Conjugates of insulin-like growth factor-1 and poly(ethylene glycol)

Abstract

A conjugate consisting of an insulin-like growth factor-1 (IGF-I) variant and one or two poly(ethylene glycol) group(s), characterized in that said IGF-I variant has an amino acid alteration at up to three amino acid positions 27, 37, 65, 68 of the wild-type IGF-I amino acid sequence so that one or two of said amino acids is/are lysine and amino acid 27 is a polar amino acid but not lysine, is conjugated via the primary amino group(s) of said lysine(s) and said poly(ethylene glycol) group(s) have an overall molecular weight of from 20 to 100 kDa is disclosed. This conjugate is useful for the treatment of neurodegenerative disorders like Alzheimer's Disease.

1 . A conjugate comprising an IGF-I variant and poly(ethylene glycol), wherein:
 said IGF-I variant is selected from the group consisting of:
 (A) a variant which differs from wild-type IGF-I in that the amino acid sequence of the variant has an alteration at residue position 27 such that lysine is replaced with arginine; 
 (B) a variant which differs from wild-type IGF-I in that the amino acid sequence of the variant has alterations at residue positions 27 and 65 such that both lysines are replaced with arginine; 
 (C) a variant which differs from wild-type IGF-I in that the amino acid sequence of the variant has alterations at residue positions 27 and 68 such that both lysines are replaced with arginine; and 
 (D) a variant which differs from wild-type IGF-I in that the amino acid sequence of the variant has alterations at residue positions 27, 65, and 68 such that the lysines are replaced with arginine and a modification at position 37 such that the arginine is replaced by lysine; and 
   said poly(ethylene glycol) is conjugated to said IGF-I variant via one or more primary amino groups.   
     
     
         2 . A conjugate according to  claim 1  wherein said poly(ethylene glycol) has an overall molecular weight of from 20 to 100 kDa. 
     
     
         3 . A conjugate according to  claim 1  wherein said IGF-I variant is additionally conjugated to poly(ethylene glycol) at the N-terminal amino acid. 
     
     
         4 . A conjugate according to  claim 1  wherein said IGF-I variant is conjugated to poly(ethylene glycol) at a member selected from the group consisting of lysine 65, lysine 68, and lysine 37 or is conjugated to poly(ethylene glycol) at both K65 and K68. 
     
     
         5 . A conjugate according to  claim 1  wherein up to three amino acids at the N-terminus are truncated. 
     
     
         6 . A conjugate according to  claim 1  wherein the poly(ethylene glycol) group(s) is/are branched poly(ethylene glycol) group(s). 
     
     
         7 . A conjugate according to  claim 1  wherein the poly(ethylene glycol) group(s) have an overall molecular weight of 20 kDa to 100 kDa. 
     
     
         8 . An IGF-I variant selected from the group consisting of:
 (A) a variant which differs from wild-type IGF-I in that the amino acid sequence of the variant has an alteration at residue position 27 such that lysine is replaced with arginine;   (B) a variant which differs from wild-type IGF-I in that the amino acid sequence of the variant has alterations at residue positions 27 and 65 such that both lysines are replaced with arginine;   (C) a variant which differs from wild-type IGF-I in that the amino acid sequence of the variant has alterations at residue positions 27 and 68 such that both lysines are replaced with arginine; and   (D) a variant which differs from wild-type IGF-I in that the amino acid sequence of the variant has alterations at residue positions 27, 65, and 68 such that the lysines are replaced with arginine and a modification at position 37 such that the arginine is replaced by lysine.   
     
     
         9 . A method for the preparation of a conjugate comprising an IGF-I variant and one or two poly(ethylene glycol) group(s), said poly(ethylene glycol) group(s) having an overall molecular weight of from about 20 to about 100 kDa, said method comprising reacting the IGF-I variant of  claim 8  with activated (polyethylene) glycol under conditions such that said (polyethylene) glycol will react with one or two primary lysine amino groups of said IGF-I variant. 
     
     
         10 . The method according to  claim 9  further comprising reacting (polyethylene) glycol with said IGF-I variant via the N-terminal amino group of said IGF-I variant to yield an IGF-I variant with poly(ethylene glycol) bound to both the N-terminal amino group and to one or two primary lysine amino groups. 
     
     
         11 . A pharmaceutical composition comprising a conjugate of  claim 1  and a pharmaceutically acceptable carrier. 
     
     
         12 . A method for the treatment of Alzheimer's Disease comprising administering to a patient in need thereof a therapeutically effective amount of a conjugate of  claim 1 . 
     
     
         13 . A composition comprising a conjugate according to  claim 3 . 
     
     
         14 . A pharmaceutical composition comprising a conjugate of  claim 3  and a pharmaceutically acceptable carrier. 
     
     
         15 . A method for the treatment of Alzheimer's Disease comprising administering to a patient in need thereof a therapeutically effective amount of a composition of  claim 13 . 
     
     
         16 . A method for the treatment of Alzheimer's Disease comprising administering to a patient in need thereof a therapeutically effective amount of a composition of  claim 14 . 
     
     
         17 . A conjugate according to  claim 1  wherein said variant is selected from the group consisting of:
 (A) a variant which differs from wild-type IGF-I in that the amino acid sequence of the variant has alterations at residue positions 27 and 65 such that both lysines are replaced with arginine; and 
 (B) a variant which differs from wild-type IGF-I in that the amino acid sequence of the variant has alterations at residue positions 27 and 68 such that both lysines are replaced with arginine. 
 
     
     
         18 . A conjugate according to  claim 1  wherein said variant differs from wild-type IGF-I in that the amino acid sequence of the variant has alterations at residue positions 27 and 65 such that both lysines are replaced with arginine. 
     
     
         19 . A pharmaceutical composition comprising a conjugate of  claim 17  and a pharmaceutically-acceptable carrier. 
     
     
         20 . A pharmaceutical composition comprising a conjugate of  claim 18  and a pharmaceutically-acceptable carrier.