Acryloyloxyethylphosphorylcholine Containing Polymer Conjugates And Their Preparation

Abstract

The present invention relates to polymeric reagents and conjugates thereof, methods for synthesizing the polymeric reagents and conjugates, pharmaceutical compositions comprising the conjugates and methods of using the polymer conjugates including therapeutic methods where conjugates are administered to patients.

1 - 40 . (canceled) 
     
     
         41 . A compound comprising a biologically active protein covalently bonded to a phosphorylcholine containing polymer, wherein said polymer is branched, having 3 or more polymer arms, wherein each polymer arm is represented by the formula: 
       
         
           
           
               
               
           
         
         wherein m is about 20 to about 2000; 
         each occurrence of E is independently selected from the group consisting of halo and a nucleophilic group; 
         each occurrence of Q is independently selected from the group consisting of H, methyl and ethyl; 
         each occurrence of T is independently —C 1-4 alkyl-phosphorylcholine; and 
         the wavy line indicates the point of attachment to the rest of the polymer. 
       
     
     
         42 . The compound of  claim 41  wherein Q is methyl. 
     
     
         43 . The compound of  claim 44  wherein T is —CH 2 —CH 2 -phosphorylcholine. 
     
     
         44 . The compound of  claim 41  wherein m is about 100 to about 500. 
     
     
         45 . The compound of  claim 41  wherein m is about 500 to about 1000. 
     
     
         46 . The compound of  claim 41  wherein the polymer has 3, 4, 5, 6, 7 or 8 polymer arms. 
     
     
         47 . The compound of  claim 46  wherein the polymer has 3 arms. 
     
     
         48 . The compound of  claim 46  wherein the polymer has 6 arms. 
     
     
         49 . The compound of  claim 41  wherein the polymer has more than 8 arms. 
     
     
         50 . The compound of  claim 41  wherein the phosphorylcholine containing polymer is covalently bonded to at least an amino group, a hydroxyl group, a sulfhydryl group or a carboxyl group of the biologically active protein. 
     
     
         51 . The compound of  claim 41  wherein the biologically active protein is a human protein. 
     
     
         52 . The compound of  claim 51  wherein the human protein is obtained by heterologous gene expression in a cell selected from the group consisting of a bacterium, a yeast cell, a mammalian cell in culture, an insect cell in culture, a plant cell in culture, an avian cell in culture, a cell of a transgenic avian, a cell of a transgenic mammal, and a cell of a transgenic plant. 
     
     
         53 . The compound of  claim 41  wherein the biologically active protein is selected from the group consisting of a cytokine, an enzyme, an antibody and an antibody fragment. 
     
     
         54 . The compound of  claim 41 , wherein the polymer portion of the compound is polydisperse. 
     
     
         55 . The compound of  claim 54 , wherein the polymer portion has a polydispersity value in the range of about 1.4 to about 1.2. 
     
     
         56 . The compound of  claim 55 , wherein the polymer portion has a polydispersity value of less than about 1.15. 
     
     
         57 . The compound of  claim 41  wherein m is about 200 to about 2000. 
     
     
         58 . The compound of  claim 41 , further represented by the following formula:
   (A-Z—(SP 1 ) n ) a -L-(K) b ;
   wherein K is the polymer arm;   each Sp 1  is a spacer group;   n is 0 or 1, wherein when n is 0, Sp 1  is a bond;   a is an integer between 1 and 8;   b is an integer selected from 3, 4, 5, 6, 7, 8 or more than 8;   L is selected from the group consisting of C 1-4 alkyl, cycloalkyl, carboxyC 1-4 alkyl, carboxycycloalkyl, C 1-4 alkoxyC 1-4 alkyl and cyclic alkyl ether;   A is a biologically active protein;   Z is the product of the reaction between a group present in said biologically active protein and a reactive group bound to said Sp 1  group when n is 1, or a reactive group bound to L when n is 0.   
     
     
         59 . The compound of  claim 58  wherein Sp 1  is selected from the group consisting of: —C 1-12  alkyl-, —C 3-12  cycloalkyl-, —(C 1-8  alkyl)-(C 3-12  cycloalkyl)-(C 0-8  alkyl)-, —(CH 2 ) 1-12 O—, (—(CH 2 ) 1-6 —O—(CH 2 ) 1-6 -) 1-12 -, (—(CH 2 ) 1-4 —NH—(CH 2 ) 1-4 ) 1-12 —, (—(CH 2 ) 1-4 —O—(CH 2 ) 1-4 ) 1-12 —O—, (—(CH 2 ) 1-4 —O—(CH 2 ) 1-4 -) 1-12 O—(CH 2 ) 1-12 —, —(CH 2 ) 1-12 —(C═O)—O—, —(CH 2 ) 1-12 —O—(C═O)—, -(phenyl)-(CH 2 ) 1-3 —(C═O)—O—, -(phenyl)-(CH 2 ) 1-3 —(C═O)—NH—, —(C 1-6  alkyl)-(C═O)—O—(C 0-6  alkyl)-, —(CH 2 ) 1-12 —(C═O)—O—(CH 2 ) 1-12 —, —CH(OH)—CH(OH)—(C═O)—O— —CH(OH)—CH(OH)—(C═O)—NH—, —S-maleimido-(CH 2 ) 1-6 —, —S-maleimido-(C 1-3  alkyl)-(C═O)—NH—, —S-maleimido-(C 1-3  alkyl)-(C 5-6  cycloalkyl)-(C 0-3  alkyl)-, —(C 1-3  alkyl)-(C 5-6  cycloalkyl)-(C 0-3  alkyl)-(C═O)—O—, —(C 1-3  alkyl)-(C 5-6  cycloalkyl)-(C 0-3  alkyl)-(C═O)—NH—, —S-maleimido-(C 0-3 alkyl)-phenyl-(C 0-3 alkyl)-, —(C 0-3  alkyl)-phenyl-(C═O)—NH—, —(CH 2 ) 1-12 —NH—(C═O)—, —(CH 2 ) 1-12 —(C═O)—NH—, -(phenyl)-(CH 2 ) 1-3 —(C═O)—NH—, —S—(CH 2 )—(C═O)—NH-(phenyl)-, —(CH 2 ) 1-12 —(C═O)—NH—(CH 2 ) 1-12 —, —(CH 2 ) 2 —(C═O)—O—(CH 2 ) 2 —O—(C═O)—(CH 2 ) 2 —(C═O)—NH—, —(C 1-6  alkyl)-(C═O)—N—(C 1-6  alkyl)-, acetal, ketal, acyloxyalkyl ether, —N═CH—, —(C 1-6  alkyl)-S—S—(C 0-6  alkyl)-, —(C 1-6  alkyl)-S—S—(C 1-6  alkyl)-(C═O)—O—, —(C 1-6  alkyl)-S—S—(C 1-6  alkyl)-(C═O)—NH—, —S—S—(CH 2 ) 1-3 —(C═O)—NH—(CH 2 ) 1-4 —NH—(C═O)—(CH 2 ) 1-3 —, —S—S—(C 0-3  alkyl)-(phenyl)-, —S—S—(C 1-3 -alkyl)-(phenyl)-(C═O)—NH—(CH 2 ) 1-5 —, —(C 1-3  alkyl)-(phenyl)-(C═O)—NH—(CH 2 ) 1-5 —(C═O)—NH—, —S—S—(C 1-3 -alkyl)-, —(C 1-3 -alkyl)-(phenyl)-(C═O)—NH—, —O—(C 1 -C 6  alkyl)-S(O 2 )—(C 1-6  alkyl)—O—(C═O)—NH—, —S—S—(CH 2 ) 1-3 —(C═O)—, —(CH 2 ) 1-3 —(C═O)—NH—N═C—S-S—(CH 2 ) 1-3 —(C═O)—NH—(CH 2 ) 1-5 —, —(CH 2 ) 1-3 —(C═O)—NH—(CH 2 ) 1-5 —(C═O)—NH—, —(CH 2 ) 0-3 -(heteroaryl)-(CH 2 ) 0-3 —, —(CH 2 ) 0-3 -phenyl-(CH 2 ) 0-3 —, 
       
         
           
           
               
               
           
         
         wherein R 1  is C 1-6  alkyl, C 3-6  cycloalkyl, or an aryl group having 5-8 endocyclic atoms; and R 2  is H, C 1-6  alkyl, C 3-6  cycloalkyl, or an aryl group having 5-8 endocyclic atoms. 
       
     
     
         60 . The compound of  claim 58  wherein Z is selected from the group consisting of: phosphate, phosphate ester, —(C═O)O—, carboxylic acid ester, thioester, amide, dislufide, amine, —NH(R 1 )—, amidine, hydrazone, —N═CH—, —NH—CH 2 —, 
       
         
           
           
               
               
           
         
         NH—C(R 1 )H—, —S—CH 2 —C(OH)(R 2 )—, —O—CH 2 —C(OH)(R 2 )—, —C(═O)O—CH 2 —C(OH)(R 2 )—, —NR 2 —CH 2 —C(OH)(R 2 )—, —S—CH 2 —C(SH)(R 2 )—, —O—CH 2 —C(SH)(R 2 )—, —C(═O)O—CH 2 —C(SH)(R 2 )—, —NR 2 —CH 2 —C(SH)(R 2 )—, 
       
       
         
           
           
               
               
           
         
         —C(R 2 )H—NH—, —(C═O)—NH—(C═O)—NH—, 
       
       
         
           
           
               
               
           
         
         C(R 2 )H—NH—(C═O)—NH—, —(C═O)—NH—(C═O)—O—, 
       
       
         
           
           
               
               
           
         
         —C(R 2 )H—NH—(C═O)—O—, (C═O)—NH—(C═S)—NH—, 
       
       
         
           
           
               
               
           
         
         —C(R 2 )H—NH—(C═S)—NH—, —(C═O)—NH—(C═S)—O—, 
       
       
         
           
           
               
               
           
         
         —C(R 2 )H—NH—(C═S)—O—, 
       
       
         
           
           
               
               
           
         
         —NH—(C═O)—NH—, —O—(C═O)—NH—, —NH—(C═S)—NH—, —O—(C═S)—NH—, —S—CH 2 CH 2 —(C═O)—, —S—CH 2 CH(CH 3 )—(C═O)— —S—CH 2 CH 2 —(C═O)O—, —S—CH 2 CH(CH 3 )—(C═O)O—, —S—CH 2 CH 2 —(C═O)NH—, —S—CH 2 —CH 2 -(pyridyl)-, —S—CH 2 —CH 2 —SO 2 —, —S—CH 2 —CH 2 —SO 2 —, —S—CH 2 —(C═O)—O—, —S—CH 2 —(C═O)—NH—, —S—CH 2 —(C═O)—, and 
       
       
         
           
           
               
               
           
         
       
       wherein R 1  is C 1-6  alkyl, C 3-6  cycloalkyl, or an aryl group having 5-8 endocyclic atoms; and R 2  is H, C 1-6  alkyl, C 3-6  cycloalkyl, or an aryl group having 5-8 endocyclic atoms. 
     
     
         61 . A pharmaceutical composition comprising the compound of  claim 41  in a liquid solution. 
     
     
         62 . A pharmaceutical composition comprising the compound of  claim 43  in a liquid solution. 
     
     
         63 . A pharmaceutical composition comprising the compound of  claim 53  in a liquid solution. 
     
     
         64 . A pharmaceutical composition comprising the compound of  claim 55  in a liquid solution.