= 7.3Hz, IH), 7.03 J=8.8 Hz, 2H), 7.14 J =8.8 Hz, 2H1), 7.79 J 8.1 Hz, 2H), 7.86 J 8.8 Hz, 2H), 7.99 J 8.3 Hz, 8.46 8.56 1H); MS (ESI) ni/z 550(MC+1); Anal. Calcd for
C,,FH
31
N
3 0 8 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)ureidoj-3-methoxyphenylacetylJ-3,4-isopropylidenedioxy- 2-pyrrolidinyl~methoxybenzoic acid I H H ooH 143 A mixture of 4-[N'-(2-chlorophenyl)ureidoJ-3-methoxyphenylacetic acid (487 mng, 1.45 mniol), methyl 4-[(2R,3R,4S)-3,4-isopropylidenedioxy-2-pyrrolidinyl~methoxybenzoate (447 mg, 1.45 mmol), EDC-HCl (418 mng, 2.18 minol) and DMAP (177 mg, 1.45 inmol) 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 the extracts were concentrated in vacuo. The residue was chromatographed on silica gel [70 g, CHCl3/acetone to give methyl 4-[(2R,3R,4S)- 1-[4-[N'-(2-clilorophenyl)ureidoJ -3methoxyphenylacetylJ-3,4-isopropylidenedioxy-2-pyrrolidinylJinetioxybeflzoate (850 mg, 94%) as a colorless amorphous solid IR (KBr) 3329, 2939, 1716, 1627, 1531, 1254 cm"; 'H-NMR (CDCI 3 8 1.33 3M1, 1.43 3H), 3.56 311), 3.61 3.64 11-1), 3.70 (in, 111), 3.79 J= 10.4 WO 01/00206 WO 0100206PCT/USOO/18079 Hz, 111), 3.88 31), 4.14 (dd, J 9.8, 2.2Hz, 111),4.40(dd, J 3.4Hz, 11), 4.67 11), 4.80 6.1 Hz, 111), 4.90 4.6 Hz, 111), 6.65 1.71-Hz, 111, 6.71-6.84(in, 311), 6.98 (dt, J 1.5 Hz, 111), 7.27 (in, 211), 7.33 (dd, J= 8.0, 1.2 Hz, 2141), 7.90-8.01 (in, 311), 8.20 (dd, J 8.3, 1.5 Hz, 111); MS (ESI) m/z 624 626 Anal. Calcd for C, 3
H,N
3
O
8 1.4H- 2
O:
C, 59.19; H, 5.71; N, 6.47. Found: C, 58.85; H, 5.35; N, 6.21.
A mixture of methyl 3R,4S)- 1-14-[N'-(2-chlorophenyl)ureido] -3 -methoxyphenylacetyl] -3,4isopropylidenedioxy-2-pyrrolidinylmethoxybenzoate (177 mg, 0.284 mmol) and g.HCI-MeOH (4 ml]) was stirred at room temperature for 2 days. The mixture was concentrated in vacuo. The residue was purified on TLC ICHCI,/MeOH to give methyl chlorophenyl)ureido]-3-methoxyphenylacetylJ-3 ,4-isopropylidenedioxy-2-pyrrolidinyl] methoxybenzoate (140 mg, 85%) as a colorless amorphous solid IR (KBr) 3338, 2949, 1712, 1623, 1604, 1533 'H-NMR (CDCI 3 5 2.27 (im, 111, 2.79 (in, 111), 3.53 (dd, J= 10.5, 5.9 Hz, 11H), 3.63 311), 3.88 311), 4.21 J =7.8 Hz, 1H), 4.31 2H), 4.43 (dd, J 4.4 Hz, 111), 4.52 J 4.6 Hz, 111), 6.71 111), 6.80 (in, 3H1), 6.99 J 7.3 Hz, 11H), 7.16 111), 7.21 111), 7.39 J= 8.1 Hz, 111), 7.91 (d,J 8.6 Hz, 211), 8.16 J= 8.3 Hz, 111); MS (ES!) m.'z 584 (Mr+ 586 To a solution of methyl 4-[(2R,3R,4.S)-1-[4-[N'-(2-chlorophenyl)ureidoJ-3-methoxypheriylacetyl]- 3,4-isopropylidenedioxy-2-pyrrolidinyl~methoxybenzoate (511 mg, 0.8 19 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 acidified with 1 N HCI and extracted with CHC1 3 -MeOH The combined extracts were dried over NaSO, and concentrated in vacuo to give 143 (504 mng, 100%) as a colorless amorphous solid. NM610.05 IR (KBr) 3330, 2983, 2937, 1711, 1689, 1604, 1533, 1252 cm-';'H-NMR (DMSO-dJ) 8 1.26 311), 1.32 311), 3.40 (in, 1H), 3.60 and.3.61 J= 2.5 Hz, 311, amide isomers), 3.62 (Mn 111), 3.78 and 3.83 311, amideisomers), 4.16 (mn, 211), 4.42-4.98 (in, 311), 6.74-7.15 (in, 611), 7.28 Ct, J 7.3 Hz, 111), 7.43 J= 8.1 Hz, 11-1), 7.78-7.97 (in, 411), 8.08 J 8.3 Hz, 111), 8.89 111), 8.92 111), 12.68 (br, 111); MS m/lz 6 10 612 Example 136 4-I(2R,3R,4S)- 1-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyl]-3,4-dihydroxy-2pyrrolidinyljmethoxybenzoic acid
.OH
rh1~1~. H Me 144ODC O WO 01/00206 WO 0100206PCT/JSOO/18079 To a solution of methyl 1-14-[N'-(2-chlorophenyl)ureido]-3 -methoxyphenylacetylJ-3,4dihydroxy-2-pyrrolidinylmethoxyjbenzoate (63 mg, 0. 108 mmnol) in THE (0.80 ml), 0.25 N NaOH (0.80 ml) was added. After stirrng at room temperature for 3 days, the mixture was acidified with 1 N HC1 and extracted with CHC1 3 -MeOH (10/ The combined extracts were dried over Na 2
SO
4 and concentrated in vacuo to give 144 (61 mg, 100%) as a colorless amorphous solid. M 569.99 IR (KBr) 3338, 1687, 1604, 1533, 1255, 1169, 1036 'H-NMR (DMSO-d, 5 8 3.59 J= Hz, 2H), 3.61 (in, 1H), 3.66 (dd, J= 10.0, 7.1 Hz, 1H), 3.80 3H), 4.00-4.33 5H), 5. 10 (hr, 114), 6.74 J 8.3 Hz, 11H), 6.87 1H), 7.03 (in, 311), 7.28 J 8.3 Hz, 11H), 7.43 J 6.6 Hz, IlH), 7.87 J 8.5 Hz, 2H), 7.95 J 8.3 Hz, IlH), 8.09 J 8.3 Hz, IHM, 8.3 2 I1H), 8.89 I1H), 8.93 1H); MS (ESI) m/z 570 572 (M 4 Anal. Calcd for C, H 2 ClNO,,- .4H 2 O: C, 57.19; H, 5.14; N, 7.15. Found: C, 57.52; HL 5.22; N, 6.76.
Examplie 137 -(2-methylphenyl)ureidoj-3-methoxyphenylacetyl]-5-(R)-phenyl-2-(S)-pyrrolidinyI methoxylbenzoic acid
Z--
Me H H Me 145 To a stirred solution of benzy N-Boc-pyrroglutarate (8.93 g, 28.0 mmol) in THF (100 ml) was added phenyllithiumn (1.0 Min Et 2 O-cyclohexane, 3 3.5 ml, 3 3.5 mmol) at -78'C, and the resulting mixture was gradually warmed up to then stirred overnight. aq.NH 4 CI was added to the reaction mixture, THE was removed in vacuo, then extracted with EtOAc. The organic layer was washed with water and, drying over anhydrous Na 2 SO., then concentrated in vacuo. The residue was chromatographed on silica gel with hexane-EtOAc 1) as eluent, then recrystallized from hexane-EtOAc to give benzyl [2-(S)-(N-Boc-amino)-5-oxo-6-phenyljpentanoate (5.02 g, as a colorless needles. mp 85-87 IC; 'H-NMvR (CDCI,) 5 1.43 91-1), 2.07-2.19 (in, 1 2.27-2.36 (in, 1I-H), 2.97-3.13 (in, 2 4.44 (brs, 1 5.19 (dd, J= 25.2, 12.0 Hz, 2 5.19 (overlap, 1 7.28-7.98 (series of m, 10 MS (ESI) m/1z, 322 (MW'AH).
To a stirred solution of benzyl [2-{S)-(N-Boc-arnino)-5-oxo-6-phenylJpentanoate (2.20 g, 5.54 mmnol) in CHCl, (50 ml) was added trifluoroacetic acid (15 ml) at rt, and the resulting mixture was stirred for 2 h. The mixture was concentrated in vacua and poured into aq.NaH-C0 3 then extracted with EtOAc. The organic layer was dried over anhydrous Na 2
SO
4 then concentrated in vacua to give benzyl 5-phenyl-5-pyrroline-2-(S)-carboxylate (1.60 g, quant.) as yellowish solid.
WO 01/00206 WO 0100206PCT/USOO/18079 The product was used for next reaction without further purification: 'H-NMR (CDCI 3 8 2.20-2.29 (in, 1 2.32-2.42 (in, 1 2.96-3.05 (in, 1 3.12-3.2 1 (in, 1 4.96-5.00 (im, 1 5.24 (s, 2 7.31-7.49 (mn, 8 7.88-7.9 1 (in, 2 MS (ESI) ni/z, 280 A mixture of benzyl 5-phenyl-5-pyrroline-2-(S)-carboxylate (1.59 g, 5.69 inmol) and Pd/C 128 mg) in MeOH (30 mid) was stirred under H, 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 inmol) and 1.0 M-NaOH (8.54 ml, 8.54 inmol), and the resulting mixture was stirred for 30 nun. The mixture was concentrated in vacuo and poured into aq.Nal-C0 3 then extracted with EtOAc. The organic layer was washed with water, saturated brine, drying over anhydrous Na 2
SO
4 then concentrated in vacua. The residue was chroinatographed on silica gel with CHC1 3 -MeOH 1) and recrystallized from hexane-EtOAc to give N-Boc-5-(R)-phenyl-(S)-proline (810 ing, 49%) as a colorless solid. Mp 113-117 0 C; 'H- NMR (CDC1 3 8 1.13 9 1.43 (brs, 1 1.96 (brs, 1 2.09 (brs, 1 2.3 1-2.34 (in, 1 H), 2.46 (brs, 1 4.52 (brs, I 4.69 (brs, 1 7.22-7.37 (in, 5 H).
To a stirred solution of N-Boc-5-(R)-phenyl-2-(S)-proline 14 g, 3.91 nunol) in THF (20 was added l0M-BH 3 -Me 2 S (780 ml, 7.82 inmol) at Mi and the resulting mixture was heated under reflux for 30 min. The mixture was poured into aq. IN-HCI and extracted with EtOAc. The organic layer was dried over anhydrous Na 2
SO
4 then concentrated in vacua. The residue was chromatographed on silica gel with CHCI 3 -MeOH (10 as eluent to give phenylpyrrolidine (1.11 g, quant.) as a colorless oil: 'H-NMR (CDCI 3 8 1.19 (brs, 9 1.65 (brs, 1 1.83-1.90 (in, 1 1.98-2.06 (in, 1 2.22-2.31 (mn, 1 3.75-3.86 (in, 2 4.164.19 (in, 1 4.83 J= 6.8 Hz, 1 4.89 (brs, 1 7.19-7.31 (in, 5 MS (ESI) m/z, 278 (MW+H).
To a stirred solution of N-Boc-2-(S)-hydoxymethyl-5-(R)-phenylpyrrolidine 10 g, 3.97 minol), triphenylphosphine (1.25 g, 4.76 inmol) and methyl 4-hydroxybenzoate (724 ing, 4.76 inmol) was added diisopropyl azodicarboxylate (955 ml, 4.76 innol) at rt and the resulting mixture was stirred at 60*C for 45 min. The mixture was concentrated in vacua, and the residue was chroinatographed on silica gel with hexane-EtOAc (4 as eluent to give methyl phenyl-2-(S)-pyrrolidinylniethoxylbenzoate (1.31 g, as a colorless oil. 'H-NMR (CDCI 3 8 1. 19 and 1.47 (brs, total 9 2.09-2.15 (in, 3 2.33-2.37 (in, 1 3.94 3 4.30 (brs, 1 4.41 (brs, 2 4.77 (brs, I 7.03 J= 8.8 Hz, 2 7.24-7.36 (in, 5 8.03-8.06 (in, 2 MIS (ESI) 412 (MC+H).
WO 01/00206 WO 0100206PCT/USOO/18079 To a stirred solution of methyl 4-[N-Boc-5-(R)-phenyl-2-(S)-pyrrulidinylmethoxy] benzoate (1.28 g, 3. 11 nunol) in CH 2
CI
2 (30 mld) was added trifluoroacetic acid (10 ml) at rt, and the resulting mixture was stirred for 45 min. The mixture was concentrated in vacua and poured into aq.NaHCO 3 then extracted with CHCl 3 The organic layer was washed with water, drying over anhydrous Na 2
SO
4 and concentrated in vacua to give methyl 4-[5-(R)-phenyl-2-(.S)-pirrolidinyl methoxyjbenzate (363 mg, quant.) as yellowish oil. The product was used for next reactions without further purification. 'H-NlvR (CDC 3 8 1.71-1.83 (in, 2 2.03-2. 10 (mn, 1 2.15-2.24 (in, 1 3.68-3.74 (in, I 3.89 3 4.01-4.09 (in, 2 4.28 J= 7.2 Hz, I 6.95 J 8.8 Hz, 2 7.22-7.27 (in, 1 7.3 3 J 8. 0 Hz, 2 7.42 J 7.6 Hz, 2 8.00 J 8.8 Hz, 2 MS (ESI) 312 (MW+H) 353 (M*+CH 3
CN).
To a stirred solution of methyl 4-[5-(R)-phenyl-2..(S)-pirrolidinylinethoxy]benzoate (13 5 mng, 0.43 minol), 4-[N'-(2-inethylphenyl)ureido-3-methoxyphenylacetic acid (136 mng, 0.43 mmol) and NNdimethylaininopyridine (52.9 mg, 0.43 inmol) in DMF (10 ml) was added EDC-HCI (90.8 mng, 0.48 nunol) at Mi 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 NaSO,, then concentrated in vacua. The residue was chromatographed on silica gel with hexane- EtOAc (1 5) as eluent to give methyl 4-[1-[4-[N'-(2-inethylphenyl)ureidoj-3-methoxyphenyl acetylj-5-(R)-phenyl-2-(S)-pyrrolidinylmethoxyjbenzoate (271 mg, quant.) as a colorless amorphous solid. 3 H-NMR (CDCI,) 8 2.00-2.18 (in, 3 2.31-2.41 (in, 1 2.27 3 3.31 2 3.67 3 3.89 3 4.35-4.48 (in, 2 4.60 (brs, 1 4.92 J 6.8 Hz, 1 H), 6.51 J= 8.4 Hz, I 6.62 1 6.96 J= 8.8 Hz, 1 7.11-7.40 (series of m, 8 7.51 J 8.0 Hz, 1 7.97-8.00 (mn, 2 MS m/z, 608 To a stirred solution of methyl 4-[l -[4-[N'-{2-methylphenyl)ureidoJ-3-inethoxypheniylacetylj-5- (R)-phenyl-2-()-pyrrolidinylmethoxybenzoate (243 mng, 0.40 inmol) in MeOH-THF (1 1, 10 nil) was added L.OM-NaOH (2.4 ml, 2.40 nunol) at Mi and the resulting mixture was heated at 60 0
C
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 and dried over anhydrous Na 2 SOI, then concentrated in vacua. The residue was chroinatographed on silica gel with CHCI 3 -MeOH (10 1) to give 145 (224 ing, 94%) as a colorless amorphous solid. MW 593.67 'H-NMvR (CD 3 OD), mixture of rotamars, 2.00-2.19 (in, 3 2.28 and 2.30 total 3 2.45-2.49 (in, 1 3.37 (dd, J 39, 16 Hz, 2 H), 3.77 and 3.80 total 3 3.92-5.18 (series of in, 4 6.48-8.03 (series of in, 16 MS (FAB) 594 (MW+H).
WO 01/00206 WO 0100206PCT/USOO/18079 Example 138 4-fl -(2-chlorophenyl)ureido]-3 -methoxyphenylacetylj-5-(R)-phenyl-2-(S)-pyrrolidinyl methoxy]benzoic acid NYNI N o-Q-COOH IHH Me 146 To a stirred solution of methyl 4-[5-(R)-phenyl-2-(S)-pirrolidinylmethoxy]benzoate (142 mg, 0.46 nmmol), 4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetic acid (153 mg, 0.46 mmol) and NNdimethylaminopyridine (55.7 mg, 0.46 mmol) in DMIF (10 ml) was added EDC-HCI (95.7 mg, 0.50 mmol) at 11, and the resulting midxture was stirred overnight The reaction mixture was poured into water and extracted with EtOAc. The organic layer was washed with brine, drying over anhydrous NaSO,, 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-methoxyphenyI acetylj-5-(R)-phenyl-2-(S)-pyrrolidinylmethoxy]benzoate (260 mg, 90%) as a colorless amorphous solid. 'H-NMR (CDCI 3 8 2.00-2.19 (in, 3 2.35-2.44 (in, I 3.35 2 3.76 3 3.89 3 1H), 4.38-4.48 (mn, 2 4.63 (brs, 1 4.94 J 7.2 Hz, 1 6.53 J =8.4 Hz, 1 H), 6.66 1 6.96-7.01 (in, 3 7.12-7.42 (series of Mn 8 7.87 J 8.0 Hz, 1 7.98 J 8.8 Hz, 2 8. 17-8.19 (in, 1 M; MS m/lz, 627(M*), 628 To a stirred solution of methyl 4-fl -[4-IIN'-(2-chlorophenyl)ureido]-3-methoxyphenylacetylj-5-(R)phenyl-2-(S)-pyrrolidinylmethoxy]benzoate (251 mg, 0.40 mmol) in MeOH-THF (1 1, 10 ml) was added 1.OM-NaOH (2.4 ml, 2.40 innol) 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-HCI, then extracted with CHCI,. The organic layer was washed with brine and dried over anhydrous Na 2 SO,, then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (10 to give 146 (181 mng, 74%) as a colorless amorphous solid. MW 614.09 IH-NUvR (CD 3 OD), mixture of rotamars, 8 1.99-2.19 (in, 3 2.42-2.53 (mn, 1 3.38 (dd J 39, 15 Hz, 2 3.79 and 3.80 total 3 3.94-5. 19 (series of m, 4 6.49-8.05 (series of m, 16 MS (FAB) mz, 614 Anal. Calcd for C,,F1,ClN 3
O
6
-H
2 O: 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)ureidol -3-inethoxyphenylacetylJ-5-(R)-phenyl-2-(S)-pyrrolidinyI ethoxylbenzoic acid WO 01/00206 PCT/US00/18079 NH H XoOH 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-methoxyphenylaetic acid (178 mg, 0.47 mmol) and NNdimethylaminopyridine (57.4 mg, 0.47 mmol) in DMF (10 ml) was added EDCHCI (99.0 mg, 0.52 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 NaSO,, then concentrated in vacuo. The residue was chromatographed on silica gel with hexane- EtOAc (1 5) as cluent to give methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl acetyl-5-(R)-phenyl-2-(S)-pyrrolidinylmethoxylbenzoate (288 mg, 91%) as a colorless amorphous solid. 'H-NMR (CDCl 3 8 2.00-2.20 3 2.34-2.43 1 3.35 2 3.72 3 3.89 3 4.38-4.49 2 4.62 (brs, 1 4.94 J= 7.2 Hz, 1H), 6.54 J= 8.4 Hz, 1H), 6.67 1 6.91-7.05 4 7.28-7.42 (series of m, 7 7.51 J= 8.0 Hz, 2 7.87 J= 8.4 Hz, 1 7.99 J= 8.8 Hz, 2 8.14 J= 8.4 Hz, 2H); MS (ESI) m/z, 672 674 To a stirred solution of methyl 4-[1-[4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetylj-5-(R)phenyl-2-(S)-pyrrolidinylmethoxy]benzoate (270 mg, 0.40 mmol) in MeOH-THF (I 1, 10 ml) was added 1.0M-NaOH (2.0 ml, 2.0 mmol) at rt, and the resulting mixture was heated at 60 0 C with stirring for 1 h. The reaction mixture was poured into IN-HCI, then extracted with CHCI 3 The organic layer was washed with brine and dried over anhydrous Na 2 SO, then concentrated in vacuo. The residue was chromatographed on silica gel with CHCI 3 -MeOH (10 1) to givel47 (212 mg, 80%) as a colorless amorphous solid. MW 658.54 'H-NMR (CD 3 OD), mixture of rotamars, 8 1.99-2.19 3 2.42-2.53 1 3.38 (dd, J= 39, 16 Hz, 2H), 3.79 and 3.80 total 3 H), 3.94-5.19 (series of m, 4 6.49-8.00 (series of m, 16 MS (FAB) m/z, 658 660 Example 140 4-[I-[4-[N'-(2,6-dichlorophenyl)ureido]-3-methoxyphenylacetyl] -5-(R)-phenyl-2-(S)-pyrrolidinyl methoxy]benzoic acid INiJH H MeCOOH 148 To a stirred solution of methyl 4-[5-(R)-phenyl-2-(S)-pirrolidinylmethoxylbenzoate (109 mg, 0.35 WO 01/00206 WO 0100206PCT/USOOI 18079 mmol), 4-IN' -(2,4-dichlorophenyl)ureidoJ-3-methoxyphenylacetic acid (129 mg, 0.35 mniol) and N,N-dimethylaminopyridine (42.8 mg, 0.35 mmol) in DMF (10 mld) was added EDC-HC1 (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 anhydrous Na 2 SO4, then concentrated in vacuo. The residue was chromatographed on silica gel with hexane-EtOAc (1 4) as eluent to give methyl 4-[l-[4-IN'-(2,6-dichlorophenyl)ureidol-3methoxyphenylacetylJ-5-(R)-phenyl-2-(S)-pyrrolidinylmethoxybenzoate (208 mg, as a colorless amorphous solid. 'H-NMR (CDCI,) 582.00-2.21 (in, 3 2.33-2.39 (in, 1 3.31 2 3.69 3 3.88 3 4.34-4.45 (in, 2 4.59 (brs, 1 4.93 (t,J 6.8 Hz, 1 6.47 (di, J 8.0 Hz, 1 6.63 1 6.68 1 6.92-6.95 (in, 2 7.12-7.41 (series of m, 9 H), 7.96-8.0 1 (in, 4 1H); MS (FAB) m/z, 662 (MW+H).
To a stirred solution of methyl 1-[4-[N'-(2,6-dichlorophenyl)ureidoj-3-methoxyphenylacetyl]-5- (R)-phenyl-2-(S)-pyrrolidinylmethoxyjbenzoate (186 mg, 0.28 mmol) in MeOH-THF (I 1, 10 ml) was added 1.0M-NaOH (1.4 ml, 1.4 inmol) at Mi and the resulting mixture was heated at 60*C with stirring for 2.5 h. The reaction mixture was poured into 1N-HCI, then extracted with CHC 3 The organic layer was washed with brine and dried over anhydrous NaSO 4 then concentrated in vacuo. The residue was chromatographed on silica gel with CHCI 3 -MeOH (10 to give 148 (166 mg, 91%) as a colorless amorphous solid. MW 648.53 'H-NMR (CDC1 3 8 2.00-2.18 (in, 3 2.34-2.40 (in, 1 3.33 2 3.68 3 4.37-4.47 (in, 2 4.61 (brs, 1 4.94 J 6.8 Hz, 1 6.48 J 8.0 Hz, 1 6.63 1 6.96 J 8.4 Hz, 3 7.12-7.38 (series of m, 9 7.95 J =8.0OHz, 1 8.01 J =8.8 Hz, 2H); MS(FAB)m/zz, 648 Example 141 4-1-4-IN' -(2-broinophenyl)ureidoJ-3-nethylphenylacetyl-5-(R)-phenyl-2-(S)-pyrrolidinyI inethoxylbenzoic acid o-HQ 149 To a stirred solution of methyl 4-[5-(R)-phenyl-2-(S)-pirrolidinylmethoxyjbenzoate (125 ing, 0.40 minol), 4-[N'-(2-broinophenyl)ureidoj-3-inethylphenylacetic acid (146 mg, 0.40 mmol) and NNdimethylaminopyridine (49.0 mg, 0.40 nunol) in DMF (10 ml) was added EDC-HCI (84.1 ing, 0.44 minol) 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 anhydrous 310 WO 01/00206 WO 0100206PCTIUSOOIlSO79 Na 2
SO
4 then concentrated in vacua. The residue was chromatographed on silica gel with hexane- EtOAc (I1: 4) as eluent to give methyl 4-(1-[4-[N'-(2-bromophenyl)ureido]-3-methylphenylacetylJ- 5-(R)-phenyl-2-(S)-pyrrolidinylmethaxy]benzoate (238 mg, 90%) as a colorless amorphous solid.
'H-NMvR (CDCI 3 5 1.92 3 2.09-2.27 (in, 3 2.42-2.50 (in, 1 3.22-3.41 (mn, 2 3.88 3 4.39 J =4.4 Hz, 1 4.64 (brs, 1 5.00 J= 6.8 Hz, 1 6.72 (s,1I 6.81- 6.93 (series of m, 8 7.22-7.42 (series of mn, 6 8.01 J 8.4 Hz, 2 8.13 J 8.0 Hz, IH MIS (FAB) m/z, 656 658 To a stirred solution of methyl -(2-bromophenyl)ureido]-3-methylphenylacetyl]-5-(R)phenyl-2-(S)-pyi-rolidinylinethoxy~bnzoate (216 mg, 0.33 mmol) in MeOH-THF (I1: 1, 10 ml) was added 1 .OM-NaOH (1.7 ml, 1.7 inmol) at Mi and the resulting mixture was heated at 60 0 C with stirring for 2.5 h. The reaction mixture was poured into 1N-HCI, then extracted with CHC 3 The organic layer was washed with brine and dried over anhydrous NaSO,, then concentrated in vacua. The residue was chromatographed on silica gel with CHCI 3 -MeOH (10: 1) to give 149 (166 mg, 9 as a colorless amorphous solid. MW 642.54 IH-NMR (CDC 3 5 2.01 3 H), 2.05-2.25 (in, 3 2.43-2.48 (in, 1 3.34 (dd, J 45, 16 Hz, 2 4.38-4.45 (in, 2 4.66 (brs, 1 4.99 J 6.8 Hz, 1 6.77 1 6.82-6.88 (mn, 2 6.94 J 8.8 Hz, 2 H), 7.15-7.55 (series of m, 10 8.00 J =8.8 Hz, 2 8.14 J 7.2 Hz, 1 MS (FAB) m/~z 642 644 Example 142 441 -(2-methylphenyl)ureidoI-3-methoxyphenylaceyl]-5-(R)-methyl-2-(S)-pyrrolidinyI methoxyjbenzoic: acid SN YN O--aCOOH Me'H H Me 150 To a stirred solution of benzy N-Boc-pyrroglutarate (7.55 g, 23.6 inmol) in THF (100 ml) was added MeLi (1.1I M in Et 2 O, 28.4 ml, 32.4 nunol) at -78"C, and the resulting mixture was gradually warmed up to rt, then stirred overnight. aq.NHCI was added to the reaction m-ixture, THF was removed in vacua, then extracted with EtOAc. The organic layer was washed with water and, drying over anhydrous Na 2
SO
4 then concentrated in vacua. The residue was chroinatographed on silica gel with hexane-EtOAc (3 as eluent to give benzyl amino)-5-oxo-6-inethyl~pentanoate (5.02 g, as a colorless needles. mp 85-87 0(2; 'H-NMR (CDCl 3 8 1.43 9 1.61-2.15 (series of in, 3 2.09 311), 2.41-2.55 (in, 2 4.30 (brs, 1 4.70 J 5.6 Hz, I 5.12-5.21 (mn, 2H), 7.29-7.37 (in, 5 MS (ESI) m/z, 336 WO 01/00206 WO 0100206PCT/USOO/18079 To a stirred solution of benzyl [2-(S)-(N-Boc-amino)-5-oxo-6-methyllpentanoate (4.46 g, 13.3 mmol) in CHCI 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 vacua, and dissolved in toluene, then evaporated to give benzyl 5-methyl-5-pyrroline-2-(S)-carboxylate trifluoroacetic acid salt (5.74 g, quant.) as a crude brown oil. This compound (1.97 g, 5.94 mniol) in MeOH (30 mlJ) was added Pd/C 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 vacua to give 5-methyl-5-pyrrolidine-2- -carboxylic acid trifluoroacetic acid salt (956 mg, as a crude white solid. To a solution of this compound (939 mg, 3.86 minol) and di-terf-buyl dicarbonate in MeCN-water (15 16 ml) was added I OM-NaOH (8.49 mmol, 8.49 mld) at Mt and the resulting mixture was stirred for Ilh.
The resultiong mixture was evaporated and poured into aq.-1N-HCI, then extracted with
CHCI
3 /MeOH The organic layer was dried over anhydrous Na 2
SO
4 then concentrated in vacua. The residue was chromatographed on silica gel with CHCI 3 -MeOH (7 to give (R)-methyl-(S)-proline (7 11 mg, 80%) as a colorless oil. 'H-NMR (CD 3 OD) 8 1.27 (di, J 6.0 Hz, 3H), 1.41-1.46 (in, 9H), 1.62-1.64 (in, 1H), 1.96-2.01 (mn, 2H), 2.22 (brs, 1H), 3.94 (brs, IH), 4.17 (brs, lIH).; MS (ESI) m/z, 230 To a stirred solution of N-Boc-5-(R)-methyl-2-(S)-proline (1.03 g, 4.49 mniol) in THF ml) was added IlOM-BH 3 -Me 2 S (1.57 ml, 15.7 minol) at rt, and the resulting mixture was heated under reflux for 5 h. The mixture was poured into aq. IN-HCl and extracted with EtOAc. The organic layer was dried over anhydrous Na 2
SO
4 then concentrated in vacua. The residue was chromatographed on silica gel with hexane-AcOEt (I1: 3) as eluent to give N-Boc-2-(S)- (838 mng, 87%) as a colorless oil: 'H-NMR (CDCI 3 8 1.17 J= 6.0 Hz, 3H), 1.48 9H), 1.48-1.64 (mn, 2H), 1.90-2. 11 (mn, 2H), 3.52-3.57 (in, 11H), 3.68- 3.70 (in, 1H), 3.94-4.13 (in, 1H).
To a stirred solution of N-Boc-2-(S)-hydoxymethyl-5-(R)-methylpyrrolidine (820 mng, 3.81 mmnol), triphenylphosphine 10 g, 4.19 mniol) and methyl 4-hydroxybenzoate (580 mng, 3.81 mmnol) was added duisopropyl azodicarboxylate (841 mlt 4.19 minol) at Mt and the resulting mixture was stirred at 60 0 C for 1 h. The mixture was concentrated in vacua, and the residue was chromatographed on silica gel with hexane-EtOAc (5 as eluent to give methyl inethyl-2-(S)-pyrrolidinylinethoxylbenzoate (1.32 g, 80%) as a colorless oil. 'H-NMR (CDCI,) 8 1.24 (brs, 3 1.49 9 M, 1.55-1.70 (in, 2 1.94-2.11 (in, 2 3.88 3 3.88 (overlap, I1H), 4.064.20 (mn, 2H), 6.93 -6.96 (in, 2H), 7.97 (di, J =8.8 Hz, 2 MS (ESI) 3 50 (MN+H).
WO 01/00206 PTU0/87 PCT/USOO/18079 To a stirred solution of methyl 4-[N-Boc-5-(R)-methyl-2-(S)-pyrrolidinylmethoxyJ benzoate (1.29 g, 3.70 mmol) in CHCl, (30 ml) was added trifluoroacetic acid (10 ml) at it, and the resulting mixture was stirred for 35 min. 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 NaSO,, and concentrated in vacuo to give methyl 4-f 5-(R)-methyl-2-(S)-pirrolidiny methoxyjbenzate (871 mg, 95%) as a colorless oil. The product was used for next reactions without further purification. 'H-NMR (CDCI 3 8 1.18 J= 6.4 Hz, 3 1.30-1.40 (in, 1 H), 1.59-1.67 (in, 1 1.87-1.97 (ms, 2 3.19-3.27 1 3.49-3.55 (in, 1 3.87 3 3.89- 4.05 (in, 2 6.89 J= 8.8 Hz, 2 7.96 J= 8.8 Hz, 2 MS (ESI)m/tz, 250 To a stirred solution of methyl 4-[5-(R)-methyl-2-(S)-pirrolidinylmethoxy]benzoate (141 ing, 0.57 nunol), 4-[N'-(2-methylphenyl)ureidoj-3-methoxyphenylacetic acid (178 mng, 0.57 usmol) and NNdiinethylarninopyridine (69.0 mg, 0.57 mmol) in DMF (10 mld) was added EDC-HC1 (120 mng, 0.62 nunol) 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 NaSO 4 then concentrated in vacuo. The residue was chroinatographed on silica gel with EtOAc as eluent to give methyl 4-[1-[4-[N'-(2-methylphenyl)ureidoJ-3-methoxyphenyl methyl-2-(.S)-pyrrolidinylmethoxylbenzoate (297 mng, 96%) as a colorless amorphous solid. IH- NMR (CDCl 3 8 1.24-1.34 (mn, 3 1.93-2.18 (series of m, 4 2.28 3 3.65 3 3.88 3 3.62-3.87 (in, 3 4.1 1-4.38 (series of m, 3 6.42-8.06 (series of M 13 MS (ESI) m/z, 546 To a stirred solution of methyl 4-fl -(2-methylphenyl)ureido]-3-inethoxyphenylacetyll-5- (R)-methyl-2-(S)-pyrrolidinylmethoxyjbenzoate (279 mng, 0.51 mmol) in MeOH-THF (1 10 ml) was added 1.OM-NaOH (2.56 ml, 2.56 innol) at rt, and the resulting mixture was heated at 600(2 with stirring for 2 h. The reaction mixture was poured into lN-HCl, then extracted with CHC3.
The organic layer was washed with brine and dried over anhydrous Na 2
SO
4 then concentrated in vacua. The residue was chroinatographed on silica gel with CHC1 3 -MeOH (15 to give 150 (269 mng, 99%) as a colorless amorphous solid. MW 531.60 'H-NMR (CD 3 OD), mixture of rotarnars, 8 1.28-1.35 (in, 3 1.74-2.21 (series of ms, 4 2.28 3 3.71-4.37 (series of mn, 6 6.76- 7.99 (series of mn, 11I MS (ESI) m/z, 532 Example 143 4-f frans-4-amino-l1-[3-inethoxy-4-[N'-(2-inethylphenyl)ureidolphenylacetyl]-(2S)-pyrrolidinyl] methoxybenzoic acid WO 01/00206 WO 0100206PCT/USOO/18079 Me H H:Me 0 C OH 151 To a solution of methyl 4-Qtrans-4-amino-1-tert-butoxycarbonyl-(2S)-pyrrolidinyl) methoxybenzoate (1.0 g, 2.86 nimol) and TEA (1.2 mil, 8.6 mmol) in C-1 2 01 2 (20.0 mil) was added trifluoroacetic anhydride (720 mg, 3.43 nunol) at 0 0 C. After stirred for 2.5 hr at room temperature, water was added to the solution and extracted with CH 2
CI
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 as eluent to give methyl 4- (trans-lI -teri-butoxycarbonyl-4-triluoroacetamido-(2S)-pyrrolidinyl)methoxybenzoate (940 mg 74%) as a colorless oil. 'H-NMR (CDC1 3 8 1.46 9B), 2.02-2.18 (in, 2.4 1-2.52 (in, 11-1), 3.30-3.45 (im, 111), 3.80-3.90 111), 3.88 3H1), 4.00-4.30 (in, 3H), 4.65-4.75 (mn, 6.50 (br s, 1H), 6.91-6.94 (in, 2H1), 7.96-7.99 (in, 2H).
To a stirred solution of methyl 4-(trans- 1 -tert-butoxycarbonyl-4-trifluoroacetamido-(2S)pyrrolidinyl)methoxybenzoate (470 mg, 1.05 inmol) in C11 2 C1 2 (10.0 nml) was added TFA (5.0 ml) at 0 The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO, was added to the residue, and extracted with CH 2 C 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, 3-inethoxy-4-[N'-(2-inethylphenyl)urediolphenylacetic acid (314 mg, 1.0 nunol), HO~t (162 mg, 1.2 mxnol), and triethylamine (417 ml, 3.0 mxnol) in TIHF (10.0 ml) and MeCN (10.0 ml) was added EDC-HCI (288 mng, 1.5 inmol) at 0 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, 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, v/v) as eluent to give methyl 4-[trans- I1- [3-inethoxy-4-[N'-(2-methylphenyl)ureidolphenylacetyl]-4-trifluoroacetaniido-(2S)-pyrrolidinylI inethoxybenzoate (350 mng, as a colorless oil. 'H-NMR (CDCI 3 8 2.01-2. 10 (in, 111), 2.31 3H), 2.42-2.48 (in, 11M1, 3.45-3.50 (in, 3.56-3.59 (in, 3.89 3H1), 4.074.14 (in, 211), 4.38-4.42 (in, 111), 4.50-4.60 (in, 111), 4.72-4.80 (in, 11-1), 6.33 11-1), 6.60-6.85 (in, 3H), 7.06- 7.26 (in, 311), 7.48-7.52 (in, 1H1), 7.93-8.05 (in, 311).
WO 01/00206 WO 0100206PCT/USOO/18079 To a stirred solution of methyl 4-[trans-1-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetylj- 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 '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 151 (100 mg, 81%) as a white crystalline solid. MW 532.59 mp 170-171 1 1 (K.Br) 3264, 2937, 1604, 1535, 1415, 1376, 1255, 1224, 1033 cmn-'; 'H-NMvR (DMSO-dQ 8 1.80-1.90 (in, JH), 2. 2.20 (in, 114), 2.24 3H), 3.55-3.80 (in, 3H), 3.57 2H), 4.08-4.18 2H), 4.36-4.60 (mn, 1H), 6.72-7. 16 (mn, 7H), 7.77-8.01 (in, 4H), 8.46 lH), 8.54 1H); MS (FAB) m/lz 532 Anal.
calcd for C H 32
N
4 6 -2.0H 2 O0: C, 61.26; H, 6.38; N, 9.85. Found: C, 61.07; H, 6.32; N, 9.58.
Example 144 methyl 4-Iltrans-4-axnino-1 -[3-methoxy-4-[N -(2-methylphenyl)ureidolphenylacetylJ-(2S)pyrrolidinyljmethoxybenzoate HCI salt JPH2 Me H 6-J-COe 152 To a stirred solution of methyl 4-[trans-1-[3-medhoxy-4-[N'-(2-methylphenyl)ureidolphenylacetyll- 4-trifluoroacetanudo-(2S)-pyrrolidinylJmethoxybenzoate (200 mg, 0.31 nunol) in MeOH (4.0 minI) was added water (2.0 ml) and K 2 C0 3 (138 mg, 1.0 inml) at room temperature. After stirred for 1 8hr at room temperature, water was added to the mixture and extracted with CH 2 Cl 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
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 mnmol) was added thereto. The mixture was concentrated in vacua to give 152 (120 mng, 63%) as an amorphous solid. MW 546.61 JR (KBr) 3382, 2948, 2879, 1604, 1533, 1286, 1255, 771 'H-NMR (DMS0-dl 6 8 2.25 3H), 2.10-2.30 (in, 2H), 3.59-3.70 (in, 3H1), 3.77-3.80 (mn, 8H), 4.00-4.24 (in, 211), 4.47-4.67 (mn, 111), 6.70-7.16 (in, 714), 7.77-8.00 8.49 1H), 8.55 111); MS (FAB)m/~z 547 Anal. calcd for CvHNN 4
O
6 -HCl-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-inethoxy-4-[N'-(2-methylphenyl)ureidolphenylacetyll-4-methylanino-(2S)pyrrolidinyljmethoxybenzoic acid WO 01/00206 WO 0100206PCT/USOO/18079 H H Me O-a153 To a stirred solution of methyl 4-(Irans-1-ier-butoxycarbonyl-4-trifluoroacetamido-(2S)pyrrolidinyl)methoxybenzoate (520 mg, 1. 17 nimol) in DMF (10.0 mil) was added K 2 C0 3 (321 mg, 2.33 mmol) and Mel (330 mng, 2.33 nimol) at room temperature. The reaction mixture was stirred at 50*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, and concentrated in vacuo. The residue was purified by column chromatography on silica gel with EtOAc-n-hexane (1:2 as eluent to give methyl 4-[trans-1-tert-butoxycarbonyl-4-(N-methyl-trifluoroacetaxnido)-(2S)-pyrrolidiny1] methoxybenzoate (390 mng, 731/) as a colorless oil. 'H-NMvR (CDCI,) 8 1.46 2.12-2.40 (in, 2H), 2.96 and 3.05 (each s, total 3H), 3.28-3.70 (mn, 2H), 3.88 3H), 3.95-4.42 (in, 3H), 5. 5.40 (in, 1H), 6.89-6.91 (in, 2H), 7.96-8.00 (in, 214).
To a stirred solution of methyl 4-[trans-1 -:ert-butoxycarbonyl-4-(N-methyl-trifluoro acetoamido)-(2S)-pyrrolidinyljmethoxybenzoate (390 mig, 0.85 nunol) in CH 2
CI
2 (8.0 ml) was added TEA (5.0 mil) at 0 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 product, 3-methoxy-4-[N'-(2-methylphenyl)uredio]phenylacetic acid (279 mng, 0.89 inmol), HOI~t (143 ing, 1. 1 innol), and triethylanine (246 ml, 1.77 minol) in THF (8.0 nil) and MeCN (8.0 mil) was added EDC-HCI (255 mg, 1.3 inmol) at 0 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-hcxane (4:1 as eluent to give methyl 4-[b'ans-1- [3mtoy4[-2mtypey~rioperlaeyl4(-ehltilooctaio-2) pyrrolidinyljmethoxybenzoate (480 ing, as a colorless oil. 'H-NUvR (CDC1 3 8 2.18-2.35 (in, 214), 2.31 3H1), 2.87 and 2.97 (each s, total 3H4), 3.45-3.46 (in, 3H), 3.47 31H), 3.49 (s, 2H4), 3.88 3H1), 4.304.70 (in, 2H), 5.20-5.40 (in, IM), 6.38-6.43 (in, 111), 6.67-6.86 (in, 4H1), 7.09-7.24 (in, 4H1), 7.5 1-7.54 (in, 1H1), 7.93-8.08 (in, 3H1).
WO 01/00206 WO 0100206PCT11JSOO/18079 To a stirred solution of methyl 4-[trans- l-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetyll- 4-(N-methyl-N-trifluoroacetylamino)-(2)-pyrrolidiyllmethoxybelzoate (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 '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 vacua to give 153 (140 mg, 70%) as a white crystall1ine solid. MW546.61 mp 162-164 IR (KBr) 3338, 1604, 1535, 1255, 1033, 755 cm-1; 'H-NMR (DMSO-dj) 8 1.85-1.95 (in, IH), 2.10-2.20 (in, 1H), 2.24 3H), 2.34 and 2.39 (each s, total 314), 3.41-3.71 (in, 311), 3.58 2H1), 3.80 3H), 4.05-4.20 (in, 2H), 4.36-4.60 (in, 114), 6.73-7. 16 (in, 7H), 7.77-8.01 (in, 4H4), 8.45 1H4), 8.53 (s, 1 MS (FAB3) m/lz 547 Anal calcd for CH}1N 4
O
6 -2.5H 2 O: C, 60.90; H, 6.64; N, 9.47.
Found: C, 6 1.0 1; H, 6.50; N, 9.3 1.
Example 146 methyl 4-[trans-l1-[3-methoxy-4-[N '-(2-methylphenyl)ureidolphenylacetyll-4-methylamino-(2S)pyrrolidinyl~methoxybenzoate To 154 Toa stirred solution of methyl 4-[trans-l1..[3-.methoxy..4-[N'-(2-methylphenyl)ureidolphenylacetyll 4-(N-methyltrifluoroacetaniido)-(2S)-pyrrolidinyllmethoxybenzoate (240 mg, 0.36 inmol) in THF mmld) and MeOH (5.0 ml) was added water (2.0 ml) and K 2 C0 3 (138 mg, 1.0 minI) at room temperature. After stirred for 1 8hr at room temperature, water was added to the mixture and extracted with CH 2 C1 2 The extract was washed with water, then dried over Na 2
SO
4 and concentrated in vacua. The residue was purified by column chromatography on silica gel with MeOH-CH 2
CI
2 (5/95 to 20/80, v/v) as eluent. The product was dissolved in EtOH (5.0 ml), and IN Hdl (in EtOH) (0.71 ml, 0.71 minol) was added thereto. The mixture was concentrated in vacua to give 154 (180 mng, 85%) as an amorphous solid. NM 560.64 IR (KBr) 3311, 2692, 2453, 1712, 1604, 153 3 cm'; 'H-NM4R (DMSO-dQ 6 2.24 3M), 2.15-2.30 (in, 2H4), 2.60 (br s, 3H), 3.60-4.20 (in, 5H1), 3.78-3.81 (in, 811), 4.47-4.70 (in, 111), 6.71-7. 16 (in, 714), 7.77-8.00 (in, 411), 8.48 (s, 111), 8.55 111), 9.21 (br s, 2H1); MS (FAB) m/z 561 Anal. calcd for C 3 ,H36N 4
O
6
-HCI-
1.41-120: C, 59.83; H, 6.45; N, 9.00; Cl, 5.70. Found: C, 60.08; H, 6.5 1; N, 8.68; Cl, 5.99.
Examp~le 147 4-[Irans-4-diinethylanmino-1 -[3-methoxy-4-[N'.(2-methylphenyl)ureido]phenylacetyl]-(2S)pyrrolidinyllmethoxybenzoic acid WO 01/00206 PCT/US00/18079 H e 'X COOMe 155 To a stirred solution of trans-i -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-CI (3.03 g, 11.0 mmol) at 0 OC. The reaction mixture was stirred at room temperature for 18 hr.
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 as eluent to give trans-l-terlbutoxycarbonyl-(2S)-(tert-butyldiphenylsilyloxy)methyl-4-hydroxypyrrolidine (1.5 g, 33%) as a white crystalline solid. 'H-NMR (CDCI 3 8 1.03 (s9H), 1.25 and 1.32 (each s, 9H), 1.90-2.10 (m, 111), 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, (993mg, 3.0 mmol) at room temperature. The reaction mixture was stirred at 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 nhexane to n-hexane-EtOAc v/v) as eluent to give cis-4-bromo-l-tert-butoxycarbonyl-(2S)- (tert-butyldiphenylsilyloxy)methylpyrrolidine (1.0 g, quant.) as a pale yellow oil. 'H-NMR
(CDCI
3 8 1.06 9H), 1.31 and 1.45 (each s, 9H), 2.63 (min, 2H), 3.49 (min, 1H), 3.89-4.14 (min, SH), 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 mi) 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 SO4, and concentrated in vacuo. The crude product was used to the subsequent reaction without further purification. The solution of the crude residue in EtOH (10 mi) 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- 1 -tert-butoxycarbonyl-(2S)-(tertbutyldiphenylsilyloxy)methylpyrrolidine (400 mg, 95%) as a colorless oil. 'H-NMR (CDCl 3 8 WO 01/00206 WO 01/1)206PCT/USOO/18079 1.06 9H), 1.32 and 1.45 (each s, total 2.20-2.35 (in, IH), 3.05-3. 18 (mn, 3.55-4.05 (mn, 6H), 7.35-7.4 1 (in, 611), 7.6 1-7.69 (in, 4H1).
To a stirred solution of trans-4 -amino- I -terI-butoxycarbonyl-(2.S)-(tert-butyldiphenysilyloxy) methylpyrrolidine (400 mg, 0.88 mmol), AcOH (120 ml, 2.0 mmol), and 37%/ HCHO aq (500 ml) in MeOfI (10 ml) was added NaBH 3 CN (111 mg, 1.76 mmol) at 0 The reaction mixture was stirred at room temperature for 18 hr. After concentrated in vacuo, water was added and extracted with CH 2
CI
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
C
2 (3:97, v/v) as eluent to give trans-I -tert-butoxycarbonyl-(2S)-QIert-butyldiphenylsilyloxy)methyl-4-dimethylaniinopyrolidine (330 mg, 78%) as a pale yellow oil. 'H-NMR (CDC1 3 8 1.06 91H), 1.33 and 1.45 (each s, total 9H), 1.80-2.25 (mn, 211), 2.23 (br s, 61H), 2.95-4.05 (mn, 611), 7.36-7.39 (in, 6H), 7.63-7.65 (in, 4H).
To a stirred solution of trans- I -tert-butoxycarbonyl-(2S)-Qtert-butyldiphenylsilyloxy)inethyl-4dimethylaminopyrrolidine (330 mg, 0.68 minol) in THF (5 mil) was added TBAF (1.0 M solution in THF, 1.0 ml, 1.0 nunol) at 0 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 gel with MeOH-CH 2
CI
2 (3:97 to 20:80, v/v) as eluent to give trans-1I-tert-butoxycarbonyl-4dimethylan-ino-(2S)-hydroxyinethylpyrrolidine (180 mg, quant.) as a pale yellow oil. 'H-NMR
(CDCI
3 8 1.47 911), 2.23 1.65-1.75 (in,2H),2.75-4. 10 (in, 3.61 J 5.6 Hz, 2H).
To a stirred solution of trans- l-terI-butoxycarbonyl-4-dimnethyLamino-(2S)-hydroxymethy pyrrolidine (180 mg, 0.73 inmol), methyl 4-hydroxybenzoate (114 mng, 0.75 minol), and Ph 3 P (296 mng, 1. 13 nunol) in THE (10 ml) was added DIAD (227mg, 1. 13 iniol) at 0 The reaction mixture was stirred at 70 *C for 18 hr. The mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc v/v) to MeOH-
CH
2 Cl 2 (5:95, v/v) as eluent to give methyl 4-ftrans-l-tert-butoxycaxtonyl-4-dinethylamino-(2S)pyrrwlidinyl~inethoxybenzoate (180 ing, 68%) as a pale yellow oil. 'H-NMvR (CDCI,) 8 1.46 (s, 9H1), 1.80-1.95 (in, 2.20-2.23 (mn, 11-1), 2.24 2.90-2.95 (in, 3. 10-3.30 1141), 3.50-3.65 (in, 3.88 3.95-4.35 (mn, 311), 6.93-6.95 (in, 7.96-7.98 (mn, 211).
To a stirred solution of methyl 4-QIrans-l-ieri-butoxycarbonyl-4-dimethylaniuno-(2S)pyffolidinyl)inethoxybenzoate (200 mg, 0.53 innol) in CH 2 C1 2 (6 ml) was added ThA (3m1) at WO 01/00206 WO 0100206PCT/USOO/18079 0 0 C. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO, was added to the residue, and extracted with CH1 2 C1 2 The extract was washed with brine ,dried over Na 2 SO1 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-[M-(2-methylphenyl)urediolphenylacetic acid (166 mg, 0.53 mxnol), HOBt (71 mg, 0.53 mmol), and triethylanune (140 ml, 1. 10 mniol) in THF (5 mil) and MeCN (5 ml) was added EDC HCI (152mg, 0.79 rnxol) 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 to
CH
2 CI,-MeOH(8:92, v/v) as eluent to give methyl 4-[trans-4-diinethylamino- 1-[3 -inethoxy-4-[N'- (2-methyiphenyl) ureidolphenylacetyl]-(2S)-pyrrolidinyllinethoxybenzoate (260 mug, 86%) as a colorless oil. 'H-NMR (CDCl 3 5 1.95-2.15 (in, 3H), 2.23 611), 2.31 311, 3.30-3.34 (in, 111), 3.57 2H), 3.61 311), 3.70-3.75 (in, 1I1), 4.11-4.15 (in, 211), 4.45-4.50 (in, 111), 6.34 111), 6.72-6.88 (in, 411), 7.08-7.24 411), 7.5 1-7.53 (in, 1IM, 7.92-8.07 (mn, 311).
To a stirred solution of methyl 4-[trans-4dimethylamino--[3-methoxy-4-[N-(2methyiphenyl) ureidol phenylacetyl]-2-pyrrolidinyljmethoxybenzoate (260 ing, 0.45 minol) in T.HF ml) and MeOH (2.0 ml) was added IN NaOH (0.90 ml, 0.90 inmol). 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 155 (200 ing, 79%) as a white crystalline solid. MW 560.64 rup 145-150 IR (kBr) 3355, 2948, 1698, 1604, 1533, 1454, 1417, 1255, 1226, 1166, 1035, 755 cm-i; '11-NMvR (DMS0dl 8 1.82-1.98 (mn, 111), 2.08-2.11 (in, 111), 2.20 6H1), 2.25 311), 3.40-3.60 (mn, 3H1), 3.64 (s, 211), 3.82 311), 4.01-4.16 (in, 211), 4.36 (in, 111), 6.74-7.15 (mn, 7.77-8.02 (in, 411), 8.44 (s, 111), 8.54 111); MS (FAB)m/tz 561 AnaI. calcd for C 3
,H
36 N0 6 1I.2H 2 0: C, 63.95; H, 6.65; N, 9.62. Found: C, 63.82; H, 6.72; N, 9.44.
Exampnle 148 methyl 4-[:rans-4-dimethylanmino-1 -[3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetylJ pyrrolidinyllmethoxybenzoate HCI salt WO 01/00206 WO 0100206PCT/U SOO/18079 N N: Q O-Cj-COOMe Me H H6e To a stirred solution of trans-4-[4-dimethyLamino-1 -methoxy-4-[N'-(2-methylphenyl) ureido] phenylacetyl-(2S)-pyrrolidinyl]methoxybenzoic acid (80 mg, 0. 14 mmol) in toluene ml) and MeOH (1.0 ml) was added TMSCHN, (2.0 M in hexane, 100 ml, 0.20 mmol) at 0 The reaction mixture was stirred at room temperature for 1.5 hr. The mixture was concentrated in vacua. The residue was purified by column chromatography on silica gel with MeOH-
CH
2 C1 2 (5:95, v/v) as eluent. The product was dissolved in EtOH (5.0 ml), and IN HCl (in EtOH-) (244 L 1, 0.244mmo1) was added thereto. The mixture was concentrated in vacuo to give 156 (72 mg, 881/6) as an amorphous solid. MW 574.67 IR (KBr) 3345, 2950, 2586, 1712, 1604, 1511, 1454, 1284, 1255, 1170, 1114, 1029, 850, 771 cm- 1 'H-NMR (DMSO-dQ 8 2.25 3H), 2.35-2.37 (in, 2H), 2.77-2.81 (in, 6H), 3.62-3.71 (in, 2H), 3.79-3.81 (in, 8H), 3.99-4.16 (in, 3H), 4.50-4.70 (in, 1H), 6.74-7.16 (mn, 7H), 7.77-8.01 (in, 4H), 8.48 lH), 8.55 IH); Anal. calcd for CHNO,,d.0HCl 1.2 H 2 0: C, 60.74; H, 6.59; N, 8.85. Found: C, 61.03; H, 6.78; N, 8.33.
Examp~le 149 4-[cis-4-dimethylanuno- l-[3-inethoxy-4-[N'-(2-inethylphenyl)ureidojpherylacetyll-(2S)pyrrolidinyljinethoxybenzoic acid Mei H H Me 0 0 a157 To a stirred solution of cis-1I -tert-butoxycarbonyl-(2S)-Qtert-butyldiphenylsilyloxy)methyl- 4-hydroxypyrrolidine (1.82 mg, 4.0 iniol), phthalim-ide (647 mg, 4.4 inmol), and Ph 3 P (1.26 g, 4.8 inmol) in TIHF (20 ml) was added DIAD (889 mg, 4.4 inmol) at 0 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 (511, v/v) as eluent to give N-[cis-l -teri-butoxycarbonyl-(2S)-(tert-butyldiphenylsilyloxy)inethyl-4pyrrolidinyljphthalimide (1.6 g, as an amorphous solid. 'H-NN4R (CDCI 3 8 1.07 9H), 1.30 and 1.44 (each s, total 9H4), 2.27-2.37 (in, lH), 2.94-2.96 (in, lH), 3.81-4.09 (in, 5H), 4.72 (in, 1H), 7.37-7.38 (in, 6H), 7.67-7.74 (in, 614), 7.84-7.86 (in, 2H).
WO 01/00206 WO 0100206PCTIJSOOI 18079 To a stirred solution of N- [cis- 1-ieri-butoxycarbonyl-(2.s)-(Iert-butyldipheniylsilyloxy) methyl-4-pyrrolidinyllphthalimide (1.60 g, 2.74 mmol) in EtOH (8 mil) was added NI-1N1-H,H0(206 mg, 4.11 mmol) at room temperature. The reaction mixture was stirred at for 1 hr. The mixture was concentrated in vacuo. The resulting solid was filtered off, and washed with CHCl 3 The filtrate was concentrated in vacuo. The resulting solid was filtered off, and washed with CHC 3 The filtrate was concentrated in vacuo to give cjs-4-arnino- 1 -lertbutoxycarbonyl-(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 (CDC1 3 5 1.06 9H), 1.30 and 1.45 (each s, total 9H4), 1.59 (in, lH), 1.85 (in, lH), 2.94 (in, 1H), 3.44 3.78-4.07 (in, 4H), 7.36-7.41 (in, 6H), 7.51-7.65 (in, 4H-).
To a stirred solution of cis-4-amino- I -tert-butoxycarbonyl-(2S)-Qteri-butyldiphenyl ilyloxy) methylpyrrolidine (1.24 g, 2.74 nunol), AcOH (374 5.48 mmol), and 37% HCHO aq nil) in MeOH (20 ml) was added NaBH 3 CN (345 mg, 5.48 minol) 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
CI
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
CI
2 (3/97, v/v) as eluent to give cis- I -tert-butoxycarbonyl-(2S')-(tert-butyldiphenylsilyloxy)methyl-4-dimethylamino pyrrolidine (1.1I g, as a pale yellow oil. 'H-NMR (CDC1 3 8 1.05 9H), 1.29 and 1.45 (each s, total 1.95-2.04 (in, 1H), 2.20-2.26 (in, 2.27 6M-1, 2.54 (in, 111I), 3.00-3.02 (in, 11-), 3.62-4.03 (in, 4H), 7.34-7.4 1 (in, 7.63-7.65 (in, 4M).
To a stirred solution of cis-lI -tert-butoxycarbonyl-2-(tert-butyldiphenylsilyloxy)methyl-4-dimethyI amino pyrrolidine (1.1I g, 2.27 mxnol) in TIHF (10 mil) was added TBAF (1.0 M solution in THF) mil, 4.5 imol) 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 gel with MeOH-CH 2
C
2 (3/97 to 20/80, vfv) as eluent to give cis-1I-tert-butoxycarbonyl-4dimethylamino-(2S)-hydroxymethylpyrrolidine (580 ing, quant.) as a pale yellow oil. 'H-NMR
(CDCI
3 8 1.47 1.25-1.96 (in, 2H), 2.25 2.53-2.58 (in, 1H), 3.174.02 To a stirred solution of cis-1I-tert-butoxycarbonyl-4-diinethylainiino-(2S)-hydroxyinethylpyffolidine (555 mg, 2.27 iniol), methyl 4-hydroxybenzoate (380 mng, 2.5 inmol), and Ph 3 P (1.07 g, 4.09 minol) in THF (10 mil) was added DIAD (826 mg, 4.09 inmol) at 0 The reaction mixture was WO 01/00206 WO 0100206PCT/USOO/18079 stirred at 70 *C for 18 hr. The mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc v/v)MeOH-CHCI 2 (5/95, v/v) as eluent to give methyl 4-(cis-l1-Iert-butoxycarbonyl-4-dimethylamino-(2S)-pyrrolidinyl)methoxy benzoate (260 mg, as a pale yellow oil. 'H-NMR (CDCI,) 8 1.45 9H1), 1.70-1.90 (in, 1H), 2.26 2.33 (in, 11-1), 2.57 (in, 111), 3.06 (in, lIM, 3.85-4.23 (in, 4H), 3.88 3H), 6.93 (in, 211), 7.95 (in, 211).
To a stirred solution of methyl 4-(cis-1-tert-butoxycarbonyl-4-dmethylainino-(2S)pyrrolidinyl) methoxybenzoate (208 mg, 0.55 minol) in CH 2 Cl 2 (6 mil) was added TFA (3m1) at 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 C1 2 The extract was washed with brine ,dried over Na 2 SO1 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)urediolphenylacetic acid (173 mng, 0.55 nunol), HOBt (74 mng, 0.55 inmol), and triethylamine (15304, 1.1 inmol) in THF (6 ml) and MeCN (6 mil) was added EDC-HCI (160 mng, 0.83 mmxol) 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
C
2 MeOH (5/95, v/v) as eluent to give methyl 4-[cis-4-dmethylainino-l-[3-methoxy-4-[N-(2methylphenyl) ureidolphenylacetylj-(2S)-pyrrolidinyllinethoxybenzoate (270 ing, 47%) as a colorless oil. 'H-NMvR (CDCI 3 8 1.95-2.04 (in, 111), 2.25 611), 2.32 311, 2.61 (in, 111), 3.21 (in, 111), 3.56-3.58 (mn, 5H), 3.80-3.83 (in, 11-1), 3.88 311), 4.184.20 (in, 114), 4.41-4.45 (mn, 2H), 6.36 6.68-6.85 (in, 4H1), 7.08-7.25 (in, 411), 7.52-7.55 (in, 111), 7.9 1-8.07 (in, 311).
To a stirred solution of methyl 4-[cis-4-dimethylamino-l-[3-methoxy-4-[N-(2methylphenyl) ureidol phenylacetylJ-(2S)-pyrrolidinyl]inethoxybenzoate (270 ing, 0.47 iniol) in THF (4.0 ml) and MCOH (2.0 nil) was added IN NaOH (1.0 ml, 1.0 iniol). The mixture was stirred at 70 '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 157 (170 ing, 65%) as a white crystalline solid. MW 560.64 mp 147-150 IR (KBr) 3353, 2952, 1700, 1604, 1533, 144, 1415, 1255, 1166, 1035, 755 'H-NMR (DMSO-d 6 :1.83-1.84 (in, 111), 2.08-2. 10 (in, 11-1), 2.21 (hr s, 6H), 2.24 311), 3.00 (mn, 211), 3.60 211), 3.78 3M1, 3.85-4.29 (in, 411), 6.7 1-7.16 (in, 711), 7.77-8.01 (in, 41-1m), 8.46 111), 8.54 111); WO 01/00206 WO 0100206PCT/USOO/18079 MS (FAB) m/z 561 Anal. calcd for C 3 ,H6N 4
O
6 e2 H 2 0: C, 62.40; H, 6.76; N, 9.39. Found: C, 62.5 1; H, 6.60; N, 9.36.
Example 150 methyl 4-[cis-4-dimethylaniino-l1-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetyll-(2S)pyrrolidinyllmethoxybenzoate HCI salt Me\ Me e e HCI 158 To a stirred solution of 4-[cis-4-dimethylamnino-1 .{3-methoxy-4-[N'-(2-methylphenyl) ureido]phenylacetylJ-(2S)-pyrrolidiniyl~methoxybenzoic acid (80 mg, 0. 14 mmol) in toluene nml) and MeOH (1.0 ml) was added TMSCHN 2 (2.0 M in hexane) (100;11, 0.20 nunol) at 0 The 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
CI
2 (5/95, v/v) as eluent. The product was dissolved in EtOH (5.0 mil), and IN HCI (in EtOH) (244gl, 0.244mxnol) was added thereto. The mixture was concentrated in vacua to give 158 (75 mg, 79 as an amorphous solid. M 574.67 IR (KBr) 3345, 2950, 2456, 1712, 1646, 1604, 1511, 1454, 1434, 1415, 1284, 1257, 1168, 1114, 1031, 771 cm-1; 'H-NMR (DMSO-) 8 2.10-2.20 211), 2.25 3H), 2.83 (in, 6H), 3.60-3.62 (in, 2H), 3.76-3.81 (in, 811), 4.204.33 (in, 4H), 6.71-7. 17 (in, 611), 7.77-7.98 (in, 5M1, 8.47 111), 8.55 MS (FAB) m/z 574 Anal. calcd for C32H 38
N
4
O
6 0 1. 0 HCl 1. 3 H 2 0: C, 60.57; H, 6.6 1; N, 8.83. Found: C, 60.80; H, 6.82; N, 8.44.
Example 151 4-I[trans- 1 -[4-[N'-(2-orophenyl)ureido -3-methoxlphenylacetyl-4-dimethylamino-(2S)pyrrolidinylimethoxybenzoic acid Me\ Me
/N
MeHP Q 159 To a stirred solution of methyl 4-QIrans-1-Iert-butoxycabonyl4-dimethylaniino-(2S)pyrrolidinyl) methoxybenzoate (430 mg, 1. 1 mxnol) in CH 2 Cl 2 (10.0 ml) was added TFA (5.0 mld) at 0 0 C. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacua. Sat. NaH-C0 3 was added to the residue, and extracted with CH 2 C1 2 The extract was washed with brine, dried over Na 2 SO,, and concentrated in vacuo. The crude product WO 01/00206 WO 0100206PCT/USOO/18079 was used to the subsequent reaction without further purification. To a stirred solution of the crude product, 4-IN'-(2-chlorophenyl)uredioJ-3-methoxyphenylacetic acid (368 mg, 1. 1 mxnol), HO~t (162 mg, 1.2 nunol), and triethylainine (417 nil, 3.0 mniol) in THFf (10.0 ml) and MeCN (10.0 ml) was added EDC-HCl (288 mg, 1. 1 mmol) at 0 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, v/v) as eluent to give 4-[frans- 1-14-[N'- (2-chiorophenyl) ureidoJ-3-methoxylphenylacetyl-4-dimethylamino-2S)-pyrrolidiriyl~methoxy benzoate (530 mig, as a colorless oil. 'H-NMR (CDC1 3 8 1.94-1.99 (in, 111), 2.48 911), 3.06-3.12 (mn, 111), 3.33-3.38 (in, 114), 3.60 2H), 3.68 3H4), 3.69-3.80 (mn, 111), 3.88 3H), 4.134.20 (in, 2H), 4.56 (mn, 111), 6.76-7.00 (mn, 5H1), 7.22-7.34 (in, 3H1), 7.92-8.00 (in, 311), 8.17- 8.19 111). For HCl salt: IR (KBr) 3324, 2950, 2454, 1710, 1604, 1511, 1284 'H-NMR (DMSO-d) 82.30-2.40 (in, 211), 2.77-2.80 (in, 611), 3.60-3.75 (mn, 211), 3.75-3.85 (in, 811), 4.00- 4.22 (in, 311), 4.504.75 (in, 111), 6.75-7.43 (in, 711), 7.89-8.09 (in, 411), 8.87 1IM1, 8.91 111; MS (FAB) Wnz 595 Anal calcd for C,,H36N 4 0 6 C1- .0HCId.0OH 2 O: C, 57.23; H, 6.04; N, 8.61; Cl, 10.90. Found: C, 57.43; K1 6.08; N, 8.38; Cl, 10.73.
To a stirred solution of methyl 4-[trans-1 -14-[N'-(2-chlorophenryl)ureidoJ-3-inethoxyl phenylacetylj-4-diinethylanino-(2S)-pyrrolidinyllinethoxybenzoate (190 mg, 0.32 rumol) in THF (3.0 ml) and MeOH (2.0 nil) 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 HCI. The resulting solid was collected, washed with water, and dried in vacuo to give 159 (150 mng, 83%) as a white crystalline solid. MW 581.06 mp 159-161*C; IR (KBr) 3318, 2938, 1604, 1531, 1438, 1340 cm'; 'H-NMR (DMSO-d) 82.10-2.40(in,811), 2.50-2.70(in, 211), 3.85-3.90 (in, 511), 4.024.18 (in, 311), 4.304.60 (in, 111, 6.75-7.43 (in, 711), 7.86-8.09 (in, 41-1), 8.86 111), 8.91 111); MS (FAB) m/z 581 Anal. calcd for C 3 ,11,NO 6 CU- 1.21-120: C, 59.79; H, 5.92; N, 9.30. Found: C, 59.69; H, 5.93; N, 9.09.
Example 152 4-[cis-l1 [4-[N'-(2-chlorophenyl)ureidoI-3-inethoxylphenylacetylJ-4-dmethylaniino-(2S)pyrrolidinyl~methoxybenzoic acid WO 01/00206 WO 0100206PCT/USOOI 18079 H H Me 160 To a stirred solution of methyl 4-(cis- 1 -tert-butoxycarbonyl-4-dimethylamino-(2S)pymiolidinyl) methoxybenzoate (1.2 g, 3.2 mmol) in CH 2
CI
2 (10.0 ml) was added TFA (5.0 ml) at 0 The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacua. Sat. NaHCO, was added to the residue, and extracted with CH 2 C1 2 The extract was washed with brine ,dried over Na 2 SO, and concentrated in vacua. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product (278 mg, 1.0 mxnol), 4-[MP-(2-chlorophenyl)uredioJ-3-methoxyphenylacetic acid (335 mg, 1.0 mmol), HOBt (135 mg, 1.0 nunol), and triethylaxnine (417 ml, 3.0 mmol) in THFf (4.0 ml) and MeCN (4.0 ml) was added EDC-HCI (288 mg, 1.5 mmol) at 0 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 NaSO, and concentrated in vacua. The residue was purified by colum chromatography on silica gel with n-hexane-EtOAc( :1 Iv/v) as eluent to give methyl 4-Icis-l1 .4-[N'-(2-chlorophenyl)ureidoj-3-methoxylphenylacetylj-4-dimethylamino-(2S)pyrrolidinyll methoxybenzoate (500 mng, 84%) as a colorless oil. 'H-NMR (CDC1 3 8 1.98-2.50 (in, 111), 2.26 3H), 2.25-2.40 (in, 1H), 2.58-2.65 (in, lH), 3.20-3.30 (in, lH), 3.60 2H), 3.64 314), 3.80-3.90 (in, 1H), 3.88 3H), 4.18-4.20 (in, IM, 4.42-4.46 (in, 2H), 6.72-7.00 (in, 4H), 7.20-7.35 (im, 5H), 7.91-7.94 (in, 3H), 8. 18-8.21 (in, lH).
To a stirred solution of methyl 4-[cis-1-[4-[N'-(2-chlorophenyl)ureidoJ-3-methoxy phenylacetyl]-4-dimethylamino-(2S)-pyrrolidinyllmethoxybenzoate (250 mg, 0.42 mmnol) in THF ml) and MeOH (3.0 ml) was added IN NaOH (1.0 ml, 1.0 mmol). The mixture was stirred at 'C for 18 hr. The mixture was concentrated in vacua, water was added thereto, and neutralized with IN HCI. The resulting solid was collected, washed with water, and dried in vacuo to give 160 (170 mg, 70%) as a white crystalline solid. MW 581.06 mp 165-167 IR (KBr) 3328, 1604, 1531, 1164, 1033 'H-NMR (DMSQ-dj) 8 1.80-1.90 (in, 1H), 2.20-2.50 (in, 7H), 3.60-3.70 (in, 2H), 3.77-3.81 (in, 5H), 4.00-4.30 (in, 4H), 6.72-7.44 (m,714), 7.86-8.10 (in, 4H1), 8.88-8.92 MIS (FAB) ni/z 581 Anal. calcd for C, 0 1-1 3
N
4 0 6 C1.1H 2 0: C, 59.87; H, 6.06; N, 9.3 1. Found: C, 59.65; H, .5.76; N, 9.09.
WO 01/00206 WO 0100206PCTUSOOII 8079 4-[cis-l -[4-[N'-(2-clorophenyl)ureido-3-methoxylpheylacetyl-4-(2-naphthalelesulfonamido)- (2S)-pyrrolidinyllmethoxybenzoic acid U NO-CJ-COOH Me~j~q161 To a stirred solution of methyl 4-(cis-l-lerI-butoxycarbonyl-4-amino-(2S)-pyrrolidinyl)methoxy benzoate (200 mg, 0.57 mmol) and TEA (317 ml, 2.3 mmnol) in CHC1 3 (10.0 ml) was added (2naphthyl)sulfonyl chloride (155 mg, 0.68 mxnol) at 0 0 C. The reaction mixture was stirred at room temperature for 1 8hr. The reaction mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc 1, v/v) as eluent to give methyl 4.-(cis-l1-tert-butoxycarbonyl-4-(2-naphthylsulfonamido)-(2S)-pyrrolidinyl) methoxybenzoate (240 mg, as apale yellow oil. 'H-NMIR (CDCI 3 8 1.25-1.45 (br s, 911), 1.70-1.80 (in, 11H), 2.20- 2.40 (in, I1H), 3.20-3.50 (in, 211), 3.90 3H1), 3.85-4.15 (in, 3H1), 4.55-4.65 (in, 111), 6.90-7. 10 (in, 211), 7.58-8.04 (in, 8H), 8.43(s, 111).
To a stirred solution of methyl 4-(cis- 1 -tert-butoxycarbonyl-4-(2-naphthylsulfonamido)- (2S)-pyrrolidinyl) methoxybenzoate (240 mg, 0.44 minol) in CH 2 Cl 2 (5.0 ml) was added TEA nil) at 0 The reaction mixture was stiffed at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaH-C0 3 was added to the residue, and extracted with CH 2
CI
2 The extract was washed with brine, dried over NaSO 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-chlorophenyl)uredio-3-methoxyphenylacetic acid (147 mg, 0.44 inmol), HOBt (59 mg, 0.44 inmol), and triethylamine (275 ml, 1.9 nunol) in THF (6.0 ml) and MeCN (6.0 ml) was added EDC-HCI (127 mg, 0.66 mmol) at 0 IC. 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 v/v) as eluent to give methyl 4-[cis-1 -14- [N'-(2-chlorophenyl)ureido]-3-methoxypheylacetyl-4-(2-naphthaleneuffonamfido)-(2S)pyrrolidinyllmethoxybenzoate (200 mg, as a colorless oil. 'H-NMR (CDCI 3 8 1.75-1.80 (in, 1IM, 2.25-2.40 (in, 111), 3.43 3.40-3.50 (in, 111), 3.60 3H1), 3.65-3.75 (in, 111), 3.90 (s, WO 01/00206 WO 011(1206PCT/USOO/18079 3H), 3.85-3.92 (mn, 1H1), 3.95-4.00 (in, 11), 4.30-4.40 4.65-4.75 (Mn 11), 6.26 9.3 Hz, 11-1), 6.50 J= 8.3 Hz, 6.23 IH), 6.86 J= 8.8 Hz, 211), 6.75-7.01 (mn, 111), 7.23- 7.36 (mn, 3H), 7.61-7.96 (in, 911), 8.20 J= 8.1 Hz, 11H), 8.43 111).
To a stirred solution of methyl methyl 4-[cis-l-[4-[N'-(2-chlorophenyl)ureidoj-3-methoxylpheny acetyl]j(2-naphthalenesulfonamide)-(2S)-pyrrolidinyl]nethoxybenzoate (200 mng, 0.26 inmol) in THF (6.0 nil) and MeOH (3.0 ml) was added IN NaOH (0.5 nil, 0.5 minol). The mixture was stirred at 70 '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 161 (2 10 mng, quant) as a white crystalline solid. MW 743.22 mp 135-142 IR (KBr) 3332, 1685, 1604, 1531, 1421,.1159 'H-NMR (DMSO-d,) 8 1.75-1.85 (mn, lH), 2.05-2.15 (in, 111), 3.05-3.15 (in, 111), 3.47 2H), 3.60-3.80 (mn, 2H), 3.73 3H), 4.05-4.20 (in, 311, 6.51 J 8.5 Hz, 111), 6.74-7.04 (in, 511), 7.27-7.31 (mn, 111), 7.43-7.45 (in, 211), 7.66-8.17 (mi, 9H), 8.46 111), 8.91 J= 9.5 Hz, I1H); MS (FAB)m/z743 (Mr+ AnaI. calcd for C 3 sH3 5
N
4 0 8 C1& 0. 5120: C, 60.67; H, 4.82; N, 7.45; Cl, 4.26. Found: C, 60.77; H, 4.84; N, 7.2 1; Cl, 4.90.
Example 154 441 -[4-[N'-(2-bronopheny1)ureido-3-nethoxypheniylacetyl]-4-(2-mesitylenesulfonanido-(2S)pyrrolidinyl]miethoxybenzoic acid N YN'n N COCH r H 6Me162 To a stirred solution of methyl 4-(cis- 1 -er:-butoxycarbonyl-4-axnino-(2S)-pyrrolidinyl) inethoxy benzoate (180 ing, 0.51 mmol) and TEA (283 nil, 2.0 inmol) in CHC1 3 (10.0 ml) was added (2-mesitylene)sulfonyl chloride (122 ing, 0.56 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 1, v/v) as eluent to give methyl 4-(cis-lI-tert-butoxycarbonyl-4-(2-inesitylenesulfonamido-(2S)-pyrrolidinyl) inethoxy benzoate (170 ing, as a pale yellow oil. 'H-NMR (CDCI 3 8 1.40 9H1), 1.85-1.95 (mn, 1H1), 2.29 311), 2.35-2.45 (mn, 111, 2.62 611), 3.804.15 (mn, 3.89 311), 3.50-3.65 (in, 111), 6.94 211), 6.94-7.00 (in, 211), 7.99 J =8.8 Hz, 211).
To a stirred solution of methyl (cis- 1 -tert-butoxycarbonyl-4-(2-inesitylenesulfonanudo)-(2S)- WO 01/00206 WO 0100206PCT/USOO/18079 pyrrolidinyl)methoxybenzoate (170 mg, 0.32 mmol) in CHCI 2 (5.0 mil) was added TFA (5.0 nil) at 0 0 C. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacua. Sat. NaHCO 3 was added to the residue, and extracted'Awith CH 2 C1. The extract was washed with brine ,dried over Na 2
SO
4 and concentrated in vacua. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product, 4-[M-(2-bromopheniyl)uredio]-3-methoxyphenylacetic acid (121 mg, 0.32 mxnol), HOBt (43 mg, 0.32 mxnol), and triethylaruine (139 ml, 1.0 mxnol) in THE (5.0 mil) and MeCN nil) was added EDC-HC1 (91 mg, 0.48 mmol) at 0 The reaction mixture was stirred at roam 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 ,vtv) as eluent to give methyl (2-bromophenyl)ureido] -3 .methoxyphenylacetylj-4-(2-mesitylenesulfonamido)-(2S)-pyrrolidinyl methoxybenzoate (210 mg, as a colorless oil. 'H-NMR (CDCI,) 6 1.85-1.90 (in, 1H), 1.95- 2.05 (in, IH), 2.32 311), 2.60 6H), 3.40-3.50 (in, 3H1), 3.60-3.70 (Cm, 211), 3.68 314), 3.89 3H), 3.96-3.99 (in, 111), 3.35-3.45 (in, 111), 3.70-3.75 (in, 111), 6.00 J 9.5 Hz, 111), 6.57- 7.08 (in, 9H1), 7.29 -7.34 (in, 7.51-7.53 (in, 111), 7.92-7.96 (Cm, 8.15 J= 6.8 Hz, 111).
To a stirred solution of methyl 4-[1-[3-inethoxy-4-[N'-(2-broinophenyl)ureido]phenylacetyl]-4-(2mesitylenesulfonanudo-(2S)-pyrrolidiniyljmethoxybenzoate (210 mg, 0.26 iniol) in T1HF (5.0 mil) and MeOH (3.0 mil) was added IN NaOH (0.47 nil, 0.47 mmol). The mixture was stirred at 70 0
C
for 24 hr. The mixture was concentrated in vacua, water was added thereto, and neutralized with IN HCL. The resulting solid was collected, washed with water, and dried in vacua to give 162 (180 mng, as a white crystalline solid. MW 779.70 mpl130-132* 0 C; IR(KBr) 3332, 1689, 1604, 1529, 1155 cm'; 'H-NMR (DMSO-d 6 8 1.70-1.85 (in, 111), 2.02-2.12 (in, 111), 2.25 3H-), 2.52 6H), 3.05-3.12 (mn, lH), 3.48 211), 3.60-3.70 (in, 2H), 3.77 311), 3.90-4.20 (mn, 3H), 6.59 J= 8.3 Hz, 1H), 6.77-6.80 (in, 11H), 6.95-7.01 Cmn, 4H), 7.3 1-7.35 (in, 111), 7.60 J= 8.1 Hz, 1H), 7.86-7.97 (in, 5H), 8.72-8.76 (in, 1H), 8.89-8.93 (in, 1H); MS (FAB) m/z 779 781 Anal. calcd for C 37 H.,NOgSBr-0.5H 2 O: C, 56.35; HK 5.11; N, 7. 10; Br, 10. 13. Found: C, 56.39; H, 5.07; N, 6.89; Br, 10.25.
Example 155 1 [P(-rmphnluedl3mehxpeyaeyl4dnyain-(S-yrldnl methoxybenzoic acid WO 01/00206 WO 0100206PCTUSOOI 18079 0 Y2 0 r M 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 mxnol) at 0 0 C. The reaction mixture was stirred at room temperature for 18hr. The reaction mixture was concentrated in vacua. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc 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. 'H-NVR (CDCI 3 8 1.34 1.50-1.60 (in, IH), 2.15-2.25 (in, IM), 2.87 611), 3.15-3.22 (in, 111), 3.35-3.45 (in, lH), 3.48-3.52 (in, 1H), 3.80-4. 10 (in, 3.91 7.01- 7.25 (in, 7.50-7.53 (mn, 8.03 J 8.7 Hz, 2H), 8.16 J 8.5 Hz, 8.28 J 7.1 Hz, 111), 8.53 J =8.5 Hz, lH).
To a stirred solution of methyl 4-(cis- 1 -tert-butoxycarbonyl-4-dansylamino-(2S)-pyrrolidinyl) methoxybenzoate (200 mg, 0.34 mxnol) in CH 2
CI
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 vacua. 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 vacua. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product, 4- (N'-(2-bromophenyl)uredioj-3-methoxyphenylacetic acid (129 mg, 0.34 minol), HOBt (46 mg, 0.34 mmol), and triethylainine (142 ml, 1.0 inmol) in THF (5.0 ml) and MeCN (5.0 ml) was added EDC-HCI (98 mng, 0.51 mmol) at 0 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 vacua. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc(l :4 as eluent to give methyl 4-[l-[4-1V'-(2-bromophenyl) ureidoj-3methoxyphenylacetylj-4-dansylanmino-(2S)-pyrolidinyllmethoxybenzoate (250 mg, 88%) as a colorless oil. 'H-NMR (CDC1 3 8 1.55-1.65 (mn, 111), 2.15-2.25 (in, 1H), 2.87 611), 3.20-3.35 (in, 3H), 3.50-3.55 (in, IH), 3.67 3H), 3.78-3.81 (mt, 1H), 3.88-3.93 (in, 3.91 3H1), 4.28- 4.31 (mt, 11-1), 4.65-4.70 (in, 111), 6.35 J= 9.5 Hz, 111), 6.54 J= 8.5 Hz, 111), 6.63 1H1), 6.90-7.13 (in, 6H1), 7.22-7.3 1 (mt, 2H1), 7.50-7.56 (in, 211, 7.88 J =8.0 Hz, 11H), 7.99 J= WO 01/00206 WO 0100206PCT/USOOI 18079 Hz, 1H), 8.13-8.16 (in, 2H), 8.27 J= 7.6 Hz, 111), 8.56 J= 8.3 Hz, 114).
To a stirred solution of methyl 4-[1-[4-IN'-(2-bromophenyl)ureido]-3-methoxyphenylacetyll-4dansylamino-2S-pyrrolidinyl~methoxybenzoate (250 mg, 0.29mmol) in THF (5.0 mld) and MeOll ml) was added IN NaOH (0.52 ml, 0.52 nunol). The mixture was stirred at 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 163 (230 mg, 94%) as a green crystalline solid. MW 830.74 mp 138-141 IR (KBr) 3340, 2940, 1604, 1527, 1421, 1162, 1145 'H-NMR (DMSO-d 6 )8 1.70-1.80 1H), 1.98-2.06 IlH), 2.81 611), 3.00-3. 10 (in, I1H), 3.39-3.40 (in, 2H), 3.50-3.80 (in, 3H), 3.76 311), 3.90-4.15 (mn, 2H), 6.52 J 9.0 Hz, 1H), 6.73-7.00 (in, 4H), 7.22-7.35 (mn, 2H), 7.55-7.64 (in, 3H), 7.83-8.48 (mn, 8H), 8.71-8.76 (in, 1H), 8.88-8.92 (in, 111); MIS (FAB) m/z 830 832 Anal calcd for
C,
4
HN
5 OBrS-0.7H 2 O: 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 4-14-methanesulfona'nido-1 '-(2-broinophenyl)ureidoJ-3-methoxyphenylacetyl-(2S)pyrrolidinyllinethoxybenzoic acid -Me r NY j-aCOOH H H O~e164 To a stirred solution of methyl 4-(cis-4-amino- 1 -ert-butoxycarbonyl-(2.S)-pyrrolidinyl)methoxy benzoate (180 mng, 0.51 mxnol) and TEA (283 ml, 2.0 mmol) in CHC1 3 (10.0 ml) was added inethanesulfonyl chloride (88 mg, 0.77 mxnol) 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 1, vA') as eluent to give methyl 4-(cis-1 -tert-butoxycarbonyl-4-inethanesulfonamido-(2S)-pyrrolidiny1) inethoxybenzoate (150 ing, as a pale yellow oil. 'H-NMR (CDC1 3 8 1.45 9H), 1.98-2.08 (in, 111), 2.52-2.65 (in, 1H), 2.99 3H), 3.40-3.50 (in, 1H), 3.55-3.80 (mn, 1H), 3.89 1H), 4.00-4.70 (in, 4H), 6.98-7.00 (in, 2H), 8.00 J= 8.8 Hz, 214).
To a stirred solution of methyl 4-(cis-1-terl-butoxycarbonyl-4-methanesulfonamido-(2S)pyrrolidinyl) methoxybenzoate (150 ing, 0.43 minol) in CH 2 Cl 2 (5.0 ml) was added TEA (5.0 ml) at 0 OC. The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was WO 01/00206 WO 0100206PCT/USOO/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 SO1 and concentrated in vacua. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product, 4-[M-(2-bromophenyl)uredio]-3-methoxyphenylacetic acid (163 mg, 0.42 mmol), HOBt (58 mg, 0.43 mmol), and triethylamine (179 ml, 1.3 nimol) in TI-F (5.0 ml) and MeCN nml) was added EDC-HCI (144 mg, 0. 75 mmol) at 0 The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacua. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO 3 2-M citric acid, and sat NaH{C0 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 v/v) to EtOH-EtOAc (I10/o, v/v) as eluent to give methyl 4-[4-methanesulfonamido- 1-[4-[N'-(2-bromophenyl)ureidoJ-3-methoxyphenyl acetyl]-(2.S)-pyrrolidinyl]methoxybenzoate (210 mg, as a colorless oil. 'H-NMR (CDC 3 1.95-2.00 (in, 111), 2.55-2.65 (in, 2.17 311), 3.55-3.70 (in, 111), 3.60 2H1), 3.67 311), 3.85-3.90 (in, 111), 3.89 311), 3.95-4.18 (in, 211), 4.45-4.55 (in, 111), 4.704.80 (in, 111), 5.87 (d, J 9.3 Hz, 11-1), 6.73-6.95 (in, 511), 7.09 211), 7.28-7.33 (in, 11-1), 7.51 J= 8.0 Hz, 111), 7.93 -7.97 (in, 311), 8.13 J 8.3 Hz, 111).
To a stirred solution of methyl 4-[4-methanesulfonamido-I-[4-[N-(2-broinophenyl) ureidoj-3-inethoxyphenylacetyl]-(2S)-pyrrolidiniylJmethoxybenzoate (210 mng, 0.3 inmol) in TI-F ml) and MeOH (3.0 ml) was added IN NaOH (0.8 ml, 0.8 mmol). The mixture was stirred at 70 'C for 24 hr. The mixture was concentrated in vacua, water was added thereto, and neutralized with IN HCl. The resuilting solid was collected, washed with water, and dried in vacuo to give 164 (170 ing, 83%) as a white crystalline solid. MW 675.55 mp 125-128 IR (KBr) 3353, 1689, 1604, 1529, 1419, 1155 'H-NM4R (DMSO-d) 8 1.88-2.00 (in, 111), 2.3 5-2.45 (mn, 111), 2.96 (mn, 311), 3.15-3.23 (in, 111), 3.60 211), 3.50-3.70 (in, 111), 3.78 311), 3.80-3.90 (mn, 111), 3.95- 4.05 (in, 111), 4.10-4.30 (in, 211), 6.71-7.03 (in, 411), 7.32 (in, 111), 7.45 J =6.8 Hz, 11-1), 7.60 J 7.8 Hz, 111), 7.87-7.95 (in, 411), 8.74 11-1), 8.92 111); MS (FAB) nl/z 675 677 AnaL. calcd for CH) 3
,N
4 OBrS-0.6HO: C, 50.75; K1,4.73; N, 8.16. Found: C, 51.04; H1, 4.62; N, 7.79.
Examp~le 157 4-1-3 -methoxy-4-[N '-(2-inethylphenyl)ureido]phenylacetyl-(2S)-octahydroindolylmethoxyI benzoic acid WO 01/00206 WO 0100206PCT/USOO/18079 Me H~ H Me-oo 165 To a stirred solution of octahydroindole-(2S)-carboxylic acid (3.00 g, 17.7 mmol) in dioxane ml) was added 1 N NaOH (45 ml) and the solution was stirred at 0 0 C. To the mixture was added (Boc0 2 0 (4.26 g, 19.5 mmol) in dioxane (25 ml) at 0 0 C and the reaction mixture was stiffed at 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 NaSO 4 and evaporated to give l-(tert-butoxy carbonyl)octahydroindole-(2S)-carboxylic acid (4.78 g, as a colorless solid. mp 130-132 *C; 'H-NMR (CDCI 3 8 1.10-1.46 (series of s and mn, total 14 1.65-1.76 (in, 3 1.90-2.18 (in, 2 2.26-2.35 (in, 1 3.77-3.86 (in, 1 4.22-4.34 (in, 1 MS (ESI) m/lz 270 To a cooled (0 0 stirred solution of l-Qtert-butoxycarbonyl)octahydroindole-(2S)carboxylic acid (1.00 g, 3.71 minol) in THF (10 ml) was added BH 3 DMS (530 ml, 5.59 mmnol) 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 CHCI 3 -MeOH (50: 1, v/v) as eluent to give 1 -QIert-butoxycarbonyl)octahydroindole-(2S)-inethano (940 ing, 99%/) as a colorless oil. 'H-NMR (CDCI 3 8 1.05-1.30 (in, 4 1.47 9 1.49-1.74 (in, 4 1.82- 1.93 (in, 3 2.19-2.26 (in, 1 3.56-3.61 (in, 1 3.70-3.75 (in, 2 3.94-3.96 (in, 1 MIS (FAR) m/z 256 (Mr+ 1).
To a cooled stirred solution of methyl 4-hydroxybenzoate (560 ing, 3.68 inol), I- (tert-butoxycarbonyl)octahydmindole-(2S)-inethano (940 mg, 3.68 nol) and Ph 3 P 16 g, 4.42 innol) in THF (20 ml) was added DIAD (870 ml, 4.42 mxnol) 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 1, v/v) as eluent to give methyl 4-[l-Qert-butoxycarbonyl)-(2S)-octahydroindolylinethoxy~benzoate (1.16 g, as a colorless oil. 'H-NMR (CDCI,) 8 1. 14-1.47 (series of s and mn, total 13 1.60-2.13 (series of mn, total 6 2.22-2.28 (in, 1 3.75-3.91 (series of s and in, total 4 4.064.18 (in, 2 4.37 (in, 1 6.94-6.96 (in, 2 7.96-7.98 (in, 2 MIS (FAR) m/z 3 90 1).
To a stirred solution of methyl 4-[1-Qtert-butoxycarbonyl)-(2S)-octahydroindolylnethoxyJ WO 01/00206 WO 0100206PCT/USOOI 18079 benzoate 16 g, 2.98 mmol) in CH 2 C1 2 (10 nil) was added TEA (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,, and extracted with CHCI 3 The extract was washed with brine, dried over K 2 C0 3 and evaporated to give methyl 4-[(2.S)-octahydioindolylmethoxylbenzoate (860 mg, as a brown oil. 'H-NMR (CDCI 3 8 1.23-1.78 (series of m, total 10 2.00-2.09 2 3.14-3.18 (in, 1 3.55-3.62 (in, 1 3.88 3 3.96-4.06 (in, 2 6.92 J 9. 1 Hz, 2 7.97 J1 =9.1 Hz, 2 MIS (FAR) m/z 290 A mixture of 3-methoxy-4-[N '-(2-methylphenyl)ureidolphenylacetic acid (298 mg, 0.95 mmol), methyl 4-[(2S)-octahydroindolylmethoxyjbenzoate (274 mg, 0.95 nunol), EDC-HC1 (218 mng, 1. 14 mmnol), HO~t (154 mg, 1. 14 inmol), Et 3 N (160 ml, 1. 15 mmol) in THE (7 ml) was stiffed at roam 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 50: 1, v/v) as eluent to give methyl 4-[I-[3-methoxy-4-[AN'-(2-methylphenyl)ureido]phenylacetyl]-(2S)-octahydroindoly methoxylbenzoate (532 mng, as a white foamn. 'H-NMR (CDCI 3 8 1.13-2.05 (series of m, total 9 2.14-2.24 (mn, 2 2.26 3 3.60 2 3.62 3 3.80-3.85 (in, 1 3.88 3 4.27-4.37 (in, 3 6.62 1 6.74-6.76 (in, 2 6.91 (d,J 8.8 Hz, 2 7.09-7.13 (in, 1 7.20-7.24 (mn, 3 7.55 J 7.8 Hz, 1 7.94 J 8.8 Hz, 2 8.04 J 7.8 Hz, 1 MS (FAR) m/z 586 1).
To a stirred solution of methyl 4-[I-[3-methoxy-4-[N'-(2-inethylpheny1)ureidolpheny acetylj-(2S)-octahydroindolylmethoxylbenzoate (532 mg, 0.91 mmol) in THF (5 ml) was added N NaOH (5 ml) and the reaction mixture was heated under reflux for 3 hr. After cooled to roam temperature, the mixture was poured into ice-l N HC1, and the resulting precipitate was collected.
The crude solid was recrystallized from MeOH-CHCI 3 -[PE to give 165 (278 mng, as a white crystalline powder. MW 571.66 mp 130-134'C; 'H-NMR (DMS0-cL) 8 1.16-2. 10 (series of m, total 9 2.15-2.30 (series of s and M total 4 3.55-3.79 (in, 3 3.81 3 3.90-3.95 (in, 1 4.17-4.23 (in, 2 4.34-4.36 (in, 1 6.72-6.74 (in, 1 6.87-6.88 (in, 1 6.91-6.95 (in, 1 7.03 J =8.8 Hz, 2 7.10-7.16 (in, 2 7.78-7.80 (in, 1 7.87 J 8.8 Hz, 2 7.98-8. 00 (in, 1 8.45 1 8.54 1 12.61 (br s, 1 MIS (FAR) m/z 572 (MW+ 1); Anal Calcd for C 33
H
3
N
3 0 6 l1/4H 2 0: C, 68.79; H, 6.56; N, 7.29. Found: C, 68.70; H, 6.82; N, 6.97.
Example 158 1-[4-IN '-(2-chlorophenyl)ureidoJ-3-methoxyphenylacetyl]-(2S)-octahydroindolylnethoxy WO 01/00206 WO 0100206PCT/USOO/18079 benzoic acid A mixture of 4-IN '-{2.chlorophenyl)ureido]-3-methoxyphenylacetic acid (307 m,0.92 mmol), methyl 4- [(2S)-octahydroindolylmethoxy]benzoate (265 mg, 0. 92 minol), EDC HCI (211 mg, 1. inmol), HOJ~t (148 mg, 1. 10 ramol), and Et 3 N (153 ml, 1. 10 mmol) in THE (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,, and evaporated. The residue was purified by column chromatography on silica-gel with CHCI 3 -MeOH (100:11to 50: 1, vfv) as eluent to give methyl 441 '-(2-chlorophenyl)ureidoj-3-methoxypheniylacetylJ -(2S)-octahydroindolyl inethoxy~benzoate (550 mg, 99%) as a white foam. 'H-NMR (CDC1 3 8 1.15-2.02 (series of in, total 9 2. 17-2.33 2 3.58 3 3.62 2 3.84-3.90 (series of s and mn, total 4 H), 4.064.40 (Cm, 3 6.71-6.74 (in, 2 6.88-7.00 (im, 3 7.2 1-7.30 (in, 2 7.62 2 7.91- 7.95 (in, 3 8.174820 (in, 1 MIS (FAB) m/z 606 To a stirred solution of methyl 441 '-(2-chlorophenyl)ureidoj-3-methoxyphenylacetylJ-(2S)octahydroindolylmethoxy]benzoate (550 mng, 0.91 nunol) in THE (5 ml) was added 0.5 N NaOH mil) 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-CHC1 3 -IIPE to give 166 (286 mg, as a white crystalline powder. NM 388.29 mp 133-136 0 C; 'H-NMR (DMSO-d 6 8 1.16-2. 10 (series of mn, total 10 H), 2.24-2.27 (in, 1 3.55-3.75 (in, 2 3.80 3 3.90-3.96 (in, I 4.174.23 (in, 2 4.34- 4.36 (in, 1 6.73-6.75 (in, 1 6.88 J= 1.5 Hz, 1 6.99-7.05 (mn, 3 7.25-7.30 (in, 1 7.43 (dd,J 1.5, 8.1 Hz, 1 7.87-7.89 (mn, 2 7.95 J= 8.1 Hz, 1 8.08 (dd, J= 8.3 Hz, 1 8.88 1 8.92 1 12.61 (br s, 1 MIS (FAR) "i/z 592 (Mf+lI); Anal Calcd for C 32
H
34 C1N 3 0 6 1/41-120: C,64.42; H, 5.83; N, 7.04; Cl, 5.94. Found: C,64.55; H,6.09; N,6.64; CI,5.93.
Example 159 4-[l '-(2-broinophenyl)ureidoj-3-methoxyphenylacetyl]-(2S)-octahydroindolylinethoxy] benzoic acid WO 01/00206 WO 0100206PCT11iSOO/18079 rI H M e167 A mixture of 4-IN '-(2-bromophenyl)ureidoJ-3-methoxyphenylacetic acid (457 mg, 1.21 nunol), methyl 4-[(2.S)-octahydroindolylmethoxy~benzoate (320 mg, 1.21 mmol), EDCHCI (277 mg, 1.44 mmnol), HOBt (196 mg, 1.45 minol), and Et 3 N (200 ml, 1.43 mniol) in THF (7 ml) was stirred at room temperature overnight The mixture was diluted with H,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, v/v) as eluent to give methyl I-[4-IN '-(2-bromophenyl)ureido]-3 -methoxyphenylacetyl J-(2.S)-octahydroindolyl methoxyjbenzoate (423 mg, 54%) as a white foam. 'H-NMR (CDC1 3 8 1.15-1.89 (series of m, total 8 1.96-2.02 (in, 1 2. 16-2.32 (mn, 2 3.63 2 3.65 3 3.82-3.86 (in, 1 H), 3.88 3 4.30-4.39 (in, 3 6.75-6.77 (in, 2 6.88-6.93 (mn, 3 7.24-7.31 (mn, 1 7.37- 7.50 (mn, 3 7.91-7.99 (in, 3 8.12-8.15 (mn, 1 MIS (FAB) m/z 650 To a stirred solution of methyl 4-[l-14-IN'-(2-bromophenyl)ureidoj-3-methoxyphenyl acetylj-(2S)-octahydroindolylinethoxyjbenzoate (420 mng, 0.65 mmol) in THE (5 nil) was added 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-CHCI 3 -IPE to give 167 (197 mng, as a white crystalline powder. MW 636.53 mp 118- 123 0 C; 'H-NMR (DMSO-d) 8 1.16-2.27 (series of in, total 10 3.56-3.75 (mn, 3 3.81 3 3.90-3.96 (mn, 1 4.174.23 (in, 2 4.34-4.36 (mn, 1 6.73-6.75 (in, 1 6.88-7.05 (mn, 4 7.30-7.34 (in, 1 7.59-7.6 1 I 7.87- 7.96 (in, 4 8.73 1 8.91 1 12.64 (br s, 1 MS (FAB) nih 636 Anal.
Calcd for C 3 2 H34BrN 3
O
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; B r, 12.47.
Examp~le 160 4-[3-[3-methoxy-4-[N-(2-methylphenyl)ureidojphenyl~acetyl-4-thiazolidinyl~methoxybenzoic acid MHH Me o j-CoH 168 To a stirred solution of thiazolidine-4-carboxylic acid (5.0 g, 37.6 iniol) in DMF (50.0 ml) was added (B00) 2 0 (9.8 g, 45.1 inmol) and TEA (8.0 ml). The reaction mixture was stirred at room 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 NaSO 4 and concentrated in vacuo. The residue was purified by column chromatography on silica gel with EtOAc-n-hexane (1:3 as eluent to give 3-tert-butoxycarbonylthiazolidine-4-carboxylic acid (6.5 g, 74%) as a white crystalline solid.
'H-NMR (CDCI 3 8 1.49 (br s, 9H), 3.20-3.30 2H), 4.09-4.87 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 0 C. 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 o0C, and 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 (CDC1) 8 1.48 9H), 2.80-2.85 1H), 3.13-3.17 1H), 3.20-3.30 1H), 3.64-3.70 2H), 4.34 (br s, 1H), 4.60 (br s, 1H).
To a stirred solution of 3-tert-butoxycarbonyl-5-hydroxymethylthiazolidine (1.9 g, 8.7 mmol), 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 v/v) as eluent to give methyl 4-(3-terlbutoxycarbonyl-4-thiazolidinyl)methoxybenzoate (1.6 g, 52%) as a pale yellow oil. 'H-NMR (CDC13) 8 1.49 9H), 3.11-3.19 2H), 3.88 3H), 4.04-4.31 3H), 4.61 2H), 6.96 (d, J 8.8 Hz, 2H), 7.98 J 8.8 Hz, 2H).
To a stirred solution of methyl 4-(3-lert-butoxycarbonyl-4-thiazolidinyl)methoxybenzoate (440 mg, 1.25 mmol) in CHCI 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 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 (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 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.
WO 01/00206 WO 0100206PCT/USOOI 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 vtv) as eluent to give methyl 4-[3-[3-methoxy-4-[N'-(2-methylphenyl)ureido] pheriyl]acetyl-4thiazolidinyl] methoxybenzoate (340 mg quant) as an amorphous solid. 'H-NMiR (CDC 3 8 2.31 3H1), 3.15-3.16 (in, 2H1), 3.67-3.69 (in, 5H1), 3.88 311), 4.094.14 (mn, 2H), 4.22-4.90 (in, 3H), 6.30 (in, 1H), 6.74-6.96 (mn, 4H), 7.11-7.25 (in, 4H1), 7.49-7.51 (in, IH), 7.95-8.12 (in, 311).
To a stirred solution of methyl 4-[3-[3-methoxy-4-[N'-(2-inethylphenyl)ureidolpheniylJacetyl-4thiazolidinyl] methoxybenzoate (340 mg, 0.62 minol) in THF (5.0 ml) and EtOH (3.0 mil) was added IN NaOH (0.62 mil, 0.62 nunol). The mixture was stirred at 70 *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 168 (290 mng, as a white crystalline solid. MW 535.62 mp 125-128 IR (KBr) 3357, 2937, 1604, 1533, 1419, 1253, 1166, 1033, 773 cm-1; 'H-NM4R (DMS0-dl 6 8 2.25 3H1), 3.05-3.20 (in, 2H1), 3.71, 3.83, and 3.85 (each s, total 511), 4.034.15 (in, 311), 4.52-4.76 (in, 2H1), 6.15-6.17 (in, 711), 7.78-8.30 411), 8.30 (in, 111), 8.56 (in, 111); MS (FAB) ,n/z 536 Anal calcd for C 2 sH29N 3 0 6 S-0.5H 2 O: C, 61.75; H, 5.55; N, 7.72. Found: C, 61.72; H, 5.55; N, 7.49.
Example 161 4-f 3-[4-[NV'-(2-chlorophenyl)ureido]-3-methoxyphenylacetyll-4-thiazolidirnyllmethoxybenzoic acid 'H H Me o-J-169 To a stirred solution of methyl 4-(3-teri-butoxycarbonyl-4-thiazolidinyl)inethoxybenzoate (600 ing, 1.7 inxol) in C11 2 C1 2 (6,0 ml) was added TFA (6.0 mil) at 0 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,
and concentrated in vacuo. The crude product was used to the subsequent reaction without futher purification. To a stirred solution of the crude product, 4-[N'-(2-chlorophenyl)uredioj-3inethoxyphenylacetic acid (570 ing, 1.7 inmol), HOBt (230 ing, 1.7 inmol), and triethylamine (709 mil, 5.1 inmol) in THF (10.0 mil) and MCCN (10.0 nil) was added EDC-HCI (490 mng, 2.55 inmol) at 0 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. Na11CO 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 WO 01/00206 WO 0100206PCT/USOO/18079 1, v/v) as eluent to give methyl 4-13-[4-IN-(2orphenyl)ureido-3-methoxyphenylacetyll-4thiazolidinyllmethoxybenzoate (900 mg, 93%) as a colorless oil. 'H-NMR (CDCI 3 8 3.15-3.18 (in, 211), 3.70 211), 3.78 3H), 3.86 311), 4.09-4.93 (in, 511), 6.80-7.01 7. 19-7.35 (mn, 4H), 7.94-8. 18 (in, 4H).
To a stirred solution of methyl 4-13-[4-IN'-(2-chlorophenryl)ureidol-3-methoxyphenylacetyll-4thiazolidinyllmethoxybenzoate (900 ing, 1.6 mmol) in THF (8.0 ml) and MeOH (4.0 ml) was added IN NaOH (3.1 ml, 3.1 inmol). 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 169 (780 mg, 89%) as a white crystalline solid. MW 556.03 mp 126-129 IC; IR (KBr) 3343, 2937, 1604, 1531, 1421, 1245, 1166, 1035, 752 ciii'; 1H-NMvR (DMSO-Q 5 8 3.06-3.24 (mn, 2H), 3.72-3.85 (mn, 511), 4.02-4.27 (in, 3H1), 4.53-4.76 (mn, 2H1), 6.74-7.44 (mn, 7H), 7.87-8.30 (in, 411), 8.89-8.95 (in, 2H1); MS (FAB) m/z 556 Anal calcd fur C27H 27
N
3 O5ClS-0.7H 2 O: 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.
Example 162 4-3[-N-2boohry~rio--ehxpeyaey]4tizldnlmtoyezi acid H 6Me170 To a stirred solution of methyl 4-(3-teri-butoxycarbonyl-4-thiazolidinyl)methoxybenzoate (560 mg, 1.6 mmol) in CH 2
CI
2 (5.0 ml) was added TFA (5.0 ml) at 0 The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat. NaHCO, was added to the residue, and extracted with CHC1 2 The extract was washed with brine ,dried over Na 2
SQ
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-[A'-(2-bromophenyl)uredio]-3-methoxy phenylacetic acid (599 mg, 1.6 inmol), HOBt (213 mng, 1.6 inmol), and triethylamine (659 ml, 4.7 nunol) in THE (10.0 ml) and MeCN (10.0 ml) was added EDC-HC1 (455 mng, 2.4 inmol) 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 2-M citric acid, and sat. NaHCO 3 then dried over Na 2
SO
4 and concentrated in vaczeo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc v/v) as eluent to give methyl 4-3[-N(-rmpey~rio--ehxpeyaey]4tizldnl inethoxybenzoate (870 mg, 89%) as a colorless oil. 'H-NMR (CDCI 3 8 3.00-3.20 (in, 3H), 3.70 (s, WO 01/00206 WO 0100206PCT/USOO/18079 2H), 3.81 3H), 3.88 3H), 4.094.23 (in, 4.42 J= 8.5 Hz, IlH), 4.59 J 8.5 Hz, 4.70-4.92 (in, 6.81-7.53 9H), 7,95-8.15 (mn, 4H).
To a stirred solution of methyl 4-[3-[4-[N'-(2-bromophenyl)ureido]-3-methoxypheniylacetylj-4thiazolidinylJmethoxybenzoate (870 ing, 1.4 mmol) in THE (8.0 ml) and MeOH (8.0 ml) was added IN NaOH (2.8 ml, 2.8 iniol). The mixture was stirred at 70 *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 vacua to give 170 (740 mg, as a white crystalline solid. MW 600.48 mp 125-133 IR (KBr) 3332, 293 5, 1604, 1527, 1421, 1245, 1166, 1027, 750 'H-NMR (DMSO-dQ 8 3.01-3.25 (mn, 2H), 3.72-3.85 (mn, 5H1), 4.02-4.30 (mn, 2H), 4.54 J 8.8 Hz, 111), 4.74-4.87 (Mn 2H), 6.76-7.07 (in, 5H), 7.30-7.34 (mn, 111), 7.59 J 8.1 Hz, 111), 7.86-7.98 (in, 4H), 8.74 11H), 8.92-8.94 lIi); MS (FAR) m/z 600 (MC+l); Anal. calcd for C 27
H-
26
N
3 O613rS0.3H- 2 O: C, 53.52; K14.43 N 6.94 Found: C, 53.54; H, 4.45; N, 6.80.
Example 163 cis-4-[[1-13 -methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetylj -2-pyrrolidinyljmethylamino] cyclohexanecarboxylic acid N'-C
OOH
~2HH Me 171 To a stirred solution o2S roi'dinemethanol (2.0 g, 20.0 nunol), 3-methoxy-4-[N'-(2-methyl phenyl)urediolphenylacetic acid (6.28 g, 20.0 nunol), HOBt (71 mg, 0.53 mmol), and triethylamine (5.5 ml, 40.0 minol) in THE (50.0 ml) and MeCN (40.0 ml) was added EDC-HCl (5.7 g, 30.0 inmol) at 0 The reaction mixture was stirred at roam temperature for 16 hr, and concentrated in vacua. 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 vacua. The residue was purified by column chromatography on silica gel with EtOAc to MeOH- CH 2
CI
2 v/v) as eluent to give l-[3-inethoxy-4-IjN'-(2-methylphenyl) ureidoJphenylacetyl]-2S-pyrrolidineinethanoI (7.0 g, 89%) as a white crystalline solid. 'H-NMvR
(CDCI
3 8 1.54-1.58 (mn, lH), 1.80-2.04 (mn, 3H), 2.27 3H1), 3.42-3.46 (in, 1H), 3.54-3.65 (in, 2H), 3.62 2H), 3.69 3H), 4.21-4.23 (in, 111), 5.04 (mn, 111), 6.68-6.79 (in, 3H), 7.09-7.3 1 (mn, 4H), 7.52 J= 7.8 Hz, 111), 8.07 J= 8.0 Hz, 11H).
To a stirred solution of oxalyl chloride (0.3 ml, 3.3 mnxol) in CH 2
CI
2 (30.0 ml) was added DMS0 (6.6 ml, 0.51 inmol) at -78 After 5 minutes, to the mixture was added 1-[3-methoxy-4-[N'-(2- WO 01/00206 WO 0100206PCT/USOO/18079 methylphenyl)ureido]phenylacetyll-2-pyrrolidinemethao (1.2 g, 3.0 inmol) in CH 2
CI
2 (5.0 mil).
The mixture was stirred for 30 minutes at -78 and triethylainine (2.1 mil, 15.0 rnnol) was added. The mixture was stirred for 30 minutes at -78 and stirred for 30 minutes at room temperature. Water was added to the midxture, and extracted with CH 2
CI
2 The extract was washed with water, then dried over Na 2
SO
4 and concentrated in vacua. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product, benzyl cis-4-aininocyclohexanecaiioxylate (769 mg, 3.3 mxnol), and AcOH (0.32 ml) in DCE (10 ml) was added NaBH(OAc) 3 (1.1 g, 5.4 minol) at 0 The reaction mixture was stirred at rn temperature for 18 hr. The mixture was concentrated in vacua. Sat. NaHCO 3 was added to the residue, and extracted with CHCI 2 The extract was washed with brine, dried over Na 2
SO
4 and concentrated in vacua. The residue was purified by column chromatography on silica gel with MeOH-CH 2
CI
2 vlv) as eluent to give benzylcis-4-[[1-[3-methoxy-4-[N'-(2-methylphenyl) ureidolphenylacetylJ-2-pyrrolidinyI~methylaminocyclohexanecaboxylate (1.5 g, 83%) as an amorphous solid. 'H-NMIR (CDCI 3 8 1.40-1.65 (mn, 6H), 1.80-1.98 (mn, 6H), 2.26 3H), 2.45- 2.65 (in, 3H), 2.81-2.86 (in, lH), 3.44-3.46 (in, 2H), 3.56 2H), 3.67 3H), 3.90-4.15 (in, 111), 5.09 and 5.11 (each s, total 2H), 6.74-6.83 (mn, 3H), 7.07-7.20 (mn, 4H), 7.3 1-7.35 (in, 5H), 7.53 7.55 (in, 11-1), 8.02-8.06 (in, 11-).
To a stirred solution of benzylcis-4-[I [.3..methoxy4-[N'-(2-inethylphenyl)ureidolpheny acetylJ-2-pyrrolidinyl~inethylaminojcyclohexanecarboxylate 5 g, 2.45 inmol) in THF (10.0 ml) and MeOH (5.0 mil) was added IN NaOH (3.68 nil, 3.68 inmol). The mixture was stirred at 70 'C for 18 hr. The mixture was concentrated in vacua, water was added thereto, and neutralized with IN HCI. The mixture was concentrated in vacuo. The residue was purified by column chromatography on silica gel with MeOH-CHCI 2 v/v) as eluent to give cis-4-[[1-[3-inethoxy- 4-[N'-(2-niethyl phenyl)ureidolphenylacetyl]-2-pyrrolidinyl~lnethylaninolCyclohexanecarboxylic acid 171 (940 mng, 73%) as an amorphous solid. MW 522.64 IR (KBr) 3283, 2945, 2860, 1534, 1453, 1415 cm-i'; 'H-NMR (DMSO-d,) 8 1.38-2.00 (in, 12H), 2.45 3H), 2.30-3.95 (in, 411), 3.22-3.75 (in, 2H), 3.58 211), 3.86 3H1), 4.12 (in, 111), 6.73-7.16 (in, 511), 7.77-7.79 (in, 111), 7.98-8.02 (in, 111), 8.51-8.52 (in, 1H), 8.57-8.59 (in, IH); MS (FAR) m/z 523 Anal. calcd for G ,iHN 4
O
5 -0.5NaCI-2.2H 2 O: C, 58.89; H, 7.23; N, 9.47. Found: C, 59.2 1; H, 7.11; N, 9. 11.
Exampule 164 methyl cis-4-[[ 1-[3-methoxy-4-[N -(2-methylphenyl)ureidojphexiylacetylJ-2-pyrrolidiniyljinethyl amidnolcyclohexaxiecarboxylate HCl salt WO 01/00206 WO 0100206PCT/USOO/18079 O H i K~ C O O M e H H 6Me 172 SOC1 2 was added to MeOH at 0 0 C. After stirred for 5 minutes, cis-4-[[ l-13-methoxy-4-IN'-(2methylphenyl)ureidolphenylacetylJ-2-pyrrolidinylmethylaminolcyclohexanecarboxylic acid (200 mg, 0.38 mnmol) was added. The mixture was stirred at room temperature for 5 hr. The mixture was concentrated in vacua. Aq. 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 vacua. The residue was purified by column chromatography on silica gel with MeOH-CH 2 Cl 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 vacua to give 172 (160 mg, as an amorphous solid. NM 536.66 IR (KBr) 3247, 2950, 2875, 1731, 1671, 1612, 1533, 1454, 1205 cm- 1 IH-NMiR (DMSO-dQ 8 1.45-2. 10 (in, 12H), 2.25 3H1), 2.60-2.70 (in, 1H), 2.90-3.20 (in, 3H), 2.50-2.55 (in, 2H1), 3.63 (in, 514), 3.86 3H), 4.154.30 (in, 111), 6.74-7. 16 5H), 7.76- 7.78 (in, I1-H), 8.00-8.09 (in, 111), 8.54-8.70 (in, 2H); MIS (FAR) m/z 537 Anal calcd for C31HINO 5 -l.0HC-1.0HO: C, 60.95; H, 7.33; N, 9.48; Cl, 6.00. Found: C, 60.87; H, 7.47; N, 8.97: Cl, 5.90.
Example 165 4-IN-[I [-ehx--N-2mtypey~riojhnlctl--yrldnlmtylN methylaminojcyclohexanecarboxylic acid H H Me L173 To a stirred solution of methyl ci--[-3mtoy4['(-etypey~riopeyaeyl 2-pyrrolidinyllmethylainiinolcyclohexanecarboxylate (300 mg, 0.55 innol), HCHO (300 ml), and AcOH (66 mng, 1. 1 mmol) in MeGH (10.0 ml) was added NaBH 3 CN (70 mg, 1. 1 mmol) at 0 0
C.
The reaction mixture was stirred at room temperature for 18 hr. After concentrated in vacua, water was added and extracted with CH 2 C1 2 The extract was washed with water, then dried over Na 2
SO
4 and concentrated in vacua. The residue was purified by TLC with MeOH-CH 2
CI
2 (3:97, vlv) as eluent to give methyl 1 [3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-2pyrrolidinylmethyl]-N-methylaminojcyclohexanecarboxylate (160 mg, 52%) as an amorphous solid. 'H-NMR (CDCI 3 8 1.30-2.20 (in, 12H), 2.27-2.37 (in, 6H), 2.50-2.60 (in, lH), 3.30-3.80 (in, 5H), 3.55 2H), 3.66-3.73 (in, 6H), 4.10-4.20 (mn, 1H), 6.60-7.55 (in, 71-i), 8.03 (in, IH), 8.15 (in, I1-H).
343 To a stirred solution of methyl 4 1 -methioxy-[N'-(2-methylphenyl)ureidoJphenylacetyl] -2pyrrolidinylmethylJ-Nf-methylaznino~cyclohcxanecarboxylate (100 mg, 0. 18 mmal) in THF (5.0 ml) and MeOH (2.5 ml) was added IN NaOH (0.36 mld, 0.36 mmol). The mixture was st-irred at 60 0
C
for 18 hr. The mi~xture was concentrated in vacuo, water was added thereto, and neutralized with I N HCI. The mixture was concentrated in vacuo. The residue was purified by TLC with MeOH-
CH
2
CI
2 vlv) as eluent to give 173 (10 mg, 10%) as an amorphous solid. MW 536.66 IR (Kl~r) 3440, 2954, 1697, 1533, 1454 1 H-NMIR (DMSO-d) 5 1,20-2.30 (in, 13H-), 2.24 (s, 3H), 2.35-4.00 (in, 13H-), 6.50-8. 10 (mn, 8.50 (in, IM); MS (FAB) m/z 537 Anal. calcd for C 3 oH40NO,2.0NaCU-0.8H 2 C, 53.94; H, 6.28; N, 8.39. Found: C, 54.08; H, 6.52; N, 8.04.
Example 166 4 -[[14[3-mtoy N-2-ehlhnluriopeyaetl 2)pridinyl]miethoxyI cyclohexanecarboxylic acid
_COOH
Me H H Me 174 :A mixture of methyl 4 -(l-1ert-butoxycarbony-(2.S)-pyrrolidinyl)methoxnrzoate (1.0 g, 2.9 :15 inmol) and 5% Rh on alumina (500 mg) in EtOH (10.0 mld) and AcOH (1.0 nil) 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 nhexane-EtOAc 1, v/v) as eluent to give methyl cis- 4 -zer:-butoxycarbonyl-(2S)-pyrrolidinyl) methoxyjeyclohexanecarboxylate (900 mng, 89% as a pale yellow oil. 'H-NMIR (CDCl 3 8 1.46 (s, 9H), 1.46-2.00 (in, 12H-), 2.34 (in, 3.20-3.55 (in, 3.67 3H), 3.84-3.92 (in, 11-4).
~To a stirred solution of methyl cis-4-[( -tert-butoxycarbony-(2S)-pyrrolidinyl)inethoxyj cyclohexanecarboxylate (900 mg, 2.6 mnmol) in CH 2
CI
2 (5.0 ml) was added TEA (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. NaJACO, was added to the residue, and extracted with CH 2
CI
2 The extract was washed with brine, dried over NaSO,, and concentrated in vacua. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product (200 mg, 0.83 inmol), 3-ehx4[,-2mtypey~rdopeyaei acid (260 ing, 0.83 343a mmol), HOBt (1 35mg, 1.0 mmol), and triethylamine 34 4 jd, 2.5 mmol) in THIF (10.0 ml) and MeCN (10.0 ml) was added EDC-HCI (238 mg, 1.24 mmol) at 0 0 C. The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacuco. Water was added to the residue, and extracted with EtOAc. The extract was wvashed 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 v/v) as eluent to give methyl cis-4-[[1-[3-methoxy-4-[AP-(2-miethylphenyl)ureido] phenylacetyl] -(2S)-pyrrolI id inyl] methoxy] cyclohexanecarboxyl ate (460 mg, quant) as amorphous solid. 'H-NMR (CDCI 3 6 1.35-2.10 (in, 12H), 2.15-2.38 (in, IH), 2.29 (in, 3H), 3.20-3.55 (in, 5H), 3.58 2H), 3.66 3H), 3.73 3H), 4.20-4.25 (in, 1H), 6.26- 6.30 (in, 1H), 6.78-6.81 (in, 2H), 7.06-7.23 (in, 3H), 7.51-7.52 (in, 8.01-8.03 (in, I1H).
[R:\LIBFFJ 10 1 WO 01/00206 WO 0100206PCT/USOOIIS079 To a stifred solution of methyl 4-[[1-[3-methoxy-4-[N'-(2-methylphenyl)ureido] phenylacetyl]-(2.S)-pyrrolidinyl]methoxyjcyclohexanecarboxylate (460 mg, 0.86 mmol) in TIHF (10.0 mil) and EtOH (5.0 ml) was added IN NaOH (1.4 mil, 1.4 mmol). The mixture was stirred at *C for 18 hr. The mixture was concentrated in vacua, water was added thereto, and neutralized with IN HCI. 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 tR (KBr) 3345, 2937, 1612, 1533, 1454 'H-NMvR (DMSO4) 8 1.00-2.00 (in, 12H), 2.24 3H), 2.20-2.30 (in, 1H), 3.20-3.80 (in, 5H), 3.55 211), 3.85 3H), 4.00-4.18 (in, IH), 6.71-7.16 (Mn 511), 7.78 J 8.0 Hz, IH), 7.90 J= 8.3 Hz, IH), 8.45 111), 8.54 MS (FAB) m/lz 524 Anal. calcd for CH 37
N
3 O&-0.2H 2 O: C, 66.07; H, 7.15; N, 7.97. Found: C, 66.02; H, 7.14; N, 7.87.
Exme167 4-[[I1-[4-[N'-(2-chlorophenyl)ureidoj-3-inethoxylphenylacetyll -(2S)-pyrrolidinyllmethoxy] cyclohexanecarboxylic acid H Me NQ .175 To a stirred solution of methyl 1-tert-butoxycarbonyl-2-pyrrolidinyl)methoxyj cyclohexane carboxylate (900 mng, 2.6 inmol) in CH 2
CI
2 (5.0 mil) was added TFA (5.0 ml) at 0 The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacua. 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 vacua. The crude product was used to the subsequent reaction without fuirther purification. To a stirred solution of the crude product (200 mg, 0.83 innol), 4-fN'-(2-chlorophenyl)uredio]-3-methoxyphenylacetic acid (277 ing, 0.83 inmol), HO~t (135 mg, 1.0 inmol), and triethylamine (344 nil, 2.48 inmol) in THE (10.0 nml) and MeCN (10.0 mil) was added EDC-HCI (238 mng, 1.24 inxol) at 0 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, and concentrated in vacua. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc vA') as eluent to give methyl 4-[j 1-14-[N'- (2-chlorophenyl)ureido]-3-inethoxylphenylacetyl]-(2S)-pyrrmlidinyl~inethoxy cyclohexane carboxylate (450 ing, 97%) as a colorless oil. 'H-NMR (CDCl 3 8 1.35-2.15 (in, 12H), 2.25-2.40 (in, 1H), 3.40-3.70 (in, 51B), 3.61 2H), 3.66 3H), 3.81 3H), 4.204.30 (in, IB), 6.81-6.99 (in, 3H), 7.17-7.34 (in, 3H), 7.92 -7.94 (in, 2H), 8. 17-8.19 (in, 2H).
WO 01/00206 WO 0100206PCT/USOOI 18079 To a stirred solution of methyl N(-hlrpey~red]3mthxlhryactl-2) pyrrolidinyllmethoxy~cyclohexanecaiboxylate (450 mg, 0.86 mmol) in THE (10.0 ml) and MeOH ml) was added IN NaOH (1.4 ml, 1.4 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 HCI.
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 tR (KBr) 3330, 2938, 1704, 1594, 1533, 1438, 1199 cm-1; 1H-NMR (DMSO-d) 8 1.00-2.00 (in, 12H), 2-20-2.30 (in, IH), 3.20-3-80 (in, 5H), 3.55 2ff), 3.85 3H), 4.00-4.20 (in, 1H), 6.73-6.75 (mn, IH), 6.87 lH), 6.99-7.03 (in, 1H), 7.25-7.29 (in, IH), 7.42 J= 7.1 Hz, IH), 7.95 J= 8.3 Hz, 1H), 8.08 J lH), 8.78 1H), 8.92 1H); MS (FAB) m/lz 544 AnaL calcd for C 2 g- 3 4NAOClO.2H1 2
O:
C, 61.41; H, 6.33; N, 7.67; Cl, 6.47. Found: C, 61.37; HK 6.32; N, 7.56; Cl, 6.55.
Example 168 1 ['(-rmohnluedl3-ehxlhnlaey](S-yroiiylehx cyclohexanecarboxilic acid &e176 To a stirred solution of methyl 4-[(l-ter1-butoxycarbonyl-2-pyrrolidinyl)methoxy]cyclohexale carboxylate (900 mg, 2.6 mmnol) in CH 2
CI
2 (5.0 ml) was added TEA (5.0 ml) at 0 The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacuo. Sat.
NaHCO, 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 (200 mg, 0.83 mmol), 4-[N'-(2-bromophenyl)uredioj-3-methoxyphenylacetic acid (314 mng, 0.83 inmol), HOBt (135 mg, 1.0 minol), and triethylanine (344 nil, 2.48 inmol) in THE (10.0 ml) and MeCN (10.0 ml) was added EDC-HCl (238 mg, 1.24 inmol) at 0 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 v/v) as eluent to give methyl [4-t l-[4-IN'- (2-broinophenyl)ureido-3-methoxylphenylacetylJ(2S)-pyrrolidinylinethoxyIcyclohexale carboxylate (450 mg, 90%) as a colorless oil. 'H-NMR (CDC1 3 8 1.30-2. 10 (in, 12H), 2.35-2.40 (in, 3.25-3.70 (in, 5H), 3.84 3H), 4.10-4.25 (in, lH), 6.81-7.06 (in, 4H), 7.25-7.32 (in, 2H), 7.50-7.52 (in. 1H), 7.90-7.92 (in, lH), 8.13-8.15 (in, IH).
WO 01/00206 WO 0100206PCT/USOO/1 8079 To a stiffed solution of methyl 4-Il-[4-['-(2-bromophenyl)ureido-3-methoxylpheniylacetylj-(2S)pyrrolidinyllmethaxycyclohexanecarboxylate (450 mg, 0.74 mmol) in TE (10.0 mil) and MeOH mil) was added IN NaOH (1.2 ml, 1.2 mmnol). The mixture was stirred at 70 'C 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 vacua to give 176 (340 mg, 77%) as a white crystalline solid. MW 588.49 mp 108-111 IR (KBr) 3328, 2938, 1702, 1594, 1529, 1434 'H-NMvR (DMSO-d) 8 1.00-2.00 (in, 12H), 2.15-2.25 (in, 111), 3.40-3.75 (in, 3.48 2H), 3.85 3H), 4.044.15 (in, 111), 6.70-6.72 (in, 111), 6.87 1H), 6.94-6.98 (in, 1H), 7.29-7.33 (in, 111), 7.59 J= 8.1 Hz, 1H), 7.93-7.95 (in, 2H), 8.73-8.74 (in, 1H), 8.91-8.92 (in, 111); MS (FAR) m/z 589 Anal. calcd for CHN 36 r0.2H 2 O: 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(1 -[4-[N'-(2-inethylphenyl)ureidophenylacetyl-(2S)-pyrrlidiny~methoxyjcyclohexane carboxylic acid ToH 177 Toa stirred solution of methyl 4-[(1-lerl-butoxycarbonyl-2-pyrrolidinyl)methoxylcyclohexane carboxylate (450 mg, 1.3 inmol) 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 vacua. 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, inethylphenyl)uredio]phenylacetic acid (375 mg, 1.3 nunol), HOBt (178 ing, 1.3 minol), and triethylainine (550 ad, 3.9 nunol) in THE (6.0 ml) and MCCN (6.0 ml) was added EDC-HCI (380 ing, 1.9 minol) at 0 The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacua. 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 nhexane-EtOAc vfv) as eluent to give methyl 4-1(1 -[4-[N'-(2-inethylphenyl)ureidoj phenylacetylJ-(2.S)-pyrrolidinyljinethoxy]cyclohexanecarboxylate (520 mg, 78%) as a colorless oil.
'H-NMR (CDCI 3 8 1.30-2.40 (in, 13H), 2.21 3H), 3.30-3.80 (in, 711), 3.65 3H), 4. 10-4.30 (in, IH), 6.90-7.20 (in, 8H), 7.40-7.70 (in, 2H-).
WO 01/00206 WO 0100206PCT/USOO/18079 To a stirred solution of methyl 4-[Il-[4-[N'-(2-methylphenyl)ureidolphenylacetyl-(2S)pyrrolidinyl]methoxy]cyclohexanecarboxylate (520 mg, 1.0 iniol) in THF (10.0 ml) arid MeOH nil) was added IN NaOH (1.5 ml, 1.5 nunol). The mixture was stirred at 70 IC 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 177 (450 mg, 91%) as a white crystalline solid. MW 493.60 mp 107-111 0 C;JR (KBr) 3353, 2938, 1704, 1540, 1454, 1240 'H-NMR (DMSOAd) 8 1.20-2.00 (mn, 12H), 2.23 3H), 2.22-2.24 (in, I1H), 3.20-3.80 (in, 711), 4.00-4.18 (in, IH), 6.90-6.94 (mn, 111), 7.10-7. 16 (in, 5H), 7.36-7.38 (in, 2H), 7.82-7.87 (in, 211), 8.89 1W), 12.0 (br s, 1W); MS (FAR) m/z 494 Anal calcd for C~sH 3
NAO
5 0.2H 2 O: C, 67.64; H, 7.18; N, 8.45. Found: C, 67.66; H, 7.19; N, 8.24.
Example 170 cis-4-11 -[4-[N'(2chlorophenyl)ureido-3-methoxylphenylacetylJ-(2S)-octahydroindolylmethoxyI cyclohexanecarboxylic acid IQ H H& C O O 178 To a stirred solution of [Il-tert-butoxycarbonyl-(2S)-octahydroindolyl]carboxylic acid (1.0 g, 3.7 nunol) in THF (10.0 mil) was added BH 3 THF(1.0 M in THF, 8.0 mil) 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 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 [Itert-butoxycarboyl-(2S)-octahydroindolyljimethanol (947 mng, quant) as a a colorless oil.
To a stirred solution of [Il-:ert-butoxycarboyl-(2S)-octahydroindolyl] methanol (947 mng, 3.7 inmol), methyl 4-hydroxybenzoate (565 ing, 3.7 inmol), and Ph 3 P (1.2 g, 4.5 minol) in THE mil) was added DIAD (984 ing, 4.5 inmol) at 0 The reaction mixture was stirred at room temperature for 18 hr. The mixture was concentrated in vacua. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc v/v) as eluent to give methyl 4- [1 -tert-butoxycarbonyl-(2S)-octahydroindolylmethoxy~benzoate (700 mg, 50%) as a pale yellow oil. 'H-NMvR (CDCI 3 8 1.10-2.25 (in, 11WH), 1.45 9W1), 3.88 3H), 3.70-4.20 (in, 3H), 4.36 (br s, I1H), 6.94 J 8.8 H z, 211), 7.96 J 8.5 H z, 2W1).
A mixture of methyl 1-terz-butoxycarbonyl-(2S)-octahydroindolylinethoxyjbenzoate (700 ing, WO 01/00206 WO 0100206PCT/USOO/18079 1.8 innol) 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 1, v/v) as eluent to give methyl cis-4-[1I-leri-butoxycarbonyl-(2S)-octaiydro indolylmethoxylcyclohexanecarboxylate (600 ing, 85%) as a pale yellow oil. 'H-NMR (CDCI 3 8 1. 10-2.35 (in, 20H), 1.44 9H), 3.45-3.90 (mn, 5H), 3.80 3H).
To a stirred solution of methyl cis-4-[I1-tert-butoxycarbonyl-(2S)-octahydroindolylmethoxy] cyclohexanecarboxylate (600 mng, 1.5 inmol) in CH 2
CI
2 (6.0 ml) was added TFA (6.0 ml) at 0 'C.
The reaction mixture was stirred at room temperature for 0.5 hr. The mixture was concentrated in vacua. 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 vacua. The crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product (221 ing, 0.75 inmol), 4-[N'-(2-chloropheniyl)uredioJ-3-methoxyphenylacetic acid (250 mg, 0.75 inmol), HO~t (101 mg, 0.75 nol), and triethylamine (312 ml, 2.3 minol) in THE (10.0 ml) and MeCN (10.0 ml) was added EDC HCI (2 16mg, 1. 1 iniol) at 0 The reaction mixture was stirred at room temperature for 16 hr, and concentrated in vacua. Water was added to the residue, and extracted with EtOAc. The extract was washed with sat. NaHCO3, 2-M citric acid, and sat.
NaHCO 3 then dried over Na 2
SO
4 and concentrated in vacua. 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- [N'-(2-chloropheniyl)ureidoj-3-methoxylphenylacetyl]-(2S)-octahydroindolylinethoxylcyclohexane carboxylate (430 mng, 949/) as a colorless oil. 'H-NMR (CDCI 3 8 1.10-2.40 (mn, 20H), 3.45 (br s, lH), 3.62 2H), 3.66 3H), 3.73 3H), 3.60-3.85 (in, 2H), 4.09-4.14 (in, 2H), 6.75-6.98 (in, 3H), 7.22-2.46 (in, 4H), 7.92 J 8.0 Hz, 1H), 8.18 J 8.3 Hz, lH).
To a stiffed solution of methyl cis-4-[I1-f4-[N'-(2-chlorophenyl)ureido]-3-inethoxylphenylacetylJ- (2S)-octahydroindolylmethoxyjcyclohexanecarboxylate (430 ing, 0.7 minol) in THE (10.0 ml) and MeOH (5.0 ml) was added IN NaOH (1.4 ml, 1.4 inmol). The mixture was stirred at 70 'C for 24 hr. The mixture was concentrated in vacua, water was added thereto, and neutralized with IN HCI. The resulting solid was collected, washed with water, and dried in vacua to givel78 (360 ing, 869/) as a white crystalline solid. MW 598.13 mp 120-121*C; IR (KBr) 3338, 2933, 2859, 1614, 1533, 1438 cm'1; 'H-NMR (CDC 3 8 1.05-2.40 (Mn 20H), 3.38-3.50 (in, 2H), 3.63 and 3.65 (each s, total 2H), 3.71 and 3.75 (each s, total 3H), 3.70-3.80 (in, 1H), 3.93-3.97 (in, IH), 4.15 (br s, 1H), 6.75-6.77 (in, 2H), 6.93-6.97 (in, lH), 7.20-7.32 (in, 3H), 7.60-7.63 (in, 1H), 7.85 J WO 01/00206 WO 0100206PCT/USOO/18079 8.3 Hz, 1H), 8.16 J 8.3 Hz, 111); MIS (FAR) in/z 598 Anal. calcd for C32HwN 3
O
6 Cl 0.51-12: C, 63.30; H, 6.81; N, 6.92; CI, 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-mehoxylphenylacetyl]-(2S)-octahydroindolylmehoxy] cyclohexanecarboxylic acid Y~ q Me 179 To a stirred solution of methyl cis-4-[ 1-teri-butoxycarbonyl-(2S)-octahydroindolylmethoxy] cyclohexanecarboxylate (600 mg, 1.5 inmol) in CH 2 Cl 2 (6.0 nml) was added TFA (6.0 ml]) at 0 CC.
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,. 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 fuirther purification. To a stirred solution of the crude product (221 mg, 0.75 mmol), 4-fN'-(2-bromophenyl)uredioJ-3-methoxyphenylacetic acid (284 mg, 0.75 inmol), HOBt (10 1 mg, 0.75 nunol), and triethylamine (312 mil, 2.3 minol) in THF (10.0 mld) and MeCN (10.0 mil) was added EDC-HCI (2 16mg, 1. 1 mmol) at 0 1C. The reaction mixture was stiffed 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., and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc(1 :3 as eluent to give methyl cis-4-j 1-[4- [N'-(2-bromophenyl)ureidoJ -3-riethoxylpheniylacetylJ-(2S)-octahydroindolylmethoxyj cyclohexane carboxylate (480 mg, 96%) as a colorless oil. 'H-NivR (CDCI 3 8 1. 10-2.40 (in, 20H), 3.45 (br s, 1H), 3.61 2H), 3.66 3H), 3.76 3H), 3.60-3.80 (Mn 2H), 4.11-4.14 (in, 2H), 6.76-6.92 (in, 311), 7.25-7.32 (in, 3H), 7.49 J 7.1 Hz, 1H), 7.90 J 8.0 Hz, 8.13 J 8.0 Hz, 111).
To a stirred solution of methyl cis-4-[l-[4-[N-(2-bromophenyl)ureidol-3-methoxylphenylacetylJ- (2S)-octahydroindolylmethoxy~cyclohexanecarboxylate (480 mg, 0.73 mrnol) in THF (10.0 ml) and MeGH (5.0 mil) was added IN NaOH (1.5 nil, 1.5 nunol). The mixture was stirred at 70 'C for 24 hr. The mixture was concentrated in vacuo, water was added thereto, and neutralized with 1IN HCI. The resulting solid was collected, washed with water, and dried in vacuo to give 179 (400 mng, 85%) as a white crystalline solid. M4W 642.58 mp 115-120 IR (KBr) 3332, 2933, 2859, 1704, 1592, 1529, 143 4 cm'; 'H-NMR (CDC1 3 8 1.10-2.40 (in, 20H), 3.40-3.50 (in, 2H), 3.61 and WO 01/00206 WO 0100206PCT/USOOII 8079 3.63 (each s, total 2H), 3.75 and 3.78 (each s, total 3H), 3.70-3.80 (in, 1H), 3.90-3.93 (in, 1H), 4.15 (br s, in), 6.76-6.92 (mn, 3H), 7.26-7.30 (mn, lH), 7.43-7.52 (in, 3H), 7.84-7.86 (mn, 8. 8.12 (in. 1I1); MIS (FAB) ni/z 643 AnaI. calcd for C 2 -1N 3 OBr-0.4H 2 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.
Example 172 3mtoy4[P(-mtypey~riophryaey]2pfoldnlcroyaio cyclohexanecarboxylic acid e H 6Me180 To a stirred solution of benzyl 4-aminocyclohexanecarboxylate (900 mg, 3.9 mmol), boc-proline (830 mg, 3.9 mmol), HOBt (521 mg, 3.9 mmol), and triethylaniine (1.6 ml, 11.6 mmnol) in CH 2 Cl 2 (30.0 ml) was added EDC-HCl (1.1I g, 5.8 mmol) at 0 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 SOI and concentrated in vacuo. The residue was purified by column chromatography on silica gel with n-hexane-EtOAc 1, v/v) as eluent to give benzylcis-4-[(ltert-butoxycarbonyl-2-pyrolidilyl)carbonylaino]Cyclohexaflecaboxylate (600 mg, as an amorphous solid. 'H-NMiR (CDCI 3 8 1.44 911), 1.50-1.90 (in, 12H), 2.20-2.26 (in, 1H), 3.25- 3.50 (in, 2H), 3.80-3.90 (in, IM, 4.10-4.25 (in, 111), 5.12 2H), 7.35-7.36 (mn, To a stirred solution of benzylcis-4-[( 1-ierz-butoxycarbonyl-2-pyrrolidinyl)carbonylaminol cyclohexanecarboxylate (600 mg, 1.4 minol) in CH 2 C1 2 (6.0 ml) was added TEA (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 CHCI 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 fiurther purification. To a stirred solution of the crude product (300 mg, 0.7 iniol), 3-ehx--A-2mtypey~rdopeyaei acid (220 ing, 0.7 minol), HOBt (94 mug, 0.7 minol), and triethylamnine (291 ml, 2.1 nunol) in THE (10.0 ad) and MeCN (10.0 ml) was added EDC-HCl (201 mg, 1. 1 iniol) at 0 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 resulting solid was collected and washed with EtOAc to give benzyl 4-[[l-[3-methoxy-4-[N'-(2-methylphenyl)ureidoI phenylacetyl]-2-pyrrolidinylcarbonylaminocyclohexanecarboxylate (380 mg, as a white WO 01/00206 WO 0100206PCT/USOO/18079 crystalline solid. 'H-NvM (CDCI,) 8 1.40-2.15 (mn, 1211), 2.28 (mn, 3H), 2.30-2.50 (in, 211), 3.40- 3.55 (in, 211), 3.61 2H), 3.71 3H), 3.82 (in, 111), 4.53 J 6.3 Hz, IH), 5. 10 2H1), 6.42 114), 6.77-6.79 (in, 211), 7.04-7.34 (in, 911), 7.50-7.52 (in, 111), 8.05-8.07 114).
To a stirred solution of benzyl 4-[I-[3-methoxy-4-[N'-(2-methylphenyl)ureido~phenylacetylJ-2pyrrolidinyllcarbonylaminolcyclohexanecarboxylate (380 mg, 0.6 minol) in THE (10.0 nil) and EtOH (5.0 ml) was added IN NaOH (0.9 ml, 0.9 inmol). 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 mng, as a white crystalline solid. MW 636.62 mp 136-142 IR (KIBr) 3345, 2940, 1650, 1625, 1535, 1454 'H-NMR (DMSO-d) 8 1.40-2.00 (in, 1211), 2.24 311), 2.30-2.40 (in, 111), 3.45-3.80 (in, 511), 3.86-3.87 (in, 311), 4.30-4.43 (in, 111), 6.65-7.30 (in, 511), 7.70-7.80 (in, 111), 7.98-8.09 (in, 111), 8.46-8.57 (in, 111); MS (FAB) ni/z 537 Anal. calcd for 119H3N 4
O
6 -0.5 112: C, 63.84; H, 6.83; N, 10.27. Found: C, 64.18; H, 6.91; N, 9.85.
Example 173 1-[4-[N-(2iclorophenyl)ureido-3-methoxylphenylacetyl-2-pyrolidinylcarbonylamino cyclohexanecarboxylic acid YI'HH Me 0181 To a stirred solution of benzylcis-4-[(l-Iert-butoxycarbonyl-2-pyrrolidinyl)carbonylamidno cyclohexanecarboxylate (600 mg, 1.4 inmol) in CH 2
CI
2 (6.0 ml) was added TFA (3.0 ml) at 0 0
C.
The reaction mixture was stiffed 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 crude product was used to the subsequent reaction without further purification. To a stirred solution of the crude product (300 mng, 0.7 mxnol), -2-chlorophenyl)uredio]-3-methoxyphenylacetic acid (237 mng, 0.7 minol), HOBt (94 ing, 0.7 innol), and triethylaxnine (291 ml, 2.1 inmol) in THE (10.0 ml) and MeCN (10. 0 ml) was added EDC-HCl (201 ing, 1. 1 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 2-M citric acid, and sat.
NaH1CO 3 then dried over Na 2 SO, and concentrated in vacuc. The resulting solid was collected and washed with EtOAc to give benzyl 1-[4-[N'-(2-chlorophenyl)ureido]-3-inethoxyl phenylacetyll-2-pyrrolidinylJcarbornylaminolcyclohexanecarboxylate (3 10 ing 68%) as a white WO 01/00206 WO 0100206PCT/1iSOO/18079 crystalline solid. 'H-NMvR (CDCI 3 8 1.40-2.15 (in, 12H), 2.30-2.60 (in, 2H), 3.42-3.55 (in, 2H), 3.64 2H), 3.84 (s,3H),4.55 (d,J=6.1Hz, 1H),5.12 6.81-7.35(m,12H),7.96-7.98 (in, lH), 8.17-8.19 IH).
To a stirred solution of methyl benzyl 4-[[1-[4-[A-(2-chlorophenyl)ureido]-3-methoxylpheny acetyl]-2-pyrrolidinyljcarbonylam-ino]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 '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 181 (260 mg, as a white crystalline solid. MW 557.03 mp 135-140 IR (KBr) 3328, 2938, 1594, 1533,1438, 1203 cm-i; 'H-NNM (DMSO-d) 8 1.40-2.20 (in, 12H), 2.30-2.40 (in, I1H), 3.40- 3.80 (in, 5H), 3.65-3.85 (in, 3H), 4.30-4.43 (in, lH), 6.66-7.31 (in, 5H1), 7.42-8.10 (in, 214), 8.89- 8.94 (mn, 2H); MS (FAB) m/z 557 Anal calcd for C 2
,H
3 3 NOsCl-0.3H 2 O: C, 59.79; H, 6.02; N, 9.96;1 Cl, 6.30. Found: C, 59.86; H, 6. 10; N, 9.60; Cl, 6.34.
Examnle 174 4-[1 -[3-inethoxy-4-[N'-(2-methylphenyl)ureidolphenylacetylj-(3S)-pyrrolidinyloxylbenzoic acid ,rg-Q-C csOOH eH H me 182 To a cooled (0 0 stirred solution of methyl 4-hydroxybenzoate (1.84 g, 12.1 iniol), benzyl-3-pyrrolidinol (2.00 ml, 12.1 minol), and P1I 3 P (3.81 g, 14.5 inmol) in THEF (25 ml) was added DIAD (2.86 ml, 14.5 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 1, v/v) as eluent to give methyl 4- 1 l-benzyl-(3S)-pyrrolidinyloxylbenzoate (3.66 g, 97%) as a pale yellow oil. IH-NMvR (CDCI 3 8 1.95-2.02 (in, 1 2.28-2.37 (in, 1 2.57-2.63 (in, 1 2.72-2.81 (in, 2 2.96-3.01 (in, 1 3.63-3.71 (in, 2 3.87 3 4.84-4.89 (in, 1 6.84 J 8.8 Hz, 2 7.23-7.34 (mn, 5 7.95 J =8.8 Hz, 2H).
A solution of methyl 4-[1-benzyl-(3S)-pyrrolidinyloxylbenzoate (3.66 g, 11.8 minol) in MeOH ml) was hydrogenated over Pd(OH) 2 /C (0.73 g, 20 overnight. The reaction mixture was filtered to remove the catalyst and the solution was evaporated to give methyl pyrrolidinyloxyjbenzoate (2.60 g, as a pale yellow oil. 'H-NMR (CDCI 3 8 1.94-2.0 1 (in, 2 WO 01/00206 WO 0100206PCT/USU0/18079 2.09-2. 18 (in, 1 2.9 1-2.97 (in, 1 3.04-3.09 (in, I 3. 16-3.23 (in, 2 3.88 3 H), 4.88-4.91 (in, 1 6.86 (in, 2 7.96-7.98 (in, 2 MS (ESI) m/lz 222 A mixture of 3-methoxy-4-[N '-(2-methylphenyl)ureidolphenylacetic acid (449 mg, 1.43 inmol), methyl 4-[(3S)-pyrrolidinyloxylbenzoate (316 mg, 1.43 nunol), EDC-HCI (330 mg, 1.72 mmol), HOBt (193 ing, 1.43 inmol), and Et 3 N (240 ml, 1.72 mmol) in THE (5 mld) was stirred at room temperature overnight. The reaction mixture was diluted with H,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 CHC1 3 -MeOH (100: 1 to 50: 1, v/v) as eluent to give methyl 1-[3-methoxy-4-IN -(2-methylphenyl)utreido~phenylacetyl-(3S)-pyrrolidinyoxyI benzoate (735 ing, as a pale yellow oil. 'H-NMR (CDC1 3 5 2.03-2.31 (series of s and mn, total 5 3.57-3.78 (series of m, total 9 3.88 3 4.95-4.99 (in, 1 6.73-7.00 (in, 5 H), 7.06-7. 10 (in, 1 7.18-7.22 (in, 2 7.42-7.46 (in, 1 7.57-7.62 1 7.95-8.08 (in, 3 MS (ESI) m/z 518 (MC+1).
To a stirred solution of methyl 4-I[-ehx4['(-ehlhnlued~hnlctl-3) pyrrolidinyloxylbenzoate (627 ing, 1.21 inmol) in THE (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-i N HCI and the resulting precipitate was collected under a reduced pressure. The crude solid was recrystallized from MeOH-CHCI 3 -IPE to give 182 (235 mg, 39%) as a white crystalline powder. MW 503.55 mp 13 1-135 0 C; 'H-NMR (DMSO-d 6 8 2.06-2.27 (series of in, total 5 3.57-3.64 (in, 4 3.71-3.88 (series of s and mn, total 5 5.11 and 5.20 (each in, total 1 6.73-6.77 (in, 1 6.88-6.95 (in, 2 7.02-7.05 (in, 2 7.11-7.17 (in, 2 H), 7.79-7.81 (in, 1 7.88-7.90 (in, 2 7.98-8.03 (in, 1 8.45-8.47 (in, 1 8.55-8.57 (m,lI H), 12.66 (br s, 1 MS (ESI) m/z 504 Anal. Calcd for C 2
,H
2 9
N
3 0 6 3/1H 2 0: C, 65.04; H 5.95; N, 8.13. Found: C, 65.11;- H, 5.99; N, 7.66.
Example 175 1-[4-IN '(2-chlorophenyl)ureido]-3-nethoxyphenylacetylJ-(3S)-pyrrolidinyloxy~belzoic acid O-O-C
OOH
H H 6Me 183 A mixture of i-2clrpeyl)ureidoJ-3-inethoxyphenylacetic acid (410 ing, 1.22 nunol), methyl 4-[(3S)-pyffolidinyloxylbenzoate (270 mng, 1.22 inmol), EDC-HCI (280 ing, 1.46 nunol), HOBt (200 ing, 1.48 inxol), and Et 3 N (205 ml, 1.47 minol) in THE (8 ml) was stirred at room WO 01/00206 WO 0100206PCT/USOO/18079 temperature overnight. The reaction mixture was diluted with H,0, and extracted with EtOAc.
The extract was washed with brine, dried over NaSO 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-(3)-pyrrolidinyloxy benzoate (652 mg, 99%) as a white solid. mp 200-203 0 IH-NMR (CDCI,) 8 2.06-2.32 (in, 2 H), 3.60-3.82 (series of mn, total 9 3.88 3 4.97-5.01 (in, 1 6.76-6.86 (in, 4 6.95-6.99 (in, 1 7.23-7.47 (in, 4 7.91-7.99 (in, 3 8. 19-8.21 (in, 1 MIS (ESI) m/z 537 To a stirred solution of methyl 4-fl -[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetylJ pyrrolidinyloxylbenzoate (650 mg, 1.21 minol) in THF (5 ml) was added 0.5 N NaOH (5 ml) and the reaction midxture 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-CHC1 3 -IPE to give 183 (443 mg, 70%) as a pale yellow crystalline powder. MW 523.97 mp 190-193*C; 'H-NMR (DMSO-d) 8 2.06-2.27 (in, 2 3.56-3.62 (in, 4 3.71-3.88 (series of s and mn, total 5 5.11 and 5.20 (each mn, total 1 6.74-6.78 (in, 1 6.89-6.91 (in, 1 7.00-7.05 (in, 3 7.26-7.30 (in, 1 7.43-7.45 (in, 1 7.88-7.98 (in, 3 8.08-8. 10 (in, 1 8.89-8.95 (in, 2 12.67 (br s, 1 MS (ESI) m/z 524 Anal Calcd for C2 H 2 CINPOAl/H 2 O: C, 61.36; H, 5.05; N, 7.95; Cl, 6.7 1.
Found: C, 61.69; H, 5.45; N, 7.29; Cl, 6.91.
Example 176 4-fl N '-(2-bromophenyl)ureidoJ-3-methoxypheniylacetylJ-(3S)-pyrrolidinyloxylbenzoic acid r H H Me184 A mixture of 4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (540 mg, 1.42 minol), methyl 4-[(3S)-pyrrolidinyloxylbenzoate (315 ing, 1.42 minol), EDCHCI (328 mg, 1.71 inmol), HOBt (230 mng, 1.70 inmol), and Et 3 N (240 ml, 1.72 inmol) in THF (8 ml) was stirred at room temperature overnight. The reaction mixture was diluted with H 2 0, and extracted with EtOAc. The extract was washed with brine, dried over Na 2 SO,, and evaporated. The residue was purified by column chromatography on silica-gel with CHCI 3 -MeOH (100: 1 to 50: 1, v/v) as eluent to give methyl 1-[4-IN -(2-bromophenyl)urcido]-3-methoxyphenylacetyl]-(3S)-pyrrolidinyloxy benzoate (620 mg, as a white foam. 'H-NMR (CDCI 3 8 2.06-2.33 (in, 2 3.60-3.82 (series of mn, total 9 3.89 3 4.97-5.01 (in, 1 6.77-7.00 (in, 5 7.27-7.40 (in, 3 H), 7.49-7.51 (in, 1 7.9 1-7.99 (in, 3 8.13-8.17 (in, 1 MIS (ESI) m/lz 583 WO 01/00206 WO 01/1)206PCTIUSOOII8079 To a stirred solution of methyl 4-ti -[4-IN '-(2-bromophenyl)ureidoJ-3-methoxypheny acetyl]-(3S)-pyrrolidinyloxyjbenzoate (620 mg, 1.06 mmol) in TI-F (5 ml]) was added 0.5 N NaOH ml]) and the reaction mixture was heated under reflux for 2.5 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-CHC 3 -TPE to give 184 (421 mg, as a white crystalline powder. MW 568.42 mp 173-175*C; 'H-NMR (DMSO-d) 2.06-2.28 (in, 2 3.56-3.65 (in, 4 3.71-3.88 (series of s and in, total 5 5.12 and 5.20 (each mn, total 1 6.74-6.78 (in, 1 6.89-7.05 (in, 4 7.3 1-7.34 (in, 1 7.59-7.61 (in, 1 7.88-7.98 (in, 4 8.73-8.74 (in, 1 8.9 1-8.93 (in, 1 12.67 (br s, 1 MS (ESI) m/z 569 Ana. Calcd for C 2
,H
2 ,BrN 3
O
6 C, 57.05; H, 4.6 1; N, 7.39; Br, 14.06. Found: C, 57.57; H, 5.12; N, 6.81; Br, 13.96.
Example 177 1-[3-inethoxy-4-[N'-(2-methylphenyl)ueidolphenylacetylj-(3R)-pyrrolidinyloxylbelzoic acid H H 6Me185 To a cooled (0 0 stirred solution of methyl 4-hydroxybenzoate (1.78 g, 11.7 inmol), l-benzyl- (3S)-pyrrolidinol (2.07 g, 11.7 iniol), and Phy 3 (3.68 g, 14.0 inmol) 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 midxture was evaporated. The residue was purified by column chromatography on silica-gel with n-hexane-EtOAc 1, v/v) as eluent to give methyl 1 benzyl-(3R)-pyrrolidinyloxylbenzoate (3.56 g, as a pale yellow oil. 'H-NMR (CDCI3) 6 1.95-2.03 (in, 1 2.28-2.37 (in, 1 2.57-2.63 (in, 1 2.73-2.81 (in, 2 2.97-3.0 1 (in, 1 3.63-3.72 (in, 2 3.87 3 4.85-4.90 (in, 1 6.83-6.85 (in, 2 7.27-7.34 (in, 5 H), 7.94-7.97 (in, 2 MS (ESI) m/~z 312 A solution of methyl 4-[l-benzyl-(3R)-pyrrolidinyloxy]benzoate (3.56 g, 11.4 nimol) in MeOH ml) was hydrogenated over Pd(OH) 2 /C (0.72 g, 20 iw%) overnight. The reaction mixture was filtered to remove the catalyst and the solution was evaporated to give methyl pyfrolidinyloxy~benzoate (2.53 g, as a pale yellow oil. 'H-NMR (CDCI 3 6 1.94-2.18 (in, 3 2.91-2.97 (in, 1 3.04-3.09 (in, 1 3. 16-3.22 (in, 2 3.88 3 4.88-4.91 (in, 1 H), 6.86-6.89 (in, 2 7-97-7.99 (in, 2 MIS (ESI) mkz 263 [WM+1+41, (+MeCN)].
WO 01/00206 WO 0100206PCT/USOO/18079 A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (460 mg, 1.46 rumol), methyl 4-[3R)-pyrrolidinyloxylbenzoate (324 mg, 1.46 mmol), EDC HCI (337 mg, 1.76 nunol), HOBt (237 mg, 1.75 mmol), and Et 3 N (245 ml, 1.76 minol) in THF (10 nil) was stirred at room temperature overnight The reaction mixture was diluted with H,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 50: 1, v/v) as eluent to give methyl 4-[l -[3-methoxy-4-[N'-(2-methylphenyl)ureidojphenylacetylj-(3R)-pyrrolidinyloxyJ benzoate (583 mg, as a colorless oil. 'H-NMR (CDCI 3 8 2.06-2.23 (in, 2 2.30 3 H), 3.58-3.89 (series of s and m, total 12 4.95-4.99 (in, 1 6.75-7.00 (in, 4 7.13-7.30 (in, 7.52-7.57 (in, 1 7.96-8.03 (in, 3 MS (ESI) m/lz 5 18 To a stirred solution of methyl 4-[l-[3-inethoxy-4-[N'-(2-inethylphenyl)ureido]phenylacetylj-(3R)pyrrolidinyloxylbenzoate (583 ing, 1. 13 minol) in THF (5 ml) was added 0.5 N NaOH (5 nil) and the reaction mixture was heated under reflux for 3 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 recistallized from MeOH-CHC 3 -Et 2 O to give 185 (297 mng, as a white crystalline powder. MW 503.55 mp 158-162 0 C; 'H-NMvR (DMSO-d) 5 2.08-2.31 (series of s and mn, total 5 3.54-3.89 (series of m, total 9 5. 11 and 5.20 (each M, total 1 H), 6.72-7.17 (series of m, total 6 7.78-7.80 (in, 1 7.87-7.90 (in, 2 7.98-8.02 (in, 2 H), 8.46-8.47 (in, 1 8.55-8.57 (mn,I 12.66 (hr s, 1 MS (ESI) nz/z 504 Anal. Calcd for C 2 sH~qN 3
O
6 1l/4HO: C, 66.19; H, 5.85; N, 8.27. Found: C, 66.12; H, 5.77; N, 8.2 1.
Example 178 4-[1 '-(2-chlorophenyl)ureidoJ-3-methoxyphenylacetylJ-(3R)-pyrrolidinyloxy~benzoic acid
,P-CJ--COOH
1H H Me 186 A mixture of 4-[N'-(2-chlorophenyl)ureidoj-3-methoxypheniylacetic acid (498 mng, 1.49 inmol), methyl 4+[3R)-pyrrolidinyloxy]benzoate (329 ing, 1.49 inmol), EDCHCI (342 mg, 1.78 minol), HOBt (241 mg, 1.78 iniol), and Et 3 N (250 ml, 1.79 minol) in THE (10 ml) was stirred at room temperature overnight. The reaction 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 50: 1, v/v) as eluent to give methyl 1-[4-IN '(2-chorophenyl)ureidoJ-3-methoxyphenylacetyll-(3R)-pyrrolidinyloxy WO 01/00206 WO 0100206PCT/USOO/18079 ]benzoate (561 mg, 70%) as a white form. 'H-NMR (CDCI 3 8 2.05-2.34 (in, 2 3.59-4.07 (series of s and mn, total 12 4.97-5.02 (in, 1 6.75-6.86 (in, 4 6.94-7.00 (in, 1 7.22- 7.33 (in, 2H), 7.59-7.66 (in, 2H), 7.92-7.99 (in, 3H), 8. 19-8.22 (in, 114); MS (ESI) mz 538 1).
To a stirred solution of methyl 4-[I-[4-IN'-(2-chlorophenyl)ureidol-3-methoxypheny acetyl]-(3R)-pyrrolidinyloxylbenzoate (561 mg, 1.04 mmol) in TI{F (5 ml) was added 0.5 N NaOH 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 under a reduced pressure. The crude solid was recrystallized from MeOH-CHC 3 -Et 2 O to give 186 (361 mng, 66%) as a white crystalline powder. MW 523.97 mp 193-194*C; 'H-NMR (DMSO-d) 8 2.06-2.28 (im, 2 3.58-3.62 (in, 4 3.7 1-3.76 (in, 1 3.83-3.89 (series of s and mn, total 4 5.12 and 5.20 (each mn, total 1 6.74-6.78 (in, I 6.90-6.91 1 7.0 1-7.05 (in, 3 H), 7.27-7.30 (in, 1 7.43-7.45 (in, 1 7.88-7.98 (in, 3 8.08-8. 10 (in, 1 8.89-8.96 (in, 2 12.67 (br s, 1 MS (ESI)mk~ 524 AnaI. Calcd for C- 2 7H26CN 3
O
6 l1/4H 2 O: 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.
Example 179 4-1-4-[N '-(2-broinophenyl)ureido]-3-methoxyphenylacetyl]-(3R)-pyrrolidinyloxylbelzoic acid 'P-Cj-OOH r H 6Me187 A mixture of 4-[N'-(2-broinophenyl)ureido]-3-inethoxyphenylacetic acid (460 ing, 1.21 inmol), methyl 4-[(3R)-pyrrolidinyloxy~benzoate (269 mg, 1.21 ininol), EDC HCI (280 mg, 1.46 minol), HOBt (197 mg, 1.46 iniol), and Et 3 N (205 ml, 1.47 iniol) in THF (10 ml) was stiffed at room temperature overnight. The reaction mixture was diluted with H 2 0, and extracted with EtOAc.
The extract was washed with brine, dried over Na 2 SO,, 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 1-[4-[N'-(2-bromophenyl)ureido-3-methoxyphenylacetyll-(3R)-pyrrolidinyloxyI benzoate (555 mg, as a white foam. 'H-NM4R (CDCI 3 8 2.05-2.3 3 (mn, 2 3.604.07 (series of s and in, total 12 4.96-5.01 (in, 1 6.76-6.93 (in, 5 7.28-7.30 (in, 1 7.48- 7.58 (in, 3 7.92-7.99 (in, 3 8.12-8.16 (in, 1 MS (ESI) m/z 582 To a stirred solution of methyl 4-Il-[4-[N'-(2-broinophenyl)ureidoJ-3-inethoxyphenylacetylJ-(3R)pyrrolidinyloxy]benzoate (555 mg, 0.95 innol) in THF (5 ml) was added 0.5 N NaOH (5 ml]) and 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-CHCI 3 -Et 2 O to give 187 (330 mg, 61%) as a white crystalline powder. MW 568.42 mp 175-177°C; 'H-NMR (DMSO-d) 5 1.99-2.27 2 3.56-3.63 4 3.71-3.76 1 3.83-3.89 (series of s and m, 4 5.12 and 5.20 (each m, total 1 6.74-6.78 1 6.89-7.05 4 7.30-7.35 1 7.59-7.61 1 H), 7.88-7.98 4 8.74-8.76 1 8.92-8.95 1 12.68 (br s, 1 MS (ESI) m/z 569 Anal. Calcd for C 27
H
2 ,BrN30 6 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.
Example 180 0 2
H
H H3 HNg 188 1 gram of Tentagel PHB resin (loading 0.29 mmol/gm) was taken up in DMF 25 mL and Fmoc-(4-carboxymethyl)-piperidine (318 mg, 0.87 mmol) was added. The resin was shaken for min then DIC (220 mg, 0.27 mL, 1.74 mmol) and DMAP (106 mg, 0.87 mmol) were added and the resin was shaken for 24 hr.
The resin was drained and washed with DMF CH 3 OH (3x) and CH 2 CI (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was taken up in piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x), CH3OH (3x) and CH 2 CI, (3x) then dried under vacuum. The resin gave a positive bromophenylblue test. The resin was taken up in 25 mL of DMF and Fmoc-L- morpholine-2carboxylic 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 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 piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x),
CH
3 OH (3x) and CH 2 CI, (3x) then dried under vacuum. The resin gave a positive 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 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
CI
2 and shaken for 4 hr. The resin was drained and the eluate collected.
The resin was taken up in fresh CH 2 CI, 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.
Example 181
OA
H H 3 189 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 min then DIC (220 mg, 0.27 mL, 1.74 mmol) and DMAP (106 mg, 0.87 mmol) were added and the resin was shaken for 24 hr.
The resin was drained and washed with DMF CH 3 OH (3x) and CH 2 Cl2 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was taken up in piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x), (3x) and CH 2 Cl 2 (3x) then dried under vacuum. The resin gave a positive 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 CH 3 OH (3x) and CH 2 CIl (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 CI, (3x) then dried under vacuum. The resin gave a positive 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 CH 3 OH (3x) and CH 2 C1I (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was then taken up in 90% TFA in CHCI 2 and shaken for 4 hr. The resin was drained and the eluate collected.
The resin was taken up in fresh CH 2 CIl 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.
Example 182 H 3 0 U190 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 min DIC (220 mg, 0.27 mL, 1.74 mmol) and DMAP (106 mg, 0.87 mmol) were added and the resin was shaken for 24 hr.
The resin was drained and washed with DMF 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 piperidine in DMF and shaken for 4 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 positive 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 CH 3 OH (3x) and CHCI, (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
CI
2 (3x) then dried under vacuum. The resin gave a positive 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 CH 3 OH (3x) and CH 2
CI
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 Cl, and shaken for 4 hr. The resin was drained and the eluate collected.
The resin was taken up in fresh CH 2
CI
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.
Example 183 0rrr C 0 2 H
COPH
Y UCO 2
H
HN 191 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 shaken for 24 hr.
The resin was drained and washed with DMF 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 piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x),
CH
3 OH (3x) and CH 2 CIl (3x) then dried under vacuum. The resin gave a positive 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 CH 3 OH (3x) and CHCI, (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 CI, (3x) then dried under vacuum. The resin gave a positive 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 CH 3 OH (3x) and CH 2
CI
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
CI
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.
Example 184 H H3 192 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 min then DIC (220 mg, 0.27 mL, 1.74 mmol) and DMAP (106 mg, 0.87 mmol) were added and the resin was shaken for 24 hr.
The resin was drained and washed with DMF 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 piperidine in DMF and shaken for 4 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 positive 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 CH 3 OH (3x) and CH 2 C1, (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), (3x) and CH 2 C12 (3x) then dried under vacuum. The resin gave a positive WO 01/00206 PCT/US00/18079 bromophenylblue test. The resin was taken up in 25 mL of DMF and Fmoc-3-amino-2-oxo-lpyrrolidineacetate (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.
The resin was drained and washed with DMF CH 3 OH (3x) and CH 2 CI, (3x) then dried under vacuum. The resin gave a negative bromophenylblue test. The resin was taken up in piperidine in DMF and shaken for 4 hr. The resin was drained and washed with DMF (3x), (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 CI1 and o-tolyl isocyanate (193 mg, 1.45 mmol, 0.18 mL) was added. The resin was shaken for 24 hr.
The resin was drained and washed with DMF CH 3 OH (3x) and CH 2 CIl (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, 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 69 mg 192.
Example 185 IH H 193 In a 250 mL round-bottomed flask was placed o-tolylisothiocyanate (10.0 g, 67.1 mmol) in 150 mL of CH 2
CI
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.
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 dissolved in methanol (100 mL) and piperidine (8.7 g, 102 mmol) at room temp. The mixture was WO 01/00206 WO 0100206PCT/USOO/18079 stirred overnight and then concentrated in vacua 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 H H194 Methyl-4-aminophenylacetate (4.0 g, 25 mmol) was dissolved in CH 2 CI (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 coaled to room temp. The solution was poured in to 1N HCI and then extracted 3x with EtOAc, dried over MgSO,, and concentrated in vacua to afford 5.2 g (67% yield) of thiourea methyl ester. The ester was saponified using LiOH to give 5.0 g of the desired 4-(otolylthioureido)phenylacetic acid 194.
Example 187
H
H
H H-(?OOH Me H Me19 A solution of ethyl 4-fl -itoy4-A-2mtyphnlued~heyaey](S 3R, 4R)-3,4-isopropylidenedioxy-2-pyrrolidiniylcarbonylJ-1-piperazinylacetate (1.27 g, 1.99 mrnol) in sat. HCl (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 CHC1 3 MeOH (4 1, The extracts were washed with brine, dried over MgSO 4 and concentrated to dryness.
Chromatography of the residue with CHC1 3 MeOH (5 vfv) as eluent gave ethyl 4-[11[3methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetylJ-(2S, 3R, 4R)-3 ,4-dihydroxy-2-pyrrolidinyl carbonylJ-1-piperazinylacetate (990 mg, 83 as a yellow amorphous solid. IR (KBr) 3338, 2937, 2830, 1743, 1625, 1600, 1532, 1454 cm-; 'H-NMR (CDCI 3 d 1.25 J =7.1 Hz, 3H), 2.20 3H), 2.46 -2.56 (in, 4H), 3.15 2H), 3.40 -3.72 (in, 9H), 3.62 3H), 4.01 4.08 (mn, 2H), 4.16 J =7.1 Hz, 2H), 4.22 (mn, 1Hf), 4. 72 J= 2.9 Hz, 111), 6.68 (d,J 7.6 Hz, 11H), 6.72 (s, 1 7.06 J 7.6 Hz, I1H), 7.15 7.18 (in, 31-H), 7.52 7.56 (in, 2H), 7.90 (di, J 8.3 Hz, IlH); MIS (FAB) m/z 598 (Mr+ 1).
To a solution of ethyl 4-[I -ehx--A-2-ehlhnluedopeyaeyj(S 3R, 4R)-3,4-dihydroxy-2-pyrrolidinylcarbonyl]-l-piperazinylacetate (870 mg, 1.46 inmol) in THF mL) was added 0.25N NaOH (7.00 mL, 1.75 nunol). After being stirred at room temp. for WO 01/00206 PCT/US00/18079 hr, the reaction mixture was concentrated. The residue was diluted with water and neutralized with IN HCI at 0 OC. The mixture was concentrated and purified by ion-exchanged resin 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 3H), 2.38 1H), 2.42 2.58 (m, 2H), 2.64 1H), 3.01 2H), 3.13 3.71 8H), 3.88 3H), 3.89 1H), 4.05 1H), 4.
58 J 3.2 Hz, 1H), 6.76 (dd, J= 8.3, 1.5 Hz, 1H), 6.91 6.95 2H), 7.10 7.16 2H), 7.79 J 8.3 Hz, 1H), 8.00 J 8.3 Hz, 1H), 8.49 1H), 8.57 1H); MS (FAB) m/z 570 Anal. Calcd for C 28
H
3 ,NsO,-2.75H 2 0: C, 54.32; H, 6.59; N, 11.31. Found: C, 54.07; H, 6.11; N, 11.00.
Example 188 S 196 To a suspension of 2-amino-4-thiazoleacetic acid (4 g, 25 mmol) in 1:1 CH 2 CI: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 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 dioxane-water and solid Na 2
CO
3 was added till the pH was 8-9. BocO (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,. The solution was filtered and the solvent was removed under reduced pressure. The product was purified by flash chromatography. (4:1 hexane-ethyl acetate) Yield 4.39g 197.
BocN "Br CH3 198 The Boc-protected benzyl amine (2.00g, 6.99mmole) was dissolved in dry THF under argon. The reaction was cooled to minus 78° 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 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 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 hexaneethyl acetate) Yield 1.68g 198.
Boc CO 2
CH
3
CH
3 CH3 199 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 (0.51g 6.16mmole), P(otolyl) 3 (0.51g, 6.16mmole), and Pd(OAc) 2 (0.25g, 1.12mmole) The tube was flushed with argon for min then methyl acrylate (0.53g, 0.53mmole, 0.55mL) was added. The tube was sealed and heated to 135° C for 24 hr. The reaction was cooled to 0°C and the tube was slowly 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,. 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
CH
3 U CH 3 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(otolyl) 3 (0.20, 0.66mmole), and Pd(OAc) 2 (0.15g, 0.66mmole). The tube was flushed with argon for 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 0° C and the tube was slowly 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 1.01g 200.
Boc' -N ,,C02CH 3 CH3 I 201 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 The a-methyl-, P-unsaturated ester (1.01g, 3.16mmole) was placed in a Paar vessel and 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 996.4 1mg 202.
3H
H
Ny N 0 O
CO
2
CH
3
CH
3 203 The Boc ester (304mg, 1.04mmole) was taken up in CH 2 Cl 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 removed under reduced pressure. The residue was taken up in CH 2
CI
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 solvent was removed under reduced pressure. The product was isolated by flash chromatography.
(ethyl acetate) Yield 380.32mg 203.
CH3 J~ N CO 2
H
ICH
3 204 The ester (380.32mg, 0.80mmole) was taken up in ethanol-water and NaOH was added. 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 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.
Example 190 CH3 3H H N N N SN C0 2
CH
3
CH
3 1
CH
3 205 WO 01/00206 PCT/US00/18079 The Boc ester (209.60, 0.65mmole) was taken up in CH 2 Cl, 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 removed under reduced pressure. The residue was taken up in CH 2 CIl-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 HCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO,. The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (ethyl acetate) Yield 237.8mg 205.
CH H H N
CO
2
H
H
3 1
CH
3 206 The ester (237.8mg, 0.49mmole) was taken up in ethanol-water and NaOH added.
The reaction was 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,. 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 2N O 207 Thel,2,3,4- tetrahydroisoquinoline (12.20g, 91.60mmole) was taken up in H 2
SO
4 and cooled to minus 100 C. Concentrated HNO, (9.0mL) was slowly added to the solution while maintaining the internal temp at minus 10° 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 added to ice and the aqueous solution was basified with NH 1 OH. The aqueous layer was extracted 4 times with CHCI 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 I-,CO 2
CF
208 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 the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were dried over NaSO, 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
CO
2
CH
3 209 The above ester (702mg, 2.81mmole)was placed in a Paar vessel and dissolved in 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.
0 N N CO2CH3 S N NN'
CH
3
OCH
3 210 The aniline (587.0mg, 2.66mmole) was taken up in dry CH 2 CIl and pyridine under argon.
The reaction was cooled to 0°C. A CH 2 ClI 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 with sat. NaHCO 3 water, brine then dried over MgSO. 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
0 N N CO2H
CH
3
OCH
3 211 The methyl ester (618.36mg, 1.20mmole) above was taken up in THF-HO0 and LiOH (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.
WO 01/00206 PCT/US00/18079 Example 192 0 2 N N OH212 212 The 3-nitro-phenyl propionic acid (1.00g, 5.12mmole) was taken up in dry THF under argon. The reaction was cooled to 0° C and BH 3 -THF (1.OM, 15.37mmole, 15.37 mL) was added over 10 min. The reaction was stirred at 0° C for 1 hr then slowly quenched with water. The solution was slowly warmed to room temp then poured into IN HC1. 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) 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
CI
2 In a second round bottom flask (COC1) 2 (700.65mg, 5.52mmole, 0.48 mL) was added to dry CH 2 Cl2 under argon. The (COC1),-CH 2 CI, solution was then cooled to minus 60° C and DMSO (862.56mg, 11.4mmole, 0.78 mL) was slowly added. The reaction was stirred at minus 600 C for 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 sat NaHCO 3 water, brine then dried over MgSO,. 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 1 CO 2
CH
2
CH
3 S CH 3 214 In a round bottom flask NaH (132.48mg, 5.52mmole) was slurried in dry THF under argon. Triethyl 2-phosphonopropionate (1.3 1g, 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 washed with sat. NaHCO 3 water, brine then dried over MgSO,. 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.
WO 01/00206 PCT/US00/18079
H
2 N. CO 2
CH
2
CH
3
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 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
0 ,NY C 0 2 C H 2
CH
3 H H
CH
3
CH
3
CH
3 216 The above aniline (850.0mg, 3.61mmole) was taken up in dry CH 2
CI
2 and pyridine under argon. The reaction was cooled to 0° C. A CH 2
CI
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,. The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography (ethyl acetate) Yield 576.0mg 216.
H
0 N C02H LN^N^ 0 CH 3 H H
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 and then poured into IN HCI. 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 ON O H 218 One gram of Wang resin (tentagel S-PHB 0.3mmole loading) was suspended in a solution WO 01/00206 PCT/US00/18079 of 3-(Fmoc-amino)phenylpropionic acid (361.31mg, 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
CI
2 The mixture was shaken for 20 hr and drained. The resin 218 was washed with DMF, MeOH, CHCl 2 and dried under reduced pressure.
H H N N 0 0 N:P'N HY HOI1~- N CH1 OCH
CH
3 219 To the above resin (500mg, 0.15mmole) was added a solution of piperidine-DMF v/v, 4 mL) and the mixture was shaken for 4 hr. The resin was washed with DMF, MeOH,
CH
2 CI,. 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 was added. The mixture was then shaken for 12 hr. The resin was drained and taken up in MeOH-1% AcOH and NaCNBH3 (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
CI
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 mL) were added. The resin was then shaken for 24 hr then drained. The resin was washed with DMF, MeOH, CH 2
CI
2 To the resin was added a solution of TFA in CHC1, (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, EtO was added to the residue and the solid was collected to afford 15mg 219 as a crystalline solid Example 194
OH
K OH 0N
CO
2
H
-N N
O
CH
3 H H CH 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 shaken for 14 hr. The resin was drained and washed with DMF CH 3 OH (3x) and CH 2 Cl 2 (3x) then dried under vacuum.
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 CH 3 OH (3x) and CH 2 C1, (3x) then dried under vacuum. The resin gave a positive bromophenylblue test.
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 CHO3H (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 elutant collected. The resin was taken up in fresh CH 2
CI
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.
Example 195 HN CO 2
H
H
3 CO H CH3 HNoN z 0HN 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 resin was shaken for 24 hr. The resin was drained and washed with DMF CH 3 OH (3x) and
CH
2 C1I (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 CH 3 OH (3x) and CH 2
CI
2 (3x) then dried under vacuum. The resin gave a positive bromophenylblue test.
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 WO 01/00206 PCT/US00/18079 hr. The resin was drained and washed with DMF CHO3H (3x) and CH 2 CI1 (3x) then dried under vacuum. The resin gave a negative bromophenylblue test The resin was then taken up in 90% TFA in CH 2 ClI and shaken for 4 hr. The resin was drained and the elutant collected. The resin was taken up in fresh CH 2 CI, 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 66mg 221.
Example 196 N
NN
H3 H H CH3
OH
o0 H 222 To a solution of oxalyl chloride (3.8 g, 30 mmol) in CH 2 CI1 (100 mL) was added DMSO (2.4 g, 31 mmol) dropwise over 30 min at minus 78° C. To this solution was added N-Bocprolinol (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 HCI, extracted 3x with EtOAc, dried, and concentrated in vacuo to afford the crude prolinal which was chromatographed 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 780 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 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 psi for 4 hr, the resulting alkane was used without purification. The Boc group was removed by the addition of 1:1 TFA/CH 2
CI
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
CI
2 (100 mL) was generated.
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-otolylureido-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 MeOH/ CH 2 C1 2 to 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 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
CH
3 H3C .N
OH
0 H3CO H CH3 HN N 223 0 U223 To a solution of methyl 8-aminooctanoate (2.0 g, 12 mmol) in 1:1 dioxane:water (100 mL) was added Boc anhydride (2.8 g, 13 mmol) and K 2
CO
3 (10 This solution was allowed to stir at room temp for 14 hr. The reaction was then poured onto 1 N HCI, extracted 3x with EtOAc, dried, and concentrated in vacuo. The crude carbamate was chromatographed 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 addition of LiHMDS (25 mL of a 2.0M soln., 50 mmol) at minus 78 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 HCI, 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 product The Boc group was removed by the addition of 1:1 TFA/CH 2
CI
2 at room temp. The reaction was stirred for 2 hr and the solvent was removed in vacuo. The crude amine 900 mg was WO 01/00206 PCT/USOO/18079 used without further purification.
A solution of the above free amine (900 mg, 4.5 mmol) in CH 2 CI, (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 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 MeOH/
CH
2
CI
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-HO0 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 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 HH H H3 224 To a suspension of 4-aminophenylacetic acid (10 g, 66 mmol) in 1:1 CH 2 Cl 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 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 224.
Example 199 0H3 H 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 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.
WO 01/00206 PCT/US00/18079 Example 200 NC r 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 reaction was heated to 500 C for 2 hr. The solution was cooled to room temperature and poured into INHCI. 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.
O
OCrN Br
H
3
C
113 227 3-Bromo-4-benzyloxybenzonitrile (1.50g, 5.21mmol) was taken up in dry THF under argon and the solution was cooled to 00 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 0° C and methanol was slowly added. When no more gas evolution was observed the solution was warmed to room temp and excess IN NaOH solution was added. Boc 2 O (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,. The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (7:1 ethyl acetate-hexane) Yield 1.80g 227 H3 Br
H
3 C CH 3 228 The Boc-protected benzyl amine (1.80g, 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 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 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 hexaneethyl acetate) Yield 1.70g 228.
H
3 C CH 3 CH3 K) 229 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 135°C for 24 hr. The reaction was cooled to 0° C and the tube was slowly 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,. 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 H3CF O 2
CH
3 H3C yH 230 The unsaturated ester (307.40mg, 0.75 mmol) was taken up in CH 2
CI
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,. 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 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 MgSO4. The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (ethyl acetate) Yield 296.30mg 230 WO 01/00206 PCT/US00/18079 u 231 The unsaturated ester (296.30mg, 0.49 mmnol) was taken up in EtOAc and Pd/C 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 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 H3c .s cy 2
H
HC H 9H3 232 The ester (233.00mg, 0.45mmol) was taken up in THF-H 2 0 and LiOH (94.41mg, 2.25mmol) was added. The reaction mixture was stirred at room temp 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,. The solution was filtered and the solvent was removed under reduced pressure. The product was washed with ether-hexane and dried under high vacuum. Yield 211.58mg 232 Example 201
H
3
C,
H3C H CH 3 233 The carboxylic acid (65.00mg, 0.13 mmol) was taken up in benzene and paratoluenesulfonic 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.
NaHCO3. 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 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 Br,02Et 234 5-Bromonicotinic acid (5.15g, 25.49 mmol) was taken up in EtOH and H 2
SO
4 (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
O.
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 234 235 The ethyl 5-Bromonicotinate (5.40g, 23.47 mmol) was taken up in 95% EtOH and NaBH, (8.31g, 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
CH
2 C1I. 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"Qci HCI 236 The benzyl alcohol (2.12g, 11.28 mmol) was taken up EtO and HCI was bubbled 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 SOCI 2 and the mixture was heated to reflux for 1.5 hr. The solution was cooled to room temp and EtO2 was added to precipitate the product. The solid was collected by filtration, washed with EtO2 and dried under vacuum. Yield 2.42g 236 Br. CH3 HCI 237 The benzyl chloride (2.42g, 9.96 mmol) was added over 1 hr to CH 3 NH, (75.9mL, 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, filtered and the solvent was removed under reduced pressure. Yield 1.19g 237 WO 01/00206 PCT/US00/18079
H
3 C, CH 3
H
3
H
3 Br 238 The 3-bromo-5-(N-methyl -aminomethyl)-pyridine(l. 19g, 5.01 mmol) was taken up in DMF and triethylamine (0.90g, 1.24mL, 8.89 mmol was added. Boc 2 0 (1.55g, 7.10 mmol) was added and the reaction mixture was stirred at room temp for 48 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 (2% methanol-CH 2 Cl 2 Yield 1.6g 238 0 d C 239 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 50° C for 24 hr. The solution was poured into IN HCI 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 solvent was removed carefully under reduced pressure. The product was isolated by flash chromatography ether-pentane) Yield 6.93g 239 H3C H 3
H
3
C\
0 H 3' 0 CH3 240 In a pressure tube was placed the (700.00mg, 2.33 mmol). The tube was then charged with DMF, triethylamine (260.05mg, 2.57mmol, 0.36mL), dppp (193.85mg, 0.47 mmol), and Pd(OAc),(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 0° C and the tube was slowly 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,. The solution was filtered and the solvent was removed under reduced pressure.
WO 01/00206 PCT/US00/18079 The product was isolated by flash chromatography. (6:1 hexane-ethyl acetate) Yield 785.23mg 240 Hz
H
C H 3
C
C
3 241 The unsaturated ester (392.61mg, 0.99 mmol was taken up in CHCI 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 was removed under reduced pressure. The residue was taken up in CH 2 ClI-DMF and HOBt 147.53mg, 1.09 mmol), 3-methoxy-4-(N'-phenylureido)phenylacetic acid 342.64mg, 1.09 mmol) 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,. The solution was filtered and the solvent was removed under reduced pressure. The product was isolated by flash chromatography. (ethyl acetate) 363.79mg 241 H3C9
HNO
H
3 C
H
CH
3 242 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 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 WO 01/00206 PCT/US00/18079 Example 203 Ph CH 3 \C CH3 NcPh 243 3-Cyanobenzaldehyde (9.41g, 71.76 mmol) was taken up in ethanol and cooled to 0° C.
The NaBH 4 (2.71g, 71.76 mmol) was added in small portions. The solutions was stirred for min at 0 C then allowed to warm to room temp and stirred for 1 hr. The reaction was slowly poured into 1NHCI and the aqueous layer was extracted 3x with ethyl acetate. The combined organic layers were washed with water, brine then dried over MgSO,. 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 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,. 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 C 3 P>H3
H
3 C CH 3 244 The silyl protected 3-cyanobenzyl alcohol (8.50g, 34.36 mmol) was taken up in ethyl acetate and Boc 2 O (8.25g, 37.79 mmol) was added. Pd/C (l.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 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 11.10g 244 HC H3 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 (42.67mL, 42.67 mmol) was added over 10 min. The reaction was stirred for 1 hr at minus 780 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. ethyl WO 01/00206 PCT/US00/18079 acetate-hexane) Yield 4.7g 245 H3 2 246 The O-silyl-Boc-N-methyl protected benzyl alcohol (4.7g, 9.60 mmol) was taken up in THF and TBAF (14.39mL, 1.OM in THF) at room temp. The solution was stirred for 4 hr. TLC 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
H
3 C H 3 H3 247 HC>.H H247 The N-methyl Boc protected benzyl alcohol (1.00g, 3.98 mmol) was taken up in dry
CH
2 CI, under argon. Triphenylphosphine (1.46g, 5.57 mmol) was added and the solution was cooled to 0° C. Carbon tetrabromide (1.85g, 5.57 mmol) dissolved in dry CH 2 CIl was added over min. The solution was stirred for I h at 0° C then the solvent was removed under reduced pressure. The residue was taken up in EtO and the resulting solid was removed by filtration and the filtrate was collected and the solvent was removed under reduced pressure. The product was isolated by flash chromatography (2%ether-pentane) Yield 1.15g 247
H
3
OCH
3
H
3
CH
3 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 over 15 min and the solution was stirred for Ihr at minus 78° 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 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 isolated by flash chromatography. (3%ethyl acetate-hexane) Yield 414mg 248.
WO 01/00206 PCT/US00/18079 U 249 The Boc ester (121.60mg, 0.36 mmol) was taken up in CH 2 C1I 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, 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 CHCl 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 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
H
HH3
H
3 C H H325 250 The ester (165.20, 0.31 mmol) was taken up in ethanol-water and NaOH was added.
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 120.00mg 250 Example 204 HaC>H 3 251 Butyrolactone (250mg, 2.90 mmol, 223.20mL) was added to LHMDS (2.90mL, 1.OM in hexane) in THF at minus 780 C under argon over 10min. The solution was stirred at minus 78°C for lhr. 3-N-methyl-N-Boc protected benzyl bromide (991.24mg, 2.90 mmol) dissolved in dry 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 1:1 ethyl acetate-hexane) Yield 501.18mg 251 HaC H H3 252 The Boc ester (250.00mg, 0.78 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 washed with water, brine then dried over Na 2 SO,. 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 (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 organic layers were washed with water, brine then dried over MgSO,. 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-[AN-(2-methylphenyl)ureido]phenylacetyl]-N-methylaminoethoxy] benzoic acid
H
3
OOH
S W 6
CH
3 CH3HH YCH 3 253 To a stirred and cooled C) solution of N-methyl ethanolamine (3.10 g, 41.27 mmol), Et 3 N (11.80 mL, 84.66 mmol) in DMF-H20 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 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,, and evaporated. The residue was purified by column chromatography on silica-gel with n-hexane:EtOAc v/v) then CHCI 3 as eluent to give 4.67 g N-methyl-N-(benzyloxy carbonyl)ethanolamine as a colorless oil. 'H-NMR (CDC 3 d WO 01/00206 WO 0100206PCT/USOO/18079 1.82 (bs, 1 3.00 3 3.46 (lbs, 2 3.77 (bs, 2 5,13 2 7.29-7.36 (in, 5 H).
To a stirred solution of ethyl 4-hydoxy-3-methoxybenzoate (2.01 g, 10.25 mmol), Nmethyl-N-(benzyloxycarbonyl)ethanolamine (2.11 g, 10.08 inmol), PPh 3 (3.26 g, 12.43 minol) in THF was added DIAD (2.65 niL, 13.46 mmol) and the reaction mixture was heated under reflux: overnight. The mixture was evaporated, and the residue was subjected to short column chromatography on silica-gel with n-hexane/EtOAc 1, vA') as eluent to give ethyl 3 -methoxy-4- [2-methyl-2-(benzyloxycarbonyl) aminoethoxylbenzoate as a crude product.
To a solution of the crude product (5.20 g, 13.42 nunol) in EtOH (50 mL) was added AcOH (5 ruL) and the solution was hydrogenated over 5% Pd/C for 4 hr. The mixture was filtered 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 NaSO 4 and evaporated. The residue was chromatographed on silica-gel with CHCI,:MeOH (10: 1, v/v) as eluent to give 5 10 mng (2 steps ethyl 3-methoxy-4-(2-methylamino ethoxy) benzoate as a yellow oil. 'H-NMR (CDC 3 d 1.39 3 H, J=7.3 Hz), 1.82 (bs, 1 2.52 3 3.04 2 H, J=5.3 Hz), 3.91 3 4.18 (t, 2 H, J=5.3 Hz), 4.36 2 H, J=7.3 Hz), 6.90 1 H, J=8.3 Hz), 7.55 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-inethylaminoethoxy) benzoate (5 10 mng, 2.01 inmol) in DMF (13 mL) was added pentafluorophenyl ester of 3-methoxy-4-1N-(2methylphenyl) ureido] phenylacetic acid (900 mg, 1.87 mmol) and Et3N (0.420 inL, 3.01 minol), and the resulting mixture was stirred for 2 days. The mixture was diluted with EtOAc, washed with 1 N HCI, 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 CHC1 3 :MeOH (50: 1, v/v) to give 880 mng ethyl 3-methoxy-4-[2-[3-inethoxy-4-[N'-(2-inethylphenyl)ureidolphenylacetyll-Ninethylaminoethoxy] benzoate as a colorless amorphous solid. 'H-NMR (CDCI,) d 1.37-1.4 1 (in, 3 2.28 3 3.03 and 3.18 3 3.56 2 3.65 2 3.75-3.87 (in, 6 4.06-4.24 (2 H, in), 4.33-4.39 (in, 2 6.68-8.08 (series of mn, 12 H).
To a solution of ethyl 3-methoxy-4-[2-[3 -methoxy-IN[A-(2-nethylphenyl)ureidoj phenylacetyl]-N-methlaininoethoxylbenzoate (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 mixture was poured into 1 N HCI (100 niL), and the solid was collected. The crude solid was WO 01/00206 WO 0100206PCT/USOO/18079 recrystallized from MeOH-CHC 3 to give 253 as a white powder. IR (KBr) 1700 cm'; 'H-NNvI (DMSO-d) d 2.25 (s,3 2.50 2 2.91 and 3.12 3 H) 3.53-3.76 (in, 2 3.80 3 H), 3.84 3 4.16-4.21 (in, 2 6.72-8.56 (series of mn, 12 12.68 (bs, 1 MS (FAB) m/lz 522 Anal. Calcd. for CH- 3
N
3
O
7 d H 2 0: C, 62.33; H, 6.16; N, 6.63. Found: C, 62.17; H, 6.05; N, 7.57.
Example 206 4-[[2-[3-methoxy-4-[AN'-(2-methylphenyl)ureidolphenylacetyl~inethylaminolethoxylisophthaic acid 9y~$Vg~COOH
CH
3 11 H OCH 3 254 To a stirred solution of N-methyl-Nr-benzyloxycarbonylethanolaxnine (1.05 g, 5.02 mmol), dimethyl 4-hydroxy isophthalate (1.05 g, 5.00 nunol), Ph 3 P (1.59 g, 6.06 mmol) in THIF (20 niL) was added DIAD (1.28 rnL, 6.50 minol) at room temp. The resulting mixture was then heated under reflux overnight. After cooling to room teinp, the mixture was evaporated. The residue was dissolved in EtOH and added 5% PdIC(200mg). The stirred resulting mixture was hydrogenated for 2 hr at Ilatmn. The mixture was filtered to remove the catalyst, and the filtrate was evaporated.
The residue was purified by column chrmatography on silica-gel with CHCI 3 -MeOH (30: 1, v/v) as eluent to give 480 ing (36% for 2 steps) dimethyl 4-(2-methylaminoethoxy) isophthalate as an oil. 'H-N1MR (CDCl 3 d 1.68 1 2.53 3 3.01-3.04 (in, 2 3.89 3 3.90 3 4.21-4.2 3 (in, 2 7.00 1 H, J=8.8 Hz), 8.14 (dd, I H, J=2.4, 8.8 Hz), 8.50 1 H, J=2.4 Hz); MS (FAB), m/z 268 To a stirred solution of dimethyl 4-(2-methylaminoethoxy)isophthalate (4 10 mig, 1.53 minol) in DMF (13 mL) was added pentafluorophenyl ester of 3-methoxy-4-[N'-(2methylphenyl)ureido] phenylaceic acid (700 mng, 1.46 inmol) and Et 3 N (340 All, 2.44 iniol), and the resulting mixture was stirred overnight. The mixture was diluted with EtOAc, washed with 1 N HCI, sat. NaHCO 3 and brine. The solution was dried over Na 2
SO
4 and evaporated to give 780 ing dimethyl 4- [2-[3-inethoxy-4-[N'-(2-methylphenyl)ureido]phenylacetyllinethylainino ethoxylisophthalate as a crystalline powder. 'H-NMR (CDCI 3 d 2.29 3 3.24 3 3.59 3 3.67-3.68 (Mn 2 3.84 3 3.91 3 3.81-3.86 (in, 2 4.25-4.28 (in, 2 H), 1-8.48 (series of in, 12 MS (FAB) m/lz 564 (Mr+l).
WO 01/00206 WO 0100206PCT/USOO/18079 To a solution of dimethyl 4- [2-[3-methoxy-4-[N'r-(2-methylphenyl)ureidolphenylacetylJ methylaininoethoxy] isophthalate (780 mg, 1. 384 nunol) in THF (30 mL) was added 0. 25 N NaOH niL). The resulting mixture was then heated under reflux overnight. The mixture was poured into ice-i N HCl (200 niL) and the solid was collected. The crude solid was recrystallized from MeOH-CHCI, to give 420 mg 4-[[2-[3-methoxy-4-[N'-(2-methylphenyl)ureido~pheny acetyllmethylaminojethoxyJ isophthalic acid 254 as a white crystalline powder. mp 139- 141 'C IR (KBr) 1700 'H-NMR (DMSO-d6) d 2.94 3 3.18 3 H) 3.62-3.86 (in, total 8 4.24- 4.28 (in, 2 6.74-8.58 (series of m, total 12 12.91 (bs, 1 MIS (FAD) m/z 536 Anal. Calcd. for Q 8 sH~N 3
O
8 -2.5HCL: C, 53.66; H, 5.07; N, 6.70. Found: C, 53.80; H, 4.64; N, 6.70.
Example 207 3-ehx--21-ehx--A-2mtypenlued~hnlctIainehxlezi acid H ~9OOH
OCH
3 PC H H3
CH
3 H 255 To a solution of 2-ethanolamine (5.16 g, 84.48 minol), Et 3 N (23.50 niL, 168.60 niL in dioxane-H 2 0 160 niL) was added dropwise (Boc0 2 0 (23.40 niL, 10 1. 86 inmol) at room temp.
The reaction mixture was stirred for 2 days at room temp. The resulting mixture was diluted with
CHCI
3 washed with 0.5 N HCI, sat. NaHCO 3 and brine. The separated organic layer was dried over Na 2 S504 and evaporated to give 11.86 g N-Boc-2-ethanolamine as an oil. 'H-NMR (CDC1 3 d 1.45 9 3.29-3.3 1 (mn, 2 3.71-3.72 (mn, 2 H).
To a stirred solution of ethyl 4-hydroxy-3-inethoxybenzoate (1.46 g, 7.44 inmol), N-Boc ethanolainine 19 g, 7.38 minol), PPh 3 (2.53 g, 9.65 minol) in THEF (30 niL) was added DIAD (1.90 niL, 9.65 inmol), 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 mQL and TEA (20 niL). 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. NaHC0 3 and extracted with
CHC
3 The extract was washed with brine, dried over Na 2
SO
4 and evaporated to give the 1.61 g for 2 steps) ethyl 3-methoxy-4-(2-antinoethoxy) benzoate as a yellow oil. 'H-NM.R(CDCI 3 d 1.39 3 H, J=7.3 Hz), 3.14-3.17 (in, 2 3.92 3 4.09-4. 11 (mn, 2 4.36 2 H, J=7.3 Hz), 6.89 1 H, J=8.3 Hz), 7.56 1 H, J=2.0 Hz), 7.66 (dd, 1 H, J=2.0, 8.3 Hz).
WO 01/00206 WO 0100206PCT/USOO/18079 To a stirred solution of ethyl 3-methoxy-4-(2-aminoetboxy)benzoate (250mg, 1.04 mmol) and pentafluorophenyl ester of 3-methoxy-4-[N'-(2-methylphenyl)ureidojphenylacetic acid (500 mg, 1.04 mmol) was added Et 3 N (2 10 p1, 3.01 nunol), and the resulting mixture was stiffed for 2 days. 0.25 N NaQH (20 mL) and THF (20 mL) was added to the mixture and the resulting mixture was heated under reflux overnight. After cooling, the mixture was evaporated and the residue was acidified by the addition of 1 N HCI. The mixture was extracted with CHCI3, and the extract was washed with brine, dried over Na 2 SO,, and evaporated. The obtained crude solid was recrystallized from CHC1 3 to give 110 mng (20% for 2 steps) 3-methoxy-4-[2-[3-methoxy-4-[N'-(2-methylphenyl) ureidolphenylacetyllaminoethoxyI benzoic acid 255 as a white crystalline powder. mp 180-181 0 C; IR (KBr) 1687 cm-1 'H-NMR (DMSO-d 6 d 2.24 3 3.37 2 3.38 2 3.4 1- 3.50 (in, 2 3.81 3 3.83 3 4.06-4.08 (in, 2 6.76-8.55 (series of mn, total 12 H); MS (FAB) m/~z 508 Anal. Calcd for C 2
,HN
3 Oj l/2H 2 0: C, 62.78; H, 5.85; N, 8.13.
Found: C, 62.46; H, 5.69; N, 8.03.
Example 208 3-methoxy-4-[2-[3-methoxy-4-IIN'-(2-methylphenyl)ureido~phenylacetyllethylamiinoethoxy benzoic acid Q-N-Z~r'
OCH
3
CH
3 H H 6H 3 256 To a cooled (00 C) solution of ethyl 3-inethoxy-4-(2-amidnoethoxy)benzoate (1.93 g, 8.07 nunol) and Et 3 N (2.00 mL, 14.35 mmol) was added TFAA (1.35 ML, 9.56 mmol) and the resulting 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 HCI, H 2 0, and brine. The extract was dried over NaSO, and evaporated to give 1.22 g ethyl 3-methoxy-4-(2-N-trifluoroacetanidoethoxy) benzoate as an oil. 'H-NNvIR (CDC 3 d 1.39 3 H, J=7.3 Hz), 3.77-3.81 (in, 2 3.92 3 H), 4.18-4.20 (in, 2 4.37 2 H, .1=7.3 Hz), 6.92 1 H, Hz), 7.59 1 H, J=2.0 Hz), 7.67 (dd, 1 H, J=2.0, 8.7 Hz); MS (FA.B) m/~z 3 35 290 (MW-OEt).
To a stirred solution of ethyl 3 -iethoxy-4-(2-N-trifluoroacetamidoethoxy)benzoate (1.20 g, 3.58 minol) in DM4F (15 mL) was added K 2 C0 3 (0.98 g, 7.09 mmol) and Etl (0.43 niL, 5.38 nunol) 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 HCI, brine, and dried over Na 2
SO
4 The solvent was evaporated and the residue was purified by column chromatography on silica-gel with n- WO 01/00206 WO 0100206PCT/LJSOO/18079 hexane-EtOAc 1, v/v) as eluent to give 990 mg ethyl 3-methoxy-4-[2-(N-ethyl-Ntrifluoroacetamido)ethoxybenzoate as a yellow crystalline solid. 'H-NMR (CDCI 3 d 1.28-1.3 1 (in, 3 1.37-1.40 (in, 3 3.64-3.69 (in, 2 3.8 1-3.84 (mn, 2 3.92 3 4.27-4.30 (mn, 2 4.34-4.39 (in, 2 6.89 1 H, J=8.3 Hz), 7.55 1 H, J=2.0 Hz), 7.66 (dd, 1 H, J=2.0, 8.3 Hz); MIS (FAB) m/z 364 To a stirred solution of ethyl 3 -methoxy-4-[2-(N-ethyl-NV-trifluoroacetamido) ethoxybenzoate (990 ing, 2.73 minol) in THF-MeOH-H 2 0 1: 1, v/v, 20 inL) 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.
NaHCO 3 brine, dried over Na 2
SO
4 and evaporated to give 800 mg ethyl 3-methoxy-4-(2ethylanunoethoxy)benzoate as an oil. IH-NMR (CDCI 3 d 1. 15 3 H, J=7.3 Hz), 1.39 3 H,J=7.3 Hz), 1. 76 (bs, 11H), 2.74 2 H, J=7.3 Hz), 3.08 2 H, J=5.4 Hz), 3.91 3 4.18 (t, 2 H, J=5.4 Hz), 4.36 2 H, J"'7.3 Hz), 6.90 1 H, J=8.3 Hz), 7.55 1 H, J=2.0 Hz), 7.66 (dd, 1 H, J-2.0, 8.3 Hz); MIS (FAB) m/z 268 1).
To a stirred solution of ethyl 3-inethoxy-4-(2-ethylaminoethoxy)benzoate (290 mng, 1.08 nunol) and pentafluorophenyl ester of 3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetic acid (502 mg, 1.05 mmol) in DMF (7 mL) was added Et 3 N (250 juI, 1.79 innol), and the resulting mixture was stirfed overnight. The mixture was diluted with EtOAc, washed with 0.5 N HCI, brine, and dried over NaSO,. The solvent was evaporated and the residue was purified by column chromatography on silica-gel with CHCl 3 -MeOH (40: 1, v/v) as an eluent to give 550 mng (93%) ethyl 3-methoxy-4-[2-[3-methoxy-4-[N' -(2-methylphenyl)ureido] phenylacetyllethylanunoethoxyj benzoate as an amorphous solid. 'H-NMR (CDCI 3 )d 1.11-1.18(in, 3H), 1.37-1.41(in, 3H), 2.30 3 3.47-3.53 (in, 2 3.61-3.75 (in, 7 3.84 3 4.03-4.27 (in, 2 4.33-4.39 (in, 2 6.34-8.07 (series of in, total 12 H).
To a solution of ethyl 3 -iethoxy-4-[2-[3-inethoxy-4-[N'-(2-methylphenyl)ureido~pheny acetyl]ethylaminoethoxylbenzoate (550 mng, 0.98 inmol) in THF (15 mL) was added 0.25 N NaOH niL). The resulting mixture was then heated under reflux for 2 days. The mixture was poured into 1 N HCl and the solid was collected. The crude solid was recrystallized from EtOH-CHCl 3 to give 182 mng (359/) 3-methoxy-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetylJ ethylaminoethoxylbeuzoic acid 256 as a white crystalline powder. mp 115-118 0 C; IR (KBr) 1707 WO 01/00206 WO 0100206PCT/USOO/18079 cnf' 'H-NMR (DMSO-d) d 1.02-1.12 (in, 3 2.25 3 2.50 2 3.35-3.89 (in, 10 H), 4.11-4.16 (in, 2H), 6.71-8.56 (series of m, total 12H), 12.65 (hr s, 11H); MS (FAB) m/z 536 1); Anal. Calcd for C 29
H
33
N
3
O
7 1-3I4H 2 O: C, 63.43; 1-16.33; N, 7.65. Found: 63.34; H, 6.28; N, 7.28.
Example 209 3-nitro-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetyllaninoethoxylbenzoic acid H 0011O 0O 2 1H H 6H 3 257 To a stirred solution of 4-hydroxy-3-nitrobenzoic acid (5.18 g, 28.29 minol) in benzene- MeOH v/v, 140 inL) was added TMSCHN 2 (14.10 mL, 28.20 mmol, 2 M solution in hexane) at room temp, and the resulting mixture was stiffed overnight. The mixture was evaporated and the residue was purified by column chromatography on silica-gel with CHC1 3 as eluent to give 4.18 g methyl 3-nitro-4-hydroxybenzoate as a yellow crystalline solid. 'H-NMvR (CDCI 3 d 3.95 3H), 7.22 IH, J=8.8 Hz), 8.24 (dd, I J=2.0, 8.8 Hz), 8.83 liI, J=2.0 Hz), 10.89 1H).
To a stirred solution of methyl 3-nitro-4-hydroxybenzoate (1.98 g, 10.04 inmol), N-Boc ethanolamnine (1.63 g, 10. 11 minol) and PPh 3 (3.43 g, 13.08 mniol) in TI-F (40 mL) was added DIAD (2.57 niL, 13.05 inmol), 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 niL) 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. NaI-C0 3 The mixture was extracted with CHCI 3 washed with brine, and dried over Na 2
SO
4 The solvent was evaporated in vacua to give the oily residue, which was purified by column chromatography on silica -gel with CHC1 3 then CHC1 3 -MeOH (20: 1, v/v) as eluent to give 930 mg (27% for 2 steps methyl 3-nitro-4-(2amrinoethoxy) benzoate as gum. 'H-NMR (CDCI 3 d 3.16-3.19 (in, 1 3.53-3.57 (mn, 1 3.90 and 3.94 3 3.95-3.98 (in, 1 4.21-4.24 (in, 1 6.89-6.91 and 7.11-7.13 (in, 1 8.03- 8.19 and 8.21 (in, I1H), 8.52 and 8.86 1 H).
To a stirred solution of pentafluorophenyl ester of 3-methoxy-4-[N'-(2-methylphenyl) ureido] phenylacetic acid (1.86 g, 3.87 inmol) and methyl 3-nitro-4-(2-aminoethoxy)benzoic acid (0.93 g, 3.87 inmol) in DMF (27 niL was added Et 3 N (0.90 niL, 6.46 minol), 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-O.25 N NaOH 20 niL) and the resulting WO 01/00206 WO 0100206PCTUSJO/18079 mixture was heated under reflux overnight. The mixture was extracted with EtOAc, washed with brine, dried over Na 2 SO,, and evaporated. The crude solid was recrystallized from CHCI 3 -EtQH to give 60 mg for 2 steps) 3-nitro-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetylI aminoethoxylbenzoic acid 257 as a yellow crystalline solid. mp 112-115 'H-NMR (DMS0c4) d 2.24 3 3.37-3.66 (in, 7 3.84 3 4.27-4.30 (in, 1 6.74-8.56 (series of m, total 12 MS (FAB) m/lz 523 Anal. Calcd for C26H 2 ,5 4
O
8 -3/2H- 2 O: 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-IN'-(2-fluorophenyl)ureido]phenylacetyllethylaminoethoxy benzoic acid 0 CH H~ H OCH 3 258 To a stiffed solution of pentafluorophenyl ester of 3-methoxy-4-[N'-(2-fluorophenyl) ureido] phenylacetic acid (135 mg, 0.28 inmol) and ethyl 3-methoxy-4-(2-ethylaxniinoethoxy) benzoate (78 mng, 0.29 mniol) was added Et 3 N lmL, 0.72 inmol), and the resulting mixture was stirred overnight. The mixture was diluted with EtOAc, washed successively with 0.5SN HCl, brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silicagel with CHCI 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]phenylacetyllethylaminoethoxy]benzoate as an oil. 'H-NMR (CDCI,) d 1. 13-1.23 (in, 3 1.37-1.40 3 2.90-3.89 (mn, 12 4.09-4.28 (in, 2 4.33- 4.39 (mn, 2 6.70-8.2 1 (series of m, total 12 H).
To a stirred solution of ethyl 3-inethoxy-4-[2-[3-methoxy-4-[N' -(2-fluorophenyl)ureido] phenylacetyl]ethylaxninoethoxy~benzoate (160 mg, 0.28 inmol) in THF (5 inL) 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- CHCl 3 -n-hexane to give 70 mg 3-methoxy-4-[2-[3-inethoxy-4-[N'-(2-fluorophenyl)ureidoI phenylacetyllethylaminoethoxy] benzoic acid 258 as a yellow crystalline powder. mp 105-110 *C; IR (KBr) 1687 'H-NMR (DMSO-d 6 d 1.00-1. 10 (in, 3 2.48 2 3.35-3.81 (mn, 10 H), 4.13-4.14 (in, 2 6.70-9.15 (series of Mn 12 MIS (FAB) m/z 540 Anal. Calcd for C~j43FN 3
O
7 -1f2H 2 0: C, 61.3 1; H, 5.82; N, 7.47. Found: C, 61.05; H, 5.82; N, 7.47.
WO 01/00206 WO 0100206PCT/USOO/18079 Example 211 4-[4-13-methoxy-4-[N'-(2-methylphenyl)ureidolphenyl]acetyl- 1 -piperazinylbenzoic acid 0 H 3,NH H CH3259 A stirred mixture of tert-butyl 1-piperazinecarboxylate (1.00 g, 5.37 mmol), ethyl 4fluorobenzoate (903 mg, 5.37 mmol), and K 2 C0 3 (1.11 g, 8.06 mmol) in DMIF (10 mL) was heated at 120*C overnight. After cooling, the mixture was diluted with EtOAc (300 mL), followed by washing with brine (2 x 200 niL, drying over MgSOA, and evaporation. The residue was chromatographed on silica-gel with CHCl 3 -EtOAc (20:110o 4: 1, v/v) as eluent to give 257 mg (140%) ethyl 4-[4-(tert-butyloxycarbonyl)-1-piperazinyIlbnzoate as a pale yellow amorphous solid.
IR (KBr) 1701, 1612 cm-1; 'H-NMR (CDCl 3 d 1.37 (3H,t, J =7.3 Hz), 1.49 (9H, 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 Anal. Calcd for C 18 H,6NO 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-Qtert-butyloxycarbonyl)-1I-piperazinyllbenzoate (240 mg, 0.7 18 mmol) in CHCI 2 (5 niL) was added TWA (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 CHC1 3 (2 x 100 niL. The combined extracts were dried over NaCO 3 and evaporated to give 168 mg ethyl 4-(1-piperazinyl)benzoate (100%) as a yellow oil. 'H-NMR (CDCl 3 )d 1.37 (3H, t, J7.3 Hz), 3.03 (4 H, t, J4.9 Hz), 3.29 (4H,1, J= 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, in).
To a stirred solution of ethyl 4-(1-piperazinyl)benzoate (170 mng, 0.730 nunol) and 3methoxy-4-[N'-(2-methylphenyl)ureido~phenylacetic acid (229 mng, 0.730 minol) in DM4F (10 niL) was added EDC-HCl (2 10 mg, 1. 10 nimol), DMLAP (catalytic amount), and HOBt (catalytic amount), and the mixture was stirred overnight. The mixture was poured into H,0 (100 niL) and the solid was collected with suction. The residue was recrystallized from CHC1 3 -n-hexane to give 290 mg ethyl 4-[4-[3-inethoxy-4-[N' -(2.inethylphenyl)ureidolphenyl]acetyl- 1-piperazinyl benzoate as a colorless crystalline powder. mp 208-2 10 JR (KBr) 1711, 1695 'H-NMR (CDC1 3 d 1.37 (3 H, t, J =7.3 Hz), 2.29 (3 H, 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, 3.72 (2 H, 3.79 (2 H, t, J 4.9Hz), 4.33 (2 H, q, J 7.3 Hz), 6.38 (1 H, 6.78-6.99 (4 H, in), 7.13-7.24 (4 H, in), 7.50 (1 H, d, J 7.8 Hz), 7.92 (2 WO 01/00206 C/S0/89 PCT/USOO/18079 H, d, J 8.8 Hz), 8.12 (1 H, d, J 7.8 Hz); MS (FAB) m/z 5 31 Anal. Calcd for C~j-l3.N 4 0 5 -0.5H 2 O: 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)ureidojphentyllacetyl-1I-piperazinylbcnzoate (290 mg, 0.547 inmol) in MeGH-THE 1, v/v, 15 mL) was added 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 inL) and the solid was collected with suction. The residue was recrystallized from CHCI 3 -MeOH to give 190 mg 4-[4-[3-methoxy-4-[N'-(2-methylphenyl) ureidolphenyljacetyl-1-piperazinylbenzoic acid 259 as a yellow crystalline powder. mp 240-245 0 C; 1H-NMR (DMS0) d 2.24 (3 H, 3.17-3.50 (8 H, in), 3.72 (2 H, 3.86 (3 H, 6.77 (1 H, d, J 8.3 Hz), 6.90 (1 H, 6.91-6.96 (3 H, mn), 7.11-7.17 (2 H, in), 7.76-7.80 (3 H, in), 8.03 (1 H, d, J 8.3 Hz), 8.47 (1 H, 8.58 (1 H, 12.30 (1 H, Anal.Calcd for C 2 {3ON,,05H 2 O: C, 64.60; H, 6.20; N, 10.76. Found: C, 64.64; H, 5.85; N, 10.51.
Example 212 (R)-3-inethoxy-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureidojphenylacetylaminoJ-l -propoxy] benzoic acid H OOH CH OCH 3 H3H H 6H 3 260 To a cooled (0 0 C) solution of (R)-2-amino-1-propanol (3.01 g, 0.04 nunol) and Et 3
N
(6.70 inL, 0.05 mmol) in DMF-11 2 0 1, vfv)(40 mL) was added (Boc) 2 0 (10.0 mL, 0.04 mmnol), and the resulting mixture was stirred at room temp for 2 days. The mixture was diluted with EtOAc, washed with H 2 0, brine, dried over Na 2
SO
4 and evaporated to give 6.91 g tert-butoxycarbonylamino-l1-propanol as a colorless oil. 'H-NMR (CDCI 3 )581.15(d, 3H, J6.8 Hz), 1.45 9 3.48-3.53 (in, 1 3.62-3.66 (in, 1 3.76-3.77 (in, 1 MS (FAB) m/~z 176 120 (MCt Su).
To a stirred solution of ethyl 4-hydroxy-3-methoxybenzoate (7.74 g, 0.04 mmol), tert-butoxycarbonylainino-1-propanol (6.91 g, 0.04 inmol) and PhjP (13.44 g, 0.05 mniol) in THF mL) was added diisopropyl azodicarboxylate(DIAD)(10.0 mL, 0.05 minol), 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.
WO 01/00206 WO 0100206PCT/USOO/18079 NaHCO 3 and extracted with CHC 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 ayellow oil. 'H-NMR (CDC 3 6 1.90 3 H, J=6.8 Hz), 1.39 3 H, J=7.3 Hz), 1.72 (bs, 2 3.42-3.47 (in, 1 3.74-3.89 (in, 1 3.91 3 H4), 3.96-4.00 (in, 11H), 4.35 2 H, J=7.3 Hz), 6.88 1 H, J=8.3 Hz), 7.55 I H, J=2.0 Hz), 7.65 (dd, 1 H, J=2.0, 8.3 Hz); MS (FAB) m/z 254 To a stirred solution of pentafluorophenyl 3-methoxy-4-[Nr-(2-methylphenyl)ureidoJ phenylacetate (459 ing, 0.96 nunol) and ethyl (R)-3-inethoxy-4-(2-antinopropoxy)benzoate (242 mg, 0.96 inmol) in DMF (5 mL) was added Et 3 N (200 MI, 1.43 mniol), and the resulting mixture was stiffed for 2 hr. The mixture was diluted with EtOAc, washed with 0.5 N HCl, 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 mng ethyl (R)-3-methoxy-4-[2-[3methoxy-4-[N '-(2-inethylphentyl)ureido~phenylacetylaminoj- 1 -propoxy~benzoate as a colorless crystalline solid. 'H-NMR (CDCl 3 8 1.23-1.28 (in, 3 1.38-1.41 3 H, J=7.3 Hz), 2.32 3 3.504. 13 (in, total 11I 4.36 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-[AN'-(2-methylphenyl) ureidol phenylacetylamino]-1-propoxyjbenzoate (360 mg, 0.66 inmol) in THF-MeOH (20 niL, 9: 1, vlv) was added 0.25 N NaOH (10 inL), and the resulting mixture was heated under reflux overnight. The mixture was poured into ice-lI N HCI, and precipitate was collected. The crude solid was recrystallized from CHCl 3 -n-hexane to give 172 mg(50%/) 260 as a white crystalline powder. mp 168-169 III (KI~r) 1687 cin-'; 'H-NMR (DMSO-d) 8 1. 18 3 H, J=6.8 Hz), 2.24 3 2.50-2.51 (in, 2 3.80 3 3.84 3 3.87-4.06 (in, 2 4.074. 14 (in, 1 6.76-8.57 (series of in, total 12 12.66 (bs, 1 MS (FAB) m/z 522 Anal. Calcd for CuHl- 31
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-inethoxy-4['-(2-methylphenyl)ureido]phenylacetyl]allylaminoethoxybenzoic acid ~H2OH k gN::o OCH 3
CH
3 H H OCH 3 261 WO 01/00206 WO 0100206PCTIUSOO/18079 To a stirred mixture of ethyl 4-(2-N-trifluoroacetylaminoethoxy)-3-methoxy benzoate 10.4mmol)and K 2 C0 3 (2.3g, 16.4mmol) in DMF (20mL) was added allyl bromidde (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 Na 2
SO
4 and evaporated in vacuo. The residue was dissolved in THF-MeOH-H 2 0 1: 1,v/v/v) mL) and added KC0 3 (2.3 g, 16.4 nunol). 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,, 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-allylanminoethoxy)-3-methoxybenzoate as a pale-yellow oil. 'H-NMR(CDC 3 400MIHz) 8 1.39 3H, J=7.3Hz), 3.07 2H, J=5.3Hz), 3.34 2H4, J=5.9Hz), 3.91 3H), 4.18 2HJ=5.4Hz), 4.35 (dd, 2H, J=7.3Hz, 14.1IHz), 5.12 1H, J= 10.3Hz), 5.22 (dd, Ili, J=15Hz, 17.1IHz), 5.92 (in, 214), 6.90 1H, J=8.3Hz), 7.55 111, 7.65 (dd, 111, J=2.OHz, 8.3Hz); MS(FAB) m/z 278, 280(M+HY+.
To a stirred mixture of ethyl 4-(2-allylamninoetlioxy)-3-methoxy benzoate (578mg, 2. 1 mmol), 3-methoxy-4-[AP"-(2-methylphenyl)ureidolphenylaceticacid (650mg,2. 1 mmol), HOBt (420mg, 3.l11mmuol), and DMAP (catalytic amount) in DMF (4 mL) was added EDC (596mg,3. 1 Iminol) 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 SO4, and evaporated in vacuo. The residue was chromatographed on silica-gel with CHC1 3 :EtOAc (95:5 to 1: 1, v/v) as eluent to give lg 3-methoxy-4-[[2-[3methoxy-4-[N'-(2-methylphenyl)ureidoJ phenylacetyl]allylaxninojethoxyJ benzoic acid as a paleyellow guni. 'H-NMR(CDCI 3 400M4Hz) 8 1.39 3H, J=7.3Hz), 2.29 311), 3.58 and 3.63 (s, total 311), 3.70-3.77 (mn, 211), 3.83 and 3.87 311), 4.05-4.13 (mn, 211), 4.25 (mn, 111), 4.36 1H, J=7.OHz), 5.04-5.22 (mn, 211), 5.73 (in, 111), 6.32 and 6.47 111), 6.69-6.85 (in, 211), 7.12 (mn, 211), 7.23 (in, 211), 7.50-7.65 (in, 211), 8.05 111, J=7.8Hz); MS (FAB) m/z 576(M+11)*.
A mixture of 3-methoxy-4-[[2-[3 -methoxy-4-[AP-(2-methylphenyl)ureidolphenylacetyll allylaminojethoxy~benzoic acid (50mg, 0.O9nunol) in THF-MeOH 1, v/v) (2niL) and IN NaOH 135mL, 0.13Snunol was stirred for 15 hr at room temp and 3 hr at 50'C. The mixture was poured into ice-IN HCI. Solid was collected, washed with water, and air-dried. The crude solid was recrystallized from CHCI 3 -n-hexane to give 38mg 261 as a white crystalline material.
nip 125-130 IR(K.Br), 3319, 2939, 1687, 1647, 1601, 1535, 1456, 1417, 1269, 1223, 1034, 760cm-1; 'H-NMR(DMSO-d, 400M1-lz) 5 2.29 311), 3.68 211), 3.75-3.85 (in, 811), 4.05 (br, WO 01/00206 WO 0100296PCT/USOO/18079 114), 4.19 (in, 314i), 5. 10-5.25 (in, 211), 5.65-5.90 (in, 111), 6.75 (in, IH), 6.85 1H1), 6.92 (in, IN), 7.02-7.20 (in, 3H1), 7.48 IN, J=10.2Hz), 7.56 (in, 111), 7.79 1H, J=6.8Hz), 8.01 (in, 11N), 8.46 IN), 8.56 I1H, J=4.4Hz), 12.7 (br, MS (FAB) m/z 548(M+N)~; Anal. calcd. for 33
N
3
O
7 O0.5H 2 O, C, 64.74; H, 6.16; N, 7.55. Found, C, 64.72; H, 6.07; N, 7.55.
Exampile 214 3-miethoxy-4-[[2-[3-methoxy-4-[iM(2-nethylphenyl)ureidojphenylacetylJ-(2-morpholino) ethylaminojethoxylbenzoic acid QN0
OCH
3 6H 3 H H CH 3 262 To a stirred solution of 3-methoxy-4-[[2-[3-methoxy-4-[N'-(2-methyiphenyl)ureidoI phenylacetyllailylaminojethoxy~benzoic acid (950mg, 1.65mmol) in THF:H 2 0 (7mL) was added N-methylmorpholine-N-oxide (579mg,4.95nmmol) and osmium tetroxide (0.2M solution in water) (0.4 l3mL, 0.O8mmol). 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 residue was dissolved in MeOH-THF-H 2 0 1: 1, v/v) (1 2mL) and added sodium periodate (318mg, 1.Snunol). 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 ethyl 3-methoxy-4-[[2-[3-methoxy-4-[M,-(2-inethylphenyl)ureido] phenylacetyl]-N-forminymethylanuino~ethoxy] benzoate as a pale-yellow gum. 'H-NMR (CDCI, 400MIHz) 8 1.39 3H, J=7.3Hz), 2.29 3H), 3.3 1-3.95 (in, 1111), 4.10-4.42 (in, 5H), 6.51-6.82 (in, 311), 7. 10-7.25 (in, 3H), 7.50 (in, 2H), 7.60 (in, 11H), 8.10 (in, IN), 9.50 (in, iH); MS (FAB) m/z 578 (M+H) 4 To a stiffed mixture of ethyl 3-methoxy-4-[[2-[3-methoxy-4-[N-(2-inethylphenyl)ureidoI phenylacetylJ-N-forinylinethylainino~ethoxylbenzoate (265mg, 0.4.6mmol), inorpholine (0.4OmL, 4.59mmol), and AcOH (0.263mL, 4.6mniol) in EtQH (3mL) was added NaBH 3 CN (288mg, 4.6nunol) 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 00 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 vacua. The residue was chromatographed on silica- WO 01/00206 WO 0/0021)6PCT/US00118079 gel with CHC1 3 :MeOH (95:5, v/v) as eluent to give 2 13mg (7 ethyl 3-methoxy-4-[[2-[3methoxy-4-[A"-(2-methylphenyl)ureidoI phenylacetylj-2-morpholino)ethylaminojethoxy]benzoate as an oil. 'H-NlvR (CDCI 3 400Mflz) 81.28 311,J=7.OHz), 2.3 1(s,311), 2.48 (brs,41-1), 2.52 (in, 211), 3.60-3.91 (in, 16H), 4. 11 and 4.28 (in, total 2H), 4.39 2H, J=7.OHz), 6.70-6.85 (in, 411), 7.15 (in, 211), 7.50-7.63 (in, 311), 8.08 I H, J=8.OHz); MS (FAB) mz 649(M+HFW.
A mixture of ethyl 3 -methoxy-4-[[2-[3-methoxy-4-[A'-(2-methylphenyl)ureidolphenyl acetyl]-(2-morpholino)ethylaminojethoxylbenzoate (265mg, 0.46mmol) in THF (4mL) and 1IN NaOH (0.984miL) 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 CHCI 3 :MeOH(9: 1, The extract was washed with brine, dried over MgSO,, and evaporated in vacua. The residue was crystallized with Et 2 O to give 160 mg(78%) 262 as a white crystalline material. mp 125-130 0 C;JR (KBr), 3346, 2956, 2937, 1705, 1622, 1599, 1537, 1456, 1417, 1299, 1114, 1032, 752cm-'; 'H-NMR (CD 3 OD, 40M&~) 6 2.29 311), 2.49-2.64 (in, 6H1), 3.65-3.85 (in, 1611), 4.13 (in, 111), 4.26 (mn, 111), 6.78-7.04 (in, 411), 7.18 (in, 211), 7.55-7.64 (in, 311), 7.99 (in, 211); MS (FAB)m~z 62l1(M+11*; Ana. Calcd. for
CBRHON
4 O0 8 -2.5H- 2 O, C, 59.54; 11, 6.81; N, 8.42. Found, C, 59.71; H1, 6.35; N, 7.98.
Example 215 3-methoxy-4-[[2-[3-methoxy-4-[A'-(2-methylphenyl)ureido~phenylacetyll-[2-[4-nethyl- 1piperazinyllethylamiino]ethoxylbenzoic acid
~~ZOH
YHW 6H 3 263 To a stiffed mixture of ethyl 3-methoxy-4-[[2-[3-methoxy-4-[A"-(2-inethylphenyl)ureido phenylacetylj-N-formylmethylainino~ethoxylbenzoate (242mg, 0.42nunol), N-inethylpiperazine (0.465inL, 4.2inmol), and AcOH (0.240miL, 4.2mmol) in EtQH (3inL) was added NaBH 3
CN
(263mg, 4.2mmol) at room temp. The resulting mixture was stiffed for 15 hr at room temp. The mixture was diuted 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 vacua. The residue was chroinatographed on silicagel with CHC1 3 :MeOH (95:5, v/v) as eluent to give 195mg ethyl 3-methoxy-4-[[2-[3methoxy-4-[N'-(2-methylphenyl)ureidoI phenylacetylj-[2-(4-methyl-l1-piperazinyllethylaminoI WO 01/00206 WO 0100206PCT/USOOI 18079 ethoxylbenzoate as an oil. 'H-NMvR (CDCI,, 400M&z) 8 1.23 3H, J=7.OHz), 2.25 3M), 2.29 3H1), 2.50 (br m, 12M1, 3.44-3.85 (in, 12H1), 4. 10 (br, 111), 4.22 (br, 111), 4.35 (mn, 6.70- 6.85 (in, 6.98 11-1), 7. 10 (in, 1H), 7.20 (in, 2M-1, 7.40 (in, 7.60-7.70 (in, 3M-1, 8.05( d, 1H, J=7.8Hz); MS (FAB) m/z 662(M+H)'.
A mixture of ethyl 3-methoxy-4-[[2-[3-methoxy-4-[AP-(2-methylphenyl)ureido] phenylacetylj-[2-(4-methyl- 1-piperazinyllethylaminolethoxylbenzoate (195mg, 0. 3Ommol) in THF:MeOH(4:I1, v/v) (SniL) and IN NaOH (0.885inL) 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 CHCI 3 :MeOH(9: 1, vlv). The extract was washed with brine, dried over MgSO 4 and evaporated in vacua. The residue was crystallized with EtO to give 141 ing(75%/) 263 as a white crystalline material. mp 155-160 0(2; IR (KBr), 2937, 1537, 783cnr'; 'H-NMR (CD 3 OD, 400MIHz) 8 2.29 3H), 2.49-2.80 (in, 151-), 3.60-3.85 (mn, 914), 3.92 11M1, 4.12 (mn, 4.25 (in, 11-1), 6.78-7.20 (in, 6H), 7.61 (in, 8.00 lH), MS (FAB) m/lz 632(M)*; Anal. Caicd. for C3,H4N5,O-2.5H 2 O, C, 60.16; H, 7.13; N, 10.32. Found, C, 59.72; H, 6.86; N, 9.97.
Example 216 3-methoxy-4-[[2-13 -methoxy-4-[A'-(2-methylphenyl)ureidolphenylacetyl]-[2-cyclopropylaminoJ ethylaminojethoxylbenzoic acid
FI
OOH
0 ~H 3 H,3H OCH3264 To a stirred mixture of ethyl 3-methoxy-4-[[2-[3-inethoxy-4-[N'-{2-methylphenyl)ureido phenylacetylj-N-forinylmethylaminolethoxy]benzoate (267mg, 0.46iniol), cyclopropylamine (0.32mL, 4.6inmol), and AcOH (0.264mL, 4.6nunol) in EtOH (3inL) was added NaBH 3
CN
(290mg, 4.6minol) 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 00(C. The resulting mixture was stirred for 0.5 hr at 00(2. The mixture was extracted with EtOAc. The extract was washed with brine, dried over MgSO 4 and evaporated in vacua. The residue was chroinatographed on silicagel with CHCI 3 :MeOH (95:5, v/v) as eluent to give 156mg ethyl 3-methoxy-4-[[2-[3inethoxy-4-[MV-(2-methylphenyl)ureido~phenylacetyl]-[2-cyclopropylanminolethylanmino ethoxylbenzoate as an oil. 'H-NN'R (CDCI 3 400Mliz) 8 0.35 (in, 1.22 (br s, 311), 2. 10 (in, 111), 2.20 2.42 (br, 2.90 (br s, 21H), 3.60-3.80 (in, 101H), 4. 10 (br, 111), 4.22 (hr. 11-), WO 01/00206 PCT/USOO/18079 4.33 (br, 2H), 6.72 3H), 7.05-7.30 4H), 7.55 4H), 8.06 (br s, MS (FAB) m/z 619(M-HW.
A mixture of ethyl 3-methoxy-4-j[2-[3-methoxy-4-IN'-(2-methylphenyl)ureido] phenylacetyl]-N-[2-cyclopropylaminolethylaminolethoxylbenzoate (195mg, 0.30mmol)) in THF:MeOH(4: 1, v/v) (5mL) and IN NaOH (0.756iL) 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 CHCI 3 :MeOH(9: 1, vfv). The extract was washed with brine, dried over MgSQ,, and evaporated in vacuo. The residue was crystallized with Et 2 O to give 57 mg (3 3-methoxy-4-[[2-[3-methoxy-4-[N'-(2methyl phenyl)ureidolphenylacetyl]-[2-cyclopropylaminoI ethylamino]ethoxylbenzoic acid 264 as a white crystalline material. mp 135-140 IR (KBr), 3324, 2937, 1535, 1032, 754cm-'; IH- NMR (CD 3 OD, 400MHz) 8 0.50-0.73 4H), 2.29 3H), 2.53 11), 2.98 lH), 3.21 (m, 11), 3.58-3.88 11), 3.91 4.09 1H), 4.25 1H), 6.76-6.92 31), 7.01 (m, 1H), 7.18 21), 7.60 311), 8.00 J=8.3Hz, 1H); MS (FAB) m/z 591(M+H); Anal. Calcd.
for C 32 HsN,O,3.0H 2 O, C, 59.62; H, 6.88; N, 8.69. Found, C, 59.25; H, 6.29; N, 8.29.
Example 217 4-[[2-[3-methoxy-4-[N '-(2-fluorophenyl)ureido phenylacetyll-N-methylaninoethoxybenzoic acid 911 3
IXCOOH
H H CH 3 265 3-chloro-4-[[2-[3-methoxy-4-[N '-(2-fluorophenyl)ureidolphenylacetyl]-N-methylaminoethoxy benzoic acid
CI
F H H OCH3 266 To a stirred cold (0 0 C) solution of 2-(N-benzyloxycarbonyl-N-methyl)ethanolamine (3.01 g, 14.4 mmol), methyl 3-chloro-4-hydroxybenzoate (2.68 g, 14.4 miol), Ph 3 P (5.65 g, 21.5 mxol) in THF (30 iL) was added diisopropyl azodicarboxylate (DIAD) (4.25 mL, 21.6 mnol), and the resulting mixture was heated under reflux overnight. The solution was evaporated off and the residue was purified by column chromatography on silica-gel with CHCI 3 as eluent to give 3.90g methyl 3-chloro-4-[2-(N-benzyloxycarbonyl-N-methylamino)ethoxylbenzoate as a pale yellow solid. 'H-NvR (CDC 3 8 3.15 3 3.74-3.76 2 3.89 3 4.17-4.27 2 5.14 2 6.81-6.94 1 7.33-7.36 511), 7.85-7.92 1 8.05 (bs, 1 H).
WO 01/00206 WO 0100206PCT/USOO/18079 A solution of methyl 3 .chloro-4-[2-(N-benzyloxycarbonyl-N-methylamino)ethoxyI 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 at 3 atm for 2 hr. The mixture was filtered and the filtrate was washed with sat.
NaHCO 3 and the basic aqueous layer was extracted with CHCI 3 washed with brine and evaporate to give unseparable mixture of methyl 3-chloro-4-(N-methylaminoethoxy)benzoate and methyl 4- [2-(N-methylamino) ethoxyjbenzoate the title compound (1.61 g) as a pale yellow oil. 'H-NMR
(CDCI
3 8 2.52-2.52 and 2.53-2.54 (each mn, 3 2.98-3.00 and 3.03-3.05 (each mn, each 2 H), 3.88 and 3.99 (each s, each 3 4.11-4.14 and 4.18-4.20 (each mn, each 2 6.91-6.96 and 7.90- 8.05 (series of in, total 7 H).
A mixture of 3-methoxy-4-[N'-(2-fluorophernyl)ureido~phenylacetic acid (392 mg), a mixture of methyl 3-chloro-4-[2-(N-methylamino)ethoxyjbenzoate and methyl 4-12-(Nmethylaniino)ethoxy] benzoate (305 mg), EDC(hydrochloride) (354 mg), HOBt (250 mg), and DMAP (250 mg) in DMIF (8 mL) was stirred at room temp for 6 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 1% MeOH in CHCI 3 as eluent to give a mixture of methyl 3-chloro-4-[[2-[3-methoxy-4-[N'-(2-fluorophentyl) ureido~phenylacetylJ-Nmethylaminoethoxylbenzoate and methyl 4-II[2-[3-methoxy-4-[N '-(2-fluorophenyl)ureidoj pheniylacetyl]-N-methylaminoethoxyjbenzoate (550 mg) as a brown amorphous solid.
To a stirred solution of this mixture (550 mg) of methyl 3-chloro-4-[[2-1j3-methoxy-4-[N'- (2-fluorophenyl)ureidojphenylacetylj-N-methylaminoethoxy]benzoate and methyl methoxy-4-[N '-(2-fluorophenyl)ureido]phenylacetyl]-N-methylaminoethoxylbenzoate in THE- MeOH (20 mL, 1: 1, vtv) 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- I N HCl, and the solid was collected. The crude solid was purified by preparative TLC with 10% MeOH in CHCI 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-[MP-(2-methylphenyl)ureido]phenylacetyljmethylamino]ethoxyjbenzoic acid 9H 3
OOH
CH
3 H H OCH 3 267 3-chloro-4-[2-3-methoxy4-[A-(2-methylphenyl)ureidophenylacetyl]methylan-inolethoxy] WO 01/00206 WO 0100206PCT/USOO/18079 benzoic acid
H
3 3 9 aQJ.COOH 26 A mixture of methyl 4-[2-(N-methyl-2-amino)ethoxyj-3-chlorobenzoate (292mg, 1 .2mmol), 3-methoxy-4-[M-(2-methylphenyl)ureidolphenylaceticacid (377mg, 1.2mmol), EDC (345mg, 1.8mmol), HOBt(243mg,l1.8mmol), and DMAP(29mg, 0.24mniol) in DMF(2.7niL) 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 CHCl 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'-(2methylphenyl)ureidolphenylacetyl] methylamino~ethoxyjbenzoate and methyl 4+-[-3-methoxy-4- [N'-(2-methylphenyl)ureido~phentyl acetyljmethylaminolethoxylbenzoate as pale-yellow oil.
A mixture (480mg as mixture) of methyl 3-chloro-4-[[2-[3-methoxy-4-[N'-(2-methyl phenyl)ureidoJ phenylacetyl~methylaxnnolethoxy~benzoate and methyl 4-[[2-[3-methoxy-4-[N'-(2methylphenyl)ureido] phenylacetyl]methylamino~ethoxyjbenzoate in THF-MeOH(4mL, 1: 1, v/v) was stirred at 500 C for 15 hr. The mixture was poured into ice-IN HCL The solid was collected, washed with wter, and air-dried. The crude solid was purified by preparative TLC CHCI 3 :MeOH (93:7, v/v) as eluent to afford 267 (180mg, 2steps 3 1% as a crystalline material) and 268 (280mg, 2steps 44% as a crystalline material). 267 mp 145-150 OC; 'H-NMR (DMSO-d, 400Mz) 862.31 3H), 3.05 and 3.19 311), 3.35 and 3.38 3.72-3.85 (in, 7H), 4.09 and 4.23 (in, total 2H), 6.79-7.20 (in, 711), 7.60 (in, IlH), 7.86-8.09(m, 3H), MS (FAB) m/z 493(M+H)~; Anal. calcd.
for C 27 H29N 3
O
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 'H-NMR (DMSO-d4 400M]Hz) 5 2.29 3M1, 3.06 and 3.26 311, 3.31 and 3.35 (s, 3M1, 3.85-3.94 (mn, 411), 4.18 and 4.32 (in, total 21M, 6.75-6.85 (in, 2M1, 6.99-7.20 411), 7.59 (in, 7.90-8.02 (in, 3H1); MS (FAB) m/z 526(M+H)*; Anal. calcd. for C',H 2 ,ClN 3
O
6 -2.0H 2 O, C, 57.70; H, 5.74; N, 7.48. Found, C, 57.99; H, 5.53; N, 7.07.
Example 219 4-[3-13-iethoxy-4-[N' -(2-methylphenyl)ureidolphenylacetylJ-N-methylaminol-l1-propyllbenzoic acid WO 01/00206 WO 0100206PCT/USOO0/18079 r-13269 To a stirred cold (minus 780 C) solution of triethyl 4-phosphonomethylbenzoate (1.22 g, 4.05 inmol) in THE (10 mL) was added NaHMDS (1.0 M in THF) (4.0 mL, 4.0 minol), and the resulting mixture was stirred for 1 hr at the same temp. A solution of 2-(N-benzyloxycarbonyl-Nmethylanuno) acetaldehyde (700 mg, 3.38 mniol) in THE (5 inL) 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 C1 (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 8 10 mg ethyl (E)-4-[3-(N-benzyloxycarbonyl-N-methylamino)-1-propenylbenzoate as a yellow oil. IH-NMvR
(CDCI
3 8 1.40 J =7.3 Hz, 3 H4), 2.95 (in, 2 4.09 (in, 2 4.3 5-4.40 (in, 2 5.17 2 H), 6.26-6.64 (series of mn, 2 7.36 (in, 7 7.99 J =8.3 Hz, 2 M).
A stirred solution of ethyl (E)-4-[3-(N-benzyloxycarbonyl-N-methylamrino)-1 -propenyl benzoate (810 mg, 2.29 inmol) in EtOH-AcOH (10:1, v/v, 22 mL) was hydrogenated over 5% Pd- C (1 g) for 3 days. The mixture was filtered and the filtrate was evaporated. The residue was made basic with sat. NaHCO, and extracted with CHCl,. The extract was dried over NaCO, and evaporated to give 438 mng ethyl 4-(3-methylanuino-l-propyl)benzoate as a yelow oil. 'H- NMR (CDC1 3 8 1.39 J=7.3 Hz, 3 1.82 (in, 2 2.43 3 2.61 (t J 7.3 Hz, 2 H), 2.72 J= 7.3 Hz, 2 3.33 (br s, 1 4.36 J= 7.3 Hz, 2 7.25 J= 8.3 Hz, 2 7.96 J =8.3 Hz, 2H).
To a stirred solution of 3-methoxy-4-[N'-(2-inethylphenyl)ureido]phenylacetic acid (456 mg, 1.45 innol) and ethyl 4-(3-methylainino- 1 -propyl)benzoate (220 mg, 1.45 mmol) were added EDC-HCI (417 mg, 2.16 mmol), HOBt and DMAP (catalytic amount) in DMF (10 inL), and the resulting mixture was stirred overnight. The mixture was diluted with EtOAc (300 mL), washed with brine, dried over MgSO, and evaporated. The residue was chromatographed on silica-gel with CHC1 3 -EtOH (10: 1) to give 503 mng (7 ethyl 4-[3-[3-inethoxy-4-[N'-(2methylphenyl) ureidojphenylacetyl-N-methylaniinoJ-l-propyl] benzoate as a yellow oil. MS (FAB) m/z 518(M+H)+.
To a stirred solution of ethyl 4-[3-[3-methoxy-4-[N'-(2-methylphenyl)ureidolpheny acetyl-N-inethylan-ino]-l-propyljbenzoate (500 mng, 0.966 inmol) in TIHE (8 mL) was added 0.25 WO 01/00206 WO 0100206PCTIUSOO/18079 N NaOH (8 and the mixture was heated under reflux overnight. The resulting solution was poured into ice- I N HCI (100 mL) and the solid was collected with suction. The solid was dissolved in CHCI 3 (100 mL.) and dried over MgSO 4 After removal of the solvent, the residue was chromatographed on silica gel with CHCI 3 -MeOH (10: 1 to 5: 1, v/v) to give 131 mg methoxy-4-[N' -(2-methylphenyl)ureido] phenylacetyl-N-methylami no] 1I -propyllbenzoic acid 269 as a pale yellow amorphous solid. 'H-NMR (DMSO-ds) 8 1.68-1.80 (in, 2 2.24 3 H4), 2.57 (in, 2 H1), 2.81 and 2.97 each, total 3 3.33 (in, 2 3.57-3.61 (in, 2 3.84 3 1H), 6.71 (dd,J =29.8, 8.3 Hz, 1 6.87 J= 11.2 Hz, 1 M,6.93 J =7.3 Hz, 1 7.15 2 H), 7.28 2 7.79 J =8.3 Hz, I 7.84-7.87 2H), 8.02 J =8.3 Hz, 1 8.49 J 7.3 Hz, 1 8.5 8 J 4.9 Hz, 1 MS (FAB) m/z 490 Anal. Calcd for
C
2 gH 3
N
3 0-1/2H,0: C, 67.45; H, 6.47; N, 8.43. Found: C, 67.27; H, 6.5 1; N, 8.02.
Example 220 '-(2-methylphenyl)ureidolphenylacetyll-N-methylaminojethoxyjbenzoic acid 911 3
COOH
3-clor-4-12-4-[ 270 3-hoo4-[-4[-(2-methylphenyl)ureidolphernylacetylJ-N-methylaminolethoxy]benzoic acid
CH
3 H H 271 A solution of pentafluorophenyl 4-[N'-(2-methylphenyl)ureidojphenylacetate (562 mg, 1 .29xniol), a midxture (304 mg) of methyl 3 -chloro-4-[2-(N-inethylamino)ethoxyjbenzoate and methyl 4-12-(N-inethyl ainino)ethoxyjbenzoate and Et 3 N (260 nl) in DMF (8 mQ. was stirred at room temp for 4 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 CHCI 3 -MeOH (50: 1, v/v) as eluent to give a mixture (670 mng) of methyl 3-chlowo-4-[[2-[4- [N '-(2-methylphenyl)ureido) phenylacetylJ-N-inethylamino) ethoxylbenzoate and methyl 4-[12-[4- IN '-(2-methylphenyl)ureido] phenylacetylj-N-inethylainino)ethoxy] benzoate as an oil.
To a stirred suspension of this mixture (670 mng) in THF-MeOH (20 mL., 1: 1, v/v) was added 0.5 N NaOH (10 inL and the resulting mixture was heated under reflux for 6 hr. The solution was poured into ice- I N HCl and the solid was collected. The crude solid was purified by WO 01/00206 WO 0100206PCT/USOO/18079 preparative thin layer chromatography (TLC) with 10% MeOH in CHC1 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 462 271 MIS (FAB) m/z 496 1).
Example 221 (S)-4-12-13-methoxy-4-[N -(2-methylphenyl)ureidolphenylacetylaminol-l1-propoxy]benzoic acid H
.~OOH
CH
3 H H OH 3 272 To a cooled C) solution of (S)-2-amino-l-propanol (2.08 g, 27.7 mrnol) and Et 3
N
(4.63 mL, 3 3.2 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 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 (N-tert-butoxycarbonylamino)-1-propanoI as a colorless oil. 'H-NMR (CDCI 3 8 1.14 3 H, J=6.8 Hz), 1.45 9 M-I, 3.51-3.52 1 3.63- 3.66 (in, 1 3.77 (in, 1 4.62 (in, 1 H).
To a cooled (00 C) solution of (S)-2-(N-tert-butoxycarborylamino)- 1 -propanol (1.02 g, 5.82 mrnol), methyl 4-hydroxybenzoate (0.89 g, 5.85 mmol), and Ph 3 P (1.98 g, 7.55 niol) in THF (20 mL) was added diisopropyl azodicarboxylate (DIAD) (1.49 niL, 7.57 innol), and the resulting mixture was heated under refiux 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 hr. The solution was concentrated in vacuo and the residue was treated with sat. NaHCO 3 The mixture was extracted with CHCI 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 methyl (S)-4-(2-amino-l-propoxy) benzoate as a pale yellow oil. IH-NMvR (CDCl 3 8 1.19 3 H, J1=6.4 Hz), 3.35-3.39 (in, 1 3.72-3.76 (in, 1 3.89 3 3.90-3.94 (in, 1 6.92 2 H, J1=8.8 Hz), 7.99 2 H, J1=8.8 Hz).
A mixture of pentafluorophenyl 3-methoxy-4-[N '-(2-methylphenyl)ureidojphenylacetate (505 mg, 1.05 minol), methyl (S)-4-(2-amino-l-propoxy)benzoate (220 mg, 1.05 minol), and Et 3
N
(0.220mL, 1.58 inmol) 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 Alter removal of the solvent, the residue was recrystallized from MeOH-CHCI 3 -n-hexane to give 290 mg methyl WO 01/00206 WO 0100206PCT/USOO/18079 (S)-4-[2-[3-methoxy-4-[N '-(2-methylphenyl) ureido]phenylacetylaminoJ- 1-propoxyjbenzoate as a white crystalline powder. 'H-NMR (DMVSO-&J 8 1. 18 3 H, J=6.8 Hz), 2,24 3 3.36 (s, 2H), 3.80 3H), 3.82 3H), 3.93-4.03 2H1), 4.09-4.14 (in, 1H), 6.75-8.57 (series of in, total 13 H).
To a stirred solution of methyl (S)-4-[2-113-methoxy-4-[N '-(2-methylphenyl)ureidoJphenyl acetylaniinoJ- 1 -propoxylbenzoate (290 mg, 0. 57 mmxol) in TBF-MeOH (20 niL, 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-lI N HCI and extracted with CHCI 3 -MeOH (10 1, vfv). The extract was washed with brine, dried over Na 2 SO, and evaporated. The residue was recrystallized from MeOH-CHC1 3 n-hexane to give 158 mg 272 as a white crystalline powder. mp 198-20 1 0 C; 'H-NMR (DMSO-d) 8 1.18 3 H, J=6.3 Hz), 2.24 3 3.36 2 3.82 3 3.87-4. 10 (in, 2 4.10-4.16 (in, 1 6.75-6.78 (in, 1 6.92-7.02 (in, 4 7.11-7.18 (in, 2 7.78-7.80 (in, 1 7.86-7.89 (mn, 2 7.98-8.00 (mn, 1 8.12-8. 14 (in, 11H), 8.46 1 8.55 1 H), 12.62 (bs, 1 MS (FAB) m/z 492 Anal. Calcd for C 27
H
9 NO,- 12H 2 0: C, 64.79; H, 6.04; N, 8.2 1. Found: C, 64.36; H, 5.85; N, 8.2 1.
Example 222 '-(2-methylphenyl)ureidolphenylacetylaninoJ- 1 -propoxylbenzoic acid 11311 H273 A mixture of pentafluorophenyl 4-[N -(2-methylphenyl)ureidojphenylacetate (560 mg, 1.24 nunol), methyl (.S)4-(2-ainino-1 -propoxy)benzoate (260 mg, 1.24 minol), Et 3 N (0.26OnmL, 1.87 mmol) in DMF (8 inL) 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-CHC1 3 -n-hexane to give 210 mng (S)-4-[2-[3-inethoxy-4-[N -(2-inethylphenyl)ureido]phenylacetylamino]- 1propoxyjbenzoic acid as a white crystalline powder. 'H-NMR (DMVSO-d) 8 1.17 3 H, J=6.8 Hz), 2.24 3 3.32 2 3.81 3 3.924.03 (in, 2 4.08-4.15 (in, 1 6.92-6.95 (in, 1 7.04-7.06 (in, 2 7.12-7.18 (in, 4 7.35-7.39 (in, 2 7.83-7.85 (in, I 7.89- 7.92 (in, 3 8.12-8.14 (in, 1 8.97 1 H).
To a stirred solution of methyl '-(2-inethylphenyl)ureido~phenylacetyl WO 01/00206 WO 0100206PCT/USOO/18079 amino] -1I- propoxy]benzoate (200 mg, 0.42 mmol) in THF-MeOH (10 mL, 1: 1, vfv) was added 0. N NaOH (10 mL), and the mixture was heated under reflux for 2 hr. The mixture was poured into ice- I N HCI, and the solid was collected. The crude solid was recrystallized from MeOH-CHCI,-nhexane to give 68 mg 273 as a white crystalline powder. mp 262-265 OC; 'H-NMR (DMSO-d,5) 8 1.17 3 H, J=6.8 Hz), 2.24 3 3.32 2 3.91-4.02 (in, 2 4.09-4.15 (in, 1 6.92-6.96 (in, 1 7.01-7.03 (in, 2 7.12-7.20 (in, 4 7.36-7.40 (in, 2 7.83- 7.95 (in, 4 8.12-8.14 I 8.99 1 12.63 (bs, 1 MIS (FAB) m/z 462 Anal.
Calcd for C26H 27
N
3 O,-1/4H 2 O: C, 67.01; H, 5.95; N, 9.02. Found: C, 67.13; H, 5.90; N, 9.02.
Example 223 (S)-3-chloro-4-12-[4-[N '-(2-inethylphenyl)ureido]phenylacetylaminoj- 1-propoxylbenzoic acid H
OOH
I H 7CI H~HH 274 To a cooled (00 C) solution of (S)-2-(N-tert-butoxycarbonylanuino)-1-propano (1.05 g, 5.99 mmol), methyl 3-chloro-4-hydroxybenzoate 12 g, 6.00 minol), and Phy (2.36 g, 9.00 mmol) in THF (20 mL) was added diisopropyl azodicarboxylate (DIAD) (1.77 niL, 8.99 mmol), and the resulting mixture was heated under refiux for 2 days. The solution was evaporated off and the residue was dissolved in CH 2
CI
2 (20 mL) and TFA (10 mQL. The resulting mixture was stirred at room temp for 1.5 hr. The solution was concentrated in vacuo, and the residue was dissolved in CHCI,. 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 washed with brine, dried over Na 2 SO, and evaporated to give 660 mg (2 steps, methyl chloro-4-(2-amino-l-propoxy)benzoate as a colorless oil. 'H-NMR (CDC1 3 )8 1.21Cd, 3H, J=6.4 Hz), 3.41-3.48 (in, 1 3.77-3.81 (in, I 3.89 3 3.98-4.01 (in, 1 6.91-6.94 (in, 1 7.90-7.93 (in, 1 8.05-8.06 (in, 1 H).
A mixture of pentafluorophentyl 4-IN'-(2-methylphenyl)ureidolphenylacetate (508 ing, 1. 13 minol), methyl (S)-3-chloro-4-(2-amino-1I-propoxy)benzoate (275 mg, 1. 13 ninol) and Et 3
N
(0.240 nil, 1.72 minol) in DMP (10 inL) 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., and evaporated. The residue was recrystallized from MeOH-CHCI 3 -n-hexane to give 240 mg methyl (.S)-3-chloro-4-[2-[4-[N -(2-inethylphenyl)ureidojphenylacetylamino-1propoxy]benzoate as a white crystalline powder. 'H-NMR (DMSO-ds) 5 1.21 3 H, J=6.4 Hz), 2.25 3 3.33 2 3.82 Cs, 3H), 4.044.14 (mn, 3H), 6.90-6.94 Cm, 1H), 7.11-7.16 (in, 4H), 7.29-7.38 (mn, 3H), 7.83-7.94 (mn, 3H), 8.13-8. 17 (in, 2H), 9.34 Cs, 1 MS(FAB) ni/z 510 WO 01/00206 WO 0100206PCT/USOO/18079 To a stirred solution of methyl (S)-3-chloro4-12-14-[N'-(2-methylphenyl)ureidol phenylacetylaininoJ-l-propoxyJbenzoate (240 mng, 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-i N HCl and the solid was collected. The crude solid was recrystallized from MeOH-CHCI 3 -n-hexane to give 98 mg 274 as a white crystalline powder. mp 228-231 0(2; 'H-NMR (DMSOAd) 8 1.20 3 H, J=6.3 Hz), 2.24 3 3.34 2 4.02-4.18 (in, 3 6.92-6.95 (in, 1 7. 12-7.42 (series of m, total 7 7.82-8.18 (series of m, total 5 9.12 1 MIS (FAB) m/lz 496 497 Anal. Calcd for C262CN 3
O
5 112H 2 0: C, 61.84; H, 5.39; C21,7.02; N,8.32. Found: C2,61.76; H,5.25; (21,7.09; N,8.25.
Example 224 -chloro.-4-[2-[3-methoxy-4-[N '-(2-methylphenyl)ureidolphenylacetylamnino]-l1-propoxy] benzoic acid H
OOH
C31 H OH 3 275 A mixture of pentafluorophenyl 3-methoxy-4-[N -(2-methylphenyl)ureido]phenylacetate (513 mg, 1.07 mmol), methyl (S)-3-chloro-4-(2-anmino-1 -propoxy)benzoate (260 mg, 1.07 nunol) and Et 3 N (220 jud, 1.58 mxnol) 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,, and evaporated. The residue was recrystalized from MeOH-CHCI 3 -EtOAc-n-hexane to give 400 mng methyl (S)-3-cloro-4-[2-[3-methoxy-4-[N '-(2-inethylpheniyl)ureidojphenylacetylaniinoJ-1 propoxyjbenzoate as pale brown crystalline powder. 'H-NMR (DM50-cl) 8 1.21 3 H, J=6.4 Hz), 2.24 3 3.37 2 3.82 3 3.83 3 4.04-4.12 3 6.75-6.77 (mn, I 6.91-6.95 2 7.11-7.17 (in, 2 7.29-7.31 (mn, 1 7.78-7.99 (mn, 4 8.12-8. 13 (mn, 1 8.46 1 8.55 1 H).
To a stiffed solution of methyl (S)-3-chloro-4-[2-[3-inethoxy-4-[N'-(2 -iethylphenyl) ureidolphenylacetylaminol-1I-propoxyJbenzoate (400 mng, 0.74 inmol) in THF-MeOH (20 m.L, 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- I N HCI and the solid was collected. The crude solid was recrystallized from MeOH-CHCI,-Et 2 O to give 200 mg (5 275 as a pale brown crystalline powder. mp 198-20 1 0(2; 'H-NMR (DMS0-l 6 5 1.21 3 H, J=6.8 Hz), 2.24 3 3.37 2 3.84 3 4.00-4.15 (mn, 3 6.76-7.28 (series of in, total 6 7.77-8. 14 (series of mn, total 5 8.46 1 8.56 1 MS (FAB) rn' 526 528 Anal. Calcd for WO 01/00206 WO 0100206PCT/USOO/18079
C
27
H
2 gClN 3
O
6 -1/4H 2 O: C,61.I3;H,5.42;CI,6.68;N, 7.92. Found: C, 60.97; H,5.48; Cl,6.86; N, 7.89.
Examplie 225 3-dimethylaxnino-4-[[2-[3-methoxy-4-[A'-(2-methylphernyl)ureidolpheniylacetylI methylamninojethoxyjbenzoic acid 11H H CH 3 9
C
3 276 To a stirred and cooled (00 C) solution of 2-(N-Boc-N-methylamidno)ethanol (3g, l7minol), methyl 4-hydroxy-3 -nitro benzoate (3.38g, l7mmol), and Ph 3 P (5.4g, 2lmxnol) in THF mL) was added diisopropyl azodicarboxylate (DIAD) (4mL, 2 1 mmol), and the resulting mixture was heated under reflux for 15 hr. The solution was evaporated off. The residue was chromatographed on silica-gel with CHCI 3 :MeOH (100:0 to 4: 1, v/v) as eluent to give 2.5g (390) methyl 4-[2-(N-methyl-2-amino)ethoxyl-3-nitro benzoate as a pale yellow oil. 'H-NMR (CDC1 3 400MIHz) 8 2.82 3H), 3.50 2H, J=4.5Hz), 3.95 3H), 4.54 2H, J=4.5Hz), 7.26 IH, J=B.8Hz), 8.25 (ci, 11, J=8.8Hz), 8.56 lH); MS (FAB) m/z 255 (Mr A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetic acid (992mg, 3.1 Immol), methyl 4-(N-methyl-2-amidnoethoxy)-3-nitro benzoate (800mg, 3.1 Inunol) and 4- DMAP (77mg, 0.63mmol), HOBt (640mg,4.7mmol), and EDC (904mg,4.7nimol) in DMF 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 CHCl 3 :EtOAc (95:5 to 0: 100, vt) as eluent to give 587mg methyl 3-nitro-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureidopheniylacetyl] methylaminojethoxy]benzoate as a pale yellow oil. 'H-NMR (CDC 3 400MfHz) 8 2.31 3M1, 3.05 111), 3.23 211), 3.71 211), 3.83 3H), 3.85 (in, 3H), 3.94 311), 4.19 and 4.39 (in, total 2H), 6.80 (in, 2H1), 7.05 (in, 111), 7.22 (in, 311), 7.62 (ci, 1H, J=8.2Hz), 8.02 (ci, 111,. J=8.2Hz), 8.21 (dci, 111, J=2.lIHz, 8.8Hz, 8.55 (ci, lH, J=2.lIHz); MS (FAB)mwz 551(Mr +l1).
A mixture of methyl 3-nitro-4-[123-methoxy-4-[M-(2-methylphenyl)ureidolphenyI acetyllmethylamidnojethoxylbenzoate (587mg, 1. lmxnol) and 5 0 /o-Pd-C (600mg) in THF-MeOH- AcOH 1: 1, v/v, 15OnmL) was hydrogenated at 45psi for l8hr. 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-[A'-(2-methylphenyl)ureido~phenylacetyllmethylaminoethoxybenzoate as a pale yellow gum. 'H-NMvR (CDC 3 400MIHz) 8 2.29 311), 3.08 (in, 111), 3.19 2H1), 3.65 (s, WO 01/00206 WO 0100206PCT/USOOI 18079 111'), 3.73-3.80 (in, 3H1), 3.84 3.87(s, 311), 4.19 and 4.40 (in, total 2H), 6.70-6.82 (mn, 211), 7.02-7.29 (in, 611), 7.60 111, J=7.8Hz), 7.92-7.99 (in, 311); MS (FAB) ink 521 (M +lI).
To a stirred solution of methyl 3-amino-4-[[2-[3-methoxy-4-[MP-(2-methylphenyl)ureidol phenylacetyllmethylaminojethoxy]benzoate (555mg, 1. 1immol), formaldehyde (lOinL), and AcOH (0.58inL, l0mniol) in MeCN (lOinL) was added NaBH 3 CN (0.67g, l1inmol) 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 CHCI 3 The extract was washed with brine, dried over MgSO 4 and evaporated in vacuo. The residue was chroinatographed on silica-gel with toluene:acetone (7:3 to 1: 1, v/v) as eluent to give 123 mg (2 methyl 3-dimethylamnino-4-[[2-[3-inethoxy-4-[A'-(2methylphenyl) ureido]phenylacetylj inethylaminojethoxyjbenzoate as an oil. 'H-NMR (CDC1 3 400MIHz) 8 2.30 311), 2.70 311), 2.75 311, 3.05 111), 3.18 211), 3.61 311), 3.70 (s, 111), 3.80 (in, 311), 3.86 311), 4.07 and 4.22 (in, total 2H1), 6.28 (in, 111), 6.70-6.80 (mn, 311), 7.03 (in, 111), 7.15-7.25 (in, 411), 7.46-7.65 (in, 211), 8.02 (mn, 111); MS (FAR) ink 548 1).
A stirred mixture of methyl 3-diinethylamino-4-[[2-[3-inethoxy-4-[N'-(2-methylphenyl) ureido] phenylacetyllinethylaminojethoxylbenzoate (123mg, O.22mmol) in THF (15mL) and IN NaOH (0.885inL, 0.885mino1) 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 CHC1 3 -MeOH(9: 1, The extract was washed with brine, dried over MgSO 4 and evaporated in vacuo. The residue was crystallized with EtO-n:hexane to give 118 mg(l00%) 276 as a white crystalline material. mp 125-130 IR (KBr) 3346, 2940, 1620, 1597, 1535, 1456, 1417, 1227, 1039, 754cm-'; 'H-NNM (CD 3
OD,
400MIHz) 8 2.29 311), 2.70 311), 2.79 211), 3.05 111), 3.22 211), 3.75 311), 3.85 (in, 411), 4.15 and 4.28 (in, 211), 6.78-7.05 411), 7. 18 (in, 211), 7.55-7.70 (in, 311), 7.98 (in, 111; MS (FAR) Pnk 535(W Anal. calcd. for C~qH3N 4
O
6 -2.0H 2 O: C, 61.04; 1,6.7 1; N, 9.82.
Found: C, 61.15; H1, 6.43; N, 8.94.
Example 226 3 -diinethylaxnino-4-[[2-[3-methoxy-4-[A"-(2-fluorophenyl)ureidolphernylacetyllinethylaminoJ ethoxyjbenzoic acid
CH
3
~JJ.OOH
H H 6H 3 277 A mixture of 3-inethoxy-4-[N'-(2-fluorophenyl)ureido~phenylacetic acid(Ig,3.l11innol), methyl 4-[2-(N-inethyl-2-ainino)ethoxy]-3-nitrobenzoate (800mg, 3.11 inmol) and 4-DMAP WO 01/00206 WO 01/11206PCT/USOO/18079 (77mg, 0.63mmol), HOBt(640mg,4.7mmol), and EDC (904mg,4.7nol) 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,, and evaporated. The residue was chromatographed on silica-gel with CHCI 3 -EtOAc (95:5 to 0: 100, v/v) as eluent to give 420mg methyl 3-nitro-4-1[2-[3-fluoro-4-[N'-(2-fluorophenyl)ureidophenylaetylJ methylamino] ethoxy~benzoate as a pale yellow oil. 'H-NMR (CDCI,, 400MIHz) 8 3.04 1 H), 3.24 2H4), 3.72 111), 3.85 3H), 3.90 (in, 3H), 3.93 3H1), 4.16 and 4.39 (2m, 3H1), 6.80 (in, 2H), 6.99 (in, 1H), 7.05 (in, 2H1), 7.22 IH, J=8.8Hz), 7.51 2H), 8.00 (mn, 111), 8.08 (in, 111), 8.21 111, J=8.6Hz), 8.51 MS (FA.B) m/z 555 (MW 1).
A mixture of methyl 3-nito-4-[[2-[3-methoxy-4-[A'-(2-fluorophenyl)ureido]phenylacetyl methylaminolethoxy~benzoate (420mg, 0.76mmol) and 51%-Pd-C (1g) in THF-MeOH-AcOH 1: 1, v/v, 15OmL) 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-I[2- [3-inethoxy-4-[M-(2-fluorophenyl)ureido]phenylacetyllinethylaminojethoxy~benzoate as a pale yellow gum. 'H-NMRv (CDCI 3 400Mz) 8 3.05 and 3.13 total 311), 3.66 311, 3.70 211), 3.65-3.90 (in, 411), 3.86 311), 4. 10 and 4.23 (in, 211), 6.70-6.83 (in, 311), 6.98-7.15 (in, 611, 7.25-7.43 (in, 211), 7.99 (in, 111), 8.13 (in, 111); MS (FAB) m/~z 525 (Mt+ 1).
To a stirred solution of methyl 3-arnino-4-[[2-[3-inethoxy-4-[N'-(2-fluorophenyl)ureido phenylacetyl~inethylaminolethoxy]benzoate (397mg, 0.76inmol), formaldehyde (lOmL), and AcOH (0.43mnL, 7.6iniol) in MeCN (lOmL) was added NaBH 3 CN (0.48g, 7.6mmiol) 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 CHCI 3 The extract was washed with brine, dried over MgSO 4 and evaporated in vacuo. The residue was chroinatographed on silica-gel with toluene:acetone (7:3 to 1: 1, v/v) as eluent to give 123mg (2 methyl 3-dimethylamino-4-[[2-[3-inethoxy-4-[A'-(2inethylphenyl)ureido~phenylacetylJ inethylaminoJethoxylbenzoate as an oil. IH-NMR (CDCl 3 400NMz) 8 2.74 311), 2.77 311), 3.08 111), 3.22 211, 3.52 31-0, 3.61 111, 3.83 (in, 311), 3.88 311), 4.12 and 4.23 (in, total 211, 6.68 111), 6.78 (in, 211), 6.98 (in, 211, 7. (in, 111), 7.55-7.68 (in, 411, 7.99 (in, 11-, 8.16 111, J=8.3Hz); MS (FAB)m./z 553 A stirred mixture of methyl 3-diinethylaniino-4-[[2-[3-inethoxy-4-[AK-(2-fluorophenyl) ureido] phenylacetyl]methylaminolethoxylbenzoate (61mg, 0. 1 Iinnol) in THE (l5mL) and IN NaOH (0.22niL, 0.22minol) was heated under reflux: for 15 hr. The pH of the midxture was adjusted to 5. 0 by the addition of IN HCI, and extracted with CHCl 3 :MeOH(9: 1, The extract WO 01/00206 WO 0100206PCT/US001 18079 was washed with brine, MeOH:acetone (93:7, vlv) as eluent to give 37 mg 277 as a white crystalline material. mp 120-125 'H-NMR (CD 3 OD, 400M]4z) 8 2.60 4H1), 2.78 2M1, 3.06 1IM, 3.22 2H1), 3.75 3H1), 3.85-3.92 (in, 4H), 4.17 and 4.29 (in, total 2M-1, 6.80-7.12 (in, 7.6 1-7.70 (in, 2M1, 8.00 (mn, 111), 8.08 (in, 111); MS (FAB) 539 Anal. calcd. for
CIH
31
FN
4
O
6 -2.75H 2 O: C, 57.18; H, 6.26; N, 9.53. Found, C, 57.20; H, 5.62; N, 9.06.
Example 227 3-dimethylamnino-4-[[2-[4-IN'-(2-methylpheniyl)ureidolphenylacetyllmethylaminolethoxyl benzoic: acid 911 3 0011
H
3
CH,
11311 H278 A mixture ofpntfurophenyl [4-[MP-(2-methylphenyl)ureido~phenyllacetate (1 .42g, 3. l5miol), methyl 4-[2-(N-methyl-2-amino)ethoxyJ-3-nitro benzoate (800mg, 3. l5minol) and triethylamine (0.66mL,4.73mxnol) in DMF (8 mL) was stirred at 50' C for 15 hr. The mixture was poured into ice-i IN HCI and extracted with CHCI,. The extract was brine, dried over Na 2
SO
4 and evaporated. The residue was chromatographed on silica-gel with CHCI 3 :EtOAc (95:5 to 0: 100, vtv) as eluent to give 1.04g methyl 3-nitro-4-[[2-[4-IN'-(2-methylphernyl)ureidoj phenylacetyl]methylaxninojethoxylbenzoate as a pale yellow oil. 'H-NMR (CDCl 3 400MHz) 8 2.30 3H), 2.90, 3.02 and 3.05 total IM, 3.22 211), 3.71 1IM, 3.85 3H), 3.93 (s,3M1, 4.19 and 4.39 (mn, 2M1, 7.02 (in, 1IM, 7.19 (in, 411), 7.35 lH, J=8.3Hz), 7.40 111, J=8.OHz), 5 211, 7.70 (in, 111), 8.22 111, .fr6.7Hz), 8.51 11H); MS (FAB) m/z 5 2 1 (v+l1).
A mixture of methyl 3-nitro-4-[[2-[4-[N'-(2-methylphenyl)ureidolpheniylacetylI methylaminojethoxylbenzoate (1 .04g, 2innol) and 5%/-Pd-C (1 .2g) in THF-MeOH-AcOH 1: 1, v/v, 15OmL) 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-amiino-4-[[2-[4-[N'-(2-methylphenyl) ureido]phenylacetyl]methylaminoethoxy~benzoate as a pale yellow gum.
To a stirred solution of 3-amnino-4-[[2-[4-[Nr-(2-inethylphenyl)ureido~phenylacetylI inethylaminoJethoxy~benzoate, formaldehyde (SmL), and AcOH l4mL, 20mniol) in MeCN 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 CHCI 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 85mg (2 WO 01/00206 WO 0100206PCT/USOO/18079 steps, methyl 3-dimethylamino-4-[[2-[4-[N'-(2-methylphenyl)ureidophenylacetyllmethy anunojethoxylbenzoate as an oil. 'H-NMR (CDCI 3 400M~z) 8 2.12 311, 2.73 31-i), 2.75 (s, 311), 3.05 111), 3.20 211), 3.60 111), 3.80 (in, 311), 3.88 311), 4.17 (mn, 21H), 6.95-7.28 (in, 811), 7.55-7.75 (mn, 311); MS (FAB) m/z 518 (W 1).
A stirred nixture of methyl 3-dimethylaniino-4-[[2-[4-[M-(2-methylphenyl)ureidol phenylacetyl] methylaininojethoxylbenzoate(ap3 15201)(85mg, 0. l6mniol) in THF (I5mL) 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 HCI, and extracted with CHC1 3 :MeOH(9: 1, The extract was washed with brine, dried over MgSO 4 and evaporated in vacuo. The residue was crystallized from Et 2 O to give 53 mg(65%) 278 as a white crystalline material. mp 110-115 0 C; '11-NMVR (CDOD, 400MFz) 8 2.29 311), 2.75 311), 2.76 311), 3.02 111, 3.20 211), 3.72 (s, 111), 3.85 (mn, 311), 4.18 and 4.28 (in, total 211), 6.95-7.03 (in, 211), 7.18 (mn, 411, 7.34 111, J=8.3Hz), 7.38 111, J=8.8H4z), 7.62 111, J=8.3Hz), 7.66 111), 7.80 (in, 111; MS (FAD) nu/z 505 1).
Examuple 228 3-spoyaio4[2[-ehx--A-2furpey~riopeyaey~ehlnio ethoxy~benzoic acid Yy'-Q&W H3%N H ~H 6CH 3 H 3 279 To a stirred cold (00 C) solution of methyl 3-ainino-4-[[2-[3-methoxy4-[Mr-(2-fluoro phenyl)ureidolphenylacetyllmethylamino~ethoxy~benzoate (I100mg, 0. l9mrnol) in acetone AcOH/DMF (13 inL, 6:6: 1, v/v/v) was added NaBH 3 CN (300 mng), 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 CHC1 3 (2OinL), and the solution was washed with brine, dried over MgSO,, and evaporated under a reduced pressure. The residue was chroinatographed on silica-gel plate with toluene:acetone 1, v/v) as eluent to give 108 mng (100%) methyl 3-isopropylainino-4-[[2-[3-methoxy-4-[N'.(2-fluorophenyl)ureidoJphenylacetylI inethylaminojethoxylbenzoate as a colorless oil. 'H-NMR (CDCI,, 40NMHz) 5 1.20 (in, 111), 2.88 611), 3.02 and 3.12 total 3H1), 3.53 211), 3.60-3.80 (mn, 7H-1), 3.85 311), 4. 10 and 4.20 (mn, 211), 6.65-675 (mn, 211), 6.90-7.08 (mn, 211), 7.22-7.35 (mn, 211), 8.02 211), 8. 10 (mn, 211), 8.21 (br, 111), 8.3 3 (br, 11H); MS (FAB) ni/z 566 (W 1).
WO 01/00206 WO 0100206PCT/USOO/18079 A stirred mixture of methyl 3-isopropylamino-4.{[2-[3-methoxy-4-[N'-(2-fluorophenyl) ureido]phenylacetyllmethylan-unolethoxylbenzoate (I116mg, 0.2mmol), 0.25 N NaOH (6 niL), and THE (6 niL) was heated under reflux: for 8 hr. The mixture was poured into water (200 mL) acidified by IN HCI and the solid was collected with suction. The solid was recrystallized from CHCI,-n-hexane-diisopropylether to give 56mg 279 as a colorless crystalline powder. mp 195-200 OC; 'H-NMR (CD 3 OD, 400Mqz) 8 1. 15-1.20 (in, 3.01 114), 3.12 211!), 3.48- 3.60 (in, 111), 3.68 3H), 3.75 2H1), 3.82 111), 3.86 (in, 3H1), 4.15-4.23 (in, 2H), 6.80 (im, 3H), 4.154.23 (in, 2H), 6.80 (in, 3H1), 6.98 (in, 111, 7. 10 (in, 2M1, 7.20 and 7.25 total 111), 7.32 (in, 111), 7.98 (in, 1IM, 8.05 (in, 111); MIS (FAB) m/lz 552 Anal. calcd. for
C
2
H
3
,FN
4 0, 6 1.0H 2 0: C, 61.04; H, 6.18; N, 9.82. Found, C, 61.36; H, 6.25; N, 9.45.
Examole 229 4-1111 r(2furpeylued]3-ehxpeylctl--mtyainj--ehl2 propoxylbenzoic acid H C3 280 To a stirred solution of 2-amino-2-methyl-1I-propanol (8.4g, 93.89mmol) and triethylainine (I11.4g, 0.l1l3mol) in DMF-water 1, vfv, lOOmL) was added di-tert-butyl dicarbontate (25g, 0. 1 I5mol) at 5 to 100 C. The resulting solution was stirred for 2 hr at room temp. The mixture was diluted with water(lOOniL) and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SO
4 and evaporated. The residue was chroniatographed on silica-gel with CH 2
CI
2 as eluent to give 12g 2-tert-butoxycarbonylamino-2-methyl-1Ipropanol as a syrup. 'H-NMR (CDCI,) 8 1.25 6H1), 1.43 9H1), 3.59 J=8.3Hz, 2H), 4.68 (br, 111).
To a stiffed suspension of 2-tert-butoxycarbonylamino-2-methyl-1-propano 7g, 30.11 minol) and powdered NaOH(6.7g, 0. 15 1 mol) in Et 2 O(200mL) was added p-toluenesulfonyl chloride (6.9g, 36. l4nol) at room temp. The stirred resulting mixture was heated under reflux for 8 hr. After cooling, ice-water(lOOmL) was added to the solution. Separated Et 2 0 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 2-tert-butoxycarbonylamino-2-methyl-lpropyl p-toluenesulfonate as a crystalline material. 'H-NMR (CDCl 3 8 1.26 611), 1.38 911), 2.37 311), 4.05 2H1), 4.49 (br, 111), 7.34 J=7.8Hz, 211, 7.78 J=7.811z, 211.
A stirred mnixture of 2-tert-butoxycarbonylamiino-2-methyl- I -propyl p-toluenesulfonate WO 01/00206 WO 0100206PCT/USOO/18079 (8.2g, 23.88mmol) and powdered NaOH (6.7g, 0. 15 Imol) in EtO (200mL) was heated under refiux for 10 hr. After coaling, the mixture was filtered. And the filtrate was washed with water, brine, dried over Na 2
SO
4 and evaporated. The residue was chroniatographed on silica-gel with nhexane:.EtOAc 1, v/v) as eluent to give 2.7g 1-tert-butoxycarbonyl-2-methylpropylene imine as an oil. 'H-NMR (CDCI 3 8 1.29 611), 1.47 9H), 2.05(s, 2M1.
To a stirred solution of 1-tert-butoxycarbony-2-methylpropyleneimnine (1.03g, and methyl 4-hydroxybenzoate (800mg, 6.26mmol) in CH 2 Cl 2 (IOmL) was added boron trifluoride diethyl ether 127mL, I mmol) 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,, and evaporated. The residue was chromatographed on silica-gel with n-hexane:EtOAc 1, v/v) as eluent to give 550mg methyl 4-(1-tert-butoxycarbonylamino-2-methyl-2-propoxy)benzoate as a gum. 'H-MIR (CDCI 3 8 1.33 611), 1.47 911), 3.36 J=6.3Hz, 211), 3.90 311), 5.05 (br, 111), 6.99 J=8.8Hz, 211), 7.97 J=8.8Hz, 2H-).
A mixture of methyl 4-(l1 -tert-butoxycarbonylamino-2-methyl-2-propoxy)benzoate (460mg, 1.42mmol) and anisole 155mL, 1.42mmol) in CH 2
CI
2 (l5mL) and TFA (3niL) was stirred for 3 hr at room temp. The mixture was evaporated off. The residue was dissolved in
CH
2
CI
2 (3OniL) and made basic by the addition of 0. 5N NaOH. The CH 2
CI
2 layer was separated, dried over Na 2
SO
4 and evaporated. The residue was chromnatographed on silica-gel with CH 2 Cl 2 as eluent to give 370mg (100%) methyl 4-(1-amnino-2-methyl-2-propoxy)benzoate as a gum. 'H- NMR (CDCl 3 8 1.34 6M1, 2.87(s,2H),3.90 10 J=8.8Hz,2H), 7.97 J=8.8Hz, 2H).
To a stirred solution of methyl 4-(1-anmino-2-methyl-2-propoxy)benzoate (370mg, 1.66mmol) and tniethylamine (0.35mL, 2.49mmrol) in C11 2
C
2 (l5mL) was added trifluoroacetic anhydride (0.3 l6nL, 2.24nunol) at 00 C. After stirred for 1 hr at the same temp, water was added to the solution. CH 2 Cl 2 layer was separated, washed with water, dried over Na 2
SO
4 and evaporated. The residue was chromatographed on silica-gel(2OmL) with CH 2
CI
2 as eluent to give 530mg (100%) methyl 4-(1.trifluoroacetamiido-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 1-trifluoroacetamido-2-methyl-2-propoxy)benzoate (530mg, 1.66mmol) and K 2 C0 3 (345mg, 2.49mmol) in DMF (lOniL) was added Mel (0.l14mL, 2.37mmol) at roam 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 WO 01/00206 WO 01/11206PCTUSOOII8079 brine, dried over Na 2
SO
4 and evaporated. The residual gum was used to the subsequent reaction without fuirther purification.
The above crude residue was dissolved in MeOH(lOniL). To the stirred solution was added water (5miL) and NaCO,(352mg, 3.32mmol), and the resulting mixture was stirred for 5 hr at room temp. The mixture was poured into water and extracted with CHCI 3 The extract was washed with water, dried over Na 2
SO
4 and evaporated. The residue was chromatographed on silica-gel with OCI 3 as eluent to give 390mg (100%) methyl 4-(1-methylamino-2-methyl-2propoxy)benzoate as a gum. 'H-NMR (CDCI 3 8 1.37 611, 2.51 311, 3.89 311, 6.92 (d, J=8.8Hz, 211), 7.97 J=8.8Hz, 211).
To a stirred mixture of methyl 4-(1-methylanmino-2-methyl-2-propoxy)benzoate (200mg, O.S4mmol), 4-[N'-(2-fluorophenyl)ureidoj-3-methoxyphenylacetic acid (268mg, 0. 84mmol), 4l.Ommol) in DMF(5niL) was added EDC(220mg, 1. l4mmol) 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 NaSO,, and evaporated. The residual gum was triturated with CHCI 2 and Et 2 O to give 200mg (44. methyl 4-[I-[4-[M,-(2-fluorophenyl)ureidoJ -3-methoxyphentylacetylJmethylamninoJ-2-methyl-2propoxylbenzoate as a crystalline material. 'H-NMR (CDCI 3 8 1.36 and 1.31 (each s, 611), 3.24- 3.88 (series of s, 1311), 6.65-8.20 (seris ofmi, 1411).
A mixture of methyl 4-(N'-(2-fiuorophenyl)ureidoJ-3-methoxyphenylacetylJ methylaminoj-2-methyl-2-propoxy]benzoate( 180mg, 0.335mmo1) in THF(3mL) and 0. NaOH(4mL) was stirred for 5 hr at room temp. The mixture was poured into ice-IN Thc solid was collected, washed with water, and air-dried. The crude solid was recrystallized from EtOH-CHCI,-n-hexane to give 70 mug (40%)280 as fine needles. rup 200-207 OC; IH-NMR (DMSO-d 6 8 1.26 and 1.33 (each s, 61H), 3.70-3.8 1 (series of s, 711), 3.83 311), 6.75-8.20 (series of mn, 1011), 8.71 111), 9.17 (br s, 111), 12.72 111).
Example 230 (.S)-3-chloro-4-12-[3-methoxy-4-[N '-(2-fluorophenyl)ureido~phenylacetylaminoJ-l1-propoxy] benzoic acid H 0 1 H HL~ CH 3 2H81J~CO A mixture of 3-methoxy-4-[N '-(2-fluorophenyl)ureido]phenylacetic acid (327 mg, 1.03 WO 01/00206 WO 0100206PCTUSOO/1 8079 mmol), methyl (S)-3-chloro-4-(2-anmino-1-propoxy)benzoate (250 mg, 1.03 mmol), EDC(hydrochloride) (295 mg, 1.54 nunol), HOBt (208 mg, 1.54 mmol), and DMAP (25 mg, 0.20 mmol) in DMF (8 niL) was stirred at room temp overnight. The mixture was diluted with EtOAc, washed with 0.5 N HCI, brine, dried over Na 2 SO, and evaporated. The residue was recrystallized from CHC1 3 -EtOAc to give 308 mg as a white crystalline powder. IH-NMvR (CDCI 3 8 1.29 3 H, J=6.8 Hz), 3.51 2 3.84 3 3.88 3 4.01-4.08 (in, 2 4.38-4.39 (in, 1 6.25 1 H, J=8.3 Hz), 6.78-6.83 (in, 2 6.90-6.95 (mn, 2 7.02-7.11 (mn, 2 7.89 (dd, 1 H, J=2.0, 8.8 Hz), 8.02 1 H, J=2.4 Hz), 8.20 1 H, J=8.3 Hz), 8.27-8.3 1 (in, 1 8.53 (s, 1 8.84 1 H).
To a stirred solution of methyl (S)-3-chloro-4-[2-[3-methoxy-4-[N'-(2-fluorophenyl) ureido]phenylacetylanuinoJ-l-propoxylbenzoate (308 mg, 0.57 nunol) in THiF-MeOH (10 miL, 1: 1, v/v) was added 0.5 N NaOH (10 niL), and the reaction mixture was heated under refL~x for 1 hr.
The mixture was poured into ice-i N HCl and the solid was collected. The crude solid was purified by recrystallization from MeOH-CHCI 3 -n-hexane to give 196 mng (651%) 281 as a white crystalline powder. mp 188-191 'H-NMR (DMSO-djj5 1.21 3 H, J=6.4 Hz), 3.38 2H), 3.83 3 4.02-4.18 (in, 3 6.78-7.29 (series of mn, total 6 7.85-8.00 (in, 3 8.12-8.19 (in, 2 8.70 1 9.17 1 12.98 (bs, 1 MIS (FAB) m/z 530 531 532 Anal. Calcd for ,HCFN 3
O
6 -l/4H 2 O: C, 58.43; H, 4.8 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.
Example 231 4-[[2-[3-methoxy-4-[AM-(2-methylphenyl)ureidolphenylacetyllmethylamino]ethylaminolbenzoic acid Hf
OOH
6HH H OCH 3 282 A stirred solution of N-benzyloxycarbonyl-N-methylaminoacetaldehyde (1.77g, 1. 1ininol) in toluene (3mL) was added methyl-4-aminobenzoate (1.29g, 8.Snunol), 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 niL). To the solution was added AcOH (2.44inL, 43imnol) and NaBH 3 CN (2.67g, 43inmol), 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 with EtOAc, washed with brine, dried over MgSO,, and evaporated. The residue was chroinatographed on silica-gel with n-hexane: EtOAc v/v) as eluent to give 1 .26g WO 01/00206 WO 0100206PCT/USOO/18079 methyl 4-[2-(N-benzyloxy carbonyl-N-methylamino)-ethylaniino] benzoate as a colorless oil. NM4R (CDCl 3 400M]Hz) 5 2.96 3.32 (br m, 2H), 3.50-3.60 (in, 211i), 3.82 3H), 5.15 (each d, 211, J= 14.61-z), 6.35 and 6.58 (in, total 2H), 7.36 511), 7.76 and 7.84 (in, total 2H); MS (FAB) m/z 342 To a stirred solution of methyl 4-[2-(N-benzyloxycaxbonyl-N-methylaniino)ethylainino benzoate (1.26g, 3.68mmol) in MeOH(20 inL) 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 methyl 4-[2-(N-methylaxnino) ethylaniinolbenzoate as a colorless oil. 'H-NMR (CDCI 3 400M41z) 8 2.46 3H), 2.88 2H, J=5.5Hz), 3.27 (br s, 3.85 3H), 6.57 2H, J=8.8Hz), 7.85 2H, J=8.8Hz); MS (FAB) m/z 209 (M +l1).
To a stirred solution of methyl 4-12-(N-methylamidno)ethylaxninolbenzoate (590mg, 2.83rrnol)3-methoxy-4-[N'-(2-inethylpheniyl)ureidojphenylacetic acid (89 lmg,2.83mmnol) in DMF(14niL) was added EDC (815ing,4.2Smmol), HOBt (574mg,4.25smnol),and 4-DMAP (5 19mg, 4.25mmxol), 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 CHC1 3 -EtOAc (10:0 to 7:3, v/v) as eluent to give 1 .25g methyl 4-[-3mtoy4[P(-ehlhnluedlhnlctl methylaminolethylaininolbenzoate as a light yellow oil. 'H-NMR (CDCl 3 400M4Hz) 83.18 (s, 2H), 3.05 and 3.32 total 2H), 3.72-3.85 (in, 911I), 4.12 and 4.23 (in, total 2H), 6.78 (in, 3H1), 7.05 IH, J=7.5Hz), 7.20 (in, 2H1), 7.43-7.50 (in, 3H), 7.62 111, J=8.8Hz), 8.05 111, J=8.8Hz); MS (FAB) m/z 505 +1 A stirred mixture of methyl 4-[[2-[3-methoxy-4-[A"-(2-methylphenyl)ureidolphenylacetylI methylaminojethylaminolbenzoate (300mg, 0.6minol) in 0.25 N NaOH (6 mL) and THF (6 mQL 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-hexanediisopropylether to give 202mg 282 as a pale yellow crystalline powder. mp 115-120 'C; IR(KBr) 3346, 2935, 1603, 1531, 1454, 1417, 1257, 1174, 1036, 754cm-'; 'H-NMvR (CD 3
OD,
400MIHz) 8 2.28 3H1), 2.98 11H), 3. 10 2H), 3.3 5-3.42 (in, 211), 3.5 3-3.65 (in, 3H), 3.70 (s, 3.80 and 3.82 311i), 6.60-6.85 (in, 411), 7.02 (mn, 7.18 (mn, 211), 7.58 (in, 11-1), 7.82 (in, 2H1), 7.96 (in, I1-H); MS (FAB) m/lz 491 (W 1).
WO 01/00206 WO 0100206PCT/USOO/18079 Example 232 (S)-3-chloro-4-[2-IN-methyl-N-[3 -methoxy-4-[N '-(2-fluoropbenyl)ureidolphenylacetylI amiino]-i1-propoxy]benzoic acid
CH
3
OOH
CWk 6H 3 I23 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 CI, (20 mnL) was added Et 3 N 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 CHCI 3 washed with 0.5 N HCI, brine, dried over Na 2
SO,
and evaporated. The residue was recrystallized from CHCI 3 -n-hexane to give 1.59 g chloro-4-(2-trifluoroacetamiido-lI-propoxy)benzoate as a white crystalline material. mp 122-125 0 C; 'H-NMR (CDC1 3 8 1.48 3 H, J6.8 Hz), 3.91 3 4.10-4.19 (in, 2 4.47-4.52 (in, 1 6.68 (bs, 1 6.92-6.94 (in, 1 7.93-7.95 (im, 1 8.07-8.08 (in, I MS (FAB) m/z 340 A nal. Calcd for C 3 H 1 3 C1F 3 N0 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.
To a stirred solution of methyl (S)-3-chloro-4-(2-tri~fluroacetamido- 1 -propoxy)benzoate (800 mng, 2.36 mmol) in DMF (5 mL) was added K 2 C0 3 (651 minol, 4.71 inmol) and Mel (0.22 mL, 3.53 inmol), and the reaction mixture was stirred at 600 C 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 chromatography on silica-gel with 5% EtOAc in CHC1 3 as eluent to give 850 ing (100%) methyl (S)-3-chloro-4-[2-(N-inethyl-Nf-trifluoroacetamido)-l-propoxy benzoate as a colorless oil. 'H-NMR (CDCI 3 5 1.43-1.46 (in, 3 3.03 and 3.21 3 3.90 (s, 3 4.04-4.22 (in, 2 4.81-4.87 (in, 1 6.91 1 H, J1=8.3 Hz), 7.93 (dd, 1 H, J=2.0, 8.3 H), 8.06 1 H, J1=2.0 Hz).
To a stirred solution of methyl (S)-3-chloro-4-12-(N-iethyl-N-trifluoroacetamido)-lpropoxyl benzoate (880 mg, 2.49 mmnol) in MeOH-H1 2 0 (10 inL, 1: 1, v/v) was added KC0 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,0, brine, dried over Na 2
SO
4 and evaporated to give 460 mng (S)-3-chloro-4-(2-inethylainiino-l-propoxy)benzoate as a colorless oil. 'H-NMR (CDCI 3 8 1.20(d, 3 H, J=6.4 Hz), 2.51 3 3.07-3.12 (in, 1 3.89 3 3.92-4.04 (in, 2 6.93- 6.95 (in, 1 7.90-7.93 (in, 1 8.05-8.06 (in, 1 H).
WO 01/00206 WO 0100206PCTIUSOO/I 8079 A mixture of 3-methoxy-4-[N 2 -fluorophenyl)ureidolphenylacetic acid (296 mg, 0.93 inmol), methyl (S)-3-chloro-4-(2-methylamino-1 -propoxy)benzoate (240 mg, 0.93 mmol), EDC(hydrochloride) (268 mg, 1.40 inmol), HOBt (189 mg, 1.40 inmol), and DMAP (23 mg, 0. 19 inmol) in DMF (8 mL) was stirred at room temp for 1.5 hr. The mixture was diluted with EtOAc, washed with 0.5 N HCI, brine, dried aver 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 520 mg (100%/) methyl (S)-3-chloro-4-2-[N-methylN-13-methoxy-4-[N'-(2-fluorophenyl)ureidolphenylacetylJ aminoj-1I-propoxyjbenzoate as a red-brown amorphous solid.
To a stirred solution of this product (520 mg, 0.93 inmol) in THF (10 mL) was added N NaQH (10 mL), and the resulting mixture was heated under reflux for 3 hr. The mixture was poured into ice-i N HCI and the solid was collected. The crude solid was recrystallized from CHCl 3 -Et 2 O to give 170 mg (2 steps, 371/) 283 as a white crystalline powder. mp 142-147 0 C; 'H- NIVR (DMSO-4) 8 1.13-1.20 (in, 3 2.74 and 2.94 3 3.65 2 3.82 and 3.84 3 4.13-4.22 2 4.53 and 4.91-4.92 (in, 1 6.71-7.29 (series of m, total 6 7.86-8.02 (in, 3 8.15-8. 19 (mn, I 8.71 1 9.17 1 13.00 (bs, 1 MS (FAB)m/Lz 544 545 Anal. Calcd for C2711 7 ClFN 3
O
6 -1/4H1 2 O: 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)ureido]phenylacetylI amino- I -propoxyjbenzoic acid %H3
OH
CH
3 H H OCH 3 284 A mixture of 3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetic acid (261 mng, 0.83 inmol), methyl (S)-3-chloro-4-(2-methylaxnino-l-propoxy)benzoate (214 mg, 0.83 rumol), EDC(hydrochloride) (239 mg, 1.25 inmol), HOBt (168 mg, 1.24 nunol), and DMAP (20 mg, 0. 16 inmol) in DMF (8 mL) was stiffed at room temp 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 CHCI,-MeOH (50: 1, v/v) as eluent to give 470 mng (100%) methyl (S)-3-chloro-4-[2-[N-methyl-N-[3-methoxy-4-[N '-(2-methylphenyl)ureido] phenylacetyllaiuinoj-1-propoxylbenzoate as a pale yellow amorphous solid.
To a stirred solution of the above product (470 mng, 0.95 mimol) in THF (10 mL) was WO 01/00206 WO 0100206PCT/USO0I 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- I N HCl and the solid was collected. The crude solid was recrystallized from CHCI 3 -Et 2 O to give 168 mg (2 steps, 33%) 284 as a pale yellow crystalline powder. mp 125-130 'H-NMR (DMSO-d) 5 1.13-1.19 3 2.24 3 2.74 and 2.94 3 3.65 2 3.83 and 3.85 3 4.144.22 (in, 2 4.91-4.93 (in, I 6.70-6.74 (in, 1 6.84 (in, 1 6.92-6.95 (in, 1 7.11-7.17 (in, 2 7.25-7.29 (in, 1 7.78-7.80 (in, 1 7.85-7.93 (in, 2 7.99-8.02 (m j1 8.46 I 8.56 1 12.99 (bs, 1 MS (FAB) m/z 540 541 Anal. Calcd for C2uH13CIN 3
O
6 -1f2H 2 C, 61.26; H, 5.69; N, 7.65. Found: C, 61.15; H, 5.58; N, 7.51.
Example 234 3-isopropylaxnino-4-[[2-[3 -methoxy-4-[AM-(2-methylphenyl)ureidolphenylacetyllmethylaminoI ethoxylbenzoic acid
?H
3 0011O 9K
H
3
%NH
611 3 11H H 3 6H 3 285 To a cold (00 stirred solution of methyl 3-aniino-4-[[2-[3-methoxy-4-[A"-(2-methyl phenyl) ureidolphenylacetyl]methylaninolethoxylbenzoate (300mg, 0.58mmol) in acetone-AcOH- DWvI (13 mL, 6:6: 1, vfvfv) 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 mng (86%) methyl 3-isopropylamino-4-[[2-[3-methoxy-4-[A'-(2-methylphenyl)ureido]phenylacetyl methylanuinolethoxyjbenzoate as a colorless oil. 'H-NMR (CDCl 3 400M4Hz) 8 1.21 (in, 6H), 2.30 3H), 3.05 and 3. 10 total 2H), 3.59 3H), 3.62-3.81 (in, 6H), 3.89 3H), 4. 10 and 4.21 (in, 2H), 6.65-6.80 (in, 414), 7. 10 (mn, 114), 7.20 314), 7.32 (in, 2H1), 7.54 lH, J=8.3Hz), 8.00 IlH), 8.07 IH, J=8.3H-z); MIS (FAB) m/z 563 (Nr+l1).
A stirred mixture of methyl 3-isopropylamino-4-[[2-[3-methoxy4-[M,-(2-methylphenyl) ureidojphenylacetyllinethylamiriolethoxylbenzoate (280mg, 0.5minol), 0.25 N NaOH (6 mL), and THF (6 mL) was heated under reflux for 8 hr. The mixture was poured into water (200 niL) acidified with IN HCI and the solid was collected wth suction. The solid was recrystallized ftrm
CHCI
3 -n-hexane-diisopropylether to give 202mg 285 as a light yellow crystalline powder.
mp 130-135 'H-NNM (CD 3 OD, 400MIHz) 5 1. 18 and 1.22 total 6H, J=6.3Hz), 2.29 and 2.32 total 3H), 3.04 and 3.14 total 2H), 3.60-3.90 (mn, 9H), 4.16 and 4.25 (in, total 2H), 6.80 (in, 314), 7.02 (mn, 7.12-7.24 (mn, 3H), 7.29-7.38 (in, 111), 7.59 (in, 114), 8.02 (in, lH); MS WO 01/00206 WO 0100206PCT/U 500/1 8079 (FAB) m/z 548 Anal. calcd. for C~jH3NO 6 C, 65.68; H, 6.61. Found: C, 65.80; H, 6.83.
Example 235 4-[[2-[3-methoxy-4-[IV-(2-methylphenyl)ureidolphenylacetyllmethylaminolethyllmethyLarnino benzoic acid H3 OOH 3 H H OCH 3 286 To a stirred cold (00 C) solution of methyl 4-[[2-[3-methoxy-4-IM-(2-methylphenyl) ureidojphenylacetyljmethylaininojethyllaminobenzoate (300mg, 0.6mxnol) in MeCNformaldehyde-AcOH (I11 mL, 8:2: 1, v/v/v) was added NaBH 3 CN (187 mg, 2.40 mmol), and the resulting mixture was stiffed for 18 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 309 mg (100%) methyl 4-[[2-[3-methoxy-4-[N'"-(2-methylphenyl)ureidolphernylacetylJ-2-N'methylaininolethyl]N-mediylaminobenzoate as a colorless oil. 'H-NMvR (CDCl 3 400M1Hz) 82.30 (in, 3H), 2.98 (in, 7H), 3.46-3.72 (in, 9H), 3.82 (in, 3H), 6.32 (in, 6.55-6.75 (in, 4H), 7.05- 7.50 (in, 2H), 7.82-8.02 (in, 4H); MS (FAB)m/z 518 (M 1).
A stirred mixture of methyl 4-[-3mtoy4[,(-ehlhnluedlhnlctl -2-N'-methylamino]ethyl]methylaminobenzoate (309mg, 0.6minol) in I N NaOH (2.4 mL), and THF (10 ml) was heated under reflux for 8 hr. The mixture was poured into water (200 inL) acidified by the addition of IN HCI. The solid was collected with suction. The crude solid was recrystallized from CHCI 3 -n-hexane-&isopropylether to give 211 mg 286 as a light yellow crystalline powder. mp 125-130 IR(KBr) 3338, 2933, 1601, 1529, 1182, 1036, 752cm-'; 'H- NMR (CD 3 OD, 400MfHz) 5 2.28 and 2.29 3H), 2.95-3.02 (in, 6H), 3.60 (br, 6H), 3.80 11H), 3.86 2H), 6.60-6.82 (mn, 4H), 7.01-7.18 (in, 3H), 7.58 (in, 11H), 7.82-7.99 (in, 3H); MS (FAB) m/~z 504 (M 1).
Example 236 (S)-3-chloro-4-[2-[N-benzyl-N-[3-methoxy-4-[N -(2-fluorophenyl)ureidoJ phenylacetyllanminoJ- 1propoxy]benzoic acid r-o COOH r J-r3287 To a stirred solution of NaBH 3 CN (985 mng, 15.68 inmol) in MeOH (5 inL) was added a WO 01/00206 WO 0100206PCT/USOO/18079 solution of methyl (S)-3-chloro-4-(2-axnino-1-propoxy)benzoate (382 mg, 1.57 mmol) and benzaldehyde 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,. The extract was washed with brine, dried over Na 2 SO, and evaporated. The residue was purified by column chromatography on silica-gel with CHCI 3 :MeOH (30: 1, v/v) as eluent to give 336 mg (64 chloro-4-(2-N-benzylainino-1-propoxy)benzoate as a colorless oil. 'H-NMR (CDCI 3 )8 1.22 3 H, J=6.4 Hz), 3.21-3.25 (in, 1 3.84-4.03 (in, total 7 6.89-6.91 (in, 1 7.23-7.37 (in, 5 H), 7.89-7.9 1 (in, 1 8.05 (in, 1 H).
A mixture of 3-methoxy-4-[N -(2-fluorophenyl)ureido~phenylacetic acid (320 mg, 1.01 mmol), methyl (S)-3-chloro-4-(2-N-benzylamino-1-propoxy)benzoate (336 mg, 1.01 nunol), EDC (hydrochloride) (289 mg, 1.51 minol), HOBt (204 mg, 1.51 mmol) and DMAP (25 mg, 0.20 mmol) in DMT (7 niL) was stirred at room temp for 2 days. The mixture was diluted with EtOAc, washed with 0.5 N HC1, brine, dried over NaSO 4 and evaporated. The residue was purified by column chromatography on silica-gel with CHCI 3 :MeOH (50: 1, v/v) as eluent to give 607 mg methyl (S)-3-chloro-4-[2-[N-benzyl-N-[3-methoxy-4-jN '-(2-fluorophenyl)ureidojphenyl acetyljaniino]-l-propoxylbenzoate as a pale yellow amorphous solid. 'H-NMR (CDCl 3 81.20- 1.27 (in, 3 3.62 2 3.73-4.16 (series of m, total 9 4.71 (bs, 2 6.67-7.38 (series of m, total 12 7.77-8.17 (series of m, total 5 H).
To a stirred solution of methyl (S)-3-chloro-4-12-IN-benzyl-N-13-methoxy-4-IN'-(2fluorophenyl) ureidojphenylacetyl~aniinoJ-1-propoxylbenzoate (607 mg, 0.96 mmol) in THE (6 mL) was added 0.5 N NaQH (6 miL), and the resulting mixture was heated under reflux overnight.
The mixture was poured into ice-i N HCI and the solid was collected. The crude solid was recrystallized to give 192 mg 287 as a white crystalline powder. mp 125-130 'H-NMR (DMSO-d) 8 1.07-1.23 (in, 3 3.76 2 3.85 3 3.90-4.26 (in, 3 4.56 2 H), 6.66-7.38 (series of mn, total 10 7.82-8.20 (series of mn, total 5 8.70-8.74 (in, 1 9.18-9.20 (in, 1 13.02 (bs, 1 MS (FAB) m/z 621 Anal. Calcd for C 3 ,lH 31
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.
Exampule 237 3-NiorplNmtyaio--[-3mtoy4[P(-ehlhnlw~djhnlctl methylaminolethoxylbenzoic acid WO 01/00206 WO 0100206PCT/USOOIISO79
H
3
OOH
H 3H H 6C3C 9 H3 288 To a stirred cold (00 C) solution of methyl 3-isopropylaxnino-4-[[2-[3-methoxy-4-[N'-(2methylphenyl)ureidolphenylacetyllmethylaminojethoxyjbenzoate (324mg, 0.S8mmol) in CH 3
CN-
formaldehyde-AcO11 (1 ImnL, 8:2: 1, v/v/v) was added NaBH 3 CN (145 mg, 2.30 mniol), and the 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 CIHC1 3 (2Oml,), and the solution was washed with brine, dried over MgSO,, and evaporated to give 317 mng methyl 3-(N-isopropyl-N-methylamino)-4-[[2-[3-methoxy-4-[A'-(2-methylphenyl)ureidolphenylacetyl methylaminojethoxy] benzoate as a colorless oil. 'H-NMR (CDC1 3 8 1.02 (in, 611), 2.29 3H), 2.63 (in, 311), 3.02-3.20 (in, 3.49-3.80 (in, 811), 3.88 311), 4.06 and 4.21 (mn, total 211), 6.59 (in, 111), 6.76 (in, 211), 7.11-7.23 (in, 311), 7.50-7.62 (in, 311), 8.05 1H, J=8.3Hz), MS (FAR) m/z 576 (NC+ 1).
A stirred mixture of methyl 3-(N-isopropyl-N-methylamino)-4-[[2-1j3-methoxy-4-[A'-(2methylphenyl)ureidojphenylacetyllmethylamnino]ethoxybenzoate(3 17mg, 0.S5inmol) in 0.25 N NaGH (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 HCl and the solid was collected with suction. The crude solid was recrystallized from CHCI 3 -n-hexane-diisopropylether to give 288 as a light yellow crystalline powder. mp 130-135 IR (KBr) 2970, 1537, 1038, 754 cm*' 2 '11-NMvR (CD 3 OD) 8 1. 12(in, 611), 2.29 311), 2.76 Cmn, 111), 2.89 211), 3.02 111), 3.21 211), 3.72 211), 3.80 311), 3.84 (in, 4.13 and 4.40 (in, total 211), 6.76 111, J=7.811z), 6.86 111), 7.00 (in, 211), 7.18 (in, 311), 7.57 I1H, J=7.8Hz), 7.95 (in, 211); MS (FAR) m/z 562 Anal. Calcd for C 3 ,1,NQ 6 2.0H 2 O: C, 62.19; H, 7.07; N, 9.36. Found: C, 62.54; 11, 6.85; N, 8.90.
Example 238 1 eiiy)4[[-3mtoy4[M(-loohnl~riopeyaetlmtyaio ethoxylbenzoic acid
CH
3 *N OOH J' H 6CH3289 To a stirred solution of methyl 3-ainino-4-[[2-[3-inethoxy-4-[N'-(2-fluorophenyl)ureido] phenylacetyljmethylamino]ethoxylbenzoate (300mg, 0.57nunol) in TBfF (2 inL) was added a solution of glutaraldehyde (50% aqueous solution) l3inL, 0.69minol) in MeO11 (1.4 mL), '[HF WO 01/00206 WO 0100206PCT/USOOI 18079 (0.993mL), and 3N H 2 S0 4 0.952 mL) at 00 C. To the solution was added NaBH 3 CN (150 mg), DMF (5 mL), and MeOH (2 raL) at room temp, and the resulting mixture was stirred for 15 hr.
The mixture was poured into sat. NaHCO 3 and extracted with C11C1 3 The extract was washed with brine, dried over MgSO 4 and evaporated under a reduced pressure. The residue was chromatographed on silica gel with toluene:acetone v/v) as eluent to give 145 mg (43%) methyl -ieiiy)4[2[-ehx--A-2-loohnluedlhnlctlmty aminolethoxy]benzoate as a colorless oil. 'H-NMR (CDCI,, 400M4Hz) 8 1.58-1.72 (in, 611), 2.80- 2.92 (in, 411), 3.26 211), 3.58 211), 3.69 211), 3.80-3.89 (in, 711), 4.22 (in, 211), 6.72 (s, 111), 6.78 (in, 6.95-7.18 (in, 211), 7.32 IH), 7.60 111), 7.63 1H, J=7.811z), 7.98 (d, IH, J=7.8Hz), 8.18 (in, 111); MS (FAB) in/z 593 (WM+1).
A stirred mixture of methyl 3-(1-piperidinyl)-4-[[2-[3-methoxy-4-[N'-(2-fluorophenyl) ureidolphenylacetyl]methylaininolethoxylbenzoate (290mg, 0.49minol), IN NaOH (1.95 mL) in and THF (10 mL) was heated under reflux for 4 hr. The miixture was poured into water (200 mnL) acidified with IN HCI (p11=4.0), and extracted with CHCI 3 -MeOH(9: 1, The combined extract was dried over MgSO 4 and evaporated. The residue was crystallized from diisopropylether-hexane to give 20 1mg (71%/)289 as a light yellow crystalline powder. mp 125- 130'C; IR (KBr) 3338, 2935, 1599, 1537, 1041, 752 'H-NMR (CDOD) 8 1.52-1.73 (in, 611), 2.88 311), 2.98 (br, 111), 3.09 111), 3.23 211), 3.66 211), 3.75 111), 3.82 411), 4.13 and 4.29 (in, total 211), 6.78 (in, 211), 6.99 (in, 211), 7. 10 (in, 211), 7.60-7.70 (in, 211), 7..96-8.07 (in, 211); MIS (FAB) ink 578 Anal. Caled for C 31
H
3
_,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-ainino-4-[[2-[3-methoxy-4-[AM-(2-nethylphenyl)ureido]phenylacetylmethylamno~ethoxy benzoic acid ?H3 ~COOH QL..
NH
2 256H3H H CH 3 290 To a stirred solution of methyl 3-amino-4-[[2-[3-inethoxy-4-[M-(2-methylphenyl)ureidoI phenylacetyljmethylamino~ethoxyjbenzoate (300 mg, 0.58 inmol) in MeOH-THF 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 niL) and acidified by the addition of IN HCI. The solid was collected with suction. The residue was recrystallized from diisopropylether to give 243 mng (83%) 290 as a yellow crystalline powder. mp 125-130 IR (KBr) 3346, 2935, 1533, 1211, 1034, 756, WO 01/00206 WO 0100206PCT/USOO/18079 637cin-'; 'H-NMR (DMSO-d) 5 2.29 3H), 3.05 1H), 3.72-3.85 (mn, 7H), 4.12 and 4.25 (in, total 2H), 6.76-6.89 (in, 3H), 7.00 (in, IH), 7.18 (in, 2H), 7.39 (mn, 2H), 7.60 lH, J=7.8Hz), 7.96 (in, 1H); Anal.Calcd for vHON 4 O,0.5H 2 O: C, 62.90; H, 6.06; N, 10.87. Found: C, 63.03; H, 6.14; N, 10.56.
Example 240 1 eiiy)4[2[3mtoy4[-2mthlhnlu-iopenlctlmtyaio ethoxylbenzoic acid
CH
3
OOH
H3& H H3 2 291 To a cooled and stirred solution of methyl 3-anuuno-4-[[2-113-methoxy-4-[M,-(2methyiphenyl) ureidolphenylacetyllmethylamino]ethoxylbenzoate (573mg, 1. 1ininol) in THE- MeOH 1, v/v, 6rnL) was added glutaraldehyde (0.423miL, 2.2mmol), 3N H 2 S0 4 1(1.83miL, 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 (lO0nmL) and the solid was collected with suction.
The crude solid was purified by column chromatography on silica-gel with toluene-acetone (10:0 to 4: 1, v/v) to give 240mg (371/) methyl 3-(1 -piperidinyl)-4-[[2-[3-methoxy-4-[A'-(2-nethy phenyl)ureidojphenylacetyl~methylaminolethoxyJ benzoate as a gum. 'H-NN4R (CD 3 OD) 8 1.52- 1.72 (in, 6H), 2.30 3H), 2.36 2H), 2.89 (br s, 3H), 2.98 (in, 1H), 3.05 IH), 3.20 2H), 3.68 2H), 3.69 (in, 2H), 3.79 (in, 2H), 3.88 2H1), 4.08 and 4.21 (in, 2H), 6.35 and 6.42 (s, total 1H), 6.70 1H), 6.70 lH), 6.79 (mn, 2H), 7.09-7.24 (in, 4H), 7.50-7.65 (in, 3H), 8.05 (d, 1H, J=8.3Hz); MS (FAB3) ink 588 1).
A stirred mixture of methyl 1-piperidinyl)-4-[[2-[3-inethoxy-4-[MP-(2-inethylphenyl) ureidolphenylacetyllinethylaniinolethoxybenzoate (240 ing, 0.41 inmol), 1 N NaOH (1.63 inL) in MeOH (6.5 inL), H 2 0 (5 mL), and THF (6.5 inL was heated under reflux for I11 hr. The mixture was poured into water (200 inL), acidified by the addition of IN HCI until pH=4.0, and the solid was collected with suction. The aqueous layer was extracted with CHCI 3 -MeOH 1, vlv, 3Oil, x The precipitate was dissolved in the combined organic extract. The organic layer was dried over MgSO, and evaporated. The residue was crystallized from diisopropylether to give 151 ing 291 as a light yellow crystalline powder. mp 120-125'C; IR (KBr) 3354, 2935, 1535, 1252, 1217, 1034, 754, 638 cm"d; 'H-NMR (CD 3 00) 6 1. 55-1.72 (in, 614), 2.30 3H), 2.89 (br, 2H), 2.98 (br, iH), 3.09 IH), 3.22 2H), 3.69 3H), 3.74 IH), 3.83 (in, 4H), 4.12 and 4.28 (in, total 2H), 6.75 (mn, 2H), 6.88-7.02 (in, IN), 7.17 (in, 2H), 7.58-7.73 (in, 414), 7.99 I H, WO 01/00206 WO 0100206PCT/USOO/18079 J=8.3Hz); MIS (FAB) m/z 574 Anal. Calcd for C 3
,H~NO
6 -0.5H 2 O: 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-[AM-(2-fluorophenyl)ureidolphenylacetyllmethylaminolethoxyI benzoic acid H rCOOH 0 NH 2 Astirred mixture of methyl 3-amino-4-[[2-[3-methoxy-4-[A'-(2-fluorophenyl)ureido phenylacetyllmethylainino]ethoxy]benzoate (250 mg, 0.48 mmol) in IN NaOH (1.91 niL), MeOH (8 niL), H 2 0 (6 mL), and THE (8 mL) was heated under reflux for 10 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 diisopropylether to give 209 mng (86 292 as a pale yellow crystalline powder. mp 125-130*C; IR (KBr) 3325, 2935, 1537, 1209, 1032, 752, 449 'H-NMR (CD 3 OD) 8 3.05 iN), 3.31 2H), 3.77 (in, 3.80 IH), 3.84 (in, 3H), 4.144.26 (in, 6.80-6.87 (in, 3H), 7.01 (in, 7. 10 (in, 2H), 7.39 (in, 7.80 (in, I H), 8.07 (in, 1I-H); MS (FAB)m/Lz 5 10 Anal. Calcd for C2H 27 FNO0 6 -0.5H,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 -iethoxy-4-[N '-(2-methylphenyl)ureido~phenylacetylamino-1-propoxy] benzoic acid H 01 QA'rq 6H1 3
NH
2 6131H 6H 3 293 To a stirred solution of (S)-2-(N-tert-butoxycarbonylainino)-lI-propanol (0.90 g, 5.13 mmol), methyl 4-bydroxy-3-nitrobenzoate (1.01 g, 5.12 minol), and Ph 3 P (1.75 g, 6.67 iniol) in THF (15 mL) was added diisopropyl azodicarboxylate (DIAD) (1.31 mL, 6.65 inmol) at room temp, and the resulting mixture was heated under reflux overnight. The solution was evaporated and the residue was dissolved in CH 2
CI
2 (20 inL) and TEA (10 mQL. The mixture was stirred at room temp for 3 hr. The mixture was concentrated in vacua. The residue was dissolved in
CHCI
3 washed with sat. NaHCO 3 brine, dried over Na 2 SO, and evaporated. The residue was purified by column chromatography on silica-gel with 5% MeOH in CHC1 3 as eluent to give 313 mng methyl(S)-3-nitro-4-(2-ainino- I-propoxy)benzoate as a pale yellow oil: 'H-NMR
(CDCI
3 8 1.22 3 H, J=6.8 Hz), 3.43-3.48 (in, 1 3.69-3.87 (in, 2 3.89 3 6.93-6.95 (in, 1 7.99-8.02 (in, 1 8.184821 (in, 1 H).
WO 01/00206 WO 0100206PCT/US00/18079 A mixture of pentafluorophenyl 3-methoxy-4-[N -(2-methylphenyl)ureidolphenylacetate (591 mg, 1.23 inmol), methyl (S)-3-nitro-4-(2-amino-1I-propoxy)benzoate (313 mg, 1.23 mmol), and Et 3 N (257 mL, 1.84 mmol) in DMIF (5 mnL) 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 5% MeGH in CHCI 3 as eluent to give 3 10 mg methyl (S)-3-nitro-4-[2-[3-methoxy-4-[N'-(2-methylphenyl)ureidolpheny acetylaminoj-1-propoxyjbenzoate as a gum. 'H-NMvR (CDCl 3 8 1.28 3 HK J=6.8 Hz), 2.31 3 3.49 2 3.71 3 3.92 3 4.14-4.16 (in, 2 4.384.41 (in, 1 5.88-5.90 (in, 1 6.49 I 6.70-6.71 (in, 1 6.77-6.79 (in, 1 7.05-7.30 (mn, 4 7.55-7.57 (in, 1 8.02-8.06 (in, 2 8.164819 (in, 1 8.51-8.52 (in, 1 MIS (FAB) m/z 551 A stirred solution of methyl (S)-3-nitro-4-[2-[3-inethoxy-4-[N '-(2-inethylphenyl)ureidoJ phenylacetylanuinol-1propoxyJbenzoate (310 mg, 0.56 inmol) in MeOH-THF (10 inL, 1:1, v/v) was hydrogenated over 5% Pd-C (50 mg, 16 overnight. The mixture was filtered to remove the catalyst and the filtrate was evaporated to give methyl (S)-3-ainino-4-[2-[3-methoxy-4-[N methylphenyl) ureidolphenylacetylamino]-1-propoxylbenzoate (240 mng) as a gum.
To a stirred solution of the above crude product (220 mg) in THF-MeOH (10 inL, 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 HCL. The solid was collected and the crude solid was recrystallized from MeOH- CHC1 3 -n-hexane to give 116 mng (2 steps, 293 as a pale yellow crystalline powder. mp 200- 204 0 C; 'H-NMR (DMSO-&J 8 1.21 3 H, J=6.8 Hz), 2.24 3 3.37 2 3.80 3 H), 3.83-3.99 (in, 2 4.10-4.19 (in, 1 4.99 (bs, 2 6.75-7.23 (series of in, total 8 7.79 1 H, J=7.8 Hz), 7.98 1 H, J=7.8 Hz), 8.17 1 H, J=8.3 Hz), 8.46 1 8.55 1 MS (FAB) m/z 507 Anal. Calcd for C2,HN 4 ,O,-/2H 2 0: C, 62.90; H, 6.06; N, 10.87. Found: C, 62.85; H, 6. 10; N, 10. 51.
Example 243 (S)-4-[2-[3-methoxy-4-[N '-(2-bromophenyl)ureidolphenylacetylaminol-l1-propoxylbenzoic acid H crCOOH Br H H 6H 3 294 To a stiffed and cooled C) solution of (.S)-2-(N-tert-butoxycarbonylainino)- 1 -propanol (6.74 g, 0.04 mol), benzyl. 4-hydroxybenzoate (8.78 g, 0.04 mol), and PhP (15.13 g, 0.06 mol) in WO 01/00206 WO 0100206PCT/USOO/18079 THE (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 CHC1 2 (30 mL) and TEA (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 CHCI 3 washed with sat. NaHCO 3 dried over NaSO, and evaporated. The residue was purified by column chromatography on sil ica-gel with CHC 3 to CHCI,:MeOH 1, v/v) as eluent to give 6.63 g (2 steps, 60%) benzyl (S)-4-(2-amino-l-propoxy) benzoate as a yellow oil. 'H-NMR (CDCI,) 8 1. 18 J 6.3 Hz, 3 3.35-3.39 (in, 1 3.71-3.75 (mn, I 3.90-3.93 (mn, 1 5.34 2 H), 6.90-6.93 (in, 2 7.32-7.46 (in, 5 8.01-8.04 (in, 2 H).
A mixture of 3-methoxy-4-[N -(2-bromophenyl)ureidolphenylacetic acid (480 mng, 1.27 inmol), benzyl (S)-4-(2-anmino-1-propoxy)benzoate (361 ing, 1.27 nunol), EDC(hydrochloride) (364 mg, 1.90 inmol), HOBt (256 ing, 1.89 inmol), and 4-DMAP (31 mg, 0.25 minol) in DMF (8 niL) was stirred at room temp. overnight. The mixture was diluted with EtOAc, washed with N HCI, brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHC1 3 to 5% MeGH in CHC1 3 as eluent to give 476 mng (5 benzyl (S)-4-[2-[3-inethoxy-4-[N'-(2-broinophenyl)ureidolphenylacetylaminoJ-1 -propoxy]benzoate as a brown solid. 'H-NMR (CDCI 3 8 1.25-1.28 (in, 3 3.51 2 3.74 3 3.95-3.97 (in, 2 4.36-4.39 (im, 1 5.33 2 6.75-6.94 (im, 5 7.26-7.70 (in, 8 7.99-8.26 (in,
H).
To a stirred solution of benzyl (S)4-[2-[3-methoxy4-[N'-(2-bromophenyl)ureido phenylacetylamiinol-1-propoxyjbenzoate (476 mg, 1.39mimol) in THE (10 rnL) 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-i N HCI, and the solid was collected. The crude solid was recrystallized from MeOH-CHC1 3 -n-hexane to give 240 ing 294 as a white crystalline powder. mp 202-205 TC; 'H-NMIR (DMSO-4) 8 1.18 J 6.8 Hz, 3 3.37 2 3.82 3 3.92-4.00 (in, 2 H), 4.034.15 (in, 1 6.77-6.79 (in, I 6.93-7.03 (in, 4 7.30-7.34 (in, 1 7.59-7.61 (mn, I 7.87-7.97 (mn, 4 8.134815 (in, 1 8.73 1 8.91 1 12.63 (bs, 1 MIS (FAB) m/z 557 (Mr+ AnaL. Calcd for C 2 4-1 2 ,BrN 3 0 6 C, 56.12; H, 4.7 1; Br, 14.36; N, 7.55. Found: C, 56. 11; H, 4.74; Br, 14.56; N, 7.49.
Exampnle 244 (S)-4-[[2-[3-methoxy-4-[N '-(2-methylphenyl)ureidolphenylacetylJ-(2-aminobenzyl)anmino1-1 propoxy] benzoic acid WO 01/00206 WO 0100206PCT/USOO/18079 %-113 J~ 13 295 To a stirred cooled (00 C) solution of benzyl (S)-4-(2-arnino-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 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 CHCI, to 5% MeOH in CHCI, as eluent to give 931 mng benzyl (S)-4-[2-(2-nitrobenzylamino) -1-propoxy~benzoate as ayellow oil. 'H-NMR (CDC1 3 1.21 J 6.4 Hz, 3 3.13-3.18 (in, 1 3.88-3.97 (in, 2 4.06-4.20 (in, 2 5.34 2 6.89-6.94 (im, 2 7.29-7.65 (in, 8 7.94-8.03 (mn, 3 MIS (FAB) m/~z 421 (MW+ 1).
A mixture of pentafluorophenyl 3-methoxy-4-[N -(2-methylphenyl)ureido~phenylacetate (460 mg, 0.96 minol), benzyl (S)-4-[2-(2-nitrobenzylainiino)-1 -propoxylbenzoate (403 mng, 0.96 minol), and Et 3 N (200 mL, 1.43 minol) 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 NaSO 4 and evaporated. The residue was purified by column chromatography on silica-gel with 1% MeGH in CHC1 3 as eluent to give 504 mg benzyl (S)-4-[[2-[3-methoxy-4-[N '-(2-methylphenyl)ureidoj phenylacyl-(2-nitrobenzy)aino--propoxylbnzoate as a brown amorphous solid. MIS (FAB) mlz 717 A stirred solution of benzyl -methoxy-4-[N'-(2-methylphenyl)ureido~phenyl acetylj-(2-nitrobenzyl)aminoJ-1-propoxyjbenzoate (504 mg, 0.70 inmol) in MeOH-T-F (I11 mL, 1, v/v) was hydrogenated over 5% Pd-C (100 mng, 20 at 3 atmn overnight. The mixture was filtered to remove the catalyst and the filtrate was evaporated. The residue was purified by preparative TIC with 5% MeGH in CHCI 3 as eluent to give 115 ing 295 as a white powder.
MS (FAB) m/z 597 1).
Example 245 (S)-4-[[2-[3-methoxy-4-[N '-(2-bromophenyl)ureido]phenylacetylj -(2-nitrobenzyl)amino]-1 propoxylbenzoic acid WO 01/00206 WO 0100206PCTIUS0OI18079 r H H 6CH 3 296 To a stirred and cooled (00 C) solution of benzyl (S)-4-(2-aniino-1-propoxy)benzoate 50 g, 5.26 mmol) and 2-nitrobenzaldehyde 87 g, 5.76 mimol) in MeOH-AcOH (16 mL, 15: 1, v/v) was added NaBH 3 CN (1.65 g, 26.3 minol), and the resulting mixture was stirred at room temp overnight. The mixture was quenched by sat. NaHCO 3 and extracted with EtOAc. The extract was washed with brine, dried over NaSO, 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 mng (42%) benzyl (S)-4-[2-(2-nitrobenzylamino) -1-propoxy]benzoate as a yellow oil. 'H-NMR (CDCI 3 8 1.21 3 H, J =6.4 Hz), 3.13-3.18 (in, 1 3.88-3.97 (in, 2 4.06-4.20 (in, 2 5.34 2 H), 6.89-6.94 (in, 2 7.29-7.65 (in, 8 7.94-8.03 (in, 3 MIS (FAB)m/iz 421 1).
A mixture of 3-inethoxy-4-[N'-(2-bromophenyl)ureidolphenylacetic acid (476 mg, 1.26 mmol), benzyl (S)-4-[2-(2-nitrobenzylamino)-1-propoxy]benzoate (528 mg, 1.26 inmol), EDC(hydrochloride) (361 mng, 1.88 nuuol), HOBt (255 mng, 1.89 minol), and DMAP (30 mng, 0.25 nunol) in DMF (10 miL) was stirred at room temp. overnight and at 600 C for 1 day. 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 coluinn chromatography on silica-gel with CHC1 3 to 2% MeOH in CHC 3 as eluent to give benzyl (S)-4-[[2-[3-methoxy-4-[N '-2-broinophenyl)ureidolphenylacetylJ-(2nitrobenzyl)aminoJ-1-propoxyjbenzoic acid as an oil, which is used to the subsequent reaction without further purification.
To a stirred solution of the above crude in THF-MeOH (10 inL, 1: 1, v/v) was added 0.5 N NaOH (10 mnL), and the resulting mixture was heated under reflux for 3 hr. The mixture was poured into ice-H,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 CHC1 3 as eluent to give 162 mng (2 steps, 19%) 296 as a white amorphous solid. MIS (FAB) m/z 692 (NC+ Anal. Calcd for C 3 3H 3 jBrN 4 ,Og-7/4H 2 O: C, 54.82; H, 4.8 1; N, 7.75. Found: C, 54.80; H, 4.6 1; N, 7.24.
Example 246 4-[2-N-[3-methoxy-4-[N '-(2-methylphenyl)ureidolphenylacetainidolethoxyjbenzoic acid WO 01/00206 PCT/US00/18079 H r C c
OOH
Me H Me 297 To a cooled solution of 2-(N-tert-butoxycarbonylamino)ethanol (3.20 g, 19.9 mmol), methyl 4-hydroxybenzoate (3.02 g, 19.9 mmol) and Ph 3 P (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 under reflux for 3 hr. The mixture was evaporated and the residue was dissolved in CHC1l 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 CHC13 and H20. The solution was made basic by sat. NaHCO 3 and extracted with CHCI,. The extract was washed with brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silicagel with CHCI,-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 (CDCI) 5 3.10-3.13 2 3.89 3 4.03- 4.06 2 6.92-6.94 2 7.98-8.00 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), 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 O. 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 8 2.23 3 3.36 2 3.42-3.45 2 3.79 3 3.81 3 4.02-4.09 2 6.73-6.75 1 6.90-6.94 2 7.02-7.04 2 7.09-7.15 2 7.77-7.79 1 7.88- 7.90 2 7.95-7.98 1 8.23-8.24 1 8.44 1 8.53 I MS (FAB), m/z 492 Anal. Calcd for C 7
H,
29
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] 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 HCI. The resulting precipitate was collected and recrystallized from Et 2 O-CHCl 3 -MeOH to give 297 (135 mg, 50%) as a white crystalline powder. MW 477.51 'H-NMR (DMSO-d) 8 2.24 3 3.38 2 3.43-3.47 (m, 2 3.82 3 4.06-4.09 2 6.76-6.78 1 6.92-7.17 6 7.79 J 8.1 Hz, 1 7.88-7.89 2 7.98 J 8.1 Hz, 1 8.24-8.27 1 8.45 1 8.55 (s, WO 01/00206 WO 0100206PCT/USOO/18079 MS (FAB) m/z 478 Anal Calcd for C26H 27
N
3
O
6 1/4 2 0.C 47;H 57;N .2 Found: C, 64.67; H, 5.63; N, 8.60.
Example 247 '-(2-bromophenyl)ureidoJ-3-methoxyphenylacetaiido]-l1-propoxyjbenzoic acid H
~COOH
r HH 6Me298 To a cooled (0 0 stirred solution of (S)-2-(N-tert-butoxycarbonylainiino)- 1-propanol (6.74 g, 0.04 mol), benzyl 4-hydroxybefizoate (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
CI
2 (30 ml) 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 CHCI 3 washed with sat. NaHCO 3 dried over NaSO, and evaporated. The residue was purified by column chromatography on silica-gel with CHC1 3 to CHCI,-MeOH 1, vlv) as eluent to give benzyl (S)-4-(2-amino-l-propoxy)benzoate (6.63 g, 2 steps, 60%) as a yellow oil. 'H-NMR (CDCI 3 8 1.18 J 6.3 Hz, 3 3.35-3.39 (im, 1 3.7 1-3.75 (in, 1 3.90-3.93 (in, 1 5.34 2 6.90-6.93 (in, 214), 7.32-7.46 (in, 1-8.04 (in, 2H).
A mixture of 4-[N'-(2-bromophenyl)ureidoj-3-methoxyphenylacetic acid (480 mg, 1.27 mmol), benzyl (S)-4-(2-aniino-1-propoxy)benzoate (361 mg, 1.27 mmol), EDCHCI (364 mng, 1.90 mmol), HOBt (256 mg, 1.89 nunol) and DM"P (31 mg, 0.25 mmol) in DMF (8 ml) was stirred at .room temperature overnight. The mixture was diluted with EtOAc, washed with 0.5 N Nd, brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silicagel with CHCI, to 5% MeOH in CHCI, as eluent to give benzyl (S)-4-[2-[4-[N'-(2-bromophenyl) ureidoJ-3-methoxyphenylacetanuidoj -1-propoxyjbenzoate (476 mg, 58%) as a brown solid.
IH-NMvR (CDC1 3 8 1.25-1.28 (in, 3 3.51 2 3.74 3 3.95-3.97 (in, 2 4.36-4.39 (in, 1 5.33 2 6.75-6.94 (in, 5 7.26-7.70 (in, 8 7.99-8.26 (in, 5 H).
The a stirred solution of benzyl (S)-4-12-14-[N'-(2-bromophenyl)ureido]-3-methoxyphenyl acetamido]-l-propoxylbenzoate (476 mng, 1.39 innol) in THF (10 ml) was added 0.5 N NaOH ml), and the reaction mixture was heated under reflux for 2 hr. The mixture was poured into ice- I N HCI, and the resulting precipitate was collected. The crude solid was purified by recrystallization from MeOH-CHCI 3 -n-hexane to give 298 (240 mg, 59%) as a white crystalline powder. MW 556.41 'H-NMR (DMSO-d 6 8 1.18 J 6.8 Hz, 3 3.37 2 3.82 3 WO 01/00206 WO 0100206PCT/USOO/18079 3.92-4.00 (in, 2 4.03-4.15 (in, 1 6.77-6.79 (in, 1 6.93-7.03 (in, 4 7.30-7.34 (in, 1 7.59-7.61 (in, 1 7.87-7.97 (in, 4 8.13-8.15 (in, 1 8.73 1 8.91 1 H), 12.63 (bs, 1 MS (FAB) m/~z 557 Anal. Calcd for C 26
H
26 BrN 3
O
6 C, 56.12; H, 4.7 1; Br, 14.36; N, 7.55. Found: C, 56.11; H, 4.74; Br, 14.56; N, 7.49.
Example 248 (S)-4-[2-N-[13-methoxy-4-[N '-(2-methylphenyl)ureidolphenyl]-N-methylacetarnido]-1 -propoxyI benzoic acid aCOH Me' H M29 To a cooled (0 0 C) solution of (S)-2-[(N-tert-butoxycarbonyl)aminoJ-1I-propanol (3.08 g, 17.6 mmol), methyl 4-hydroxybenzoate (2.67 g, 17.6 inmol) and Ph 3 P (5.53 g, 21.1 mmol) in THF mil) 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 1, v/v) as eluent to give methyl [(N-tert-butoxycarbonyl)amino]-1-propoxyjbenzoate (1.24 g, as a white solid. IH-NMR (CDCI,) 8 1.30 J 6.8 Hz, 3 1.45 9 3.89 3 3.98-3.99 (in, 2 4.07 (in, 1 H), 4.76 (in, 1 6.91-6.93 (in, 2 7.98-8.00 (in, 2 MIS (FAB) m/~z 3 10 1).
To a stirred solution of methyl (S)-4-[2-[(N-tert-butoxycarbonyl)aininoJ-1-propoxyj benzoate (1.24 g, 4.01 minol) in CH 2
CI
2 (10 nil) was added TFA (10 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 CHC1 3 washed with brine, dried over
K
2 C0 3 and evaporated to give methyl (S)-4-(2-ainino-l-propoxy) benzoate (790 mg, 94%) as a yellow oil. 'H-NMR (CDC1 3 8 1.19 J 6.7 Hz, 3 3.34-3.42 (in, I 3.70-3.77 (in, I H), 3.86-3.94 (series of s and mn, total 4 6.92 J 9.0 Hz, 2 7.98 J 9.0 Hz, 2 H).
To a cooled (0 0 C) solution of methyl (S)-4-(2-ainino-1-propoxy)benzoate (790 mg, 3.78 inmol) and Et 3 N (630 mil, 4.52 minol) in TIHF (10 ml) was added TFAA (640 ml, 4.53 inmol) and the reaction mixture was stirred at room temperature for 4 days. The mixture was diluted with N HCI and extracted with CHC1 3 The extract was washed with brine, dried over K 2 CO3, and evaporated. The residue was purified by recrystallization from n-hexane-CHC13 to give methyl 4-[2-(trifluoroacetamido)-l-propoxyJbenzoate (790 mg, 69%) as a white crystalline material. IH- NMR (CDCl 3 6 1.42 J 6.8 Hz, 3 3.89 3 4.01-4.13 (in, 2 4.44-4.50 (in, 1 H), 6.57-6.61 (in, 1 6.92 J =9.0Hz, 2 8.00 J 9.0 Hz, 2 MIS (FALB) ,n/z 306 WO 01/00206 WO 0100206PCT/US00118079 Anal. Calcd for C 1 3
H,,F
3 N0 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-(trifluoroacetaxnido)-1-propoxyJbenzoate (695 mg, 2.28 mmnol) and K 2 CO3 (630 mg, 4.56 mniol) in DM4F (10 ml) was added MeI (210 ml, 3.37 mmol) and the reaction mixture was stirred at room temperature for 2 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 EtOAc-CHC1 3 (1:19, v/v) as eluent to give methyl (S4-[2-[(N-methyl)trifluoroacetamiddo]-1propoxy~benzoate (720 mg, as a white solid. mp 73-75*C; 'H-NMR (CDC13)8 1.36-1.39 (in, 3 2.96 and 3. 10 (each s, total 3 3.89 3 3.994.14 (in, 2 4.84-4.92 (in, 1 6.88- 6.92 (in, 2 7.98-8.00 (in, 2 MS (FAB) m/z 320 Anal. Calcd for C 1 4HI 6
F
3 N0 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)trifluoroacetaniidoj-l-propoxy] benzoate (710 mg, 2.22 mmol) in MeOH-H 2 0 (10 ml, 1:1, v/v) was added K 2 C0 3 (460 mg, 3.33 minol) and the reaction mixture was stirred at room temperature overnight. The mixture was diluted with H,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, as a colorless oil. 'H-NMR (CDCI,) 5 1. 17 J 6.6 Hz, 3 2.48 3 2.98-3.06 (in, 1 3.86-3.96 (series of s and mn, total 5 6.92 J 9.0 Hz, 2 7.98 J 9.0 Hz, 2 H).
A mixture of 3-inethoxy-4-[N-(2-inethylphenyl)ureidolphenylacetic acid (605 mg, 1.93 rniol), methyl (S)-4-[2-(N-methylamino)-l1-propoxylbenzoate (430 mg, 1.93 inmol), EDCHCI (444 mng, 2.32 mmol), HOBt (313 mg, 2.32 mniol), Et 3 N (320 ml, 2.30 inmol) in THE (13 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 CHC1 3 -MeOH (100: 1 to 75: 1, v/v) as eluent to give methyl (S)-4-[2-N-[[3-methoxy-4-[N -(2-inethylphenyl)ureido~phenylj-N-inethylacetaniido -l-propoxyjbenzoate (953 ing, 95%) as a white foam. IH-NMIR (CDC1 3 8 1.12-1.13 and 1.25-1.27 (each mn, total 3 2.27 3 2.84 and 2.92 (each s, total 3 3.63 3 3.67 2 H), 3.71-4.05 (series of s and mn, total 5 4.39-4.44 and 4.96-5.01 (each mn, total 1 6.66-6.85 (in, 5 7.09-7.27 (in, 4 7.53-7.55 (mn, 1 7.92-7.98 (in, 2 8.04-8.08 (in, 1 MS (FAB) ,n/z 520 Anal. Calcd for C 29 Hn3N 3
O
6 I 1/4H,0: C, 61.20; H, 6.82; N, 7.38. Found: C, 61.14; H, 5.86; N, 7.16.
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]-l-propoxy]benzoate (663 mg, 1.28 mmol) in THF (5 ml) was added 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 under a reduced pressure. The crude solid was dissolved in CHCI, and evaporated. The residue was washed with EtO2 to give 299 (465 mg, 72%) as a white amorphous solid. MW 505.56 'H-NMR (DMSO-d) 8 1.11-1.15 3 2.25 3 2.73 and 2.88 (each s, total 3 H), 3.60-3.76 2 3.83 3 4.03-4.12 2 4.41-4.50 and 4.48-4.94 (each m, total 1 H), 6.71-6.76 1 6.84-6.86 1 6.91-7.01 3 7.11-7.12 2 7.78-7.80 1 7.86-7.90 2 8.01-8.03 1 8.45 1 8.54 1H); MS (FAB) m/z 520 (Ml+1); Anal. Calcd for C 8
H
3
,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 rCOOH Me H H 0Me 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 ml) was heated at 100 OC for 14days under a current of nitrogen. The mixture was poured into icewater, and extracted with EtOAc. The extracts were washed with brine, dried over Na 2
SO
4 and 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 (CDCI) 5 1.37 6 3.89 3 4.03 2 6.92 J=9 Hz, 2 7.98 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 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 NaSO 4 and evaporated. The residue was chromatographed on silica gel with toluene-EtOAc (10:1, v/v) as eluent to give methyl 4-[l-(2-amino-2-methyl)prpoxy]benzoate as a gum (520 mg), which was used to the subsequent reaction without further purification. 'H- NMR (CDCl 3 5 1.41 9 3.89 3 4.04 2 4.69 (br s, 1H), 6.94 (dd, J =2 and 7 Hz, 2 7.98 (dd, J =2 and 7 Hz).
WO 01/00206 WO 0100206PCTI USO0118079 A solution of methyl 1-(2-methyl-2-terl-butoxycarbonylamino-)prpoxyJbenzoate (520 mg) and anisole 175 ml, 1.61 mmnol) in CH 2 C1 2 (5 ml) and TFA (3 mld) was stirred at room temperature for 18 hr. The mixture was evaporated off. The residue was dissolved in CH 2
CI
2 and the mixture was made basic by the addition of sat. NaHiCO 3 Separated CH 2
CI
2 layer was dried over NaSQ 4 and Na 2
CO
3 and evaporated. The residue was chromatographed on silica gel with
CHCI
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. 'H-NMR (CDCI 3 5 3.75 2 3.89 3 6.93 J =8.8 Hz, 2H), 7.98 J=8.8 Hz, 2 H).
To a stirred mixture of methyl 4-f 1-(2-amino-2-methyl)prpoxy~benzoate (250 mg, 1. 12 mmol), 3-methoxy-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (352 mg, 1. 12 mmol), 4- DMAP (165 mg, 1.34 minol) in DMIF (10 ml) was added EDC-HCI (290 mg, 1.51 nol) 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 CHC1 3 -EtOH 1, v/v) as eluent to give methyl 4-[2-N-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetamidoJ-2-methyl-1propoxylbenzoate (580mg, as a crystalline material. IR (KBr) 3350, 3286, 1712, 1687, 1637, 1606cm-'; 'H-NMIR (CDCI 3 5 1.39 6 2.33 3 3.41 2 3.63 3 3.88 3H), 4.05 2 5.44 (br s, 1 6.3 3 (br s, 1 6.79 J 8.3 Hz, 2 7.12 1 7.18 J Hz, 1 7.53 J =7.8 Hz, 1 7.94 J =7.8 Hz, 2H), 8.14 (d J 8.3 Hz, 1 MS (FAB) m/z 520 (M 4 Anal. Calcd for C 2 11H 33
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-[AP-(2-methylphenyl)ureidolphenyI acetamido]-2-methyl-2-propoxyjbenzoate (5 10 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 hr. The mixture was pored into ice-IN HCl (5 ml). the solid was collected, washed with water and air-dried. The crude solid was recrystal1lized from CHCI 3 -EtOH-Et 2 O to give 300 (480 mg, as fine needles. MW 505.56 IR n 3346, 3294, 1687, 1637, 1604 cm-1; 'H-NMvR (DMSO-d) 8 1.35 6 2.24 3 3.33 2 3.80 3 4.15 2 6.75 J 8.3 Hz, 1 6.88 1 6.95-6.99 (in, 3 7.11-7.17 (in, 3 7.80 J 8.3 Hz, I 7.82 1 7.87 J 8.8 Hz, 2 7.98 J 7.8 Hz, 1 8.45 1 8.54 1 12.62 (br s, 11H); MS (FAB) m/z 506 Anal. Calcd for CaH 3
N
3 0 6 C, 66.52; H, 6.18; N, 8.3 1.
Found: C, 66.22; H, 6.28; N, 8. 11.
Example 250 3-amiino-4-[ 2 -N-t4-IN'-(2-chiloropllenyl)ureido.3 -methoxyphenylacetamidoj -2-methyl-IpropoxyJbenzoic acid H <JOCOH ~NN N:Q I H H 6Me 301 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. HS0 4 at 0-5 0 C. To a stirred this solution was added dropwise a solution of NaNO, (5.4 g, 77.9 mmol) in water (5 ml) below 20 'C for over 10 min. The resulting mixture was further stirred for 0.5 hr at 5 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 min. The resulting mixture was for a further 1 hr at an ambient temperature. The mrixture was extrcte wih C 2
CI
2 The extracts were washed successively with sat. NaS 2
O
3 and brine. The :0 seeorganic layer was dried over Na 2
SO
4 and evaporated. The residue was chromatographed on silica- :gel (50mI) with CHCI 3 -EtOAc 1, v/v) as eluent to give 4-iodo-2-nitrophenol (2.5 g, 15%) as a yellow crystalline material. 'H-NMR (CDCI 3 856.94 J 8.8 Hz, I 7.82 (dd, J =2 and 8.8Hz, 1 8.42 J 2 Hz, 1H), 10.49 1 H).
To a stirred solution of 4-iodo-2-nitrophenol (2 g, 7.75 mmol), hydroxypivalic acid be* methyl ester (1.05 g, 7.92 mmol) and PPh 3 (2.3 g, 8.68 minol) in THF (10 nil) was added dropwise solution of DlAD (1.77 g, 8.30 mmnol) in THF (2 ml) under ice-water bath cooling. The resulting mixture was then heated under reflux for l8hr. After cooling, the mixture was .006 *5*20 evaporated off. The residue was chromatographed on silica gel (100 m.1) with with toluene-EtOAc 1, v/v) as eluent to give methyl 3 4 -iodo- 2 -nitro)phenoxy-2,2..iimethylpropionate (2.9 g, q.y.) as a crystalline material. 'H-NMR (CDCI 3 8 1.34 6 3.71 3 4.08 2 6.86 J sees 8.8 Hz, 1 7.78 (dd, J= 2 and 8.8Hz, 1 8.12 J= 2 Hz, I1H).
A mixture of methyl 3-( 4 -iodo-2-nitro)phenoxy2,2-dimethylpropionate (2.8 g, 7.38 mmol) in THF (15 mld) and 0.25 N NaOH (60 mlJ, 15 mnmol) was stirred 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 CHCI 3 -EtOH-IPE to afford 3-(4-iodo-2nitro)phenoxy-2,2..dimethylpropionic acid (2.0 g, 74%) as a crystalline material. Mp 165-182'C; I-R (KBr) n 1716, 1525, 1344 'H-NMR (CDC1 3 8 1.38 6 4. 10 2 6.86 J 8.8 Hz, I 7.79 (dd, J= 2.2 and 8.8H4z, 1 8.12 J= 2 Hz, I1H); MS (FAB) m/z 366 1); Anal. Caled for C,,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.
WO 01/00206 WO 0100206PCT/U S018079 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 mnnol) in iert-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 HCI (5mi) was added to the mixture and extracted with toluene. The extracts were washed with brine, dried over NaSO,, and evaporated. The residue was chromatographed on silica gel 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, as a gum. 'H-NMVR (CDCI,) 8 1.38 9 1.39 6 H), 4.19 2 4.67 (br s, 1 6.88 J= 8.8 Hz, 1 7.77 (dd, J= 2.O0and 8.8Hz, 1 8.12 (d, J =2.0OHz,1IH).
A mixture of 3-nitro-4-(2-:er:-butoxycarbonylamidno-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 v/v, 48 ml) was stirred under a current of CO (gas) at 70 0
C
for 6 hr. After cooling, the mixture was poured into water and extracted with EtOAc. The extracts were washed with brine, dried over Na 2
SO
4 and evaporated. The residue was chromatographed on silica gel (50mI) with toluene-EtOAc 1, v/v) as eluent to give methyl 4-{2tert-butoxycarbonyl amino-2-methyl-1-propoxy)-3-nitrobenzoate (820 mng, 51%) as a gum. 'H- NMR (CDCI 3 5 1.38 9 1.42 6 3.93 3 4.29 2 4.67 (br s, 1 7.15 J =8.8 Hz, I1H), 8.18 (dd, J= 1. 7and 8.8Hz, 1 8.52 J= 1.7 Hz, 1H).
A stirred mixture of methyl 4-(2-teri-butoxycarbonylainino-2-methyl- l-propoxy)-3nitrobenzoate (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)-l -propoxy-3-aminobenzoate as a gum, which was used to the subsequent reaction without further purification. 'H-NMR (CDCI 3 8 1.41 9 1.43 6 3.86 3 4.07 2 4.67 (br s, 1 6.80 J 8.5 Hz, 1 7.39 J =2.2 Hz, I H), 7.44 (dd, J 2.2 and 8.5Hz, 1 H).
To a stirred mixture of the above methyl 4-(2-tert-butoxycarbonylanmino-2-methyl)- I1propoxy-3-aminobenzoate and triethylamine (0.20 ml, 1.43 mmol) in CH 2
CI
2 (10 ml) was added a solution of trifluoroacetic anhydride 182 mil, 1. 28 nunol) in CH 2
CI
2 (3 ml) at 0-5 The resulting mixture was stirred at room temperature for I hr. Ice-sat. NaHCO, was added to the mixture, and extracted with CH 2
CI
2 The extracts were washed with brine, dried over Na 2 SO,, and WO 01/00206 WO 0100206PCT/USJO/18079 evaporated. The residue was dissolved in CH 2
CI
2 (5 nil) and added anisole 105 ml, 0.95 mmol) and TEA (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
CI
2 and made basic by the addition of sat. NaHCO 3 The separated CH 2
CI
2 layer was dried over Na 2
SO
4 and evaporated. The residue was chromatographed on silica gel (50mI) with CHCI,-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.
IH-NMvR (CDCI 3 5 1.29 9 H, lerl-Bu), 3.86 2 H, CH-I), 3.91 3 H, 6.99 J =8.5 Hz, 1 7.91 (dd, J 2.0 and 8.5 Hz, 11H), 8.83 J =8.5Hz, 1 H).
To a stirred mixture of methyl 4-(2-anmino-2-methyl)-1-propoxy-3-trfluoroacetanmido benzoate (200 mug, 0.598 mmol), 4-[N'-(2-chlorophenyl)ureidoJ-3-methoxyphenylacetic acid (210 mug, 0.598 mniol), 4-DMAP (90 mug, 0.72 mxnol) in DMIF (7 ml) was added EDC-HCI (160 mg, 0.81 mmrol) at room temperature. The resulting midxture was stirred at room temperature for 20 hr.
The midxture 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 CHC1 3 EtOH (98:2, v/v) as eluent to give methyl 4-[2-N-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylacetamidoj-2methyl-l-propoxyJ-3-trfiuoroacetamidobenzoate (580mg, as a crystalline material. 'H-NvfR (CDC1 3 5 1.42 6 3.42 2 3.69 3 3.89 3 4.23 2 5.33 (br s, 1 H), 6.66 1 6.71 (in, 1 6.92 J 8.5 Hz, 1 7.02 (mn, 1 7.09 (mn, 2 7.29 (mn, 1 H), 7.37 J= 8.0 Hz, IlH), 7.87 (dd, J= 2 and 8.5Hz, 1 7.97 J= 8.0 Hz, 1 8.19 J 8.2 Hz, 1 8.59 (br s, 1 8.74 J =2.0 Hz, 1 H).
To a stirred solution of methyl 4-[2-N-[4-[AIM-(2-chlorophenyl)ureidoJ-3-methoxypheny acetamidoJ-2-methyl-1-propoxyJ-3-trfluoroacetanidobenzoate (320 mg, 0.492 ruiol) in 11-F (2 ml) was added 0.25N NaOH (6 ml, 1.5 inmol) at an ambient temperature. And the resulting mixture was stirred for 20 hr. The mixture was poured into ice-IN HCI (2 Ml). The solid was collected, washed with water and air-dried. The crude solid was recrystallized from CHC1 3 EtOH- Et 2 O to give 301 (240mg, 89%) as fine needles. MW 541.00 IR (KBr) n 3338, 3296, 1691, 1641 cml']; 2 H-NMR (CDCI 3 8 'H-NMdR (DMS0 d 6 8 1.37 6 3.35 2 3.77 3 4.05 (s, 2 4.96 (br s, 1 6.75 (br d, J= 8.3 Hz, I 6.78 J 8.3 Hz, I 6.87 (d,J 1. 7Hz, 1 7.01 (mn, 1 7.15 (dd, J= 2 and 8.5 Hz, lH), 7.24 J 2 Hz, 1 7.27 (dt,J= 2.0 and 8.5 Hz, 1 7.43 (dd, J= 2 and 8.0 Hz, 1 7.78 (br s, 1 7.90 J= 8.0 Hz, 1 8.08 (dd, J 2 and 8.3 Hz, 1 8.85 1 8.89 1 12.23 (br s, 1 MS (FAB) m/z 541 Anal. Calcd for C, 7
HCIN
4 O0 6 C, 58.00; H, 5.59; N, 10.02. Found: C, 57.97; H, 5.39; N, 10.01.
WO 01/00206 WO 0100206PCT/US0O/1 8079 Example 251 2-acetytamino-4-[2-N-[3-methoxy-4-[N'-(2-methylphenyl)ureidolphenyll-N-methylacetamido] ethylamninobenzoateic acid 9 M----J(HAc me 6e 302 To a stirred solution of 2-acetylamino-4-nitrobenzoic acid (1.28 g, 5.71 mmol) in benzene-MeOH 1, v/v, 25 niL), was added trimethylsilyldiazomethane (2.0 M solution in nhexane, 4.28 ml, 8.56 mmol) at 0 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 2acetylamino-4-nitrobenzoate (1.32 g, 97%) as a white solid; mp no data; IH-NMR (400 MIHz,
CDCI
3 8 2.30 3 4.00 3 7.88 (in, 1 8.18 (dd, J 2.0 Hz, 8.8 Hz, 1 9.60 J 2.2 Hz, 1 11. 10 1 MIS (ESI) mnk 238 To a solution of methyl 2-acetylamino-4-nitrobenzoate (1.31 g, 5.50 ninol) in MeOH 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 crude solid was recrystallized with CHCI 3 -MeOH-hexane to give methyl 2-acetylamino-4aminobenzoate (1.03 g, as awhite solid. 'H-N4R (400 MIHz, CDCI 3 )8 2.23 3 3.82 3 4.20 2 6.30 (dcl, J 2.5 Hz, 8.8 Hz, 1 7.80 J 8.8 Hz, 1 8.06 1 H), 11.26 1 MS (FAB), m/z 208 (MW).
To a cooled solution of methyl 2-acetylamino-4-aminobenzoate (300 mg, 1.44 mmnol) and N-teri-butoxycarbonyl-N-methylglycinaI (499 mg, 2.88 nunol) in 1,2-dichloroethae (30 ml), was added NaBH(OAc) 3 (964 mg, 4.32 inmol) and the stirring was continued for 64 h at 0 0 C. The mixture was poured into sat. NaHCO 3 and was extracted with CHCI 3 (50 ml x washed with brine, and dried over MgSO 4 After removal of the solvent in vacuc, the residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC- 5404-FC, linear gradient of hexane-EtOAc from 9:1 to 2: 1) to give methyl 2-acetylamino-4-[2-(Niert-butoxycarbonyl-N-methylanino)ethylaminolbenzoate (451 mg, 86%) as a colorless oil. 'H- NMR (CDCI 3 400 MHz) 8 1.48 9 2.22 3 2.90 3 3.32 (in, 2 3.50 (in, 2 H), 3.80 3 6.20 (dd, J 2.2 Hz, 8.8 Hz, 1 7.80 (in, I 7.95 (in, 1 11.30 (brs, 1 H); MIS (FAR) m/z 366 1).
To a stirred solution of methyl methyl 2-acetylaniino-4-[2-(N-Ier-butoxycaibonyl-N- WO 01/00206 WO 0100206PCTIIJSOOII8079 methylamino)ethylaminolbenzoate (450 mg, 1.23 minol) in dichloromethane (5 miL), was added TFA (5 mL) and the stirring was continued far 18 h at rt. After removal of the solvent in vacuo, the residue was dissolved in CHCI 3 (200 washed with brine, sat. NaHCO 3 and dried over MgSO 4 The solvent was removed to give methyl 2-acetylarnino-4-[2-(N-methylamino) ethylamino] benzoate (298 mg, 88%) as a colorless oil. IH-NMR (CDC 3 400 Mffz) 8 2.21 3 2.46 3 2.88 (in, 2 3.31 (in, 2 3.83 3 4.85 (br, 1 6.24 (dci, J 2.5 Hz, 8.8 Hz, 1 7.80 J 8.8 Hz, 1 7.99 (di, J 2.5 Hz, 1 MIS (FAB), m/z 266 A mixture of methyl 2-acetylamino-4-[2-(N-methylamino)ethylamiino~benzoate (145 mng, 0.55 inmol), 3-methoxy-4-[N'-(2-methylphenyl)ureido~phenylacetic acid (172 mg, 0.55 mmol), EDC-HCI (158 mng, 0.83 inmol), HOBt (141 mg, 1.05 mmol), 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 ftrm 100:0 to 70:30) to give methyl 2-acetylamino-4-[2-N-[3-methoxy-4-[N'-(2methylphenyl)ureidolphenylJ-N-methylacetan-idojethylaminobenzoate (309 ing, 100%) as an amorphous foam. 'H-NMvR (CDCI 3 )5 2.22 3 2.30 3 3.02 3 3.35 (in, 2 H), 3.58 3 3.50-3.74 (in, 4 3.85 3 6.20 (in, 1 6.58 1 6.65-6.75 (in, 3 H), 7.13 (in, 2 7.40-7.50 (in, 2 7.75 (in, 2 7.90 (in, 1 8.00 (in, 1 11.32 1 MIS (FAB) m/z 562 (Mr+ 1).
To a solution of methyl 2-acetylainino-4-[2-N-[3-inethoxy-4-[N'-(2-inethylphenyl)ureido] phenylj-N-inethylacetamido~ethylanuinobenzoate (309 mg, 0.55 innol) in THF-MeOH 1, v/ v, 9 ml), was added 0.25 N NaQH (4.4 ml, 1. 1 inmol) 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 with 1.0 N HCI. The resulting precipitate was recrystallized with hexane-diethylether to give 302 (175 mng, 58%) as a pale red powder. NM 547.60 'H-NMR (CD 3 OD) 5 2.16 (di, J 4.8 Hz, 3 2.28 J 4.2 Hz, 3 2.98 and 3.10 (2 s, total 3 3.35 (in, 2 3.68 (in, 4 3.81 and 3.84 (2 s, total 3 6.30 (in, 1 6.60-6.82 (in, 2 7.00 (in, 1 7.15 (in, 2 7.53 (in, 1 7.77-7.96 (in, 3 MIS (FAB) mnk 548 Anal. Calcd for C29H 3 3 N5O 6 -0.5 H,0: C, 62.58; H, 6.16; N, 12.58. Found: C, 62.55; H, 6.3 1; N, 12.15.
Example 252 2-acetylainino-4-[2-N-[4-[N -(2-bromophenyl)ureido]-3 -methoxyphenylj-N-methylacetainidoJ ethylaminobenzoic acid WO 01/00206 WO 0100206PCT/USOOI 18079
.~COOH
Br H H 6Me 303 A mixture of methyl 2-acetylamino-4-(2-N-methylamino-l1-ethylamino)benzoate (145 mg, 0.55 mmol), 3-methoxy-4-[N'-(2-bromophenyl)ureidolphenylacetic acid (209 mg, 0.55 mmol), EDC-HCI (158 mg, 0.83 minol), HOBt (141 mg, 1.05 mmol), and DMAP (13 mg, 0. 11 mmol) in DMIF (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)ureidoJ-3-methoxypbenyl]-N-methylacetamidolethylaminobenzoate (294 mg, 85%) as an amorphous foam. 'H-NMR (CDCI 3 8 2.21 3 3.05 3 3.40 (in, 2 3.65-3.70 (in, 4 3.78 3 3.86 3 6.21 (in, 1 6.79 (in, 2 6.93 (in, 1 7. 10 J 10. 3 Hz, 1 7.30 (in, 1 7.42 (in, 1 7.61 (mn, I 7.78-7.85 (in, 2 7.93 (in, 2 8.13 (in, 1 H); MIS (FAB), m/lz 627 To a solution of methyl 2-acetylamino-4-[3-methoxy-4-[N'-(2-broinophenyl)ureidoI phenylacetainido]-2-N-inethylamino-1-ethylaminobenzoate (294 mg, 0.47 iniol) in THF-MeOH 1, v/ v, 8 ml), was added 0.25 N NaOH (3.8 ml, 0.94 inmol) 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 hexanediethylether to give 303 as a white powder (210 mg, MW 612.47 mp 155-160 0 C; 'H- NMvR (CD 3 OD) 6 22.18 J 5.5 Hz, 3 3.00 and 3.12 (2 s, total 3 3.39 (in, 1 3.60 (in, 4 3.70 1 3.85 and 3.86 (2 s, total 3 6.31 (in, 1 6.68 (in, 1 6.78 (in, 1 H), 6.78 and 6.85 (2 m, total 1 6.97 (in, 1 7.30 (in, 1 7.56 (in, 1 7.80-7.95 (in, 4 H); MIS (ESI) ,n/z 613 Anal. Calcd for CH 3 ,Br,N 5
O
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.
Example 253 4-[2-N-[[4-[N'-(2-chlorophenyl)ureidoJ-3-inethoxyphenyl]-N-phenylacetamidojethoxylbenzoic acid y?
~COOH
I H 6Me304 To a solution of methyl 4-[2-(methanesulfonyloxy)ethoxy~benzoate (2.74 g, 10 nunol) in MeCN (50 ml), was added aniline (9.1 ml, 100 minol) at it. The reaction was stirred for 64 hours WO 01/00206 WO 0100206PCT/USOO/18079 at reflux. The mixture was poured into H 2 0 (200 mL), extracted with EtOAc (100 mL x dried over MgSO,. After removal of the solvent in vacua, the unreacted aniline was removed in vacua by co-evaporation with toluene (10 mL x 3) at 80 TC. The residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC-5404-FC, f750 mm x 150 mm, CHCI,) to give methyl 4-[2-(N-phenylamino)ethoxy~benzoate (2.23 g, as a colorless oil. 'H- NMR (CDC 3 8 3.56 J =5.1 Hz, 2 3.90 3 4.21 J= 5.1 Hz, 2 6.68 (dd, J= Hz, 8.6 Hz, 2 6.75 J= 7.3 Hz, 1 7.20 (AB type d, J =7.3 Hz, 2 8.00 J= 9.1 Hz, 2 MIS (ESI) m/z 272 (M 1 A mixture of methyl 4-[2-(N-phenylarnino)ethoxy~benzoate (136 mg, 0.5 mmol), 3methoxy-4-[N'-(2-chlorophenyl)ureidolphenylacetic acid (167 mg, 0.5 mmol) and PyBOP (781 mg, 0.75 mmol), i-PrNTEt 2 (261 ml1, 0.96 mxnol) in DMF (10 ml) was stirred for 18 hours. The mixture was diluted with EtOAc (100 mL) washed with 1 N HCI, brine and dried over MgSO 4 The residue was co-evaporated with toluene (10 ml x 3) to remove DM[F. The residue was chromatographed on TLC (MERCK, silicagel 60, 2 mm, 2 plates, CHCI 3 -MeOH, 20: 1) to give methyl 4-[2-N-[[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylj-N-phenylacetamidoJethoxy] benzoate (129 mg, 44%) as a white amorphous foamn. 'H-NMR (CDCl 3 5 3.42 1 3.69 J 8.3 Hz, 1 3.74 3 3.87 3 4. 10 (in, 2 4.23 (mn, 2 6.48-7.44 (mn, 13 7.93 J= 9.3 Hz, 2 8.18 (dd, J= 1. 5Hz, 8.3 Hz, I MIS (ESnIk lz588 To a solution of methyl 4-[2-N-[[4-[N'-(2-chlorophenylI)ureidoJ-3-methoxyphenylJ -Nphenyl acetamidojethoxy~benzoate (124 mg, 0. 19 inmol) in THF-MeOH 1, v/ v, 3 nil), was added 0.5 N NaOH (2 ml, 1 mmol) at Mi and heated to reflux in a sealed bottle. The stirrng was continued for 15 hours at reflux. The reaction mixture was poured into water, acidified with N HCl, extracted with CHC1 3 -MeOH 20 mL x and dried over MgSO,. After removal of the solvent, the residue was crystallized with CHCl 3 -hexane-diethylether to give 304 (77 mg, 64%/) as a white powder. MW 574.02 'H-NMR (CDOD) 8 3.45 2 3.79 3 4.12 (in, 2 H), 4.22 2 6.48 (dd, J =2.0OHz, 8.3 Hz, 1 6.61 (d,J =2.0Hz, 1 6.87 J =8.8 Hz, 2 7.00 (in, 1 7.22 (in, 3 7.36 (in, 1 7.43 (in, 3 7.90 J =8.3 Hz, I 7.95 J 8.8 Hz, 2 8.02 (dd, J 1.5 Hz, 8.3 Hz, I MIS (ESI) ,n/z 574 Anal. Calcd for
C
3
,H
2 C1N 3 01 6 0.5 H 2 0: C, 63.86; H, 5.01; N, 7.21. Found: C, 63.67; H, 4.91; N, 6.99.
Example 254 -methoxy-4-[N '-(2-methylphenyl)ureido~phenylj-N-(2-aminobenzyl)acetamidoJ- 1propoxylbenzoic acid WO 01/00206 WO 0100206PCT/USOO/18079 Me H H OMe 305 To a cooled (0 0 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, was added NaBHCN (1.65 g, 26.3 mmol) and the reaction mixture was stirred at room temperature overnight. The mixture was quenched by sat. NaHCO 3 and extracted with EtOAc. The extract was washed with brine, dried over Na 2
SQ
4 and evaporated. The residue was purified by column chromatography on silica gel with CHCl 3 to 5% MeGH in CHC1 3 as eluent to give benzyl (2-nitrobenzylamino) -1-propoxy~benzoate (931 mg, 42%) as a yellow oil. IH-NMR (CDC1 3 8 1.21 J 6.4 Hz, 3 3.13-3.18 (in, 1 3.88-3.97 (in, 2 4.06-4.20 (im, 2 5.34 2 6.89-6.94 (in, 2 7.29-7.65 (in, 8 7.94-8.03 (in, 3 MS (FAB) m/z 421 A mixture of pentafluorophenyl 3-inethoxy-4-[N'-(2-methylphenyl)ureidolphenylacetate (460 mg, 0.96 iniol), benzyl (S)-4-[2-(2-nitrobenzylamino)-l-propoxy]benzoate (403 mg, 0.96 minol) and Et 3 N (200 ml, 1.43 innol) in DMVF (8 ml) was stirred at room temperature overnight.
The mixture was diluted with EtOAc, washed with 0.5 N HCl, brine, dried over NaSO 4 and evaporated. The residue was purified by column chromatography on silica-gel with 1% MeOH in CHC1 3 as eluent to give benzyl (S)-4-[2-N-[[3-methoxy-4-[N -(2-methylphenyl)ureidojphenylJ-N- (2-nitrobenzyl) acetamnido]-1-propoxy~benzoate (504 mg, 73%) as a brown amorphous solid. MIS (FAB), ,n/z 717 A stirred solution of benzyl (S)-4-[2-N-[[3-inethoxy-4-[N'-(2-methylphenyl)ureido] phenylJ-N-(2-nitrobenzyl)acetaniidoJ-1-propoxylbenzoate (504 mg, 0.70 mxnol) in MeOH-THF (I11 ml, 10: 1, v/v) was hydrogenated over 5% Pd-C (100 mng, 20 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 CHCI, as eluent to give 305 (115 ing, 27%) as a white powder. MW 596.67 MS (FAB), mnk 597 Example 255 '-(2-bromophenyl)ureido]-3-inethoxyphenyl-N-(2-nitrobenzyl) acetaniido]- 1propoxy]benzoic acid WO 01/00206 WO 0100206PCTIUSOO/18079 r ~,COOH rH H OMe 306 To a cooled solution of benzyl (S)-4-(2-amino-l-propoxy)benzoate (1.50 g, 5.26 mmol) and 2-nitrobenzaldehyde (0.87 g, 5.76 mxnol) 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 overnight. The mixture was quenched by sat. NaHCO 3 and extracted with EtOAc. The extract was washed with brine, dried over NaSO, 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 benzyl (2-nitrobenzylamino) -1 -propoxylbenzoate (931 mg, 42%) as a yellow oil. 'H-NdR (CDCI 3 8 1.21 J 6.4 Hz, 3 3.13-3.18 (in, 1 3.88-3.97 (in, 2 4.06-4.20 (mn, 2 5.34 2 6.89-6.94 (in, 2 7.29-7.65 (mn, 8 7.94-8.03 (in, 3 FAB-MAS, ,n/z 421 1).
A mixture of 4-[N'-(2-bromophenyl)ureidoj-3-methoxyphenylacetic acid (476 mng, 1.26 inmol), benzyl (S)-4-[2-(2-nitrobenzyLamino)-l -propoxylbenzoate (528 mg, 1.26 nunol), EDCHCI (361 mg, 1.88 nunol), HOBt (255 mg, 1.89 mmol) and DMAP (30 mg, 0.25 inmol) in DMIF ml) was stirred at room temperature overnight. And the reaction could not be completed, so the reaction mixture was stirred at 60'C for 1 day. The mixture was diluted with EtOAc, washed with N HCI, brine, dried over Na 2
SO
4 and evaporated. The residue was purified by column chromatography on silica-gel with CHCI, to 2% MeOH in CHCI, 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.
The mixture was poured into ice-H 2 0 and the basic aqueous layer was acidified (pH 4.3) with I N HCL. The resulting precipitate was collected and the crude solid was purified by preparative thin layer chromatography with 5% MeGH in CHC1 3 as eluent to give 306 (162 mg, 2 steps, 19%) as a white amorphous solid. NM 691.53 MIS (FAB), mnk 692 AnaL. Calcd for
C
33 11 3 1BrN 4
O
8 -7/4H 2 0: C, 54.82; H, 4.8 1; N, 7.75. Found: C, 54.80; H, 4.6 1; N, 7.24.
Exampgle 256 4-[2-N-cyclopropyl-N-[4-IIN'-(2-chlorophenyl)ureidoJ-3-inethoxyphenylJacetainido ethoxybenzoic acid 7 rYCOOH I H H A mixture of methyl 4-(2-cyclopropylaminoethoxy)benzoate (290 mg, 1.23 minol), WO 01/00206 WO 01/11206PCT/USOO/18079 chlorophenyl)ureidoJ-3-methoxyphenylacetic acid (412 mg, 1.23 mmol), EDC-HCl (354 mg, 1.85 mmol), HOBt and DMAP (cat.) in DMF (10 ml) was stirred overnight. The mixture was partitioned between EtOAc (300 ml) and H,0 (100 ml). The organic phase was separated, washed with brine (2 x 100 mil), 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-[2-N-cyclopropyl-N-[4-[N'-(2chlorophenyl)ureido]-3-methoxyphenyllacetamido]ethoxybenzoate (506 mg, 75%) as a yellow viscous oil. 'H-NMR (CDC1 3 8 0.90-0.97 (in, 4 2.75 (in, 1 3.61 3 3.79 J 5.4 Hz, 2 3.87 3 3.88 2 4.16 J 5.4 Hz, 2 6.76-6.80 (mn, 4 6.95 (dt, J 7.8, 1.5 Hz, 1 7.21-7.3 1 (mn, 2 7.53 1 7.56 1 7.9 3 J 8.3 Hz, 3 8.19 (dd, J= 8.3, 1.5 Hz, 1 H).
To a stirred solution of methyl 4-[2-N-cyclopropyl-N-[4-[N'-(2-chlorophenyl)ureidoJ-3inethoxyphenyl~acetamido~ethoxybenzoate (506 mng, 0.9 17 nunol) in THF (7 ml) was added 0.25 N NaOH (7.3 ml, 1.83 minol). After stirring overnight, the mixture was poured into 1 N HC1 ml) and extracted with CHCI 3 -MeOH 2 x 200 ml). The combined extracts were dried over MgSO 4 and evaporated. The residue was chromatographed on silica gel with CHCI 3 -MeOH (20:1 to 10: 1) as eluent to give 307 (403 mg, as a colorless amorphous solid. MW 537.99 1- NMvR (DMS0) 8 0.86-0.91 (in, 4 2.75 (in, 1 3.69 J 5.5 Hz, 2 3.81 3 3.84 (s, 2 4.16 J= 5.5 Hz, 2 6.76 J= 8.3 Hz, 1 6.88 1 6.97-7.04 (in, 3 7.28 (t, J =7.8 Hz, 1 7.44 J= 7.8 Hz, 1 7.88 J =8.8 Hz, 2H), 7.96 J =8.1 Hz, 1 H), 8. 10 J =8.3 Hz, I 8.89 1 8.93 1 12.65 br MIS (FAB), ink 538 Anal. Calcd for CgH 28
CIN
3
O
6 C, 62.51, HK 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-inethylphenyl)ureido]phenyl] acetamidolethoxybcnzoic acid 9 0,,fCOOH efHNH M 308 To a solution of methyl 4-[(2-methanesulfonyloxy)-1-ethoxy]benzoate (2.74 g, 10 nunol) in MeCN (50 ml), was added cyclohexylamine (5.72 ml, 50 inmol) at rt. The reaction was stirred for 18 hours at reflux. The mixture was poured into H 2 0 (200 inL), extracted with EtOAc (100 nil x dried over MgSO 4 After removal of the solvent, residue was chroinatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC-5404-FC, f5O mun x 150 mun, linear gradient of CHC1 3 -EtOAc from 10:-0 to 1: 1) to give methyl 4-(2-N-cyclohexylanuino)ethoxy WO OIA)0206 WO 01/)0206PCT/USOO/1 8079 benzoate (2.43 g, 88%) as a colorless oil. 'H-NN4R (CDCI 3 5 1. 10 (in, 2 1.25 (in, 2 1.60 (br, 2 1. 73 (in, 2 1. 90 (br, 2 2.49 (in, 1 3.02 J 5.2 Hz, 2 3.88 3 4.12 J 5.2 Hz, 2 6.90 J =6.90 Hz, 2 7.99 J =7.99 Hz, 2 MS (ESI) m/z 278 1).
A mixture of methyl 4-(2-N-cyclohexylamiino)ethoxybenzoate (139 mg, 0.5 mmol), 3methoxy-4-[N'-(2-methylphenyl)ureido~phenylacetic acid (157 mg, 0.5 inmol), EDC-HCI (144 mg, 0. 75 mmol), HOBt (128 mg, 0. 95 minol), and DMA (12 mng, 0. 1 inmol) in DMFf (2.5 mil) was stirred for 18 hours. The mixture was diluted with EtOAc (200 mil), 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 CHCI 3 EtOAc 10:0 to 1:4) to give methyl 4-[2-N-cLyclohexyl-N-[3-inethoxy-4-[N'-(2-methylphenyl) ureidolphenylJ acetamido] ethoxybenzoate (247 mg, as an amorphous foam. 'H-NMR (CDCI,) 8 1.08-1.80 (mn, 10 2.30 3 3.60-3.79 (in, 8 3.88 3 4.16 (in, 2 6.30 1 6.70-6.83 (in, 2 6.88 2 H, J= 9.0 Hz), 7.12 (in, 2 7.23 (in, 1 7.60 1 H, J 8.3 Hz), 7.92 2H,J =9.0 Hz), 8.10 1 H, J =8.0 Hz);MNS (ESI)mz 574 To a solution of methyl 4-[2-N-cyclohexyl-N-[3-methoxy-4-[N'-(2-inethylphenyl)ureidoJ phenyl] acetamido]ethoxybenzoate (247 ing, 0.43 minol) in THF-MeOH 1, v/ v, 7 mil), was added 0.5 N NaOH (3.4 mld, 0.84 inmol) 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 1.0 N HCI, extracted with CHC1 3 -MeOH 20 mL x and dried over MgSO 4 After removal of the solvent, the residue was crystallized with CHCI 3 -hexane-diethylether to give 308 (196 ing, 8 1%O) as a white powder. NM 559.62 'H-NMR (CD 3 OD) 8 0.90-1.82 (in, 10 2.29 3 H), 3.62 (in, 2 3.78 3 3.80 (in, 3 4.12 (in, 2 6.82 (in, 2 6.96 (in, 3 7.16 (in, 2 7.58 J =7.7 Hz, 1 7.92 (in, 3 MS m/z 560 Anal. Calcd for
C
32
H
37
N
3 0 6 *0.5 H 2 0: 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-methoxyphenylJ-N-propargylacetamido] ethoxybenzoic acid
HCOOH
1 H 6Me309 To a solution of methyl 4- [(2-methanesulfonyloxy)-1I-ethoxylbenzoate (2.74 g, 10 minol) WO 01/00206 WO 01/(i206PCT/USOO/18079 in MeCN (50 ml), was added propargylamine (3.43 mil, 50 mmol) at rt. The reaction was stirred for 18 hours at reflux. The mixture was poured into H,0 (200 mL), extracted with EtOAc (100 mL x dried over MgSO 4 After removal of the solvent in vacua, the residue was chromatographed on silica gel (middle pressure chromatography system: YAMAZEN YFLC- 5404-FC, f50 mm x 150 mm, linear gradient of CHC1 3 -EtOAc from 10:0 to 9: 1) to give methyl 4- (2-N-propargylamino) ethoxybenzoate (2.33 g, 100%) as a colorless oil. 'H-NMR (CDCI 3 8 2.28 J= 2.4 Hz, 1 3.11 J =5.1 Hz, 2 3.52 J= 2.4 Hz, 2 3.88 3 4.15 J 1 Hz, 2 6.90 J 8.8 Hz, 2 7.98 J =8.8 Hz, 2 MS (ESI) m/~z 234 A mixture of methyl 4-(2-N-propargylamino)ethoxybenzoate (117 mg, 0.5 mmol), 3methoxy-4-[N'-(2-chlorophenyl)ureidojphenylacetic acid (167 mg, 0.5 mmol), EDC-HCl (144 mg, 0.75 mmol), HOBt (128 mg, 0.96 mmol), and DM[AP (12 mg, 0.1 mmol) in DMF (10 mld) was stirred for 18 hours. The mixture was diluted with EtOAc (100 mL), washed with 1 N HCI, brine and dried over MgSQ 4 The residue was co-evaporated with toluene (10 ml x 3) to remove DM[F.
The residue was chromatographed on TLC (MERCY, silicagel 60, 2 mm, 2 plates, CHCl 3 -MeOH, 20: 1) to give methyl 4-[2-N-[4-[N'-(2-chlorophenyl)ureido]-3-methoxyphenylJ-N-propargyI acetamido] ethoxybenzoate (244 mg, 89%) as a white amorphous foam. 'H-NMR (CDCl 3 5 2.20 and 2.32 (2 m, total I 3.72 2 3.83 (in, 5 3.88 3 4.094.35 (in, 4 6.77-6.86 4H), 6.99 1 7.11 (in, 2 7.24 1 7.34 J =7.9 Hz, I1H), 7.96 3 H), 8.18 (dd, J 1.5 Hz, 8.3 Hz, 1 MS (ESI) m/lz 550 (MW).
To a solution of methyl 4-[2-N-[4-[N'-(2-chlorophenyl)ureidoJ-3-methoxyphenylJ-Npropargyl acetamidojethoxybenzoate (240 mng, 0.44 inmol) in THF-MeOH-H 2 0 1, v/v. 10 ml), was added NaOH (500 mng, 12.5 inmol) at rt. The stirring was continued for 2 hours at rt. The reaction midxture was poured into water, acidified with 1.0 N HCI, extracted with CHCl 3 -MeOH 20 mL, x and dried over MgSO,. The residue was chmomatographed on TLC (Whatman, 1 mm, 3 plates, CHCI 3 -MeOH, 92:8) to give 309 (202 mng, 86%) as a white solid. MW 535.98 'H- NMR (CD 3 OD) 8 2.60 and 2.81 (2d, J 2.5 Hz, total 1 3.79-3.94 (mn, 4 3.85 3 4.15 (in, 1 4.24 (mn, 1 4.32 (in, 2 6.80 J 8.3 Hz, 1 6.85 J 4.3 Hz, I 6.94 (in, 2 7.02 (in, 1 7.25 (in, 1 7.38 I 7.87-8.02 (in, 4 MS (ESI) m/lz 536 Anal. Calcd for C 3 g H, 6 C1N 3
O
6 -2.25 H 2 0: C,58.33; H,5.33; N,7.29. Found: C,58.23; H,4.77; N,6.91.
Examples 259 and 260 4-2NallN[4[P(-hoopey rio--ethoxyphenyl~acetainidolethoxybenzoic acid WO 01/00206 WO 0100206PCT/USO0II 8079 I H0 M 310 A mixture of methyl 4-(2-N-allylamino)ethoxybenzoate (118 mg, 0.5 mmol), 3-methoxy- 4-[N'-(2-chlorophenyl)ureidolphenylacetic acid (167 mg, 0.5 mniol), EDC-HCI (144 g, 0.75 mmol), HOBt (128 mg, 0.95 mmol), and DMAP (12 mg, 0. 1 mniol) in DMF (2.5 mnl) was stirred 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 CHCI 3 EtOAc 100:0 to 85: 15) to give methyl 4-[2-N-allyl-N-[4-[N'-(2-chlorophenyl)ureidoJ-3methoxyphenylJacetaxnidoJ ethoxybenzoate (253 mg, 92%) as an amorphous foam. 'H-NMR
(CDCI
3 8 3.65-3.85 (in, 4 3.73 3 3.88 3 5.08 (in, 2 4.22 (mn, 2 5. 10-5.24 (in, 2 5.76 (mn, I 6.77 (mn, 2 6.85 (in, 2 6.99 (mn, 1 7.06 (in, 2 7.26 (in, 1 H), 7.34 1 H, J 8.1 Hz), 7.94 (di, 2 H, J 8.8 Hz), 7.98 (in, 1 8.18 (di, 1 H, J 6.9 Hz); MS (FAB) m/lz 552 To a solution of methyl 4-[2-N-allyl-N-[4-[A'-(2-chlorophenyl)ureido]-3-methoxyphenyl] acetamidojethoxybenzoate (250 mg, 0.45 inmol) in TH F-MeOH (1L 1, v/ v, 8 ml), was added 0.25 N NaOH (3.6 ml, 0.91 nunol) 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 Nd. The resulting precipitate was collected by filtration. The precipitate was recrystallized with hexanediethylether to give 310 as a white powder (195 mng, MW 537.99 'H-NMR (CD 3 OD)8 3.61 I 3.76 3 3.82 (in, I 3.85 1 3.88 (in, 1 4.11-4.25 (in, 4 5. 12-5.25 (in, 2 5.81 (in, 1 6.78 1 H, J 8.3 Hz), 6.82 (in, 1 6.92 (in, 2 7.01 (in, 1 H), 7.26 1 7.37 (in, 1 7.96 (mn, 3 8.02 (in, 1 MS (FA.B) m/z 537 (M 4 Anal. Calcd for C~gH 28
CIN
3
O
6 1/4 H 2 0: C, 61.99; H, 5.30; N, 7.75. Found: C, 62.00; H, 5.56; N, 7.76.
Example 261 4-2NallA-4[V(-rmpeiluedo--ehxpeylctmd~toyezi acid K, rY 0 (CO0H A mixture of methyl 4-(2-N-allylamino)ethoxybenzoate (118 ing, 0.5 inmol), broinophenyl)ureidoJ-3-inethoxyphenylacetic acid (190 mg, 0.5 innol), EDC&HCI (144 mng, 0.75 innol), HOBt (128 ing, 0.95 iniol), and DMAP (12 ing, 0. 1 nunol) in DMF (2.5 ml) was stirred WO 01/00206 WO 0100206PCT/USOO/18079 for 18 hours. The mixture was diluted with EtOAc (300 mla), washed with IN HCI and brine, and dried over MgSO,. After removal of the solvent, residue was chromatograph ed on silica gel (middle pressure chromatography system: YAMAZEN YFLC-5404-FC, linear gradient CHCI 3 EtOAc 100:-0 to 70:30) to give methyl 4-[2-N-allyl-N-[4-[N'-(2-bromopheny1)ureidoJ-3methoxyphenyli acetamido]ethoxybenzoate (251 mg, 84%) as an amorphous foam. 1
H-NNMR
(CDCI
3 )8 3.65-3.85 (in, 4 3.73 3 3.88 3 4.08 (in, 2 4.22 (in, 2 5. 10-5.25 (in, 2 5.78 (in, 1 6.79 (in, 1 6.85 (in, 3 6.93 (in, 1 7.02 (in, 2 7.30 (mn, 1 H), 7.51 (in, 1 7.94 2 H, J 8.8 Hz), 7.97 1 8.14 (in, 1 MS (FAB)m/tz 596 To a solution of methyl 4-[2-N-allyl-N-[4-[N'-(2-broinophenyl)ureidoJ-3-methoxyphenylI acetamido~ethoxybenzoate (251 mng, 0.42 mmol) in THF-MeOH 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 HCI. The resulting precipitate was collected by filtration. The precipitate was recrystallized with Hexanediethylether to give 311 (192 ing, 78%) as a white powder. MW 582.44 'H-NMR (CD 3 OD) 5 3.61 3 3.77 3 3.80 (in, 1 3.85 I 3.88 1 4.12-4.25 (in, 4 5. 12-5.23 (in, 2 5.81 (in, I 6.76 (in, 1 6.82 (mn, 1 6.93 3 7.29 (in, 1 7.56 (mn, 1 H), 7.94 (mn, 4 MS (FAB), ,n/z 582 Anal. Calcd for CgH,8BrN 3
O
6 C, 57.74; H, 4.85; N, 7.2 1.
Found: C, 57.40; H, 5.07; N, 7.04. For HCI salt of 311: Anal. Caled for C28H2BrN 3
O
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.
Example 262 4-[2-N-allyl-N-[3-inethyl-4-[Nf'-(2-methylphenyl)ureido~phenyljacetanmidoethoxybenzoic acid e H HOO 312 A mixture of methyl 4-(2-N-allylainino-1-ethyl)ethoxybenzoate (87 ing, 0.37 minol), 3inethyl-4-[N'-(2-inethylphenyl)urido~phenylacetic acid (100 mg, 0.37 inmol), EDC-HCI (105 mg, 0.56 inmol), HOBt (95 mg, 0.70 inmol), and DMAP (9 mng, 0.07 inmol) in DMF (7.4 ml) was stirred for 18 hours. The mixture was diluted with EtOAc (100 nil), washed with IN HCI and brine, and dried over MgSO 4 The residue was co-evaporated with toluene (10 ml x 3) to remove DMT. The residue was chroinatographed on TLC (Whatman, PLK-5F, 2 plates, CHCI 3 -MeOH, 97:3) to give methyl 4-[2-N-allyl-N-[3-inethyl-4-[N'"-(2-inethylphenyl)ureido]phenyllacetamidoj ethoxybenzoate (190 ing, 100%) as a white amorphous foam. 'H-NMIR (CDC1 3 )8 2.05 and 2.09 (2s, total 3 2.20 and 2.21 total 3 3.62 2 3.76 (in, 2 3.90 3 3.88 (in, 1 WO 01/00206 WO 0100206PCT/USOO/1 8079 4.08 (in, 2 4.11 (in, 1 6.28 (mn, 2 5.78 (mn, 1 6.88 J =8.8 Hz, 2 7.05 (mn, I 7.12 (in, 1 7.21 (mn, 1 7.58 (in, 2 7.95 J 8.8 Hz, 2 8.02 (in, 1 MS (FAB), m/z 516 (Mir+l).
To a solution of methyl 4-f 2-N-allyI-N-[3 -methyl-4-[N'-(2-methylphenyl)ureido]phenylJ acetamido]ethoxybenzoate (217 mg, 0.43 minol) in THF-MeOH 1, v/ v, 7 ml), was added 0. 5 N NaOH (1.9 ml, 0.86 nunol) 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 CHCI 3 -MeOH 20 mL x and dried over MgSO 4 After removal of the solvent, the residue was crystallized with CHCI,-hexane-dethylether to give 312 (84 mng, as a white powder. MW 501.57 'H-NMR (CD 3 OD)8 2.18 and 2.24 total 3 2.30 J= 4.9 Hz, 3 3.70 I 3.78 (mn,2H), 3.88 I1H), 4.12 (mn,4H), 5.20 (mn,2H), 5.81(n, I1H), 6.92- 7.20 (in, 7 7.58 (in, 2 7.96 (in, 2 MIS (ESI) m/lz 502 Anal. Calcd for C,H,,N 3 C, 69.44; H, 6.23; N, 8.38. Found: C, 68.99; H, 6.39; N, 8.03.
Example 263 4-[2-N-allyl-N-[3 -chloro-4-[N'-(2-methylphenyl)ureidoJphenylacetamido]ethoxybenzoic acid ,Or cCOOH e H H313 To a stirred solution of methyl 4-(2-N-allylaminoethoxy)benzoate (141 ing, 0.60 inmol) and 3-chloro-4-[N'-(2-methylphenyl)ureido]phenylacetic acid (191 mg, 0.60 iniol) in DMF mL) were added EDC-HCI (172.5 mg, 0. 90 inmol), HOBt (154 mg, 1. 14 nunol), and DMAP mg, 0. 12 mxnol), and the stirring was continued overnight at rt. The mixture was diluted with EtOAc (50 mL) and washed with IM HCI (x IM NaOH (x and brine. The mixture was dried over anhydrous MgSO 4 and concentrated under a reduced pressure to give methyl 4-12-Nallyl-N-[3-chloro-4-[N'-(2-inethylphenyl)ureido] phenyljacetamidoJethoxybenzoate (350 mg, 109%) as a white powder. 'H-NN4R (CDCI 3 8 2.35 3 3.60 1 3.75 (in, 2 3.90 (s, 3 4. 10 (in, 4 4.21 (in, 1 5.20 (in, 2 5.80 (in, 1 6.50 1 H) 6.85 (in, 2 7.08 2H), 7.20 4H), 7.50 J =8.1 Hz, 1 7.95 J =8.1 Hz, 2 8.12 J =8.1 Hz, I MS (ESI) tn/z: 536 (MW+H).
To a stirred solution of methyl 4-[2-N-allyl-N-[3-chloro-4-[N'-(2-inethylphenyl)ureidoj phenyllacetaniidolethoxybenzoate (321 ing, 0.6 mmnol) in THF-MeOH-H 2 0 1, vfv, 30 ml), was added NaOH (500 ing, 12.5 minol) at rt. The stirring was continued for 18 hours at rt. The WO 01/00206 WO 0100206PCT/USOO/18079 reaction mixture was poured into water, washed with diethyl ether, acidified with IM HCI, extracted with CHCT 3 -MeOH 20 ml x dried over anhydrous MgSO,, and concentrated under a reduced pressure. The residue was solidified with CHC13Vn-hexane to give 313 (283 mg, as a white solid. IR (KBr): 3345, 1581, 1529, 1243, 1167 cm"; 'H-NMR (CD 3 OD)8 2.30 (s, 3H1), 3.71 IH), 3.78 (m,1IH), 3.82 (m,1IH), 3.89 (s,1IH), 4. 10(m,1IH), 4.19 2H), 4.21 J 5.4 Hz, 1 5.20 (in, 2 5.82 (in, 1 6.95 (in, 2 7.03 (in, 1 7.18 (in, 3 7.28 1 7.60 J 8.1 Hz, 1 7.99 (in, 3 MS (ESI) tn/z 522 Anal. Calcd for
C
2 aHC1N 3 5 i1.75 H 2 0: C, 60.76; H, 5.74; N, 7.59. Found: C, 60.43; H, 5.34; N, 7.17.
Examle 264 methyl 4-[2-N-[2-(4-morpholiny1)ethylJ-N-Ij3-methoxy-4-[N'-(2-methylphenyl)ureidophenyI acetamido]ethoxybenzoate e H 6Me314 To a stirred solution of methyl 4-[2-N-(2,3-dihydroxy- 1-propyl)-[3-methoxy-4-[N'-(2methyiphenyl) ureidolphenylacetaniido~ethoxy~benzoate (1.83 g, 3.24 minol) in THF-MeOH-H,0 1: 1, v /vf/v, 15 mL) was added sodium periodate (2.08 g, 9.71 minol), and stirred for 18 hours at rt. A saturated Na 2
S
2
O
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 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-NMvR (CDCI,) 8 2.32 3 H, J 2.8 Hz), 3.33-4.30 (in, 8 3.72 3 H), 3.86 3 6.20 (in, 1 6.70 (in, 1 6.80 (in, 4 7.06 (in, 1 7.18 (in, 1 7.26 (in, 1 7.49 1 H, J 7.4 Hz), 7.96 (in, 2 8. 10 (in, I 9.57 and 9.63 (2 s, total 1 MS (FAB), ni/z 534 1).
To a stirred solution of methyl 4-[2-N-formylinethyl-N-[3-inethoxy-4-[N'-(2-methyl pheniyl)ureido]phenylacetainido]ethoxylbenzoate (400 mg, 0.75 nimol) in EtOH (7.5 ml), were added inorpholine (654 ml, 7.5 inmol) and acetic acid (429 ml, 7.5 mmol) at it. The reaction was stirred for 5 min. at rt, then cooled to 0 To the cooled solutionl, was added NaBH 3 CN (471 mg, 7.5 minol) 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 washed with brine, and dried over MgSO,.
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 100:0 to 1: 1) to give 314 (346 mng, as a white amorphous foam. 'H-NMR (CDC 3 400 WO 01/00206 WO 0100206PCT/USOO/18079 MII-z) 8 2.31 3H), 2.46 (in, 4 3.52-3.79 (in, 12 3.70 3 H, J 2.7 Hz), 4.05 and 4.22 (in, total 2 6.24 and 6.29 total 1 6.73 (in, 2 6.85 (in, 2 7.07 1 7.17 (in, 1 7.25 (in, 2 7.50 I H, J =7.3 Hz), 7.96 (in, 2 8.08 (in, 1 MS (FAB), m/.z 605 (Mr+ Anal. Calcd for C3 HINO, 1/2 H 2 0: C, 64.58; H, 6.73; N, 9.13. Found: C, 64.95; H, 6.88; N, 8.82. HC1 salt of 314: Anal. Calcd for C3H 4
,CINO
7 2.5 H 2 0: C, 57.76; H, 6.76; N, 8.16; CI, 5.17; Found: C, 58.29; H, 6.81; N, 7.42; Cl, 5.05.
Example 265 4-12-N-12-(4-morpholinyl)ethyl]-N-13-methoxy-4-IN '-(2-methylphenyl)ureido]phenyl]acetanido ethoxybenzoic acid
-~COOH
Me H6M 315 To a solution of methyl 4-[2-N-[2-(4-morpholinyl)ethyl]-N-[3-methoxy-4-[N-(2-methylphenyl) ureidolphenyl]acetamidolethoxybenzoate (146 mng, 0.24 iniol) in THF-MeOH 1, v/ v, 6 ml), was added 0.25 N NaOH (1.9 ml], 0.48 minol) 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 HCI. The mixture was extracted with CHCI,-MeOH 1, v/ v, 30 mld x The combined organic solvent was dried over MgSO 4 After removal of solvent, the residue was crystallized with diethylether to give 315 (102 mg, 7 as a white powder. 'H-NMR (CD 3 QD) 8 2.28 J Hz, 3 2.46 (in, 1 2.40 (in, 1 2.56 (in, I 2.63 (in, 1 3.62-3.80 (in, 12 3.85 (s, 3 4.12 (in, 1 4.26 (in, 1 6.82 (in, 2 6.96 (in, 2 7.01 (in, 1 7.17 (in, 2 H), 7.58 J 7.8 Hz, 1 7.93 (in, 3 MS (FAB) m/z 591 Anal. Calcd for CH~N 4
O
7
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-inethoxy-4-[N'-(2-nethylphenyl)ureidoJpheniyl]acetamidoI ethoxybenzoic acid Y( 0 yCOOH 31 Me Hi 1 To a stirred solution of methyl 4-(2-hydroxyethyloxy)benzoate (5.00 g, 25.5 inmol), DMSO (18.1 ml, 255 inmol), Et 3 N (17.7 ml, 127.5 inmol) in CH 2
CI
2 (200 ml]) was added SO 3 -Py (12.2 g, 76.5 inmol). After stirring for 5 h, the mixture was concentrated in vacuo and the residue was diluted with H 2 0 (100 mrl). The mixture was extracted with EtOAc (2 x 200 ml). The combined extracts were washed with brine (100 ml), dried over (MgSO, 4 and evaporated. The WO 01/00206 WO 0100206PCT/USOO/18079 residue was chromatographed on silica gel with hexane-EtOAc 1) to give 4:1 mixture of methyl 4-formyl methyloxybenzoate and methyl 4-hydroxybenzoate (2.00 g) as a white solid. 'H-NMR (CDCl 3 )8 3.90 3 4.64 J= 1.0 Hz, 2 6.92 J= 9.0 Hz, 2 8.02 J =9.0 Hz, 2 9.86 J =1.0 Hz, 1 H).
To a stirred solution of 4:1 mixture of methyl 4-fonmylmethyloxybenzoate and methyl 4hydroxybenzoate (1.00 g) and cyclopropylanune (425 mL, 6.18 minol) in MeOH-AcOH (10: 1, 11 was added NaBH 3 CN (681 mg, 10.3 inmol). After stirring overnight, the mixture was quenched by addition of sat. NaHCO 3 (50 ml) and extracted with CHC1 3 (2 x 200 ml). The combined extracts were dried over MgSO, and evaporated. The residue was chromatographed on silica gel with CHCI 3 -MeOH (20: 1) to give methyl 4-(2-cyclopropyl aminoethoxy)benzoate (595 mg, as acolorless oil. 'H-NMR (CDCl 3 )8 0.37-0.49 (in, 4 1.91 (in, 1 2.18-2.23 (in, I 3.11 J =5.2 Hz, 2H), 3.88 3H), 4.12 J =5.2 Hz, 2H), 6.92 J =8.8 Hz, 2H), 7.98 J 8.8 Hz, 2 H).
A mixture of methyl 4-(2-cyclopropylaminoethoxy)benzoate (290 mg, 1.23 mmol), 3methoxy-4-[N'-(2-methylphenyl)ureidophenylacetic acid (387 mg, 1.23 inmol), EDC-HCI (354 mg, 1.85 inmol), HOBt and DMAP (cat.) in DMff (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, 65%) as a yellow viscous oil. 'H-NMR (CDC1 3 )S 0.90-0.97 (in, 4 2.28 3 3.60 3 H), 3.77 J 5.4 Hz, 2 3.85 2 3.87 3 4.15 J 5.4 Hz, 2 6.60-6.81 (mn, 5 H), 7.09-7.23 (in, 4 7.57 J= 8.3 Hz, 1 7.92-7.95 (in, 2 8.04 J =8.3 Hz, 1 H).
To a stirred solution of methyl 4-[2-[N-cyclopropyl-[3-methoxy-4-[N'-(2-inethylphenyl) ureidolphenyl]acetamidolethoxylbenzoate (426 mg, 0.80 1 iniol) in THF (7 mld) was added 0.25 N NaOH (6.4 ml, 1.60 inmol). After stirring overnight, the mixture was poured into 1 N HCl ml) and extracted with CHC1 3 -MeOH 2 x 200 ml). The combined extracts were dried over MgSO, and evaporated. The residue was chromatographed on silica gel with CHCI 3 -MeOH (20:1 to 10: 1) as eluent to give 316 (333 mg, as a colorless amorphous solid. 'H-NMR (DMSO) 8 0. 86-0.91 (in, 4 2.25 3 2.74 (in, 1 3.69 J= 5.5 Hz, 2 3.81 3 3.83 2 4.15 J= 5.5 Hz, 2 6.75 J= 8.3 Hz, 1 6.87 1 6.92-6.99 (in, 3 7.11-7.17 (in, 2 7.81 J =8.1 Hz, 1 7.88 J =8.5 Hz, 2H), 8.01 J=8.1 Hz, 1 8.47 I 8.55 1 12.96 br MS (FAB), m/~z 518 Anal. Calcd for C291 31
N
3 0 6
C,
WO 01/00206 WO 0100206PCT/USOO/1 8079 67.30; H, 6.04; N, 8.12. Found: C, 66.7 1; H, 6.26; N, 7.82.
Example 267 4-12-N-12-(N', N' -dmethylamino)-1-ethyl)-N-[-4-IM '-(2-chlorophenyl)ureido]-3-methoxphenylJ acetamido]ethoxybenzoic acid 1 HH 6Me317 To a solution of methyl N'-dimethylamino)- I -ethyl)-N- chiorophenyl) ureido] -3 -methoxphenyllacetanidolethoxybenzoate (100 mg, 0. 17 mmol) in THE- MeOH v/ v, 3 ml), was added 0.25 N NaOH (2.0 mil, 0.5 nunol) at rt, and heated to reflux.
The stirring was continued for 3 hours at reflux. The reaction mixture was poured into water, and acidified to pH 5.0 with 1.0 N HCL 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 mng, 64%/) as a white powder. 'H-NMR (CD 3 OD) 8 2.41 2 2.65 3 2.69 3 3.02 (in, 2 H), 3.62-3.85 (in, 4 3.84 3 4.05 and 4.22 (in, total 2 6.82-6.88 (in, 4 7.02 (in, 1 H), 7.25 (in, 1 7.3 8 (in, 1 7.92 (in, 2 8.01 (in, 2H); MS (FAB), ,n/z 569 Anal. Calod for C2,F1 33 C1N 4
O
6 3.0 H 2 0: C, 55.90; H, 6.3 1; N, 8.99. Found: C, 56.40; H, 6.50; N, 8.08.
Example 268 isopropyl 4-[2-N-[2-(4-morpholinyl)ethylj-N-[3-methoxy-4-[N'-(2-methylphenyl)ureidoj phenyll acetamidolethoxybenzoate e H H Me318 To a stirred solution of 4-[2-N-f2-(4.morpholinyl)ethyl]-N-[3-methoxy-4-[N-(2methyiphenyl) ureidolphenyl]acetamidolethoxybenzoic acid (250 mng, 0.42 inmol) in DMF (2 mL), were added isopropyl iodide (264 nml, 2.53 inmol) and K 2 C0 3 (88 mng, 0.64 mmol) at rt. The reaction was stirred for 2 hours at 50 CC. The mixture was poured into brine and was extracted with CHC1 3 (50 mil x washed with brine, and dried over MgSO 4 After removal of the solvent in vacua, 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 mng, as a white amorphous foam. 'H-NMIR(CDC 3 400 Miz) 81.35 3H), 1.36 3H), 2.31 3 2.45 (in, 4 2.50 (in, 2 3.55-3.78 (in, 10 3.70 3 4.05 and 4.20 J 5.2 Hz, total 2 5.22 (in, 1 6.24 and 6.33 (2 s, total 1 6.70-6.83 (in, 4 7.08 I H), 7.15 (in, 1 7.22 (in, 1 7.40 J= 9.0 Hz, 1 7.93 J= 8.8 Hz, 1 7.97 J= 8.8 WO 01/00206 WO 0100206PCT/USOO/1 8079 Hz, 1 8.07 J 7.8 Hz, 1 MS (FAB), m/z 63 3 Anal. .Calcd for C,,FIN 4 Oj.
H
2 0: 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)ureidoj-3 -methoxyphenyl] acetamidolethoxybenzoic acid sodium salt
F
yCOONa r H 6Me319 To a stirred solution of methyl 4-[2-N-formylmethyl-N-[3-methoxy-4-[N'-(2bromophenyl)ureido] phenyllacetamido]ethoxybenzoate (300 mg, 0.5 nunol) in 1 ,2-dichloroethane (3.6 ml), was added 3,3-difuoropyrrolidine AcOH salt (420 mg, 2.5 mmnol) at rt. The reaction was stirred for 5 mlin. at Mt then cooled to 0 To the cooled solution, was added NaBH(OAc) 3 (530 mg, 2.5 mmol), and the stirring was continued for 4 It at rt. The mixture was poured into sat. NaHCO 3 was extracted with CHC1 3 (50 mL x washed with brine, and dried aver 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 CHC1 3 -MeOH 10:0 to 97:3) to give methyl 4-[2-N-[2-(3,3-diluro- l-pyrrolidinyl)ethyl] -N-[4-IN '-(2-bromophenyl) ureido]-3-methoxyphenylj acetamuido]ethoxybenzoate (345 mg, 100%/) as a white amorphous foam.
'H-NMR (CDC 3 400 MFz) 8 2.20-2.80 (in, 6 3.48 (in, 2 3.70-3.80 (in, 9 3.89 3 H), 4.09 (in, I 4.21 (in, I 6.79-6.85 (in, 4 6.93 (in, 1 7.08 (in, 2 7.30 (in, I 7.50 (in, 1 7.96 (in, 3 8.13 (dd, J 1.5 Hz, 8.3 Hz, 1 MS (ESI) m/z 689 To a solution of methyl 4-[2-N-[2-(3,3-difluoro-l -pyrrolidinyl)ethyl]-N-[4-[N bromophenyl) ureidoj-3-methoxyphenyljacetamidojethoxybenzoate (345 mg, 0.5 nunol) in THE- MeOH (1L 1, v/ v, 8 ml), was added 0.25 N NaOH (4 ml, 1.0 inmol) at rt, and heated to reflux.
The stirring was continued for 6 hours at reflux. The solvent was removed, and the residue was chroinatographed on IHP-20 (H 2 0-MeOH, 0: 100 to 100: 0) to give 319 (306 mg, 9 as a pale red powder. 'H-NM1R (CDOD, 400 M1Hz) 8 2.20-2.90 (in, 6 3.60 (in, 2 3.70-3.92 (in, 9 4. 10 (in, 1 4.22 (in, 1 6.84 (mn, 4 6.96 (mn, 1 7.30 (in, 114), 7.55 (in, 11H), 7.93 (mi, 3H), 7.97 (in, 1 MS (ESI) m/z 676 Anal. Calcd for C 3 1 H1 32 Brf 2
NIO
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 4-[2-N-(Mmethoxy-'-methylanino)ethyl-N-[3-nethoxy-4-[N' '-(2-inethylpheniyl)ureido] WO 01/00206 PTU0/87 PCT/USOO/18079 phenyljacetamido]ethoxybenzoic acid sodium salt Na Me H H 6Me 320 To a stirred solution of methyl 4-[2-N-formiylmethyl-[3-methoxy-4-[N'-(2-methylphenyl) ureidol phenylacetamidojethoxy]benzoate (3 50 mg, 0.66 mmol) in EtOH (13 ml), was added Nmethoxy-N-methylamine hydrochloride (637 mg, 6.6 mmol) at rt. The reaction was sonicated for min. at i, then cooled toO 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, and was extracted with CHCl 3 (50 ml x washedi with brine, and dried over MgSO,. 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 2: 3) to give the title compound (344 mg, 91%) as a white amorphous foam. 'H-NMR (CD Cl 3 400 MHz) 8 2.29 3 2.52 and 2.58 total 3 2.79 (in, 2 3.49.3.76 (in, 9 3.88 3 H), 4.05 and 4.20 (mn, total 2 6.69-6.86 (in, 5 7. 10 (in, 1 7.20 (in, 2 7.29 (mn, 1 7.44 (in, 1 7.94 and 7.99 J 8.6 Hz, total 2 8.06 (in, 1 MIS (FAB), ni/z 579 1); Anal. Calcd for C 3 jH~sNIO 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 -methoxy-N'-methylamiino)ethyl-N113-methoxy-4-[N''- (2-inethylphenyl)ureidojphenyllacetamidolethoxybenzoate (138 mg, 0.24 mmcl) in THF-MeOH 1, v/ v, 4 ml), was added 0.25 N NaOH (1.9 mld, 0.48 minol) at rt, and heated to reflux. The stirring was continued for 18 hours at reflux. The solvent was removed, and the residue was chroinatographed on HP-20 (H 2 0-MeOH, 100: 0 to 0: 100) to give 320 (140 mg, 100%) as a white powder. 'H-NMR (CD 3 OD) 5 2.29 3 2.54 and 2.56 (2 s, total 3 2.82 (mn, 2 3.48 (in, 2 3.65-5.80 (in, 9 4.09 and 4.21 (2 m, total 2 6.80 (in, 4 7.00 J 7.5 Hz, I H), 7.18 (in, 2 7.57 J= 7.8 Hz, I 7.88 (mn, 2 7.99 (mn, I MIS (FAB), m/z 565 1); Anal. Calcd for CH33lNO 7 Na- 1.0 H 2 0: C, 59.59; H, 6.17; N, 9.27. Found: C, 59. 10; H, 6.28; N, 8.86.
Example 271 4-[2-N-(N'-inethoxy-N'-inethylamfino)ethyl-N-[4-[N '-(2-bromopheniyl)ureido]-3-inethoxyphenyI acetaniido]ethoxylbenzoic acid WO 01/00206 WO 0100206PCT/USOO/18079 I
OH
-H H 6Me 321 To a stirred solution of methyl 4-[2-N-formylmethyl-N-[3-methoxy-4-[N'-(2bromophenyl)ureido] phenyljacetamido]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 for 5 min. at xl, then cooled to 0 To the cooled solution, was added NaBH(OAc) 3 (370 mg, 1.75 mmol) and the stirring was continued for 18 hat rt. The mixture was poured into sat.
NaHCO 3 extracted with CHC1 3 (50 ml x and dried over MgSO 4 After removal of the solvent, the residue was chromatographed on TLC (Whatman, PLK-5F, 2 plates, CH-C1 3 -MeOHi, 98:2) to give methyl 4-[2-N-(N'-methoxy-N'-methylamino)ethyl-N-[4-[N -(2-bromophenyl)ureidoJ -3methoxyphenyl acetamidolethoxylbenzoate (89 mg, as a white amorphous foam. 'H-NMR (CDCl 3 400 M&z) 8 2.52 and 2.60 (2 s, 3 2.80 (in, 2 3.48 and 3.50 (2 s, total 3 3.65 (in, 2 3.72 3 3.77 (in, 4 3.90 3 4.08 (in, 1 4.22 (in, 1 6.82 (in, 5 H), 7.12 2 7.30 1 7.52 J 8.1 Hz, 1 7.94 (mn, 3 8.15 J= 8.3 Hz, 1 H);MNS (ESI) m/~z 643 (hr).
To a solution of methyl 4-[2-N-Amethoxy-'-ethylanino)ethyl--[4-[N-(2bromophenyl)ureidoj-3-methoxyphenylacetamidolethoxyjbenzoate (89 mg, 0. 14 inmol) in TI-F- MeOH 1, v/ v, 6 ml), was added 0.5 N NaOH (1.4 ml, 0.7 inmol) 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 I N HCI to pH 5, extracted with CHCI,-MeOH 1, v/ v, 30 mL x dried over MgSO,. The solvent was removed in vacuo to give 321 (53 mg, 60/o) as a white powder. 'H-NMR (CD 3 OD, 400 MHz)8 2.62 and 2.64 (2 s, total 3 2.80 (in, 2 3.50 J 7.3 Hz, 3 3.63-3.88 (in, 6 4.12 (in, 1 4.25 (in, 1 6.82-7.00 5 7.30 (in, 1 H), 7.58 (in, 1H), 7.95 (in, 4 MS (FAB), mhz 629 Anal. Calcd for C 3 J4 33 BrNO 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.
Exampole 272 4-f N'"-diallyl)ethyl-N-[3-inethoxy-4-[N' '-(2-inethylphenyl)ureido]phenyljacetainidoI ethoxybenzoic acid sodium salt 0e H H6M e322 WO 01/00206 WO 0100206PCT/USOO/18079 To a stirred solution of methyl 4-[2-N-formylmethyl-N-[3-methoxy-4-[N'-(2methylphenyl)ureidoJ phenyl~acetamidolethoxybenzoate (400 mg, 0.75 ramol) in EtOH (15 ml), were added diallylamnine (926 ml, 7.5 nimol) and acetic acid (429 ml, 7.5 mmol) at rt. The reaction was stirred for 5 min. at it, then cooled to 0 To the cooled solution, was added NaBHCN (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 washed with brine, and dried over Mg SO 4 After removal of the solvent in vacua, 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 4-112-N-(N', N'"-diallyl)ethyl-N-[3-methoxy-4-(N' methylphenyl)ureido] phenyllacetamido] ethoxybenzoate (385 mg, 84%) as a white amorphous foam. 'H-NNM (CDCI 3 400 MHz) 8 2.30 3 2.60 (in, 2 3.09 (in, 4 3.49 (in, 2 H), 3.60 2 3.70 (in, 5 3.89 3 4.02 and 4.19 (2 mi, total 2 5.01-5.20 (in, 4 4.79 (in, 2 6.30 and 6.32 (2 s, total 1 6.70-6.84 (in, 5 7.08 (in, 1 7.14 (in, 1 7.25 (in, 1 7.60 (in, 1 7.93 and 7.98 (2 d, J= 8.8 Hz, 8.03 and 8.06 (2 d, J= 8.3 Hz, 1 MS To a stirred solution of methyl N'-diallyl)ethyl-N-[3-methoxy-4-[N' methylphenyl) ureidojphenyl~acetamido~ethoxybenzoate (385 mg, 0.63 mmol) in MeOH (3 ml), was added IN HCl (756 ml, 0.76 mmol) at rt. The reaction was stirred for 5 min. at it, then evaporated to give methyl 4-f N'-diallyl)ethyl-N-[3-methoxy-4-(N' '-(2-methylphenyl) ureidojphenyl] acetamidojethoxybenzoate HCI salt (385 mig, 99%1/) as an amorphous foam. Anal.
Calcd for C 3 5H3CIN1O 6 0.5 H,0: C, 63.67; H, 6.72; N, 8.49. Found: C, 63.67; H, 6.69; N, 8.43.
To a solution of N'-diallyl)ethyl-N-[3-methoxy-4-[N' '-(2-methylphenyl) ureidolphenyllacetarnido]ethoxybenzoate (175 mig, 0.29 inmol) in THF-MeOH 1, v/ v, 20 ml), was added 0.25 NaOH (2.5 ml, 0.63 mmol) at it, and heated to reflux. The stirring was continued for 1 hours at reflux. The solvent was removed, and the residue was chromatographed on HP-20 (H,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 3 2.60 J 6.9 Hz, 1 2.67 J= 7.0 Hz, 1 3. 10 J= 6.6 Hz, 2 3.14 J 6.6 Hz, 2 H4), 3.59 (mn, 2 3.69-3.80 (in, 4 3.80 3 4.06 J 5.2 Hz, 4.21 J= 5.1 Hz, 1 5.15 (in, 4 5.80 2 6.79 (in, 2 6.84 J =8.8 Hz, 2 H), 7.00 J= 7.5 Hz, 1 7.14 (in, 2 7.48 (in, 1 7.91 (dd, J= 6.1 Hz, 8.8 Hz, 2 8.00 (in, MIS (FAB), m/z 601 Anal. Calcd for C3,1 39
N
4
O
6 Na -0.5 H 2 0: C 64.65; H, 6.38; N, 8.87. Found: C, 64.53; H, 6.58; N, 8.78.
WO 01/00206 WO 0100206PCT/USOOI 18079 Example 273 N'-diallyl)ethyl-N-[4-[N '-(2-chlorophenyl)ureido]-3-methoxyphenylacetamido~ethoxyI benzoic acid sodium salt KJ~j> COONa 32 To a stirred solution of methyl 4-[2-N-formylmethyl-N-[4-[N'-(2-chlorophenyl)ureido]-3methoxy phenylacetamidolethoxy]benzoate (100 mg, 0. 18 minol) in EtOH (3.6 ml), was added diallylamine (223 ml, 1.81 mmol) at rt. The reaction was stirred for 5 mn. at rt, then cooled toO0 To the cooled solution, were added AcOH (104 ml, 1.81 nimol) 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 was extracted with CHC1 3 (30 niL x 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 N'-diallyl)ethyl-N-[4-[N'-(2-chlorophenyl)ureidoJ-3-methoxyphenyl acetamidolethoxy] benzoate (96 mg, as a white amorphous foam. 'H-NMR (CDC1 3 400 MI-z) 82.60 2H), 3.09 J =6.4 Hz, I1H), 3.11l(d, J =6.4 Hz, 3H), 3.52 2H), 3.61 2 3.70-3.80 5 3.86 3 4.05 and 4.20 (2 m, total 2 5.06-5.21 (in, 4 5.80 (in, 2 6.71-6.85 (in, 4 6.98 (in, 1 7.22 (in, 1 7.32 (in, 3 7.93 J 7.8 Hz, 2 H), 7.98 (mn, 1 8.18 (dd, J 1.5 Hz, 8.2 Hz, 1 MS (ESI) m/lz 635 To a solution of methyl N'-diallyl)ethyl-N-[4-[N'-(2-chlorophenyl)ureidoJ -3methoxy phenylacetamido]ethoxy]benzoate (96 mg, 0. 15 minol) in THF-MeOH 1, v/ v, 8 ml), was added 0.25 N NaOH (3.91 mil, 0.98 mmol) at rt, and heated to 50 The stirring was continued for 6 hours at reflux. The solvent was removed, and the residue was chromatographed on HP-20 (H2O-MeOH, 0: 100 to 100: 0) to give 323 (88 mng, 94%) as a white powder. 'H-NMR
(CD
3 OD, 400 MHz) 8 2.61 (mn, 2 3. 10 2 3.12 2 3.59 (in, 2 3.70 2 3.78 (mn, 2H), 3.82 3 4.10 1 4.21 (in, 1 5.18 (mn,4H), 5.81 2H), 6.82 4H), 7.01 Ct, J 7.8 Hz, 1 7.25 (in, 1 7.39 J 7.8 Hz, I 7.90 (in, 2 8.02 (in, 2 MS (ESI) m/z 621 Anal. Calcd for C 3 3fICINO 6 Na .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 4-2N[-ehx--A-2mtypey uedjhnl--ehlctaioeh1piperazinylacetic acid WO 01/00206 WO 0100206PCT/USOO/18079
NKN..COOH
To a stirred suspension of 1-(2-hydroxyethyl)piperazine (5.2 1ig, 40.Onunol) and K 2 C0 3 (8.76g, 63.4mmol) in CH 3 CN (lO0mI) was added ethyl bromoacetate (5.60m], 50.5nunol) at 0 0
C.
The reaction mixture was heated under reflux for 5h, diluted with EtOAc, and washed with water and brine. The extract dried over Na 2
SO
4 concentrated to dryness and afforded ethyl 4-(2hydroxyethyl)-l-piperazinylacetate (9.65g, 100%) as a yellow oil. 'H-NMR (CDC 3 )8 1.23 3H Jr7.3Hz), 2.5 I-2.61(m,I1 3.22(s, 2H), 3.6 1(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 34m1, 4.2Ommol) in THF (19m1) was added dropwise methylamine (2.OM THF solution, 2.3ml, 4.60mmol) at 0 0 C. The reaction mixture was stirred for Ilhr, 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 NaSO, and concentrated to dryness. The residue was recrystallized from EtOAc-Et 2
O
to give methyl 2,4-dinitrobenzenesulfonamiide (546mg, 56%) as a colorless solid. 'H-NMR (DMSO) 8 2.60 3H,.PJ=4.9Hz), 8.22 lH, J=8.8Hz), 8.31 1H, J=4.9Hz), 8.66 (dd, lH, J=8.8, 2.0Hz), 8.90 1H, Jr2.OHz).
To a solution of ethyl 4-(2-hydroxyethyl)-l-piperazinylacetate (452mg, 2.O9mmoI), methyl 2,4-dinitrobenzenesulfon-amide (546mg, 2.09mmol) and PPh 3 (658mg, 2.5 1immol) in THF was added DIAD (0.50ml, 25 1minoI) at 0 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 afford ethyl 4-[2-[N-(2,4-dinitrobenzensulfonyl)-N-methylanminolethyl]l1-piperazinylacetate (864mg, 90%) as a reddish oil. 'H-NMR (CDC 3 8 1.27 3H, J=6.8Hz), 2.35-2.63 (in, 101-), 2.98 3H), 3.20 2H), 3.41 2H, .1=6.8H1z), 4.17 2H, T-=6.8Hz), 8.33 1H, J=8.3Hz), 8.46 1H, J=2.OHz), 8.50 (dd, I1H, J1=8.3, A solution of ethyl 4-[2-[N-(2,4-dinitrobenzensulfonyl)-N-methylaminoethyl-1 piperazinylacetate (864mg, 1.88 mmol) ,mercaptoacetic acid (0.1l7ml, 2.44minol) and Et 3
N
(0.53ml, 3.76mmoI) in CH 2
CI
2 (25m1) was stirred at rt for 3hr. The reaction mixture ethyl 4-(2methylaminoethyl)-1-piperazinylacetate (388mg, 90%) as reddish oil. 'H-NMR (CDC 3 8 1.27 (t, 3H, .1=6.8Hz), 2.50 3H), 2.53-2.60 (in, 8H), 2.75 2H, J=5.9Hz), 3.20 2H), 4.18 2H, .1=6.8Hz).
To a solution of ethyl 4-(2-methylami noethyl). I-piperazinylacetate 3 88mg, 1 .69minol), EtN (0.32m1, 2.25mmol) and DMAP (46mg, 0.38mrnol) in DMF (15m1) was stirred for 15mmd at room temperature, then 3 -methoxy- 4 -[j'h(2.methylphenyl)ureido~phenylacetic acid (532mg, l.69rnmol), HOBt (103mg, 0.76mmol) and EDC-HCT (486mg, 2.53 mrnol) was added to the reaction mixture which was stirred for 15h at room temperature The reaction mixture was diluted with EtOAc, which was washed with brine, dried over Na,S0 4 and concentrated to dryness. Chromatography of the residue with CHCI 3 -MeOH (10: 1, v/v) to afford ethyl -methoxv- 4 [A(2methylp henyl)u reidophenyl Nm ylacetoaidthl-l pi perazi nyl acetate (889mg, mixture of DMF-) as a reddish oil. 'H-NlvIR (CDC 3 5 1.25-1.29 (in, 3M-I, 2.29 3M-1, 2.42-2.63 (mn, I01OM, 3.20, 3.18 (each s, total 31-1, 3.55, 3.40 (each t, total 2H, J=6.8Hz), 3.65, 3.69 (each s, total 2H), 3.72 31M1, 4.154.21 (mn, 2M-1, 6.50 (in, 6.77-6.81 (in, 8M, 7.11-7.24 (in, 31-M, 7.53 IH, J=8.3Hz), 8.02 1IM1, 8.06 IH, J=7.8Hz).
V se To a stirred solution of ethyl 4 2 3 -methoxy-4-[iN-(2-methylphenyl)ureido]phenyl]> 0* so soo*o A'-methylacetoamidoj ethyl- I -piperazinylacetate (889mg, 1 .69mmol) in THF-EtOH 1, v/v, 18 15 ml) was added 4N NaOH (0.84m], 3.38inmol). The reaction mixture was stirred at iifor 4h, adjusted to pH 7.5 with IN HCI and extracted with CHCI,-MeOH 1, The combined 0 extracts were dried over MgSO, and concentrated to afforded 324 (2 18mg, 26% 2steps) as a brown amorphous foam. IR (KBr) n 3299, 3004, 1700, 1627, 1598, 1536cm- 1 'H-NMR (DMSO) 862.25 2.36-2.62 (in, IO,2.4 29 (each s, totl 3, .14 (each s, total 2Mi~, 3.834 20 (mn, 2H), 3.61, 3.65 (each s, total 2H), 3.86 6.74 1H, J=7.8HZ), 6.87 IN)f, 6.93 I1H, looses: 1=7.8H4z), 7 .11- 7 .17(m, 2M-1, 7.79 IH, J=7.8Hz), 8.01 I1H, J=7.8Hz), 8 4 7 8.57 (s, MS (FAB) ,n/z 498 Anal. Calcd for C1HAjN,S2HCl.HO: C, 53.06; H, 6.67; N, 11.89. Found: C, 53.04; H, 6.15; N, 11.09.
'*elb* Exanmle 275 1 -fN-methyl-N-[3 -mtoy4[P-2mtypey~ edlhnl ctnLd]ehl 4 piperidinylacetic acid NYW COCH MeH H OMe 325 To a stirred solution of 2 -(N-benzyloxycarbonyl..N.methylanmdno)acetaidehyde (2.07 g, 10.0 mimol) and ethyl 4 -piperidinylideneacetate (1.69 g, 10.0 rnmiol) 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 mixture was quenched by addtion of sat. NaHCO 3 (200 ml) and extracted with CHC1 3 (3 x 150 ml).
The combined extracts were dried over MgSO, and evaporated. The residue was crhomatographed on silica-gel with CHC1 3 -EtOH (40: 1, v/v) to give ethyl I 2 -(N-benzyloxycarbonylNAf WO 01/00206 WO 0100206PCTIUS0OII 8079 methylamino) ethyl] -4-piperidinylideneacetate (1.71 g, 47%) as a colorless oil. 'H-NMR (CDCI 3 8 1.25 J 7.3 Hz, 3 2.16 (in, 2 2.57 (in, 4 2.95 (in, 7 3.44 (m,2 4.13 J= 7.3 Hz, 2 5.12 2 5.49-5.53 (in, 1 7.35 (in, 5 M).
A solution of ethyl 1 -[2-(N-benzyloxycarbonyl-N-methylamiino)ethyl]-4-piperidiniylidene acetate (1.70 g, 4.72 mmol) in EtOH-AcOH (20: 1, vfv, 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 CHC1 3 (300 mld). The extract was dried over Na 2
CO
3 and evaporated to give ethyl l-(2-methylaminoethyl)-4-pipenidinylacetate (813 mg, 75%) as a yellow oil. 'H-NMR (CDCl 3 8 1.25 J 7.3 Hz, 3 1.68-1.81 (in, 5 H), 1.97 J= 11.2 Hz, 2H), 2.22 J =7.3 Hz, 2H), 2.43-2.47 5H), 2.66 J =6.4 Hz, 2H), 2.85-2.90 (in, 2 4.13 J 7.3 Hz, 2 H).
To a stirred solution of 3-methoxy-4-[N'-(2-methylphenyl)ureido~phenylacetic acid (550 mg, 1.75 iniol) and ethyl l-(2-methylaminoethyl)-4-piperidinylacetate (400 mng, 1.75 iniol) in DMF (10 ml) were added EDC-HCI (503 mg, 2.63 mmol), HOBt and DMAP (cat.) and the stirring was continued overnight. The mixture was diluted with EtOAc (300 ml), washed with brine (200 ml), dried over MgSO,, and evaporated. The residue was crhoinatographed on silica gel with CHCI 3 -EtOH (10: 1, v/v) to give ethyl 1-[2-[N-methyl-N-[3-methoxy-4-[N'-(2inethylphenyl) ureidojphenyl]acetainido]ethyl]-4-piperidinylacetate (697 mg, 76%) as a yellow gum. 'H-NMR (CDCI,) 8 1. 19-2.06 (series of in, 12 2.21 J 7.8 Hz, 2 2.28 3 H), 2.41 J 7.3 Hz, 1 2.46 J 7.3 Hz, 1 2.80-2.89 (in, 2 2.95 and 3.01 each, total 3 3.40 and 3.50 J 6.8 Hz, each, total 2 3.64-3.75 (mn, 5 4.09-4.16 (in, 2 6.59 (s, 1 6.77-6.79 (in, 2 7.12 J= 7.3 Hz, 1 7.21-7.27 (in, 3 7.54 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-inethoxy-4-[N'-(2-methylphenyl) ureidolphenyllacetainidolethylj-4-piperidinylaoetate (690 ing, 1.32 mmol) in THF (1 Iml-) was added 0.25 N aq. NaOH (1 Imln, 2.75 minol) and the stirring was continued overnight. The mixture was diluted with H 2 0 (50 ml), neutralized with 1 N HCI, and extracted with CHCI 3 -MeOH 1, v/v, 3 x 100 ml). The combined extracts wre 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 suspension was refluxed for 30 min with stirring and filtered through Celite. The filtrate was evaporated and the residue was triturated by taking up CHC, and adding hexane until a precipitate foined. This precipitate was collected and dried in vacuo to give 325 (75 mg, as a white WO 01/00206 WO 0100206PCT/USOO/18079 amorphous solid. 'H-NMR (DMSO) 8 1. 19-2.99 (series of m, total 17 3.32-3.43 (in, 4 H), 3.62-3.65 (in, 2H), 3.86 3 6.73 J 8.3 Hz, 1 6.87 1 6.93 J =7.8 Hz, 1 H), 7.11-7.17 (in, 2 7.79 J= 8.3 Hz, 1 8.01 J= 8.3 Hz, 1 8.47 1 8.57 1 H); MS-FAB m/z 497 Anal. Calcd for 2 7 H36NO 5 -HCI: C, 60.84; HK 7.00;1 N, 10.51. Found: C, 60.97; H, 7.14; N, 10.17.
Example 276 1-[2-[N-methyl-N-[4-[N'-(2-methylphenyl)ureidojphenyl~acetamidolethyl]-4-piperidiniylacetic acid Me H H326 To a stirred solution of ethyl 1 -(2-methylaminoethyl)-4-piperidiniylacetate (400 mg, 1.75 minol) and Et 3 N (366 ul, 2.63 minol) in DMF (10 ml) was added pentafluorophenyl methylphenyl)ureido)phenylacetate (788 mg, 1.75 inmol) 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 CHCI 3 -EtOH (10: 1, v/v) to give ethyl 1 -[2-[N-methyl-Nr-[4-[N'"-(2-methylphenyl)ureidolphenyllacetamidolethylJ-4-piperidiny acetate (630 ing, as a colorless oil.
To a stirred solution of ethyl 1 -[2-[N-methyl-N-[4-IN' .<2-methylphenyl)ureido~phenyl] acetamzddojethylJ-4-piperidinylacetate (630 ing, 1.27 nunol) in THE (10 ml) was added 0.25 N aq.
NaOH (10 ml) and the stirring was continued overnight. The reaction nixture was diluted with
H
2 0 (100 ml), neutralized with 1 N HCI, and extracted with CHCI 3 -MeOH 1, v/v, 3 x 100 ml).
The combined extracts were dried over MgSO 4 and evaporated. The residue was triturated by taking up CHCI 3 and adding hexane until precipitate formed. This precipitate was collected and dried in vacuo to give 326 (20 ing, as a white amorphous solid. 'H-NMR (DMSO)B 1.69 (mn, 2.15 (in, 4 2.24 2 2.50 (mn, 2 2.83 and 2.99 each, total 3 3.32-3.49 (mn, 4 3.63 J= 6.8 Hz, 2 6.91-6.95 (in, 1 7.13 (mn, 4 7.39 J =8.3 Hz, 2 7.83 (d, J 7.3 Hz, 1 7.97 (in, 1 9.12 (in, 1 MS (FAB): m/z 467 Example 277 4-f 2-N-[4-[N'-(2-inethylphenyl)ureidojphenyl]-N-methylacetamido]ethyl-l1-piperaziniylacetic acid ~N40O0CCOH e H H327 To a solution of ethyl 4-(2-inethylaminoethyl)-l1-piperazinylacetate (700mg, Et 3 N (0.64m1, 4.58inmol) and DM4AP (75mg, 0.61Inuol) in THE (15m1d) was stirred for 30min at WO 01/00206 WO 0100206PCT/USOO/18079 rt, then 4-IN'-(2-methylphenyl)ureidolphenylacetic acid (917mg, 3.O5nunol), HO~t (82mg, 0.6 immol) 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 CHCI 3 -MeOH (10: 1, vfv) afforded ethyl 4-[2-N-[4-[N'-(2-methylphenyl)ureido] phenylj-N-methylacetamidojethyl- 1-piperazinylacetate (996mg, 66%) as a yellow amorphous foam. 'H-NMR (CDCI 3 8 1.25-1.29 (in, 3H), 2.20 3H), 2.47-2.58 (in, 101-), 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=6.8H-z), 3.64, 3.68 (each s, 2H), 4.15-4.21 (in, 2H), 7.01-7.19 (in, 8H), 7.48 (in, 1H), 7.64 (in, 1H); MIS (FAB) rn/z 496 (Mr+1).
To a stirred solution of ethyl 4-[2-N-[4-[N'-(2-methylpheniyl)ureidolphenylJ-N-methy acetamido] ethyl-lI-piperazinylacetate (996mg, 2.0 1Immol) in THF-EtOH 12 ml) was added 4N NaOH (1.Oml, 4.00mmol). The reaction mixture was stirred at At for 4h, adjusted to pH with IN HCI and extracted with CHCI 3 -MeOH 1, The combined extracts were dried over MgSO, and concentrated to afforded 327 (73mg, as a yellow amorphous foam. [R (KBr) n 3338, 2925, 2850, 2821, 1704, 1627, lS4Ocnf'; 'H-NMR (DMSO) 8 2.24 3H), 2.33-2.61 (mn, 1OH), 2.82, 2.95 (each s, total 3H), 3.00, 3.02 (each s, total 2H), 3.39 2H, J=6.8Hz), 3.60, 3.62 (each s, total 2H), 6.92 1H, J=7.8Hz), 7.09-7. 16 (in, 4H), 7.4 1-7.44 (in, 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/lz 468 Anal.
Calcd for C,3H3N 5 52HCI: C, 55.56; H, 6.53; N, 12.96. Found: C, 54.99; H, 6.45; N, 11.58.
Example 278 '-(2-fluorophenyl)ureidoj-3-methoxyphenylj-N-methylacetamidolethyl- 1piperazinylacetic acid H M 328 To a solution of ethyl 4-(2-methylanunoethyl)-l-piperazinylacetate (695mg, 3.O3mxnol), Et 3 N (0.64m1, 4.58mmol) and D"A (75mg, 0.6lmmol) in DMF (I5mI) was stirred for 15min at rt, then 4-[N -(2-fluorophenyl)ureido]-3-methoxyphenylacetic acid (965mg, 3 .O3nuol), HOBt (82mg, 0.6linmol) 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 NaSO 4 and concentrated to dryness. Chromatography of the residue with CHCI 3 MeOH (10: 1, v/v) afforded ethyl -(2-fluorophenyl)ureido]-3-methoxyphenyl]-Nmethyl acetamido]ethyl-1-piperazinylacetate (1.21ig, ixture of DMF) as a black oil. 'H-NMIR
(CDCI
3 8 1.24-1.29 (in, 314), 2.45-2.59 (in, 101-), 2.98, 3.05 (each s, total 3H), 3.17, 3.20 (each s, WO 01/00206 WO 0100206PCT/USOO/18079 total 2H4), 3.44, 3.52 (each t, total 2H, J6.8H-z), 3.66 4.15-4.21 (in, 2H), 6.77-6.78 (in, 2H), 6.79-7. 11 (in, 3H), 7.68-7.95 (in, 2H), 7.64 (broad s, 1H), 8.20 I H, fr7.8Hz); MS (FAB) m/z 530 To a stirred solution of ethyl 4-[2-N-[4-[N'-(2-fluorophenyl)ureido-3-methoxyphenyl]-Nmethyl acetamidolethyl- 1-piperazinylacetate (1.21g, mixture of DMF) in THF-EtOH 1, v/v, 12 ml) was added 4N NaOH (l.Oml, 4.O0inmol). The reaction mixture was stirred at rt for 4h, adjusted to pH 7.5 with IN HCl and extracted with CHC1 3 -MeOH 1, The combined extracts were dried over MgSO 4 and concentrated to afforded 328 (78mg, 5% 2steps) as a brown amorphous foam. IR(KBr) n3299, 2940, 2830, 1704, 1627, 1598, 1536cm'; 'H-NMR (DMSO) B 2.36-2.61 (in, 1OH), 2.83, 2.98 (each s, total 3H), 3.11 2H), 3.37-3.43 (in, 2H4), 3.62, 3.65 (each s, total 2H), 3.85 3 6.75 (in, IlH), 6.87 I 6.98 I1H), 7.12 I H, J=7.8Hz), 7.20, 7.2 3 (each d, 2H, J7.8Hz), 8.01 lHJ=7.8Hz), 8.17 IH, J=7.8Hz), 8.72 IH), 9.19 1H); MS (FAR) m/z 502 Anal. Calcd for C 5
H
32
FNO
3 -2HCl-0.5HO: C, 51.46; H, 6.05; N, 12.00. Found: C, 51.08; H, 5.69; N, 11.27.
Example 279 3 -fluoro-l1-[2-N-methyl-N-[3-inethoxy-4-[Nf'-(2-inethylphenyl)ureidojphenyllacetanido~ethyl-4piperidinylacetic acid c NN Q cQ O H Me H H6Me 329 To a stirred solution of 1-tert-butoxycarbonyl-4-piperidone (14.9 g, 74.8 mmol) in DMF (35 inL) was added TN4SCI (11.4 iL, 89.7 inmol) and then EtN (25.0 mL, 179 nnol) dropwise at room temperature, and the reaction mixture was heated at 80 *C for 18 hr. Hexane was added to the reaction mixture, and the resulting mixture was washed with sat. NaHCO 3 and brine, dried over NaSO,, and concentrated to dryness. Chromatography of the residue with hexane EtOAc v/v) as eluent gave I -tert-butoxycarbonyl- 1,2,3,6-tetrahydro-4-(trimethylsilyloxy)pyridine (20.4 g, as a yellow oil. 'H-NMR (CDC1 3 8 0.19 1.46 9H), 2.05 2.15 (in, 2H), 3.48 -3.56 (mn, 2H), 3.83 3.91 (in, 2H), 4.79 (broad s, I1H).
To a solution of 1 -tert-butoxycarbonyl-1,2,3,6-tetrahydro-4-(trimethylsilyloxy)pyridine 4 g, 75.0 mmol) in CH 3 CN (500 mL) was added SelectfluorTm (29.2 g, 82.5 minol) at room temperature, and the reaction mixture was stirred for 2 hr. EtOAc was added to the reaction 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- WO 01/00206 WO 0100206PCT/USOO/18079 butoxycarbonyl-3-fluoro-4-piperidone (14.5 g, 89 as a colorless oil. 'H-NMR (CDCI 3 5 1.50 9H), 2.44 J =6.9 Hz, IH), 2.48 -2.63 (in, 2H), 3.26 (ddd, J 13.5, 10.5, 3.9 Hz, 1H), 3.72 (J J= 6.9 Hz, I1H), 4.16 (in, 114), 4.42 (in, IlH), 4.83 (dt, 49.2, 6.9 Hz, I H).
To a solution of triethyl phosphonoacetate (3.72 g, 16.6 minol) in THF (70 mL) was added lithium bis(trimethylsilyl)amide (1.OM THF solution, 15.5 mL, 15.5 mmol) at -78 IC. After being stirred at the same temperature for 1 hr, 1-tert-butoxycarbonyl-3-fluoro-4-piperidone (3.02 g, 13.9 inmol) was added to the reaction mixture. The mixture was stirred for 30 min at the same temperature, quenched by the addition of sat. NHC1 solution and extracted with EtOAc. The extracts were washed with brine, dried over NaSO,, and concentrated to dryness.
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. IH-NMR 8 1.30 J= 7.1 Hz, 3H), 1.48 9H), 2. 10 (in, 1H), 2.56 (in, 1H4), 2.77 (in, 1H), 3.13 -3.54 (mn, 2H), 3.70 (in, lH), 4.17 4. 18 (each q, J =7.1 Hz, total 2H4), 5.82 5.98 (each s, total 1H), 6.41 (each d, J 46.9 Hz, total IlH); MS (FAB) m/z 288 (M+l A solution of ethyl (1-butoxycarbonyl-3-fluoropipenidin-4-yliden)acetate (1.32 g, 4.59 minol) in THE (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 I -tert-butoxycarbonyl-3 -fluoro-4-piperidinylacetate (653 mg, 73 as a colorless oil.
'H-NMR (CDCI,) 8 1.26 J= 7.1 Hz, 3H), 1.45 9H), 1.53 1.79 (in, 2H), 1.92 -2.09 (in, 2H4), 2.31 (dd, J= 16.4, 6.9 Hz, I 2.52 (dd, J =16.4, 7.3 Hz, I 2.61 -3.06 (in, 2H4) ,4.14 (q, J 7.1 Hz, 2H), 4.28 -4.77 (in, 2H); 3 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; FAR-MS m/z 290 To a solution of ethyl 1 -tert-butoxycarbonyl-3-fluoro-4-pipenidinylacetate (653 mg, 2.26 inmol) in CH 2
CI
2 (10 mL) was added TEA (5 mL) at 0 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 THE EtOAc (I 1, vtv). The extracts were dried over MgSO 4 and concentrated to afford ethyl 3-fluoro-4-pipenidinylacetate (420 mg, 98 as a yellow oil. 'H-NMR
(CDCL
3 8 1.26 J 7.4 Hz, 314), 1.68 1.80 (mn. 2H), 2.18 (mn, 1H), 2.32 (dd, J 16.6, 6.8 Hz, 1 2.54 (dd, J 16.6, 7.5 Hz, I 2.82 (in, 11H), 2.90, 3.00 (each d, J 14.4 Hz, total I 3.3 1 (in, IH), 3.51 (in, 1H), 4.15 J= 7.4 Hz, 214), 4.82, 4.71 (each broad s, total 1H).
WO 01/00206 WO 0100206PCT/USOO/18079 To a solution of ethyl 3-fluoro-4-piperidinylacetate (230 mg, 1.22 mmol) and 2-(AT-tertbutoxy carbonyl-N-methylamino)acetaldehyde (211 mg, 1.22 mmol) in THE (5 mnL) was added NaBH(OAC) 3 (386 mg, 1.82 rnmol) and acetic acid (70.0 niL, 1.22 mmol) at room temperature After being stirred for 24 hr, the reaction mixture was quenched by the addition of sat. NaHCO, solution and extracted with EtOAc. The extracts were washed with brine, dried over NaCO 3 and concentrated to dryness. Chromatography of the residue with CH-C1 3 MeOH (6 1, vlv) as eluent gave ethyl 3-fluoro-l 1 -etbtxyabnlNmtylmn~ty]-iprdnlctt (236 mg, 56%O/) as a reddish oil. 'H-NMR (CDCI 3 8 1.25 J 7.1 Hz, 3H1), 1.45 9H1), 1.53 1.79 (in, 2H), 1.90 2.09 211), 2.15 (dd, J 16.4, 7.1 Hz, 111), 2.45 2.58 (in, 2H1), 2.87 3H1), 2.90 2.99 (mn, 211), 3.20 (in, 111), 3.24 -3.45 (in, 2H1), 4.14 J =7.1 Hz, 2H1), 4.61, 4.73 (each broad s, 111); ESI-MS m/z 347.
To a solution of ethyl 3-fluoro- 1-[2-(N-tert-butoxycarbonyl-N-methylamiino)ethyll-4piperidinylacetate (236 mg, 0.68 mniol) in CH 2
CI
2 (10 m.L) was added TEA (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 CHCI 3 The extracts were dried over MgSO, and concentrated to afford ethyl 3-fluoro-1-[2-(N-methylamino)ethy1J4-piperidinylacetate (117 mg, 70 as a reddish oil. 'H-NMIR (CDCI 3 )8 1.26 J= 7.1 Hz, 3M-1, 1.58 IMH, 1.70 (in, 111), 1.99 (in, 111), 2.14 (m,1E1), 2.27 2.33 (in, 1H1), 2.47 311), 2.48 -2.56 (in, 411), 2.72 (t J 6.3 Hz, 2H), 2.90 (in, 1H1), 3.16 (in, 111), 4.14 J =7.1 Hz, 2H1), 4.67 J =48.3 Hz, 11H); ESI-MS m/z 247 (Mr+1).
To a solution of 3-methoxy-4-[N'-(2-inethylphenyl)ureidolphenylacetic acid (164 ing, 0.52 nol), ethyl 3-fluoro-1-[2-(N-methylamino)ethyl]-4-piperidinylacetate (117 mg, 0.47 innol), Et 3 N 10 niL, 0.71 nimol), and HOBt (13.0 mg, 0.09 mmol) in THE (5 mL) was added EDC-HCl (137 ing, 0.71 minol). After being stirred at room temperature for 8 hr, the reaction mnixture 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 toluene acetone (1: 2, v/v) as eluent gave ethyl 3-fluoro-l1-[2-N-methyl-N-[3 -iethoxy-4-[AN'-(2-inethylphenyl)ureido phenyljacetamtidolethyl-4-piperidinylacetate (160 ing, 62 as a yellow amorphous solid. 'H- N1'MR (CDC1 3 8 1.24 1.28 (in, 3H), 1.51 2.23 (in 711), 2.26 311), 2.35 2H1), 2.39 -2.56 (in, 311), 2.88 (in, 111), 2.95, 3.03 (each s, total 3.11 (in, 1H1), 3.44 (in, 11H), 3.56 (in, 111), 3.64 211), 3.68 311), 4.11 4.17 (in, 2H1), 4.57, 4.69 (each s, total 111), 6.72 6.82 (in, 211), 7.10 -7.33 (in, 5H), 7.54 J 8.1 Hz, 111), 8.06 J 8.1 Hz, 111); ESI-MS m/z 543 WO 01/00206 WO 0100206PCT/USOO/18079 To a solution of ethyl 3-fluoro-1 -[2-N-methyl-N-[3-methoxy-4-[N'-(2-methylphenyl) ureidojphenyllacetamidolethyl-4-piperidinylacetate (160 mg, 0.29 minol) in TI-F (3 niL) 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 HCI at 0 The mixture was concentrated and purified by ion-exchanged resin Mitsubishi Chemical) to give 329 (110 mg, 74 as a yellow amorphous solid. IX (KBr) 3343, 2937, 1700, 1617, 1589, 1535, 1486, 1455, 1417 'H-NMR(CD 3 OD)8 1.57- 1.72Cm ,2H), 1. 98 (in, 111), 2.26 1I1), 2.28 311), 2.37 2.59 Cm, 311), 2.66 2.89 (in, 2H), 2.95, 3.09 (each s, total 311), 3.14 (mn, 111), 3.40 (mn, 111), 3.51 (in, 111), 3.59 (in, 111), 3.71 211), 3.78 (in, 111), 3.89 311), 4.69, 4.81 (each s, total 11-1), 6.79 (dd, J= 8.1, 1.5 Hz, 111), 6.90 (broad s, 111), 7.01 J =7.8 Hz, 1H), 7.13 -7.19 (mn, 211), 7.58(d, J =7.81-z, 1H),8.0O (dJ= 8.1 Hz, 111); ESI-MS m/z 515 Anal. Calcd for C2 7
H
3
FN
4
O,H
2 O: C, 60.89; H, 7.00; N, 10.52. Found: C, 61.09; H, 6.80; N, 9.87.
Example 280 4-[2-N-[2-(4-fluorophenoxy)ethyl-N-[3 -methoxy-4-[N'-(2-inethylphenyl)ureidophenylJ acetamidojethylpiperazinyl- 1 -acetic acid 0
JZXF
Me H 6Me330 To a solution of 2-(N-benzyl-N-terf-butoxycarbonylamino)ethanol (6.85 g, 27.3 inmol), 4fluorophenol (3.07 g, 27.3 mmol) and PPh 3 (7.83 g, 30.0 iniol) in TUF (100 inL) was added DIAD (6.00 inL, 30.0 nunol) at room temperature After being stirred for 3 hr, the reaction mixture was concentrated. Chromatography of the residue with hexane EtOAc 1, v/v) as eluent gave 1 -[2-(N-benzyl-N-tert-butoxycarbonylaxnino)ethoxyj-4-fluorobenzene (8.19 g, 64 as a yellow oil. 'H-NMR (CDC13) 1.42 1.50 911), 3.41 3.67 (in, 211), 3.92 -4.11 211), 4.51 4.63 (in, 211), 6.73 6.85 211), 6.89 -6.98 (mn, 211), 7.24 (m 5H); FAR-MS inz 346 To a solution of 1 -[2-(N-benzyl-N-tert-butoxycarbonylaniino)ethoxy]-4-fluorobenzene (8.19 g, 23.7 inmol) in CH 2
CI
2 (50 niL) was added TFA (40 inL) at 0 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 C11C1 3 The extracts were washed with brine, dried over Na 2 SO,, and concentrated to give 1 -(2-N-benzylaniinoethoxy)-4-fluorobenzene (4.38 g, 75 as a reddish oil. 'H-NMiR (CDC1 3 )8 3.00 J 5.2 Hz, 2H1), 3.87 211), 4.04 J 5.2 Hz, 211), 6.80 WO 01/00206 WO 0100206PCT/USOO/18079 6.85 (in, 2H), 6.92 6.98 (in, 2H), 7.23 7.36 (in, 511); FAB-MS m/z 246 I).
A mixture of 1-(2-N-benzylaininoethoxy)-4-fluorobenzene (1.08 g, 4.40 minol), ethyl 4- (2-bromoethyl)piperazinyl-1 -acetate (1.23 g, 4.40 inmol), and K 2 C0 3 (0.61 g, 17.9 inmol) in
CH
3 CN (50 niL) was heated under reflux for 8 hr. The resulting mixture was filtered and the filtrate was concentrated to dryness. Chromatography of the residue with toluene acetone (3 1, vlv) as eluent gave ethyl 4-[2-N-benzyl-N-[2-(4-fluorophenoxy)ethyl]amiinolethylpiperazinyl-1acetate (1.51 g, 77 as a reddish oil. 'H-NMlR (CDClI)61.27 J=7.1 Hz, 2.31 -2.66 (in, 1011), 2.75 J =6.6 Hz, 2H), 2.90 J =6.1 Hz, 2H), 3.18 211), 3.72 211, 3.97 J 6.1 Hz, 2H), 4.17 J 7.1 Hz, 2H), 6.75 -6.79 (in, 2H), 6.91 6.96 (in, 2H), 7.21 7.35 (in, 5H); FAR-MS m'z 444 (Mr+l).
A solution of ethyl 4-[2-N-benzyl-N-[2-(4-fluorophenoxy)ethyl]amino]ethylpiperazinyl- 1acetate (1.50 g, 3.38 inmol) in EtOH (30 mL) was hydrogenated over 5 Pd-C (53.1 wet, 0.73 g) under hydrogen atmosphere for 4 hr. Thie catalyst was filtered off and the filtrate was concentrated. The residue was taken up with sat. NaHCO 3 solution and extracted with CHC 3 The extracts were washed with brine, dried over Na 2
SO
4 and concentrated to afford ethyl 4-[2-N- [2-(4-fluorophenoxy)ethyllanuno]ethylpiperazinyl-1I-acetate 12 g, 94%) as a reddish oil. 'H- NMR (CDC1 3 8 1.27 J 7.1 Hz, 311), 1.81 (broad s, 111), 2.47 2.66 (in, 1211), 3.00 J 5.4 Hz, 211), 3.20 211), 4.03 J 5.4Hz, 211), 4.18 J 7.1 Hz, 211), 6.81 6.85 (in, 211), 6.90 6.99 (in, 211; FAR-MS m/z 3 54 1).
To a solution of 3-methoxy-4-[N'-(2-methylphenyl)ureidolphenylacetic acid (485 mng, 1.54 inmol), ethyl 4-[2-N-[2-(4-fluorophenoxy)ethyl~aminolethylpiperazinyl-1 -acetate (545 mg, 1.54 inmol), Et 3 N (0.32 mL, 2.32 minol), and HOBt (41.5 mng, 0.31 mmol) in THF (15 mL) was added EDC-HCl (883 mg, 2.32 minol) at room temperature After stirring for 24 hr, the reaction mixture was diluted with water and extracted with CHC1 3 MeOH (10: 1, vfv). The extracts were washed with brine, dried over NaSO,, and concentrated to dryness. Chromatography of the residue with CHCI 3 MeOH 1, v/v) as eluent to give ethyl 4-[2-N-[2-(4-fluorophenoxy)ethylj- N-[3-mtoy4[ (-ehlhny~riopeylcmtd~ehlieaiy--ott (532 mng, 53 as a yellow amorphous solid. 'H-NMR (CDCI 3 )8 1.26, 1.27 (each tL J 7.1 Hz, total 3H1), 2.27 311), 2.51 2.63 (in, 101-1), 3.16, 3.19 (each s, total 211), 3.51 3.56 (in, 211), 3.63, 3.67 (each s, total 211), 3.69 3.81 (in, 511), 3.95, 4. 10 (each t, J Hz, total 211), 4.16, 4.18 (each q, J 7.1 Hz, total 211), 6.56 J 8.1 Hz, 111), 6.72 6.78 (in, 411), 6.89 -6.98 (in, 2H-), 7.12 J= 7.6 Hz, IM1, 7.20 7.23 (mn, 3H1), 7.51 J= 7.6 Hz, 11-1), 8.04 J= 8.1 Hz, 111); WO 01/00206 WO 0100206PCT/USOO/1 8079 FAB-MS m/z 650 (Nr+ 1).
To a solution of ethyl 4-[2-N-[2-(4-fluorophenoxy)ethylj-N-[3-methoxy-4-[N'-(2methylphenyl) ureidoJphenyllacetamido]ethylpiperazinyl-1I-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 the reaction mixture was stirred for 1 hr. The resulting midxture was concentrated, diluted with water, and neutralized with IN HCI at 0 The mixture was extracted with CHCl 3 MeOH (4: 1, vfv). The extracts were washed with brine, dried over Na 2
SO
4 and concentrated to dryness.
Chromatography of the residue with CHC1 3 MeOH (3 v/v) as eluent gave 330 (168 mg, 33 1/)as a pale yellow amorphous solid. IR (KBr) 3338, 2938, 2829, 1635, 1533, 1506, 1454, 1415 crl; 'H-NMR (DMSO-d 6 )8 2.25 311), 2.37 2.48 (in, 611), 2.53 2.67 (in, 611, 3.09 (in, 2H), 3.44 3.49 (in, 2H), 3.64 3.69 (in, 2H), 3.72 211), 3.80, 3.84 (each s, total 3H), 4.06 4.09 (mn, 211, 6.73 (in, 1H1), 6.85 lH), 6.91 6.97 (m ,3H4), 7.07 7.18 (in, 4H), 7.78 J 8.1 Hz, 1H), 8.00 (dd, J 8.1, 2.7 Hz, 111), 8.53 (in, 111, 8.59 (in, 111); FAB-MS m/z 622 (Mr+ Anal.
Calcd for CH1-1 4 YNO4HCl: C, 57.06; H, 6.09; N, 10.08. Found: C, 56.83; H, 6.05; N, 9.90.
Example 281 4-[2-N-[2-(4-acetylphenoxy)ethylJ-N-[3 -iethoxy-4-[Nl'-(2-inethylphenyl)ureidojphenylJ acetamidolethylpiperazinyl- 1 -acetic acid r'aAc M4e H H OMe 331 To a solution of 2-(N-benzyl-N-:ert-butoxycarbonylamino)ethano (5.90 g, 23.5 inmol), 4hydroxyacetophenone (3.18 g, 23.5 mmol) and PPh 3 (6.74 g, 25.8 mmol) in TH-F (100 niL) was added DMAD (5.20 niL, 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-butoxycarbonylamidno)ethoxyjacetophenone (3.64 g, 42 as a yellow oil. 'H-NMR (CDC 3 8 1.43 1.65 (in, 911), 2.55 311), 3.41 3.69 (in, 2H), 4.02 4.24 (in, 4.49 -4.66 (in, 2H), 6.81 6.93 (mn, 2H), 7.19 -7.37 (mn, 511), 7.91 J 8.8 Hz, 211); FAB-MS ni/z 370 (Mr+1).
To a solution of 4-[2-(N-benzyl-N-tert-butoxycarbonylamino)ethoxylacetophenone (3.05 g, 8.26 nunol) in CHCl 2 (30 niL) was addTFA (20 inL) at 0 After being stirred at room temperature for 2 hr, the reaction mixture was concentrated. The residue was taken up with sat.
NaHCO 3 solution and extracted with CHCI 3 The extracts were washed with brine, dried over WO 01/00206 WO 0100206PCTIIJS0OII8079 Na 2
SO
4 and concentrated to give 4-(2-N-benzylaniinoethoxy)acetophenone (2.21 g, 99 as a reddish oil. 'H-NMiR (CDCl 3 6 2.04 (broad s, 111), 2.55 3M), 3.05 J 5.4 Hz, 2H), 3.88 (s, 2H), 4.15 J= 5.4 Hz, 2H), 6.92 J= 8.9 Hz, 2H), 7.24 -7.36 5H), 7.91 J= 8.9 Hz, 2H); FAB-MS m/z 270 1).
To a solution of ethyl 4-(2-hydroxyethyl)piperazinyl- 1 -acetate (11. 3 g, 52. 1 mmol) and CBr, (20.7 g, 62.5 mmol) in CH 2 Cl 2 (200 niL) was added PPh 3 (19.2 g, 73.0 mmnol) portionwise at 0 'C 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-1I-acetate (13.7 g, 94 as a yellow oil. 'H-NMR (CDCl 3 )8 1.27 J= 7.1 Hz, 3H), 2.61 2.78 (in, 8H), 2.81 J= 7.6 Hz, 2H), 3.20 2H), 3.42 J= 7.6Hz, 311), 4.18 J 7.1 Hz, 211).
A mixture of 4-(2-N-benzylaminoethoxy)acetophenone (681 mng, 2.52 nimol), ethyl 4-(2bromoethyl)piperazinyl-1 -acetate (705 mng, 2.52 mniol), and K 2 C0 3 (349 g, 2.52 mmol) in CH 3
CN
mL) was heated under reflux: for 22 hr. The resulting mixture was filtered and the filtrate was 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)ethyljaniino~ethylpiperazinyl-1 -acetate (920 mg, 78%I/) as a reddish oil. 'H-NMR (CDC1 3 8 1.27 J= 7.1 Hz, 2.45 -2.53 (in, 1011), 2.55 3H), 2.76 J 6.8 Hz, 2H), 2.94 J 6.1 Hz, 211), 3.18 2H), 3.73 2H), 4.06 J =6.1 Hz, 2H), 4.17 J 7.1 Hz, 211), 6.85 J 7.1 Hz, 2H1), 7.22 7.35 (in, 7.90 J= 7.1 Hz, 2H); FAB-MS m/z 468 (MX+ 1).
A solution of ethyl 4-[2-[N-benzyl-N-[2-(4-acetylphenoxy)ethyl]axninolethylpiperazinyl- I acetate (920 mg, 1.97 mmol) in EtOH (30 inL) was hydrogenated over 5 Pd-C (53.1 wet, 5 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 CHCI 3 The extracts were washed with brine, dried over Na 2 SO,, and concentrated to afford ethyl 4-[2-N- [2-(4-acetyl phenoxy)ethyl]aminolethylpiperaziniyl-1I-acetate (690 mg, 93%) as a reddish oil. 'H- NMvR (CDC1 3 8 1.27 J 7.1 Hz, 3H), 2.55 2.46 2.54 (in, 1011), 2.78 J 6.2 Hz, 2H1), 3.04 J =5.2 Hz, 2H), 3.20 211), 4.13 J= 5.2 Hz, 2H1), 4.18 J= 7.1 Hz, 6.92 J 8.8 Hz, 2H1), 7.92 J 8.8 Hz, 211); FAB-MS m/lz 3 78 1).
To a solution of 3-methoxy-4-[NV'-(2-inethylphenyl)ureido]phenylacetic acid (558 ing, 1.77 inmol), ethyl 4-[2-N-[2-(4-acetylphenoxy)ethyl]arninolethylpiperazinyl- 1-acetate (670 mng, WO 01/00206 WO 0100206PCT/USOO/18079 1.77 mmol), Et 3 N (0.38 mL, 2.66 mmol), and HO~t (50.0 mg, 0.35 mmnol) in THF (10 mL) was added EDC-HC1 (883 mg, 2.32 mmol) at room temperature After stirring for 15 hr, the reaction mixture was diluted with water and extracted with CHC1 3 -MeOH (10: 1, vlv). The extracts were washed with brine, dried over Na2S0 4 and concentrated to dryness. Chromatography of the residue with CHC1 3 MeQH (4 v/v) as eluent to give ethyl 4-[2-N-[2-(4-acetylphenoxy)ethyl]- N-[3-methoxy-4-[N'"-(2-methylphenyl)ureidolphenyl~acetaniido]ethylpiperazinyl-l1-acetate (724 mg, 61 as ayellow amorphous solid. 'H-NMR (CDCI3) 8 1.25, 1.27 (each t J =7.1 Hz, total 3H), 2.29 3H), 2.45 2.51 (in, 811), 2.54 3H), 2.55 2.63 (in, 2H), 3.17, 3.19 (each s, total 2H), 3.51 3.55 (mn, 2H1), 3.60 2H1), 3.69 3H), 3.71 3.78 (in, 211), 4.04 4.23 (mn, 4H), 6.47 (in, 1H, J =8.1 Hz), 6.72 -6.76 (in, 211), 6.85 J =8.8 Hz, 2H), 7.12 -7.19 (in, 2H), 7.20 7.24 2H), 7.51 8.1 Hz, IH), 7.88 J =8.8 Hz, 2H4), 8.04 J =8.1 Hz, 1H); FAB-MS m/z 674 1).
To a solution of ethyl 4-[2-N-[2-(4-acetylphenoxy)ethyl]-N-[3-methoxy-4-[N'-(2inethylphenyl) ureido~phenyl]acetamidolethylpiperazinyl-1 -acetate (724 mg, 1.07 minol) in THF (10 inL) was added dropwise 0.25N NaOH (6.50 mL, 1.61 iniol) 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 The mixture was extracted with CHCI, MeOH 1, vlv).
The extracts were washed with brine, dried over Na 2
SO
4 and concentrated to dryness.
Chromatography of the residue with CHCI 3 MeOH (3 1, v/v) as eluent gave 331 (450 mng, as apale yellow amorphous solid. IR (KBr) 3345, 2938, 2821, 1673, 1631, 1598, 1533, 1455, 1417 cm-'1; 'H-NMR (DMSO-d 6 8 2.25 311), 2.35 -2.47 (in, 811), 2.51 3H), 2.53 2.58 (in, 2H), 2.96 2H1), 3.46 3.50 (in, 211), 3.69 211), 3.73 3.77 (in, 211), 3.80, 3.83 (each s, total 3H1), 4.19 -4.22(n, 2H), 6.73 (mn, 6.85 1H), 6.92 (t,J =7.3 Hz, IH), 7.30 J= 7.3 Hz, 211), 7.10 7.16 (in, 2H), 7.79 J 8.3 Hz, 11H), 7.90 7.95 (in, 2H), 8.00 J =8.3 Hz, 11H), 8.55 (mn, 111), 8.61 (in, 111); FAD-MS niz 646 (M 4 Anal. Calcd for C 35 F 3
NO
7 -2HCIldHO: C, 57.06; H, 6.43; N, 9.5 1. Found: C, 59.17; H, 6.32; N, 9.6 1.
Example 282 4-[2-N-[4-[N'-(2-bromophenyl)ureido]3-nethoxyphenyl]-N-benzylacetamido ethyl-ipiperidinylacetic acid 'M N N'COOH TrH H6e332 methyl 4-12-N-[4-[N'"-(2-broinophenyl)ureido] -3-methoxyphenyl]-N-benzylacetainiidolethy1- 1- WO 01/00206 WO 0100206PCT/USOO/18079 piperidinylaceate N NN'COOMe A mixture of piperidine ethanol (10.0 g, 77.4 minol), benzyl 2-bromoacetate (17.8 g, 77.6 nunol) and KIC0 3 (21.4 g, 155 ntmol) in CH 3 CN (200 ml) was heated under reflux with 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 lN-HCI and extracted with CHCI 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-(2hydroxyethyl)-1-piperidinylacetate (16.3 g, as a colorless oil. 'H-NMR (CDCl3)8 1.3 1-1.40 (M 3 1.52 (dd, J= 12.8, 6.3 Hz, 2HM, 1.68 (brd, J= 10.0 Hz, 3 2.16 11.6 Hz, 2 H), 2.92 (brd, J= 11.6 Hz, 2 3.24 2 3.69 J= 6.8 Hz, 2 5.16 2 7.35 (in, 5 H).
To a stirred solution of benzyl 4-(2-hydroxyethyl)-1-piperidinylacetate (14.9 g, 53.8 mmol) in CHCI 2 (150 ml) was added Et 3 N (37.5 ml, 269 minol) and DMSO (41.6 ml, 538 minol).
The reaction mixture was cooled to 0 0 C and S0 3 -Py (25.7 g, 161 inmol) was added portion wise, 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 EtO. 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 as eluent to give (4-N-carbobenzyloxy methylpiperidinyl)acetaldehyde (4.63 g, 3 as a colorless oil. 'H-NMR (CDC 3 8 1.36-1.46 (in, 2 1.69 (br s, 2H), 1.72 (br s, 1 1.90 I1H), 2.21 (dt J 11.6, 2.4 Hz, 2H), 2.37 (dd,J 6.8, 1.6 Hz, 2 2.92 J= 11.6 Hz, 2 3.25 2 5.16 2 7.35 (in, 5 9.77 (t,J Hz, 1 H).
To a stirred solution of N-benzylanune (1.06 ml, 9.75 innol) in MeOH (20 was added AcOH (560 ml, 9.75 mmol) and (4-N-carbobenzyloxymethylpiperidinyl)acetaldehyde (1.79 g, 6.50 mmol) in MeOH (5 ml) and cooled to 0 0 C. NaBH 3 CN (645 mg, 9.75 minol) 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 CHC1 3 -MeOH-EtQAc (10 1: 1) as eluent to give Nbenzyl-2-[4-(N-carbobenzyloxymethylpiperidinyl)]ethylamine (872 mng, 37%) as a colorless oil.
WO 01/00206 WO 0100206PCT/USOO/18079 'H-NMR (CDC1 3 8 1.36-1.46 (in, 2 1.69 (brs, 2 1.72 (Ins, 1 1.90 (in, 1 2.21 (dt, J 11.6, 2.4 Hz, 2 2.37 (dd, J 6.8, 1.6 Hz, 2H), 2.92 J 11.6 Hz, 2H), 3.25 2 5.16 2 7.35 (in, 5 9.77 J =2.0 Hz, 1 H).
To a stirred solution of N-benzyl-2-[4-(N-caibobenzyloxymethylpiperidinyl)Jethylainine (356 mg, 0.972 mmol), 4-[N'-(2-bromophenyl)ureido]-3-methoxyphenylacetic acid (369 mg, 0.972 rnmol) and NN-dimethylanuinopyridine (119 mg, 0.972 inmol) in DMF (15 ml) was added EDC-HCI (372 mg, 1.94 minol) at rt, and the resulting midxture 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 chroinatographed on silica gel with CHCI 3 -MeOH (10 as eluent to give benzyl (2-bromophenyl )ureido]3-methoxyphenyl]-N-benzylacetamidojethyl-1-piperidinylacetate (611 mg, as a colorless oil. 'H-NMR (CDCI 3 mixture of rotamars; 8 1. 15-2.09 (series of Rn 7 2.06- 2.14 (in, 2 2.84-2.96 (in, 2 3.17-3.23 total 2 3.21 and 3.23(s, total 2 3.38-3.42 (mn, 1 3.65 and 3.73 total 2 3.83 and 3.84 total 3 H),4.49 1 4.60 1 5.15 J 2.8 Hz, 2 6.74-7.83 (series of m, 16 7.51-7.53 (in, 1 7.94-8.02 (in, 1 8.18 (d, J 8.4 Hz, 2 MIS (FAB) m/z 727 729 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 nunol) in MeOH-H 2 0 1, 6 ml) was added LiOH (13.6 mg, 0.57 minol) at rt, and the resulting mixture was stirred for overnight.
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 CHCI,-MeOH as eluent to give 332 (97.3 mg, as a colorless amorphous solid, and 333 (168 mng, 54%) as a colorless amorphous solid, respectively. 332 'H-NMvR (CD 3 OD), mixture of rotaniars 1.29-1.90 (series of in, 7 M, 2.75-2.86 (mn, 2 3.28-3.51 (series of mn, 6 3.74 and 3.78 (s, total 2 3.87 and 3.89 total 3 4.66 I 4.85 1 6.79-7.00 (series of m, 3 H), 7.16 J =7.2 Hz, I 7.23-7.37 5 7.57 (dd,J 1.2 Hz, 1 7.88-8.00O(series of mn, 2 MIS (FAB) m/z 637 639 333 'H-NMR (CDC 3 mixture of rotamars 8 1. 15-2. 11 (series ofin, 7 2.89-2.97 (mn, 2 3.17-3.49 (series ofim, 6 3.17 and 3.18 (s, total 3 3.70 and 3.71 total 3 3.83 and 3.84 total 2 4.49, 4.60 and 4.71 total 2 6.75-8.16 (series of mn, 14 MIS (FAB) m/z 651 653 (Mr+2).
Example 283 4-[2-N-[4-[N'-(2-chlorophenyl)-3-methoxyureido]phenyl-N-benzylacetamidojethyl- 1-piperidinyl WO 01/00206 WO 0100206PCT/USOO/18079 acetic acid
ON
To a stirred solution of Nr-benzyl-2-[4-(N-carbobenzyloxyinethylpiperidinyl)]ethylamine (372 mg, 1. 11 mmol), -(2-chlorophenyl)ureidoj-3-methoxyphenyl]acetic acid (758 mg, 1. 11 mniol) and N,NV-dimethylanuinopyridine (136 mg, 1. 11 minol) in DMff (I15m1) was added EDC-HCI (426 ing, 2.22 minol) 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 NaSO 4 then concentrated in vacuo. The residue was chromatographed on silica gel with CHCl 3 -MeOH (10 as eluent to give benzyl 4-[2-N-[4-[N'-(2-chlorophenyl)-3methoxyureidojphenyl]-N-benzylacetamidoJethyl-1-piperidinylacetate (668 mg, 88%) as a pale yellowish oil. 'H-NMR (CDCI 3 mixture of rotaniars 6 1. 15-1.83 (series of in, 7 2.04-2. 13 (in, 2 2.83-2.95 2 2.88 and 2.99 total 2 3.20 and 3.23 total 2 3.20-3.23 (overlap, 1 3.38-3.42 (mn, 1 3.65-3.74 (mn, 3 4.50 I 4.61 1 5.14 J 4.4 Hz, 2 6.72-6.82 (series of Mn 2 6.94-6.99 (mn, 1 7.13-7.50 (series of in, 14 7.94-8.02 (in, 1 8.18 J =8.0 Hz, 2H);MNS (FAB) mz683 To a stirred solution of benzyl 4-[2-N-[4-[N'-(2-chlorophenyl)-3 -iethoxyureidojphenyl]- N-benzylacetamnidolethyl-l-piperidinylacetate (633 ing, 0.93 minol) in MeOH-H 2 0 (10: 1, 10 MIr) was added LiOH (24.4 mng, 1.02 inmol) at rt, and the resulting mixture was stirred overnight. The reaction mixture was poured into water and the solution was neutrallized with aq. lN-HCI, then extracted with CHC1 3 The organic layer was washed with brine and dried over anhydrous NaSO, then concentrated in vacuo. The residue was chromatographed on silica gel with CHCI 3 -MeOH as eluent to give methyl 4-[2-N-[4-[N'-(2-chlorophenyl)-3-inethoxyureidolphenyll-Nbenzylacetamido~ethy1- 1-piperidinylacetate (504 mng, 89%) as a colorless amorphous solid. 'H- NMR (CDCI 3 mixture of rotamars 8 1.26-1.64 (series of in, 7 2.02-2. 10 (in, 2 2.86 J 10.4 Hz, 2 3.17 J= 8.0 Hz, 2 3. 16-3.22 (in, overlap, 1 3.40 J =7.6 Hz, 1 M), 3.48 1 3.65 and 3.73 total 2 3.70 and 3.71 total 3 3.79 and 3.80 total 3 H), 4.50, 4.60 and 4.70 total 2 6.73-6.85 (series of in, 2 6.98 J =7.6 Hz, 1 7.13-7.37 (series of in, 9 7.96 (in, 1 8.18 J= 8.0 Hz, 2 H1); NIS(FAB)m/z 607 (MW+H).
To a stirred solution of methyl 4-[2-N-[4-[N'"-(2-chlorophenyl)-3-methoxyureidolphenyl]- N-benzylacetan-idol ethyl-1I-piperidinylacetate (177 mng, 0.292 inmol) in MeOH-H 2 0 1, 6 ml) 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. IN-HCI, then extracted with CHCI 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 mixture of rotamars 8 1.28-1.90 (series of m, 7 2.84 (brs, 2 3.30-3.52 (series of m, 6 H), 3.74 and 3.82 total 2 3.87 and 3.88 total 3 4.63 1 4.66 1 6.79-7.05 (series of m, 3 7.16 J 7.2 Hz, 1 7.24-7.40 5 7.88 1 7.98-8.04 (series of m, 2 MS (ESI) m/z 593 615 (NM+Na).
It should be understood that while this invention has been described herein in terms of 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.

Claims (22)

1. A compound represented by Formula I, or a salt thereof, A N N 1 H H I wherein W is a substituted or unsubstituted phenyl W 1 is a substituted or unsubstituted divalent group of phenyl, pyridine, pyrrolidine or thiazole; io A is =S or =NH; R is -(CH 2 wherein "n is 1 or 2; X is 5 M is chosen from the following groups Q R 2 R 3 R R4 S. N Y-Z-A R wherein N is a divalent 5- or 6- membered heterocyclic radical, with the nitrogen atom being in the position of its attachment to X; wherein Q represents -CH 2 or R 2 and R 3 are independently chosen from the group consisting of -OH, quinolinyloxy, -NH 2 mono- or dialkylamino, alkylsulfonylamino, arylsulfonylamino, naphthalylsulfonylamino, dialkylamino substituted naphthalylsulfonylamino, CI-C 6 alkyl, benzyloxymethyl, halogen, phenyl, CI-C 4 alkoxy, phenyloxy, naphthyloxy, and phenyloxy substituted with COOH or halogen; or [R:\LIBUU102848.docHJG 481 two of adjacent R 2 and R 3 taken together may form an alkylene group having 1 to 5 carbon atoms or alkylenedioxy group having 1 to 3 carbon atoms optionally substituted with from 1 to 3 alkyl group(s); R 4 represents hydrogen or lower alkyl; s Y is chosen from a bond, a C 2 -C 8 alkenylene group, alkynylene group, C(O)-NH- and-(CH 2 )k-Y 2 wherein k is chosen from 0, 1, 2 and 3; and Y2 represents a direct bond or a divalent radical chosen from -S(O) 2 and -NY 3 wherein Y 3 is independently chosen from hydrogen and lower alkyl; Z represents, C 3 -C 8 cycloalkylene, phenylene, pyridylene optionally substituted with halogen, piperidine, piperazine, phenyl, or phenyl independently substituted with 15 one or two group(s), chosen from alkyl, alkoxy, halogen, amino, mono- and dialkylsubstituted amino, acylamino, nitro, carboxy; A' is a direct bond or represents bond, (CH 2 or alkynyl, wherein t is chosen from 1, 2, 3; NH\ NH NH .NH N 20 N- or \N N; 20 R 5 represents OH or lower alkoxy; and -Rl--Z Q2- L i wherein N N R" is chosen from and-NR12 wherein R' 2 is chosen from lower alkyl optionally substituted with alkyl, 3-
6-cycloalkyl,hydroxy, phenyl (optionally substituted with amino or nitro), amino, cyclopropylamino, dialkylamino, diallylamino, methoxymethylamino, morpholino, dihalogen substituted [R:\LIBFF] 12694.doc:HJG 482 pyrrolidino or 4-alkylpiper-idino; lower alkenyl; lower alkynyl; and phenyl; Z 3 is chosen from a divalent aliphatic hydrocarbon moiety having I to 12 carbon atom, wherein x is 0Oor 1; y is 1, 2or 3; R 4 s-r -OH; N is and provided that Z' schosen from the following three formulated divalent grusin the limited case of R' I is -NqR 12 14 Ra wherein 14 Ra is -H or halogen; N N- ,and Q 2 represents 17 R R wherein R 1 7 and R 18 are independently -H or a lower alkyl; [R:\LIBFF] I 2694.doc:HJG 483 or phenylene which may be substituted with halogen, alkoxy, amino, mono or dialkylamino, acylamino, piperidino, nitro or carboxy; L' is chosen from -COOH and -COOR' 9 wherein R 1 9 is a lower alkyl. 2. A compound according to claim 1, or a salt thereof, wherein M is 1 O R2R3 R Y-Z-AR hN 3. A compound according to claim 1, or a salt thereof, wherein M is 1 -R1-Z Z3Q2 -L *fe* 4. A compound according to claim 2, or a salt thereof, wherein at least one radical ofR', R 2 and R 3 is -OH or halogen atom. 15 5. A compound according to claim 4, or a salt thereof, wherein A is R is S* -(CH 2 and X is 6. A compound according to claim 5, or a salt thereof, wherein Y is chosen from alkenylene group, alkynylene group and -(CH 2 )kY 2 Y 2 is chosen from a direct bond, and -NY3-; and Y3 is -H. 20 7. A compound according to claim 6, or a salt thereof, wherein Y 2 is chosen from and -NY 3 S• 8. 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.
9. A compound according to claim 8, or a salt thereof, wherein W' 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. A compound according to claim 2, or a salt thereof, wherein W' 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. [R:\LIBFF] 2694.doc:HJG 484
11. A compound according to claim 2, or a salt thereof, wherein A' is a direct bond or -(CH2)t-.
12. A compound according to claim 11, or a salt thereof, wherein A' is a direct bond.
13. A compound according to claim 12, or a salt thereof, wherein R 5 is -OH.
14. 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' 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 and R 5 is -OH. A compound according to claim 14, or a salt thereof, chosen from the group consisting of *oo*: s* 0 H 0 COOH H N H HNH H °-0-CO2H NYN O^O2H H H H H M f S S S S. Se *5 S* S S S. p. 5 5 5 S *5b5 S 4 6 ~*SS S S 51 S. Y~H~Y 02H H H YN-JN S<XCO 2 H HHe P: NY-(N-O-o-C02 H HH 9 N5N/O aQC2H H H C02 H 486 ON N 0 0 2 H C0 2 H H H -O H0 2 C ciC02H 9iCO-9 C 0 H 9 N Q 2 HH 0* Me j02H H H0 H H H *0JOH OH 2 487 o N a 2 Q,.lC 2 9N-4Q1 02H t&CO2 O1--N ZJ C0Oc~ -N O C2H H H 9 0 Me M (K kq' N~ r C 2 H 1 Q&CO 2 H HH ~N Q NH 9- N C02H F N* H Me rH H eH 2 'o 2H~e 0 2 H H 2 H I~fOOH or Me HH e0 H H- 9 9. iH~e~r~~ e 0 -Z OH 'NYW 0 H,2OOH ~HH Me QC O H 7H 6H. H2 H Me ,JXO S. Yk p Oc OH 'Z 0H r Me QNN qQ OH iiH 6Me QN'p---OOH M, CkOOH H H HC)--COH H HYe00,OOH 489 H 6OH 'H H 6e4 OOH JH2 OOHO m6e OOH OQW O H HrH M H M H Me rO H M OOH MeOH Me 0, ,OOH rHHeH 6MOe 0"OOH TeHH Me ~NNI T) Q q C WH OOH 490 HHGOOH 9 H 6 CoOH(P H I0 ?QHw YL::xAOO H H'OMe ZD,_COOHr O OOM H H NOOH -NAJY OH (e NY Me I'?e HH Me H H 0H H N)NOH NJqOH H H H H ?eH Me Me O-j-OOH -CCOOH l u n lOOH w *TOOH H M OOH OOH OOH H H JMe OOH N'OOH 6Me I H H Me 491 I OOH LYNOOH .M WeH H M eH He -OH ?HHeOOH OOH '~NN OOH OH r H'H Me 6jHH M .*OH 'YN MeOOH Me *OOH OOH HH Me K M OMe Me *OOH OOH lH 6Me r H H Me Yjp-r OOH ?HMeOOH 492 NAN Y 6 OOH OOH I H H6MeHH M 0OC q-0-Me M M eY N Q O HM l O O H MeUH Me :H H 6Me H H 'Vi wx)o- Y-6 4 4 4 4**4 4 *44* *t4. 4 4 4 4 4.. 4* 4. 4. 4 4** 4 4. 0 4. 4 4. 4 *446 4 *4*4 4 4 4* 44 4. 4 .4.4.4 HOH 0) L *~li i H H.H H H003 %9. HOG HOO k HOO-1~ HOG OIAA HOH HLJL HOO Hb H o o a -C Y jS-) H o a 0 0 *1 A 4* 495
16. A compound according to claim 15, or a salt thereof, chosen from the group consisting of 02 H H CO 2 H H H 12H 2 H H C02H (>NYNAXN 02H SHO CI 496 N rrN?( N -O2 c G -S 0 C 2 H 0 2 H H N N-q" o 02 NXN- N0 0 2 H-O2 N 0 H HF *Me H* iii'02K H 0~~X OH 0 2 C 0 2 F 02H HH0C2 N-N 02 Z 9 _yt-Q- C 2H H A r) NY N? -CO 2H H H N IN 2 H 'H H 02H I Hr 02H I NH2 H 'r H H M le Tr OOH 8 Me M 0OH H OOH 0 0 NY H2 OMe eH H Me N- ry y OHOOH OMe F OH OH H H H SMe H Me IH H RA OH3J -r Y a-lO M e H H M ?'wVY pyOOOH OOH Me O H H Me Q CO, ?OHMe OH F NHm M OOH N~H y'Hvr OOH Me Me Q~ OO 'N OOH 9, 'H'Q OOH 0 -Q--OOH 499 H Me H H O e Me OOH OH H H Me e H H Me O O OMe IJ H e O *H H H OH OH H H Me leH Me H H Me H H 6 M e O O H MO H e M HOOH 0 H H H Me K)CH e H H M e H H I OOH OOH H Me TeH Me 500 OO OOH OOH H NOO H H M Q, JRX OOH 0 0* ~H~M )S COOH CY '-J/4C&COOH mM OOH 502 M M u OOH l OOH W 'yQONHW OOH M F 00H OOH F tJ~JJ~OOH OOH *H H :F F .*OO f"IN.NKJ 9O OOH M Me M Me Ar H e ILPe H HMe 503 F /-Jwf o- -OOH OOH F HOO HO F I~ J-QI AOOH ~jHHH FF *OOHB OOH FF .*H OOH Tr0N90l, ;&JtI 41:4 e OOH 4.00. Me H H ;)AWHj .*ego: F Meb~~ F F OOH0 OOH 1 1 Hr rH H F \COOH
17. A compound according to claim 2, or a salt thereof, wherein R 5 is lower alkoxy group. 504
18. 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 one of claims 1 to 17.
19. A method of treating a condition associated with VLA-4 mediated cell adhesion in a mammal, said method comprising administering to said mammal an effective amount of a compound according to any one of claims 1 to 19. A method according to claim 19, wherein the condition associated with VLA- 4 mediated cell adhesion is chosen from inflammatory responses, autoimmune responses and tumor metastasis.
21. A method according to claim 19, 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.
22. Use of a compound according to any one of claims 1 to 17 for therapy. 15 23. Use of a compound according to any one of claims 1 to 17, in the manufacture of a medicament for the treatment of a condition associated with VLA-4 mediated cell adhesion.
24. A pharmaceutical composition comprising as a therapeutic agent, a compound according to any one of claims 1 to 17 and a pharmaceutically acceptable carrier or 20 excipient. A pharmaceutical composition according to claim 24, further comprising one or more additional therapeutic agents.
26. A pharmaceutical composition according to claim 25, wherein said one or more additional therapeutic agents are chosen from the group consisting of S 25 antiinflammatory, antirheumatic, corticosteroid, immunosuppressive, antipsoriatic, bronchodilator, antiasthmatic and antidiabetic agents.
27. A pharmaceutical composition according to claim 26, wherein one of said one or more additional therapeutic agents is an antiinflammatory agent.
28. A pharmaceutical composition according to claim 27, wherein said antiinflammatory agent is chosen from a steroid and an NSAID.
29. A compound according to claim 1, substantially as hereinbefore described with reference to any one of the Examples. A process for preparing a compound according to claim 1, said process substantially as hereinbefore described with reference to any one of the Examples.
31. A compound according to claim 1, prepared by the process of claim [R:\LIBUU102848.doc:HJG 505
32. A pharmaceutical composition comprising a compound according to claim 29 or 31, together with a pharmaceutically acceptable carrier or excipient.
33. A pharmaceutical composition, substantially as hereinbefore described with reference to any one of the Examples.
34. Use of a compound according to claim 29 or 31, in the manufacture of a medicament for the treatment of a condition associated with VLA-4 mediated cell adhesion. A method of inhibiting cell adhesion in a mammal, said method comprising administering to said mammal an effective amount of a compound of claim 29 or 31, or a composition according to any one of claims 24 to 28, 32 or 33. Dated 21 January 2005 Daiichi Pharmaceutical Co., Ltd. Patent Attorneys for the Applicant/Nominated Person SPRUSON FERGUSON 0:00 S te *j [R:\LIBUU]02848.doc:HJG
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