AP737A

AP737A - New cryptophycins from synthesis.

Info

Publication number: AP737A
Application number: APAP/P/1997/001069A
Authority: AP
Inventors: Richard E Moore; Marcus A Tius; Russell A Barrow; Jian Liang; Thomas H Corbett; Frederick A Valeriote; Thomas K Hemscheidt; Trimurtulu Golakoti
Original assignee: Univ Hawaii; Univ Wayne State
Priority date: 1995-03-07
Filing date: 1996-03-07
Publication date: 1999-03-15
Also published as: BG101875A; HUP9801880A2; HUP9801880A3; NO974105L; MD2196B2; ATE279935T1; PL185949B1; NO326685B1; JPH11510476A; SK119997A3; MD970313A; ES2230561T3; FI973591A0; NZ305571A; GEP20002206B; CN1183726A; AU723652B2; CZ278197A3; WO1996040184A1; HK1009747A1

Abstract

The present invention provides novel cryptophycin compounds having the following structure: The present invention further provides total synthetic methods for producing cryptophycins. The present invention also provides for the use of cryptophycins in pharmaceuticals, to inhibit the proliferation of mammalian cells and to treat neoplasia.

Description

NEW CRYPTOPHYCINS FROM SYNTHESIS

This invention was made in pan with U.S. Government support under Grant Nos. CA12623 and CA53001 from The National Cancer Institute, Department of Health and Human Services. Accordingly, the U.S. Government may have certain rights in this invention.

Background of the Invention

Neoplastic diseases, characterized by the proliferation of cells not subject to the normal control of cell growth, are a major cause of death in humans. Clinical experience in chemotherapy has demonstrated that new and more effective drugs are desirable to treat these diseases. Such experience has also demonstrated that drugs which disrupt the microtubule system of the cytoskeleton can be effective in inhibiting the proliferation of neoplastic cells.

The microtubule system of eucaryotic cells is a major component of the cytoskeleton and is in a dynamic state of assembly and disassembly; that is, heterodimers of tubulin are polymerized to form microtubules, and microtubules are depolymerized to their constituent components. Microtubules play a key role in the regulation of cell architecture, metabolism, and division. The dynamic state of microtubules is critical to their normal function. With respect to cell division, tubulin is polymerized into microtubules that form the mitotic spindle. The microtubules are then depolymerized when the mitotic spindle’s use has been fulfilled. Accordingly, agents which disrupt the polymerization or depolymerization of microtubules, and thereby inhibit mitosis, comprise some of the most effective chemotherapeutic agents in clinical use.

Such anti-mitotic agents or poisons may be classified into three groups on the basis of their molecular mechanism of action. The first group consists of agents, including colchicine and colcemid, which inhibit the formation of microtubules by sequestering tubulin. The second group consist of agents, including vinblastine and vincristine, which induce the formation of paracrystalline aggregates of tubulin. Vinblastine and vincristine are well known anticancer drugs: Their action of disrupting mitotic spindle microtubules preferentially inhibits hyperproliferative cells. The third group consists of agents, including taxol, which promotes the polymerization of tubulin and thus stabilizes microtubule structures.

However, merely having activity as an antimitotic agent does not guarantee efficacy against a tumor cell, and certainly not a tumor cell which exhibits a drug-resistant phenotype. Vinca alkaloids such as vinblastine and vincristine are effective against neoplastic cells and tumors, yet they lack activity against some drug-resistant tumors and cells. One basis for a neoplastic cell displaying drug resistance (DR) or multiple-drug resistance (MDR) is through the over-expression of P-glycoprotein. Compounds which are poor substrates for transport of P-glycoprotein should be useful in circumventing such DR or MDR phenotypes.

Accordingly, the exhibition of the DR or MDR phenotype by many tumor ceils and the clinically proven mode of action of anti-microtubule agents against neoplastic cells necessitates the development of anti-microtubule agents cytotoxic to non-drug resistant neoplastic cells as well as cytotoxic to neoplastic cells with a drug resistant phenotype. Agents which have shown promise in this regard include a class of compounds known as cryptophycins.

With respect to methods of producing cryptophycins, no method for total synthesis of cryptophycins is currently known. Cryptophycin compounds are presently produced via isolation from blue-green alga or are semi-synthetic variations of such naturally produced compounds. The lack of a total synthetic method necessarily makes it difficult to produce stereospecific cryptophycins which can maximize activity and increase the stability of the compound. For example, research has shown that cryptophycins with an intact macrocyclic ring are more active. Accordingly, a total synthetic method which could produce cryptophycins with a macrocyclic ring that is more stable than naturally derived cryptophycins would be desirable. The present invention solves these problems.

Disclosure of the Invention

The present invention provides novel cryptophycin compounds having the following structure:

wherein

At is methyl or phenyl or any simple unsubstituted or substituted aromatic or heteroaromatic group;

Rj is a halogen, SH, amino, monoalkylamino, dialkylamino, trialkylammonium, alkylthio, dialkylsulfonium, sulfate, or phosphate; R2 is OH or SH; or R, and R2 may be taken together to form an epoxide ring, an aziridine ring, an episulfide ring, a sulfate ring or a monoalkylphosphate ring; or

Ri and R2 may be taken together to form a double bond between Clg and C,9; R3 is a lower alkyl group; R< and Rj are H; or R, and Rj may be taken together to form a double bond between C,3 and C,4; R* is a benzyl, hydroxy benzyl, alkoxybenzyl, halohydroxybenzyl, dihalohydroxybenzyl, haloalkoxybenzyl, or dihaloalkoxybenzyl group; R7f Rg, R, and R10 are each independently H or a lower alkyl group; and X and Y are each independently Ο, NH or alkylamino.

The present invention further provides total synthetic methods for producing cryptophycins. The present invention also provides for the use of cryptophycins in pharmaceuticals, to inhibit the proliferation of mammalian cells and to treat neoplasia.

Brief Description of the Drawings

Figure 1 provides a general structure of selected cryptophycin compounds of the present invention and a numbering system for the hydroxy acid units A and D and amino acid units B and C in selected embodiments;

Figures 2A and B graphically depict the effects of cryptophycin compounds and vinblastine on Jurkat cell proliferation and cell cycle progression. Jurkat cells were incubated with the indicated concentrations of cryptophycin compounds (A) or vinblastine (B) for 24 hours. For each sample, the number of viable cells (H) and the mitotic index (□) were determined as described in the Experimental Section. Values represent the means ± standard deviation (sd) for triplicate samples in one of three similar experiments;

Figure 3 graphically depicts the reversibility of the effects of vinblastine, cryptophycins and taxol on cell growth. SKOV3 cells were treated with O.lnM vinblastine (□), O.lnM cryptophycins (B) or InM taxol (S) at time = 0. These concentrations inhibited cell growth by 50% for each compound. After 24 hours the cells were washed and incubated in drug-free medium for the time indicated. The cell density was determined by sulforhodamine B (SRB) staining as described in the Experimental Section, and is expressed as the mean ± sd absorbance at 560 nm for triplicate samples in one of three experiments;

Figure 4 provides Isobolograms for combinational effects of vinblastine and cryptophycins on cell proliferation. SKOV3 cells were treated with vinblastine (0-600pM) and/or cryptophycins (l-100pM) for 48 hours. Cell numbers were then determined by SRB staining as described in the Experimental Section, and the ICjqS (B) and the line of additivity (—) were determined for combinations of vinblastine and cryptophycin compounds. Values represent means for two experiments each containing triplicate samples;

Figure 5 provides a fust scheme for synthesizing cryptophycins in accordance with the present invention;

Figure 6 provides a scheme for producing a hydroxy acid unit A;

Figure 7 provides a scheme for producing the subunit of a cryptophycin comprising a hydroxy acid unit A and amino acid B;

Figure 8 provides a scheme for producing the subunit of a cryptophycin comprising an amino acid' unit C and hydroxy acid D;

Figure 9 provides a first scheme for syn±esizing selected cryptophycins in accordance with the present invention;

Figure 10 provides a second scheme for synthesizing selected cryptophycins in accordance with the present invention;

Figure 11 provides a scheme for synthesizing a subunit of a cryptophycin comprising a hydroxy acid D;

Figure 12 provides a third scheme for synthesizing selected cryptophycins in accordance with the present invention;

Figure 13 provides a fourth scheme for synthesizing selected cryptophycins in accordance with the present invention; and

Figure 14 provides a fifth scheme for synthesizing selected cryptophycins in accordance with the present invention.

Detailed Description of the Invention

wherein

Ar is methyl or phenyl or any simple unsubstituted or substituted aromatic or heteroaromatic group; R, is a halogen, SH, amino, monoalkylamino, dialkylamino, trialkylammonium, alkylthio, dialkylsulfonium, sulfate, or phosphate; R2 is OH or SH; or R, and R2 may be taken together to form an epoxide ring, an aziridine ring, an episulfide ring, a sulfate ring or a monoalkylphosphate ring; or R, and R2 may be taken together to form a double bond between C,g and C,9; R3 is a lower alkyl group; R< and Rj are H; or R< and Rj may be taken together to form a double bond between C,3 and C,«;

Re is a benzyl, hydroxy benzyl, alkoxybenzyl, halohydroxybenzyl, dihalohydroxybenzyl, haloalkoxybenzyl, or dihaloalkoxybenzyl group; R7, Rg, R$ and R,o are each independently H or a lower alkyl group; and X and Y are each independently Ο, NH or alkylamino.

In one aspect of the present invention, novel cryptophycin compounds are provided having the following structure:

Wherein

Ri is H, OH, a halogen, O of a ketone group, NH2, SH, a lower alkoxyl group or a lower alkyl group; R2 is H, OH, O of a ketone group, NH2, SH, a lower alkoxyl group or a lower alkyl ' group; or R, and R2 may be taken together to form an epoxide ring, an aziridine ring, an episulfide ring or a double bond between C10 and CH; or Ri and R, may be taken together to form a tetrahydrofuran ring; r3 is H or a lower alkyl group; R< is OH, a lower alkanoyloxy group or a lower α-hydroxy alkanoyloxy group;

Rj is H or an OH group; is H; or

Rj and R^ may be taken together to form a double bond between C5 and C6; R7 is a benzyl, hydroxybenzyl, methoxy benzyl, halohydroxybenzyl, dihalohydroxybenzyl, halomethoxybenzyl, or dihalomethoxybenzyl group;

Rg is OH, a lower /3-amino acid wherein C, is bonded to N of the /3-amino acid, or an esterified lower /3-amino acid wherein C, is bonded to N of the esterified lower /3-amino acid group; R< and Rg may be taken together to form a didepsipeptide group consisting of a lower /3-amino acid bonded to a lower α-hydroxy alkanoic acid; and

Rj and Rg may be taken together to form a didepsipeptide group consisting of a lower /3-amino acid bonded to a lower a-hydroxy alkanoic acid; and

with the following provisos: R, is H, a lower alkyl group, or a lower alkoxyl group only if R2 is OH, O of a ketone group, NH2, SH; R, is H, a lower alkyl group, or a lower alkoxyl group only if R, is OH, O of a ketone group, NH2, SH; when R, is OH, R2 is OH, R3 is methyl, R5 and R* are taken together to form a double bond between C5 and C6, R^ and R8 are taken together to form the didepsipeptide group with the structure X:

wherein Oj of X corresponds to R^, Ng of X corresponds to Rg, R$ is methyl, and R10 is isobutyl, R7 is not 3-chloro-4-methoxybenzyl; when Rt and R2 are taken together to form an epoxide ring, R3 is methyl, Rs and R* are taken together to form a double bond between Cs and C6, R* and Rg are taken together to form a didepsipeptide with the structure X, R$ is methyl, and R10 is isobutyl, R7 is not 3-chloro-4-methoxybenzy 1; when R, and R2 are taken together to form a double bond between C,o and Cn, R3 is methyl, R5 and R* are taken together to form a double bond between C5 and C6, R< and Rg are taken together to form a didepsipeptide with the structure X, R, is methyl, and R10 is isobutyl, R7 is not 3-chloro-4-methoxybenzyl; and when R, and R2 are taken together to form an epoxide group, R3 is methyl, Rj and R* are taken together to form a double bond between C5 and C6, R, is bonded to the carboxy terminus of leucic acid, and Rg is bonded to the nitrogen terminus of either 3-amino-2-methylpropionic acid or 3-amino-2-methylpropionic acid methyl ester, R7 is not 3-chloro-4-methoxy benzyl.

The invention further provides cryptophycin compounds wherein at least one of the groups attached to C2, Cg, C9, C10, and C,, has R stereochemistry. In a further

embodiment of the invention, at least one of the groups attached to C2, Cg, C9, C10, and C,, has 5 stereochemistry.

The invention further provides cryptophycin compounds in accordance with the above structure where the structure of the didepsipeptide that is formed when R^ or Rj is taken together with Rg is the following structure X:

wherein Oj of X corresponds to R, or Rj, Ng of X corresponds to Rg, R, is H or a lower alkyl group, and R10 is H or a lower alkyl group.

As used herein, the following terms have the indicated meanings unless a contrary meaning is clearly intended from the use in context: "lower /3-amino acid" means any /3-amino acid having three to eight carbons and includes linear and non-linear hydrocarbon chains; for example, 3-amino-2-methylpropionic acid. "esterified lower /3-amino acid" means any /3-amino acid having three to eight carbons where the hydrogen of the carboxylic acid group is substituted with a methyl group; for example, 3-amino-2-methylpropionic acid methyl ester. "lower alkanoyloxy group" means an alkanoyloxy group of one to seven carbons and includes linear and non-linear hydrocarbon chains. "lower α-hydroxyalkanoyloxy group" means an α-hydroxyalkanoyloxy group of two to seven carbons and includes linear and non-linear hydrocarbon chains; for example, 2-hydroxy-4-methylvaleric acid. "lower alkoxy 1 group" means any alkyl group of one to five carbons bonded to an oxygen atom. "lower alkyl group" means an alkyl group of one to five carbons and includes linear and non-linear hydrocarbon chains including, for example, methyl, ethyl, propyl,

isopropyl, butyl, isobutyl, tert-butyl, sec-butyl, methylated butyl groups, pentyl, and tertpentyl groups. "allylically substituted alkene" means any alkene which contains an alkyl substitution. "epoxide ring" means a three-membered ring whose backbone consists of two carbons and an oxygen atom. "aziridine ring" means a three-membered ring whose backbone consists of two carbons and a nitrogen atom. "episulfide ring" means a three-membered ring whose backbone consists of two carbons and a sulfur atom. "sulfate ring" means a five-membered ring consisting of a carbon-carbon-oxygen-sulfur-oxygen backbone with two additional oxygen atoms connected to the sulfur atom. "monoalkylphosphate ring" means a five-membered ring consisting of a carbon-carbon-oxygen-phosphorus-oxygen backbone with two additional oxygen atoms, one of which bears a lower alkyl group, connected to the phosphorus atom. "simple unsubstituted aromatic group" refers to common aromatic rings having t 4n+2 pi electrons in a monocyclic conjugated system (for example, furyl, pyrrolyl, thienyl, pyridyl) or a bicyclic conjugated system (for example, indolyl or naphthyl). "simple substituted aromatic group" refers to a phenyl group substituted with · single group (e.g. a lower alkyl group or a halogen). "heteroaromatic group" refers to aromatic rings which contain one or more noncarbon substituents such as oxygen, nitrogen, or sulfur. "halogen" refers to those members of the group on the periodic table historically known as the halogens. Methods of halogenation include, but are not limited to, the addition of hydrogen halides, substitution at high temperature, photohalogenation, etc., and such methods are known to those of ordinary skill in the art.1·2

One embodiment of a cryptophycin compound of the present invention is when R, and R2 are taken together to form an epoxide group, R3 is methyl, Rj and Rj are taken together to form a double bond between Cs and C6, R7 is 4-methoxybenzyl, and R, and RB are taken together to form the didepsipeptide with the structure X where R, is methyl and R,o is isobutyl. The structure of this compound, Cryptophycin 2, is as follows:

CRYPTOPHYCIN 2 A further embodiment of a compound of the present invention is when R, and R2 ’ are taken together to form a double bond between the C10 and Cn carbons, R3 is methyl,

Rj and Rj are taken together to form a double bond between Cs and C6, R7 is 4-methoxybenzyl, and R< and Rg are taken together to form the didepsipeptide with the structure X where R, is methyl and R10 is isobutyl. The structure of this cryptophycin compound, Cryptophycin 4, is as follows:

1 CRYFTOPHYCJN 4

A further embodiment of a compound of the present invention is when R, and R„ are taken together to form a tetrahydrofuran ring, R2 is an OH group, R3 is methyl, R5 and R^ are taken together to form a double bond between C5 and C6, R7 is 3-chloro-4-methoxybenzyl, and Rs is a (2-carbomethoxypropyl)amino group. The structure of this compound, Cryptophycin 6, is as follows:

A further embodiment of a compound of the present invention is when R, and R, are taken together to form a tetrahydrofuran ring, R2 and Rg are OH groups, R3 is methyl, Rj and R are taken together to form a double bond between Cs and C6 such that there is a double bond, and R7 is 3-chloro-4-methoxybenzyl. The structure of this compound, . Cryptophycin 7, is as follows:

CRYPTOPHYCIN 7

A further embodiment of a compound of the present invention is when R, is a chloro group, R2 is an OH group, R3 is methyl, R5 and Ri are taken together to form a double bond between C5 and C6, R, is 3-chloro-4-methoxybenzyl, and R< and Rg are taken toge±er to form the didepsipeptide with the structure X where R, is methyl and R10 is isobutyl. The structure of this compound, Cryptophycin 8, is as follows:

1

I A further embodiment of a compound of the present invention is when R, is a methoxy group, R2 is an OH group, R3 is methyl, Rj and R* are taken together to form a double bond between Cs and C6, R7 is 3-chloro-4-methoxybenzyl, and R, and Rg are taken together to form the didepsipeptide with the structure X where R, is methyl and R10 is isobutyl. The structure of this compound, Cryptophycin 9, is as follows:

A further embodiment of a compound of the present invention is when R, is a methoxy group, R2 and R, are OH groups, R3 is methyl, R5 and R* are taken together to form a double bond between C3 and C6, R7 is 3-chloro-4-methoxybenzyl, and R8 is a (2-carboxypropyl)amino group. The structure of this compound, Cryptophycin 10, is as follows:

A further embodiment of a compound of the present invention is when Rj and R« are taken together to form a tetrahydrofuran ring, R2 is an OH group, R3 is methyl, Rs and Rj are taken together to form a double bond between C5 and C6 R7 is 3-chloro-4-methoxybenzyl, and Rg is a (2-carboxypropyl)amino group. The structure of this compound, Cryptophycin 12, is as follows:

A further embodiment of a compound of the present invention is when R, and R; are taken together to form a double bond between the C10 and CH carbons, R3 is methyl, R is an OH group, R5 and R are taken together to form a double bond between C5 and C6, R7 is 3-chloro-4-methoxybenzyl, and R8 is a (2-carboxypropyl)amino group. The structure of this compound, Cryptophycin 14, is as follows:

A further embodiment of a compound of the present invention is when R, and R2 are taken together to form an epoxide group, R3 is methyl, R5 and R are taken together to form a double bond between C5 and C6, R7 is 3-chloro-4-hydroxybenzyl, and R and RB are taken together to form the didepsipeptide with the structure X where R is methyl and R10 is isobutyl. The structure of this compound, Cryptophycin 16, is as follows:

A further embodiment of a compound of the present invention is when R, and R2 are taken together to form a double bond between C10 and C„ carbons, R3 is methyl, Rs and R are taken together to form a double bond between Cs and C6, R7 is 3-chloro-4-hydroxy benzyl, and R and Rg are taken together to form the didepsipeptide with the structure X where R, is methyl and R,o is isobutyl. The structure of this compound, Cryptophycin 17, is as follows:

A further embodiment of a compound of the present invention is when R, and R2 are taken together to form a double bond between CI0 and CH carbons, R3 is methyl, Rs and R* are taken together to form a double bond between C5 and C6, R7 is 3-chloro-4-methoxybenzyl, and R, and Rg are taken together to form the didepsipeptide with the structure X where R is methyl and R10 is sec-butyl. The structure of this compound, Cryptophycin 18, is as follows:

A further embodiment of a compound of the present invention is when R, and R; are taken together to form a double bond between C,o and CH carbons, R3 is methyl, R5 and R are taken together to form a double bond between Cj and C6, R7 is 3-chloro-4-methoxybenzyl, and R and Rg are taken together to form the didepsipeptide with the structure X where R is methyl and R]0 is isopropyl. The structure of this compound, Cryptophycin 19, is as follows:

A further embodiment of a compound of the present invention is when R, and R2 are taken together to form an epoxide group, R3 is methyl, Rs and R are taken together to form a double bond between C5 and C6, R7 is 3-chloro-4-methoxybenzyl, and R and Rg are taken together to form the didepsipeptide with the structure X where R is hydrogen and R10 is isobutyl. The structure of this compound, Cryptophycin 21, is as follows:

A further embodiment of a compound of the present invention is when R, and R, are taken together to form an epoxide group, R3 is methyl, Rs and R^ are taken together to form a double bond between C5 and C6, R7 is 3,5-dichloro-4-hydroxybenzyl, and R< and Rg are taken together to form the didepsipeptide with the structure X where R, is methyl and R10 is isobutyl. The structure of this compound, Cryptophycin 23, is as follows:

A further embodiment of a compound of the present invention is when R, and R2 are taken together to form an epoxide group, R3 is methyl, Rs and R* are taken together to form a double bond between Cs and C6, R7 is 4-methoxybenzyl, and R< and Rg are taken together to form the didepsipeptide with the structure X where R, is hydrogen and Rj0 is isobutyl. The structure of this compound, Cryptophycin 24, is as follows:

Claims

Original document published without claims.