Enzyme and receptor modulation

Abstract

Covalent conjugates of an α,α-disubstituted glycine ester and a modulator of the activity of a target intracellular enzyme or receptor, wherein the ester group of the conjugate is hydrolysable by one or more intracellular carboxylesterase enzymes to the corresponding acid and the α,α-disubstituted glycine ester is conjugated to the modulator at a position remote from the binding interface between the inhibitor and the target enzyme or receptor pass into cells and the active acid hydrolysis product accumulates within the cells.

1 . A covalent conjugate of an α,α-disubstituted glycine ester and a modulator of the activity of a target intracellular enzyme or receptor, wherein:
 the ester group of the conjugate is hydrolysable by one or more intracellular carboxylesterase enzymes to the corresponding acid; and either
 a) the α,α-disubstituted glycine ester is conjugated to the modulator at a position remote from the binding interface between the inhibitor and the target enzyme or receptor or 
 (b) the α,α-disubstituted glycine ester is conjugated to the modulator such that the binding mode of the conjugated modulator and the said corresponding acid to the target enzyme or receptor is the same as that of the unconjugated modulator. 
   
     
     
         2 . (canceled) 
     
     
         3 . A method of increasing or prolonging the intracellular potency and/or residence time of modulator of the activity of a target intracellular enzyme or receptor comprising structural modification of the modulator
 a) by covalent attachment thereto of an α,α-disubstituted glycine ester at a position remote from the binding interface between the inhibitor and the target enzyme or receptor, the ester group of the conjugate being hydrolysable by one or more intracellular carboxylesterase enzymes to the corresponding acid; or   (b) by covalent attachment thereto of an α,α-disubstituted glycine ester such that the binding mode of the conjugated modulator and the said corresponding acid to the target enzyme or receptor is the same as that of the unconjugated modulator, the ester group of the conjugate being hydrolysable by one or more intracellular carboxylesterase enzymes to the corresponding acid.   
     
     
         4 . (canceled) 
     
     
         5 . The conjugate of  claim 1  wherein the α-substituents of the α,α-disubstituted glycine ester conjugated to the modulator are independently selected from phenyl and groups of formula —CR a R b R c  in which:
 each of R a , R b  and R c  is independently hydrogen, (C 1 -C 6 )alkyl, (C 2 -C 6 )alkenyl, (C 2 -C 6 )alkynyl, phenyl(C 1 -C 6 )alkyl, (C 3 -C 8 )cycloalkyl; or   R c  is hydrogen and R a  and R b  are independently phenyl or heteroaryl such as pyridyl; or   R c  is hydrogen, (C 1 -C 6 )alkyl, (C 2 -C 6 )alkenyl, (C 2 -C 6 )alkynyl, phenyl(C 1 -C 6 )alkyl, or (C 3 -C 8 )cycloalkyl, and R a  and R b  together with the carbon atom to which they are attached form a 3 to 8 membered cycloalkyl or a 5- to 6-membered heterocyclic ring; or   R a , R b  and R c  together with the carbon atom to which they are attached form a tricyclic ring (for example adamantyl); or   R a  and R b  are each independently (C 1 -C 6 )alkyl, (C 2 -C 6 )alkenyl, (C 2 -C 6 )alkynyl, phenyl(C 1 -C 6 )alkyl, or a group as defined for R c  below other than hydrogen, or R a  and R b  together with the carbon atom to which they are attached form a cycloalkyl or heterocyclic ring, and R c  is hydrogen, —OH, —SH, halogen, —CN, —CO2H, (C1-C4)perfluoroalkyl, —CH2OH, —O(C 1 -C 6 )alkyl, —O(C 2 -C 6 )alkenyl, —S(C 1 -C 6 )alkyl, —SO(C 1 -C 6 )alkyl, —SO2(C 1 -C 6 )alkyl, —S(C 2 -C 6 )alkenyl, —SO(C 2 -C 6 )alkenyl, —SO2(C 2 -C 6 )alkenyl or a group -Q-W wherein Q represents a bond or —O—, —S—, —SO— or —SO2- and W represents a phenyl, phenylalkyl, (C 3 -C 8 )cycloalkyl, (C 3 -C 8 )cycloalkylalkyl, (C 4 -C 8 )cycloalkenyl, (C 4 -C 8 )cycloalkenylalkyl, heteroaryl or heteroarylalkyl group, which group W may optionally be substituted by one or more substituents independently selected from, hydroxyl, halogen, —CN, —CONH2, —CONH(C 1 -C 6 )alkyl, —CONH(C 1 -C 6 alkyl) 2 , —CHO, —CH2OH, (C 1 -C 4 )perfluoroalkyl, —O(C 1 -C 6 )alkyl, —S(C 1 -C 6 )alkyl, —SO(C 1 -C 6 )alkyl, O 2 (C 1 -C 6 )alkyl, —NO2, —NH2, —NH(C 1 -C 6 )alkyl, —N((C 1 -C 6 )alkyl) 2 , —NHC 0 (C 1 -C 6 )alkyl, (C 1 -C 6 )alkyl, (C 2 -C 6 )alkenyl, (C 2 -C 6 )alkynyl, (C 3 -C 8 )cyclo alkyl, (C 4 -C 8 )cycloalkenyl, phenyl or benzyl.   
     
     
         6 . The conjugate of  claim 1  wherein the α-substituents of the α,α-disubstituted glycine ester conjugated to the modulator, taken together with the α-carbon itself, form a 3-6 membered saturated spiro cycloalkyl or spiro heterocyclyl ring; or wherein at least one of the said α-substituents is a C 1 -C 6  alkyl substituent. 
     
     
         7 . The conjugate of  claim 1  wherein one of the α-substitutents of the α,α-disubstituted glycine ester conjugated to the modulator is a C 1 -C 6  alkyl substituent, and the other is selected from the group consisting of methyl, ethyl, n- and iso-propyl, n-, sec- and tert-butyl, phenyl, benzyl, thienyl, cyclohexyl, and cyclohexylmethyl. 
     
     
         8 . The conjugate of  claim 1  wherein one of the α-substitutents of the α,α-disubstituted glycine ester conjugated to the modulator is methyl. and the other is methyl, ethyl, or n- or iso-propyl; or the said α-substitutents taken together with the carbon to which they are attached form a cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl ring. 
     
     
         9 . The conjugate of  claim 1  wherein the α,α-disubstituted glycine ester is covalently conjugated to the modulator via the amino group of the α,α-disubstituted glycine ester. 
     
     
         10 . The conjugate of  claim 1  wherein the α,α-disubstituted glycine ester is covalently conjugated to the modulator via one of the α substituents of the α,α-disubstituted glycine ester. 
     
     
         11 . The conjugate  claim 1  wherein the ester is selectively hydrolysable to the corresponding α,α-disubstituted glycine conjugate by cells containing the intracellular carboxylesterase enzyme hCE-1 relative to cells containing hCE-2 or hCE-3 but not hCE-1. 
     
     
         12 . A pharmaceutical composition comprising the conjugate of  claim 1  and one or more pharmaceutically acceptable carriers or excipients. 
     
     
         13 . A method of identifying a covalent conjugate of an α,α-disubstituted glycine ester of a given modulator of the activity of a target intracellular enzyme or receptor, said conjugate having increased or prolonged cellular potency relative to the given modulator, which method comprises:
 (i) identifying a position or positions on a given modulator of the activity of a target intracellular enzyme or receptor, or on a plurality of given modulators of the activity of a target intracellular enzyme or receptor sharing the same binding mode for the target enzyme or receptor, said position or positions being remote from the binding interface between the modulators and the target enzyme or receptor;   (ii) covalently modifying the modulator(s) by attachment of an α,α-disubstituted glycine ester radical, or a range of different α,α-disubstituted glycine ester radicals at one or more of the positions identified in (i);   (iii) testing the alpha amino acid-conjugated modulator(s) prepared in (ii) to determine their activity against the target enzyme or receptor; and   (iv) from data acquired in (iii), selecting one or more of the tested α,α-disubstituted glycine ester-conjugated versions of the given modulator(s) which cause modulation of enzyme or receptor activity inside cells, are converted to and accumulate as the corresponding carboxylic acid inside cells, and which show increased or prolonged cellular potency.   
     
     
         14 . The method of  claim 3  wherein the α-substituents of the α,α-disubstituted glycine ester conjugated to the modulator are independently selected from phenyl and groups of formula —CR a R b R c  in which:
 each of R a , R b  and R c  is independently hydrogen, (C 1 -C 6 )alkyl, (C 2 -C 6 )alkenyl, (C 2 -C 6 )alkynyl, phenyl(C 1 -C 6 )alkyl, (C 3 -C 8 )cycloalkyl; or   R c  is hydrogen and R a  and R b  are independently phenyl or heteroaryl such as pyridyl; or   R c  is hydrogen, (C 1 -C 6 )alkyl, (C 2 -C 6 )alkenyl, (C 2 -C 6 )alkynyl, phenyl(C 1 -C 6 )alkyl, or (C 3 -C 8 )cycloalkyl, and R a  and R b  together with the carbon atom to which they are attached form a 3 to 8 membered cycloalkyl or a 5- to 6-membered heterocyclic ring; or   R a , R b  and R c  together with the carbon atom to which they are attached form a tricyclic ring (for example adamantyl); or   R a  and R b  are each independently (C 1 -C 6 )alkyl, (C 2 -C 6 )alkenyl, (C 2 -C 6 )alkynyl, phenyl(C 1 -C 6 )alkyl, or a group as defined for R c  below other than hydrogen, or R a  and R b  together with the carbon atom to which they are attached form a cycloalkyl or heterocyclic ring, and R c  is hydrogen, —OH, —SH, halogen, —CN, —CO2H, (C 1 -C 4 )perfluoroalkyl, —CH2OH, —O(C 1 -C 6 )alkyl, —O(C 2 -C 6 )alkenyl, —S(C 1 -C 6 )alkyl, —SO(C 1 -C 6 )alkyl, —SO2(C 1 -C 6 )alkyl, —S(C 2 -C 6 )alkenyl, —SO(C 2 -C 6 )alkenyl, —SO2(C 2 -C 6 )alkenyl or a group -Q-W wherein Q represents a bond or —O—, —S—, —SO— or —SO2- and W represents a phenyl, phenylalkyl, (C 3 -C 8 )cycloalkyl, (C 3 -C 8 )cycloalkylalkyl, (C 4 -C 8 )cycloalkenyl, (C 4 -C 8 )cycloalkenylalkyl, heteroaryl or heteroarylalkyl group, which group W may optionally be substituted by one or more substituents independently selected from, hydroxyl, halogen, —CN, —CONH2, —CONH(C 1 -C 6 )alkyl, —CONH(C 1 -C 6 alkyl) 2 , —CHO, —CH2OH, (C 1 -C 4 )perfluoroalkyl, —O(C 1 -C 6 )alkyl, —S(C 1 -C 6 )alkyl, —SO(C 1 -C 6 )alkyl, -502(C 1 -C 6 )alkyl, —NO2, —NH2, —NH(C 1 -C 6 )alkyl, —N((C 1 -C 6 )alkyl) 2 , —NHCO(C 1 -C 6 )alkyl, (C 1 -C 6 )alkyl, (C 2 -C 6 )alkenyl, (C 2 -C 6 )alkynyl, (C 3 -C 8 )cycloalkyl, (C 4 -C 8 )cycloalkenyl, phenyl or benzyl.   
     
     
         15 . The method of  claim 3  wherein the α-substituents of the α,α-disubstituted glycine ester conjugated to the modulator, taken together with the α-carbon itself, form a 3-6 membered saturated spiro cycloalkyl or spiro heterocyclyl ring; or wherein at least one of the said α-substituents is a C 1 -C 6  alkyl substituent. 
     
     
         16 . The method of  claim 3  wherein one of the α-substitutents of the α,α-disubstituted glycine ester conjugated to the modulator is a C 1 -C 6  alkyl substituent, and the other is selected from the group consisting of methyl, ethyl, n- and iso-propyl, n-, sec- and tert-butyl, phenyl, benzyl, thienyl, cyclohexyl, and cyclohexylmethyl. 
     
     
         17 . The method of  claim 3  wherein one of the α-substitutents of the α,α-disubstituted glycine ester conjugated to the modulator is methyl. and the other is methyl, ethyl, or n- or iso-propyl; or the said α-substitutents taken together with the carbon to which they are attached form a cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl ring. 
     
     
         18 . The method of  claim 3  wherein the α,α-disubstituted glycine ester is covalently conjugated to the modulator via the amino group of the α,α-disubstituted glycine ester. 
     
     
         19 . The method of  claim 3  wherein the α,α-disubstituted glycine ester is covalently conjugated to the modulator via one of the α substituents of the α,α-disubstituted glycine ester. 
     
     
         20 . The method of  claim 3  wherein the ester is selectively hydrolysable to the corresponding α,α-disubstituted glycine conjugate by cells containing the intracellular carboxylesterase enzyme hCE-1 relative to cells containing hCE-2 or hCE-3 but not hCE-1.