Modified peptides as therapeutic agents

Abstract

The present invention concerns fusion of Fc domains with biologically active peptides and a process for preparing pharmaceutical agents using biologically active peptides. In this invention, pharmacologically active compounds are prepared by selecting a peptide that modulates the activity of a protein of interest and preparing a pharmacologic agent having an Fc domain covalently linked to the peptide. Linkage to the Fc domain increases the half-life of the peptide, which otherwise would be quickly degraded in vivo. The peptide can be selected, for example, by phage display, E. coli display, ribosome display, RNA-peptide screening, yeast-based screening, chemical-peptide screening, rational design, or protein structural analysis.

1 . A process for preparing a pharmacologically active compound, which comprises: 
 (a) selecting from a peptide phage display library at least one peptide sequence that bind to NGF;    (b) preparing a compound incorporating at least one said selected peptide sequence, wherein the compound has the formula      (X 1 ) a —F 1 (X 2 ) b      and multimers thereof, wherein:    F 1  is an Fc domain;    X 1  and X 2  are each independently selected from -(L 1 ) c -P 1 , -(L 1 ) c -P 1 -(L 2 ) d -P 2 , -(L 1 ) c -P 1 -(L 2 ) d -P 2 -(L 3 ) e -P 3 , and -(L 1 ) c -P 1 -(L 2 ) d -P 2 -(L 3 ) e -P 3 -(L 4 ) f -P 4      P 1 , P 2 , P 3 , and P 4  are each independently the selected peptide sequences;    L 1 , L 2 , L 3 , and L 4  are each independently linkers; and    a, b, c, d, e, and f are each independently 0 or 1,    provided that at least one of a and b is 1.    
     
     
         2 . The process of  claim 1 , wherein the compound prepared is of the formulae  
         X 1 —F 1    or  F 1 —X 2 .  
     
     
         3 . The process of  claim 2 , wherein the compound prepared is of the formulae  
         F 1 -(L 1 ) c -P 1    or  F 1 -(L 1 ) c -P 1 -(L 2 ) d -P 2 .  
     
     
         4 . The process of  claim 1 , wherein F 1  is an IgG Fc domain.  
     
     
         5 . The process of  claim 1 , wherein F 1  is an IgG1 Fc domain.  
     
     
         6 . The process of  claim 1 , wherein F 1  comprises the sequence of SEQ ID NO: 2.  
     
     
         7 . The process of  claim 1 , wherein a is 1 and b is 0.  
     
     
         8 . The process of  claim 6  wherein X 1  is -(L 1 ) c -P 1 -(L 2 ) d -P 2 .  
     
     
         9 . The process of  claim 1 , wherein a is 0 and b is 1.  
     
     
         10 . The process of  claim 8  wherein X 2  is -(L 1 ) c -P 1 -(L 2 ) d -P 2 .  
     
     
         11 . The process of  claim 1 , wherein step b of  claim 1  is carried out by: 
 (a) preparing a gene construct comprising a nucleic acid sequence encoding the peptide selected in step a of  claim 1  adjacent to either the N-terminus or the C-terminus of a nucleic acid sequence encoding an Fc domain; and    (b) expressing the gene construct.    
     
     
         12 . The process of  claim 10 , wherein the gene construct is expressed in an  E. coli  cell.  
     
     
         13 . The process of  claim 1 , wherein step (a) of  claim 1  is carried out by a process comprising: 
 (a) preparing a gene construct comprising a nucleic acid sequence encoding a first selected peptide and a nucleic acid sequence encoding an Fc domain;    (b) conducting a polymerase chain reaction using the gene construct and mutagenic primers, wherein 
 (i) a first mutagenic primer comprises a nucleic acid sequence complementary to a sequence at or near the 5′ end of a coding strand of the gene construct, and  
 (ii) a second mutagenic primer comprises a nucleic acid sequence complementary to the 3′ end of the noncoding strand of the gene construct.