ZA200208186B - Synthetic process for the manufacture of an ecteinaschidin compound. - Google Patents

Description

C.

SYNTHETIC PROCESS FOR THE MANUFACTURE OF AN ECTEINASCHIDIN COMPOUND

The present invention relates to synthetic processes, and in particular it relates to synthetic processes for producing ecteinascidin compounds.

BACKGROUND OF THE INVENTION :

European Patent 309,477 relates to ecteinascidins 729, 743, 745, . 759A, 759B and 770. The ecteinascidin compounds are disclosed to have antibacterial and other useful properties. Ecteinascidin 743 is ; now undergoing clinical trials as an antitumour agent.

Ecteinascidin 743 has a complex tris(tetrahydroisoquinolinephenol) structure of the following formula (I):

OCH;

HO CH;

OCOCH;

H

CH, :

N——CH;

NV

0 i 0 OH (0) o=< J

CH30 . .

NH

HO

It is currently prepared by isolation from extracts of the marine tunicate Ecteinascidin turbinata. The yield is low, and alternative preparative processes have been sought.

A synthetic process for producing ecteinascidin compounds is described in US Patent 5,721,362, see also WO 9812198 which is incorporated herein by reference in full. The claimed method is long and complicated, there being 38 Examples each describing one or more steps in the synthetic sequence to arrive at ecteinascidin 743.

In the known synthetic process, a 1,4 bridge is formed using a 1- labile, 10-hydroxy, 18-protected hydroxy, di-6,8-en-5-one fused ring compound. As shown in Example 33, a compound (13) is converted to compound (14):

OMe o MOMO. 7) Me AllocHN one

Me oH hd 1) DMSO, Ti,0 ox po Me 1 ® r N-|-me 2 DIPEA AO) s 4d ZL BOR mel AT ove ® 9g Neu N \ Ps ) Duo Nm, \g en

ANON NH c “ T & ) 5) Acp0, CH,Cly . 13

According to the known synthetic process, a spiroquinoline is then formed in the 1,4 bridge by the steps of Examples 34 to 36, and the 18-MOM protecting group is removed to give ecteinascidin 770 which can then be converted to ecteinascidin 743.

Claim 25 of US 5,721,362 is directed at an intermediate phenol compound of a given formula (11), which we refer to also as

Intermediate 11 or Int-11. It has the following bis(tetrahydroisoquinolinephenol) structure (II):

OCH,

MOMO CH,

OH

H

CH; H

N——CH;

Nn lo) ; \-o CN

OTBDPS where MOM is a methoxymethyl substituent and TBDPS is a tert- butyldiphenylsilyl substituent.

From Intermediate 11 it is possible to synthesise another interesting antitumour agent, phthalascidin, see Proc. Natl. Acad. Sci.

USA, 96, 3496-3501, 1999. Phthalascidin is a bis(tetrahydroisoquinolinephenol) derivative of formula (III):

OCH, )

HO CH,

OCOCH;, }

CH, H r N——CH;,

NH

0] :

Lo EN

N

HH

In ecteinascidins 743 and 770, the 1,4 bridge has the structure of formula (IV): 4

Ho \ ai

CH;0

NH

HO

Other known ecteinascidins include compounds with a different bridged cyclic ring system, such as occurs in ecteinascidin 722 and 736, where the bridge has the structure of formula (V): { \ = J

HN NH ecteinascidins 583 and 597, where the bridge has the structure of formula (VI): & \ 4 0 ~ 7

H “NH, and ecteinascidin 594 and 596, where the bridge has the structure of formula (VII):

A

= J 0

The complete structure for these and related compounds is given in J. Am. Chem. Soc. (1996) 118, 9017-9023. This article is incorporated by reference.

Other literature on the ecteinasdin compounds includes: Corey,

E.J., J. Am. Chem. Soc., 1996, 118 pp. 9202-9203; Rinehart, et al.,

Journal of Natural Products, 1990, “Bioactive Compounds from Aquatic and Terrestrial Sources”, vol. 53, pp. 771-792; Rinehart et al., Pure and

Appl. Chem., 1990, “Biologically active natural products”, vol 62, Pp. 1277-1280; Rinehart, et al., J. Org. Chem., 1990, “Ecteinascidins 729, 743, 745, 759A, 759B, and 770: potent Antitumour Agents from the

Caribbean Tunicate Ecteinascidia turninata”, vol. 55, pp. 4512-4515;

Wright et al., J. Org. Chem., 1990, “Antitumour Tetrahydroisoquinoline

Alkaloids from the Colonial ascidian Ecteinascidia turbinata”, vol. 55, pp. 4508-43512; Sakai et al., Proc. Natl. Acad. Sci. USA 1992, “Additional anitumor ecteinascidins from a Caribbean tunicate: Crystal structures and activities in vivo”, vol. 89, 11456-11460; Science 1994, “Chemical

Prospectors Scour the Seas for Promising Drugs”, vol. 266, pp.1324:

Koenig, K.E., “Asymmetric Synthesis”, ed. Morrison, Academic Press,

Inc., Orlando, FL, vol. 5, 1985, p. 71; Barton, et al., J. Chem Soc. Perkin

Trans., 1, 1982, “Synthesis and Properties of a Series of Sterically .

Hindered Guanidine bases”, pp. 2085; Fukuyama et al., J. Am. Chem.

Soc., 1982, “Stereocontrolled Total Synthesis of (+)-Saframycin B”, vol. } 104, pp. 4957; Fukuyama et al., J. Am. Chem. Soc., 1990, “Total

Synthesis of (+) — Saframycin A”, vol. 112, p. 3712; Saito, et al., J. Org.

Chem., 1989, “Synthesis of Saframycins. Preparation of a Key tricyclic

Lactam Intermediate to Saframycin A”, vol. 54, 5391; Still, et al., J Org.

Chem., 1978, “Rapid Chromatographic Technique for Preparative

Separations with Moderate Resolution”, vol. 43, p. 2923; Kofron, W.G.;

Baclawski, L.M., J. Org. Chem., 1976, vol. 41, 1879; Guan et al., J.

Biomolec. Struc. & Dynam., vol. 10, pp. 793-817 (1993); Shamma et al., “Carbon-13 NMR Shift Assignments of Amines and Alkaloids”, p. 206 (1979); Lown et al., Biochemistry, 21, 419-428 (1982); Zmijewski et al.,

Chem. Biol. Interactions, 52, 361-375 (1985); Ito, CRC Crit. Rev. Anal.

Chem., 17, 65-143 (1986); Rinehart et al., “Topics in Pharmaceutical

Sciences 1989”, pp. 613-626, D. D. Breimer, D. J. A. Cromwelin, K. K.

Midha, Eds., Amsterdam Medical Press B. V., Noordwijk, The

Netherlands (1989); Rinehart et al., “Biological Mass Spectrometry”, 233-258 eds. Burlingame et al., Elsevier Amsterdam (1990); Guan et al.,

Jour. Biomolec. Struct. & Dynam., vol. 10 pp. 793-817 (1993); Nakagawa et al., J. Am. Chem. Soc., 111: 2721-2722 (1989);; Lichter et al., “Food and Drugs from the Sea Proceedings” (1972), Marine Technology

Society, Washington, D.C. 1973, 117-127; Sakai et al., J. Am. Chem.

Soc., 1996, 118, 9017; Garcia-Rocha et al., Brit. J. Cancer, 1996, 73: 875-883; and Pommier et al., Biochemistry, 1996, 35: 13303-13309.

Further compounds are known which lack a bridged cyclic ring system. They include the bis(tetrahydroisoquinolinequinone) antitumor-antimicrobial antibiotics safracins and saframycins, and the marine natural products renieramicins and xestomycin isolated from cultured microbes or sponges. They all have a common dimeric tetrahydroisoquinoline carbon framework. These compounds can be . classified into four types, types I to IV, with respect to the oxidation & ; pattern of the aromatic rings.

Type 1, dimeric isoquinolinequinones, is a system of formula (VIII) : most commonly occurring in this class of compounds, see the following table I.

Table [ :

Structure of Type I Saframycin Antibiotics.

OCH, o CH, o ' H

CH, Po 0]

N—CH,

Pa

CH;0 > 7” 5, Rub

H : H [e) R21

HN

PN

R258" ‘Rasp

Compound Rida Ri14b R21 R25a R2sb R25¢ saframycin A H H CN (0) (0) CHs saframycin B H H H 0 0] CHj3 saframycin C H OCHa3 H 0) 0 CH3 - saframycin G H OH CN 0) 0] CHj3 saframycin H H H CN OH CH2COCH CHa 3 saframycin S H H OH 0) Oo CH3 saframycin Y3 H H CN NH: H CH3 saframycin Yd; H H CN NH: H C2Hs saframycin Ad; H H CN 0) 0 C2Hs saframycin Yd2 H H CN NH2 H H saframycin Yao H Q?b CN NH2 H CHj3 saframycin Yab-q H Qb CN NH. H C2Hs . saframycin AH> H H CN Ha OHa CH3 saframycin AH2Ac H H CN H OAc CH3 : saframycin AH; H H CN OHa He CH3 saframycin AHi1Ac H H CN OAc H CHj saframycin AR3 H H H H OH CH3 @ assignments are interchangeable. b where the group Q is of formula (IX):

OCH, 0) CH,

O H

CH PB 0 : ” “N——CH,

NJ

: CHO H : 'H

Oo CN

HN }

NH— be

CHs

Type I aromatic rings are seen in saframycins A, B and C; G and

H; and 8 isolated from Streptomyces lavendulae as minor components.

~

A cyano derivative of saframycin A, called cyanoquinonamine, is known from Japanese Kokai JP-A2 59/225189 and 60/084288. Saframycins

Ya, Ydi, Adi, and Yd2 were produced by S. lavendulae by directed biosynthesis, with appropriate supplementation of the culture medium.

Saframycins Y2» and Y2b-a dimers formed by linking the nitrogen on the

C-25 of one unit to the C-14 of the other, have also been produced in supplemented culture media of S. lavendulae. Saframycins AR, (=AH3,), a microbial reduction product of saframycin A at C-25 produced by Rhodococcus amidophilus, is also prepared by nonstereoselective chemical reduction of saframycin A by sodium borohydride as a 1:1 mixture of epimers followed by chromatographic separation [the other isomer AH, is less polar]. The further reduction product saframycin ARs, 21-decyano-25-dihydro-saframycin A, (= 25- 8 dihydrosaframycin B) was produced by the same microbial conversion.

Another type of microbial conversion of saframycin A using a Nocardia o species produced saframycin B and further reduction by a

Mycobacterium species produced saframycin AH!Ac. The 25-O-acetates of saframycin AH2 and AH: have also been prepared chemically for . biological studies.

Type I compounds of formula (X) have also been isolated from marines sponges, see Table II.

Table II

Structures of Type I Compounds from Marine Sponges.

OCH, 0) CH; 0) H nA Re

CH;0 i" 7 Re 0 R21

Oo or

TTT suwbstmens ‘Risa Rw Ra RT renieramycin B OC2Hs H H -C(CH3)=CH-CH3; renieramycin C OH 0) 0) -C(CH3)=CH-CH3 renieramycin D OC2Hs Oo (0) -C(CH3)=CH-CH3 renieramycin E H H OH -C(CH3)=CH-CH3 renieramycin F OCH3 H OH -C(CH3)=CH-CHs3; xestomycin OCHs3; H H -CH3 _—_—mm

Renieramycins A-D were isolated from the antimicrobial extract of a sponge, a Reniera species collected in Mexico, along with the biogenetically related monomeric isoquinolines renierone and related compounds. The structure of renieramycin A was initially assigned ‘ with inverted stereochemistry at C-3, C-11, and C-13. However, careful examination of the !H NMR data for new, related compounds - renieramycins E and F, isolated from the same sponge collected in

Palau, revealed that the ring junction of renieramycins was identical to that of saframycins. This result led to the conclusion that the formerly assigned stereochemistry of renieramycins A to D must be the same as that of saframycins.

Xestomycin was found in a sponge, a Xestospongia species collected from Sri Lancan waters.

Type II compounds of formula (XI) with a reduced hydroquinone ring include saframycins D and F, isolated from S, lavendulae, and saframycins Mx-1 and Mx-2, isolated from Myxococcus xanthus. See table III.

Table III

Type II Compounds

OCH,

HO CH, lo) ” H

CHa : ~ OH

N——CH; a i eee = R14b

H : H : lo) R21

HN oP

R258" Yq25n : Substituents

Compound Rl4a R14b R21 R25a R25b R25¢ saframycin D 0] 0) 'H 0 Oo ~~ CHs saframycin F O 0) CN 0 ©) CHs saframycin Mx-1 H OCH3 OH H CHs NH: a saframycin Mx-2 H OCHs H H CHs NH2 ? "The type III skeleton is found in the antibiotics safracins A and B, ~ isolated from cultured Pseudomonas fluorescens. These antibiotics of formula (XII) consist of a tetrahydroisoquinoline-quinone subunit and a tetrahydroisoquninolinephenol subunit. :

OCH;

HO CH; 0 H

H=

CH, HP

Y N——CHa,

CHsO Ng 3 na BH J o R21 i;

HN

PL

: NH, where R2! is -H in safracin A and is -OH in safracin B.

Saframycin R, the only compound classified as the Type IV skeleton, was also isolated from S. lavendulae. This compound of

SUBSTITUTE SHEET (RULE 26)

formula (XIII), consisting of a hydroquinone ring with a glycolic ester sidechain on one of the phenolic oxygens, is conceivably a pro-drug of saframycin A because of its moderate toxicity. 0 OCH, o) CH,

HO

CH, 5% 0

N—CH;

CH;0 ~ DH : OH CN

HN

CH

AY 3 oO

All these known compounds have a fused system of five rings (A) . to (E} as shown in the following structure of formula (XIV): 17 } 18 16 4 11 > O 15 5 10 3 XN D 20

B Cc 1a 7 (4) N 13 8 i 2

The rings A and E are phenolic in the ecteinascidins and some other compounds, while in other compounds, notably the saframycins, the rings A and E are quinolic. In the known compounds, the rings B and D are tetrahydro, while ring Cis perhydro.

OBJECT OF THE INVENTION

The need remains for alternative synthetic routes to the ecteinascidin compounds and related compounds. Such synthetic routes may provide more economic paths to the known antitumour agents, as well as permitting preparation of new active compounds.

SUMMARY OF THE INVENTION

This invention relates to synthetic processes for the formation of intermediates, derivatives and related structures of ecteinascidin or other tetrahydroisoquinolinephenol compounds.

In one aspect, the present invention provides a process for preparing an ecteinascidin product with a spiroamine-1,4-bridge. The process involving forming a 1,4 bridge using a 1-labile, 10-hydroxy, 18- protected hydroxy, di-6,8-en-5-one fused ring compound, where the ¥ fused ring is the formula (XIV). In the present invention, the C-18 ) protection is removed before spiroamine introduction.

Suitable starting materials for the new synthetic processes include compounds related to the natural bis(tetrahydroisoquinoline) alkaloids. Such starting materials may be prepared either from the different classes of saframycin and safracin antibiotics available from different culture broths as detailed in WO 0069862 or by other synthetic or biochemical processes. In this respect, WO 0069862 is incorporated herein in full by reference. The present PCT application claims priority from application PCT/GB 00/01852 which was published as WO 0069862. We incorporate that text by reference to the extent that there is disclosure therein which is not in the present specification.

PREFERRED EMBODIMENTS OF THE INVENTION

In one particular aspect, the present invention is directed at the use of the compound Intermediate 21 in a number of new synthetic processes for the preparation of ecteinascidin 743 and related compounds, xn OMe

HO Me

Me @® N=|-Me

N

=

[0] NH 21 }

The Intermediate 21 has a 5-allyloxy group, where the allyl group serves to protect the 5-hydroxy group. It will be understood that other protecting groups can easily be employed, and that the present invention extends generally to the use of other such S-protected hydroxy compounds.

FORMATION OF ECTEINASCIDIN 743 AND RELATED COMPOUNDS

In general, the conversion of Intermediate 21, or a related compound, to an ecteinascidin product involves the following key transformations: (a) Conversion of the NH2 to OH by reaction, for example with sodium nitrite in acetic acid. (b) E-ring phenol protection. (c) Esterification by protecting the primary 1-hydroxy function with a protected cysteine sidechain. (d) Deprotection of allyl group and oxidation.

(e) Creation of the bridged ring by cyclization reaction. (f) Deprotections of E-ring phenol and the cysteine moiety (8) Quinoline Introduction by Trans-amination and Petter Spengler reactions.

The high functionality of the intermediate compounds necessitates the use of protecting groups for the E-ring phenol and for the cysteine sidechain in order to prevent unwanted side reactions.

As such, a number of alternative intermediates can be generated : dependent on the particular selection of protecting groups. . Different possible sequences are possible for combining these transformations dependent primarily on the protecting groups selected for the phenol ring and for the amine of the cysteine sidechain.

The total number of synthetic transformations is also a function of the protecting groups selected.

By way of illustration, the use of different combinations of protecting groups is described below for six typical routes for the preparation of

ET-743 from Intermediate 21, also referred to herein as SF21.

Route Phenol Protection Cysteine Protection Number of steps 1 MOM Boc 12 2 MEM Boc 10

3 MEM Cbz 11 4 MOM Alloc 13

S MEM Alloc 13 6 MOM Cbz 15

As the skilled artisan will readily appreciate, the reaction schemes described herein may be modified and/or combined in various ways, and the alternative sequences of steps and the compounds generated therefrom are part of this invention.

Additionally, the use of other protecting group strategies not : detailed is also part of this invention.

PROCESS DETAILS OF Six TYPICAL SYNTHETIC ROUTES

Full reaction schemes for each route are in the following Schemes 1 to 6.

Scheme 1 - ET-743: Hemisynthetic Alternative Route 1 o 2 2 2 ) 2 3 = 1 ~ as; THA : i 9 iQ 2d a %o EVP $

Fa o Q ~~ 2 bi = [1 0 2 3 74 2 [= = 5 . Q, & 5 ul $ £82 £ . al z 955 |%% 3 #3 258 Id a : TESA § @ + 2 o [eo] ES) ° E £ o

W, Z2 —a/ 2A EN 4-4 AN : CI \ A & os I Og 2 = ° =-0 = Zz ! a oO © [»] 3 © Ws: 2 a 8 3 < I Bi

O qn - o [o] = ~~ ( bi 2d = > = Tr 8 74 3 [eo] 5 o £ z g o_o 8 . tls d oP 1 : 3 = do |£ : o I 2 [7] : gg ° = 3 oN [] . SWS £10 o | SN: lo] ° /—° BE PY } =-0 Zz 3 ~ 2 . C1) d Z “ZF o = >, 5 3 = "8 2 oy —o / 3 ° 2d’ Z 3 02 v © 3 ? . [eo] $ o’ = [+] [ x oN } 3 ad 3 2 @ 2 © L ou z © 2 VAR o |Z x ° 3 [4] o +=2.5 0% ine g 1 5 © 28 ¢ ig d|£ <) S a&

Fe 3% 2 = © o 2 2 = ats SYENIDT i We 74 o =z @ Z > $ © ils = Ss Q GQ o 2-0 Z © q ha jutl z 3

I] 2.5 . Ll a o yo peas Es

Cot ares d © g 8 - = zZ WN o [4] 2 -

Yd (2 Bi g | JS 3 2 © 8 %%

I

SUBSTITUTE SHEET (RULE 26)

Scheme 2 ~ ET-743: Hemisynthetic Alternative Route 2 © [+]

PS

Zz . 9 ¢ () : so N ONL : z$ a 2 Ea 0

So RT al

I [1] [o} g 2] Zz = g MAY 8 5g hat >. 9 2 z 4 2 [} =

Zz 32 2 518 2 $ $ = ot 1 Le

Sim 1 & = la = wu 4 o o ns

Ys 8 i, a ° ©, Zz hy 0 ry 2 -} Vo § = = = é 9 0 zz [+] §) 7 wg < J 2 Q *

AN =z 2 5 g © 5

I] i] z =

T Zz © So re) 9 [+] oN 5 = = a Zz = 2) ' a ° 553 z= pe . 2 2 - ib a® sii] 2 5) = cael § @ ZL [o] Ll] g 3 I: 1%, Qu =a 5| »8 2 2 og © 2 | oot ODT Pa 2 [o] z= z oc [o] [o} = : = : oJ 8B v2 ° = g Z O S -— £ = T

HY : :

Zz z oO CY To] — fo] 941), =

I = °c @ o” 3 $ = 28a 21 ed @ S 93 2, 4 oO [Ve = = = . g|° Se °\ ¢% ¢ = [y [eo] ne [TH =z [} - £2 z /S\F (S ON é|3 ve C8 = 2 < g " z [3 re Ng © iu Zz © 3 oz ?

H p = Ye) dg 5 2 = ° “Jt 5 ) 8 § X 2% {or 5 - on o =e “ ™ x | 5d Y=-2 or 0 {Y o &| 2 SS 3d od 5 ole ae 4 [+) @ 5 - ~~ £ c - Ne

SUBSTITUTE SHEET (RULE 26)

Scheme 3 — ET-743: Hemisynthetic Alternative Route 3 [ov] o 5

IN ui © ]

Z 4 [o] oui 2 oe Za 2 33 3 8 °% =X J p-4 IT [] O g [o] “iQ => = = © ri Z 0 b 3 £ = (o) - > S : a SRE “\ z [o} [23 i of [o] ol 7 3 E % z Za )2 ; 2 2 © o

AE : CE )=C3 518 2 2 8 o g © gs © J 2 gs z3\ N = [=] 5 § Ts z z se 2 Fal 5 i 2 @ No ES o 2 . 1] = | Oo 3 [=] Oo Oo = . [eo] | wv N [4 J I [4 172]

Az (2 3 HA = 9 “ng 4 - [-]

Z © & oo 2 a 0 2 2 = 2 2 Zz it 87 5 © ] a gs % a Ssi gfs| 3 § g -& an < / gg © S85] & & LN 8 - OA o 1%, Qo R542 a J = ® 2 [=} 2 2 = ES = o

B| ot © \ J ! » 8 2 ul Q FN =z 2 5

E 0 ni; Z © he 2 H z - ® o n Ww 2 oS: Z 5 ) 5 2-2 ES +] i Cl = a9

Q

=z 2 ° (CD ° ~ o> °] “oid J - Nm

Zz 5 dg © © = o hot = a o 2 = as HE: HR g © £15 05 ® 9 8 g 2 © =z £0 3 £3 2 =F FN 3 HAN. C8 ° ENF t g Q wl z ] oO . iE 2 0 = 2 d= 2 = °o 8

EO z ”

Ww, x = > >, o z 2 "Oe o 0 3) 4 z Zz = T Q 3 g1= hes i|2 : 7 J Fl o 4 2 Oo g o.

SUBSTITUTE SHEET (RULE 26)

Scheme 4- ET-743: Hemisynthetic Alternative Route 4 2 g @ — = 55 [o] \ y, i N $ $ JE « "NE. Sz g z i, 2" 8 22

E $ ° x é = 8 do Neo 2) 2 [ 0” 2 © 3 ? © 7 = © 0 z z - = g "QO °o § 2 $3 ENS 3 A = 87 5 \ © i 2 Zz 2 thy 5 Tz = o 2 (*] © \ 0% zg FS ca $2 & < © o- d og: EE EE © 9g = . 3 Z © =z 5

Rcd Jk ~ - ota, 0 jh 74 0 [+] 2 2 Ho br 2

ES 5 §) \ " ; “N\ =z x 8 2 g Fr) g [od NO Zz © 2 21/538 3 LN =o 3 : o = z © 9 J 2 - = “og . £ os: " o< 9 { aE "

Z 85 2 © °=y—o fo] a fo x Z 1) g o 4 bi 8 5 £ °

P & id FH o =

PB ° TI/ Ne EJ 7 { o : ] 5-{ = 28 ; © 2 —- a

Eig 4 0% Te FF ol = - ° o gg g Xa Wai © g e 2 - Zz = = 5 e 0 rE) o 2 IT z CO) 7 o {) o —On/ 4 8 2 z 2 0 yo el p "Oz 2 J 3 9

EK] oO a o ed a 3 I|3 74 ° [=) 2 o 3, z w g © A) I = 2 2 0

Fl3 0 8 2 7

Q > = Z Bb o 23 ] Zz 2 £ = a 2 0 i o 2 2 I © (o] TO « as Pa yo ° 2 Be 5 = . Zz 02 bi 2s £ °

Q ™ ea @, bi £ 5 81s / 4 Yo FR

SUBSTITUTE SHEET (RULE 26)

Scheme 5- ET-743: Hemisynthetic Alternative Route 5 o £ 3 2 3 = Z © © i &) = i z 52 5 1 S 2 g = A\ z 2 |Z 3] Zz Oo § irs] = < 2 = 5 0 Z - 2 5

T - Ny ®

Yo ir Wek:

Pe o 0% (e] 1 s J z

N g oO Q “0g

T|o I 2 ~ 8% ° § 5 2a = © a £ oN 3 Z 2 lo] w = 5 o 2 © 9 o 2 2g a 2R3 Zz o °g J 5 I =L4 5 a & o [=] 1 29 EE = go o =

F\P8 cam| § @ 2 8 }

Q - . T fr] Z 0 © o 2 £ 2 <) ii po 0) © 2 CI) g

Vd % 2 = 0 0 4 do 0 2 = CHEE = © WV, ! P ° — = ~ 2 . 81x Io! 5 z - 2 2 ils & Zz 0 =o ° : = = T OQ 2 = 2 © fol iG 0 2 2 o (CY 0 { xz z = : Bi Sa 8 : © CH 22 = oO yo g X OB A) cg o

ES Zz 5 2 CI p, J 2 5 £ - [7] . ° &, o 5 » 2 Me S

V4 do a &: 5 3x2 2 3 = © 3 =o oO of k @ X 2 g Q = 8 & 28 Sh Cd : ja] ° ® . mE AN =z = = - - [o) ~Qp [+] i o z Z [o] a 4 > © = $e = T - 2) “i = ' fo] : * Zz © Ws o CY (o} oF * 0

A =z J 2 yO ” g "By go = Iq ° 2° = °& J $ E o @ [=] 4 Fld g ad 1 ; 7 J = | £ 2 2 [o) 2 I. 4, pe ’

Q = r-) © = i a (<] o ° o - = [a] om [] = 28 g % 5 5 < 2 4 2 > 2 oO - [eo] wg) 2 £ ° (LY 0 Zo) . © 5: > od o =o 3 = ii] o Oe 8 z Q o

T Zz = bf o < < J

HN Q [o]

Waly: £5 s| 8 2s zg

SUBSTITUTE SHEET (RULE 26)

Scheme 6 — ET-743 Hemisynthetic Alternative Route 6 2 a @ = 2 2 2 ] o| = [1 e =

Zz = 219 3 3 "oO o 13 8 Eo 3 R o 3 2 -y = =

Dre © i °o ‘ 2 © < J [eo] wd 4 2 © z To z =

Ne F iw 2 2 & a

Pa I o x = © °< 3 , a i o ES % © [o] iii 33 gg f°

Ne & I) - z i wv, F 2 CT) 512% 2 $ @ = [«] Q ) i » & O = 2 4 2 I oN z ] fof ne) z BK 1 2 z 0 z . -—- to) g io o — 0 a 2 ° CY Q oo

Q - o 2 (o] . = < o J o g 2 fo) e 5-{ =| 2

J = Fe) a of 2 u = ae © 2 2 a 2%

Zz z 4 J o Gp 2 = 3 =z =] e "iQ

Zz 5 ® — Z a o - aS o o

C3 Dra o [+] . © < J = ] [o x = 21¢ =| 2 : = 2

Wan th Ne $ See Ht: [3

[1] = 2 = 4 1 ” 8 x 8 =

A\ =z I 8 a 3 © g "wo Z z F ]

Zz ©0 o g - ° 3 Te ° < on = (Y bi = g © 5 2/2 [ond

SUBSTITUTE SHEET (RULE 26)

In route 1, protection of the E-ring phenol is achieved in three steps involving protection /deprotection of the amine of SF21 with Troc.

For routes 1 and 2, protection of the cysteine sidechain with Boc allows the phenol and cysteine groups to be deprotected in a single step rather than as two separate steps. For the rest of the routes, an additional deprotection step is required.

For route 2, Intermediate 25 is avoided through the use of the direct esterification methodology and the subsequent protection of the phenol with the MEM group.

In routes 2 and 3 protection of the E-ring phenol is delayed until after the diazotisation and esterification steps thereby allowing the phenol to be protected in a single step rather than by the three step : sequence of route 1. :

For routes 1, 2 and 3, direct esterification of the primary alcohol with the cysteine derivative eliminates the unproductive protection/deprotection steps of the primary alcohol with a silyl group (routes 4 and 5) thereby shortening the sequence by two steps.

Route 6 only contemplates herein the last steps from intermediate 161, which can be easily obtained from intermediate 21.

In routes 4 and 5 the primary alcohol produced by the initial diazotisation step is protected with silicon to allow selective protection of the E-ring phenol and avoiding intermediate 25. Following

Claims (29)

Claims

1. A process for preparing an ecteinascidin product with a spiroamine-1,4-bridge, the process involving forming a 1,4 bridge using a 1-labile, 10-hydroxy, 18-protected hydroxy, di-6,8-en-5-one fused ring compound, wherein C-18 protection is removed before spiroamine introduction.

2. A process according to claim 1, wherein the ecteinascidin product has a 21-hydroxy group, the process including converting a 21-cyano group to the 21-hydroxy group.

; 3. A process according to claim 1 or 2, wherein the spiroamine is a spiroquinoline.

4. A process according to any preceding claim, wherein the 18- protected group of the 1-labile, 10-hydroxy, 18-protected hydroxy, di- 6,8-en-5-one fused ring compound is protected with: MOM, methoxymethyl; or MEM, methoxyethoxymethyl group.

5S. A process according to any preceding claim, wherein the 1-labile group is an N-protected cysteinyloxymethylene group of the formula -CH2-O-CO-CNHProt!-CH>-S-H.

6. A process according to claim 5, where Prot! is: Bog, t- butyloxycarbonyl; Troc, 2,2,2-trichloroethyloxycarbonyl; Cbz, benzyloxycarbonyl; or Alloc, allyloxycarbonyl.

7. A process according to claim 5 or 6, wherein Prot! is removed in the same step as C-18 protection.

8. A process according to claim 5, 6 or 7, wherein the 1-labile group is generated from a 1-substituent of the formula: -CH2-O-CO-CNHProt1-CH,-S-Prot2.

9. A process according to claim 8, wherein Prot? is Fm, O- fluorenylmethyl. ’

10. A process according to claim 8 or 9, wherein the 1-substituent of the formula: -CH2-O-CO-CNHProt!-CH»-S-Prot2. is formed by esterification of a -CH2-O-H substituent.

11. A process according to claim 10, wherein the esterification is carried out before formation of the 10-hydroxy, di-6,8-en-5-one structure.

12. A process according to claim 10, wherein the esterification is carried out after introduction of the 10-hydroxy, di-6,8-en-5-one structure.

13. A process according to any preceding claim, which starts from a l-aminomethylene, 5-protected hydroxy, 7,8-dioxymethylene, 18- hydroxy, 21-cyano fused ring compound

14. A process according to claim 13, where the l-aminomethylene group is temporarily protected to allow protection at the 18-hydroxy group, and the temporary protection is removed.

15. A process according to claim 13, wherein the C-18 hydroxy group . is protected after formation of a 1-ester function.

16. A process according to claim 13, wherein the l-aminomethylene group is converted to a 1-hydroxymethylene group and the 1- hydroxymethylne group is temporarily protected, to allow protection at the 18-hydroxy group, and the temporary protection is removed.

17. A process according to claim 1, wherein the 1-labile, 10-hydroxy, 18-protected hydroxy, di-6,8-en-5-one fused ring compound is prepared by steps starting from a 21-Nuc compound with a structure of formula (XIV): 17 18 16 4 11 0 15 6 10 3 5 D 20 B C 14 7 (4) N 13 8 ] 71 where at least one ring A or E is quinolic, and where Nuc Indicates the residue of a nucleophilic agent.

18. A process according to claim 17, wherein the compound of formula (XIV) is cyanosafracin B.

19. A process according to any preceding claim, wherein the product is of formula (XXIIb): R'® CH, R® RR HC gy R' El SF : N R12 R42 14b (o) i R . AN Rr! g21 where: Rl and R* together form a group of formula (IV), (V), (VI) or (VII): . . 4 1 4 4 / ( . 0] 1 1 0 S$ ®) 7 ( ( CH,0 Co! HNN o % o\ H 7 HO , , Ne or © ; RS is -OH or a protected or derivatised version of such a group; R142 and R14" are both -H or one is -H and the other is -OH or a protected or derivatised version of such a group, -OCH3 or -OCH,CHs, or R142 and R14b together form a keto group; R12 js -NCHza-; RIS is -OH or a protected or derivatised version of such a group; and R!8 is -OH or a protected or derivatised version of such a group.

20. A process according to claim 19, wherein R5 is alkanoyloxy of 1 to S carbon atoms.

21. A process according to claim 20, wherein RS is acetyloxy.

22. A process according to claim 19, 20 or 21, wherein R42 and R14b are hydrogen. -

23. A process according to any of claims 19 to 22, wherein R!5 is hydrogen.

24. A process according to any of claims 19 to 23, wherein R2! is -OH or -CN.

25. A process according to claim 1 1, wherein R7 and R® together form a group -O-CH2-O-.

26. A process according to any of claims 19 to 25, wherein R! and R* together form a group of formula (IV):

( 0) oJ CH;0 2 . NH HO

27. A process according to any preceding claim, wherein the ecteinascidin product is ecteinascidin 743.

28. A process step in the manufacture of an ecteinascidin comopund, the step comprising removing both protecting groups in a single step, in accordance with the following scheme: . NHProthH NH Proto OMe 2 OMe O \ Me O=¢" "| HO Me AcO 5 S AcO S me N—J-Me Me YT N-]-Me N 0] i (0) N z 0 CN 0 EN where ProtNH is amino protecting group, and ProtOH is a hydroxy protecting group.

29. A process according to any of claims 1 to 27, which includes the process step according to claim 28.