Use of transthyretin peptide/protein fusions to increase the serum half-life of pharmacologically active peptides/proteins

Abstract

The present invention provides a means for increasing the serum half-life of a selected biologically active agent by utilizing transthyretin (TTR) as a fusion partner with a biologically active agent. Specifically, the present invention provides substantially homogenous preparations of TTR (or a TTR variant)-biologically active agent fusions and PEG-TTR (PEG-TTR variant)-biologically active agent fusions. As compared to the biologically active agent alone, the TTR-biologically active agent fusion and/or PEG-TTR-biologically active agent fusion has substantially increased serum half-life.

What is claimed is:  
     
         1 . A method for increasing the serum half-life of a biologically active agent comprising fusing the biologically active agent to transthyretin (TTR) or a TTR variant.  
     
     
         2 . The method of  claim 1  where said TTR or TTR variant is chemically modified with a chemical selected from the group consisting of dextran, poly(n-vinyl pyurrolidone), polyethylene glycols, propropylene glycol homopolymers, polypropylene oxide/ethylene oxide co-polymers, polyoxyethylated polyols and polyvinyl alcohols.  
     
     
         3 . The method of  claim 2  where said TTR or TTR variant is chemically modified with polyethylene glycol.  
     
     
         4 . The method of  claim 3  wherein said polyethylene glycol has a molecular weight of between about 1 kD and 100 kD.  
     
     
         5 . The method of  claim 4  wherein said polyethylene glycol has a molecular weight of between about 5 kD and 30 kD.  
     
     
         6 . The method of  claim 1  wherein said TTR is encoded by the nucleic acid molecule of SEQ ID NO:2.  
     
     
         7 . The method of  claim 1  wherein the biologically active agent is a protein.  
     
     
         8 . The method of  claim 1  wherein the biologically active agent is a peptide.  
     
     
         9 . The method of  claim 8  wherein the peptide is a TPO mimetic peptide (TMP).  
     
     
         10 . A substantially homogenous preparation of a TTR-biologically active agent fusion, optionally in a pharmaceutically acceptable diluent, carrier or adjuvant.  
     
     
         11 . A substantially homogenous preparation of a PEG-TTR-biologically active agent fusion, optionally in a pharmaceutically acceptable diluent, carrier or adjuvant.  
     
     
         12 . The preparation of  claim 11  wherein the biologically active agent is a protein.  
     
     
         13 . The preparation of  claim 11  wherein the biologically active agent is a peptide.  
     
     
         14 . The preparation of  claim 13  wherein the peptide is a TPO mimetic peptide (TMP).  
     
     
         15 . A substantially homogenous preparation of a TTR variant-biologically active agent fusion, optionally in a pharmaceutically acceptable diluent, carrier or adjuvant.  
     
     
         16 . A substantially homogenous preparation of a PEG-TTR variant-biologically active agent fusion, optionally in a pharmaceutically acceptable diluent, carrier or adjuvant.  
     
     
         17 . The preparation of  claim 16  wherein the biologically active agent is a protein.  
     
     
         18 . The preparation of  claim 16  wherein the biologically active agent is a peptide.  
     
     
         19 . The preparation of  claim 18  wherein the peptide is a TPO mimetic peptide (TMP).  
     
     
         20 . The preparation of any of claims  10 - 19  wherein the fusion contains a linker peptide.  
     
     
         21 . A process for preparing a substantially homogenous preparation of a TTR-biologically active agent fusion comprising: (a) fusing said TTR to a biologically active agent to provide a TTR-biologically active agent fusion; and (b) isolating said TTR-biologically active agent fusion.  
     
     
         22 . A process for preparing a substantially homogenous preparation of a TTR variant-biologically active agent fusion comprising: (a) engineering a cysteine residue into a specific amino acid position within the amino acid sequence of said TTR to provide a variant of said TTR; (b) fusing said TTR variant to a biologically active agent to provide a TTR variant-biologically active agent fusion; and (c) isolating said TTR variant-biologically active agent fusion.  
     
     
         23 . A process for preparing a substantially homogenous preparation of a PEG-TTR-biologically active agent fusion comprising: (a) conjugating a polyethylene glycol to said TTR to provide a PEG-TTR; (b) fusing said PEG-TTR to a biologically active agent to provide a PEG-TTR-biologically active agent fusion; and (c) isolating said PEG-TTR-biologically active agent fusion.  
     
     
         24 . A process for preparing a substantially homogenous preparation of a PEG-TTR variant-biologically active agent fusion comprising: (a) engineering a cysteine residue into a specific amino acid position within the amino acid sequence of said TTR to provide a variant of said TTR; (b) conjugating a polyethylene glycol to said TTR variant at said cysteine residue to provide a PEG-TTR variant; (c) fusing said PEG-TTR variant to a biologically active agent to provide a PEG-TTR-biologically active agent fusion; and (d) isolating said PEG-TTR-biologically active agent fusion.  
     
     
         25 . A method of treating thrombocytopenia comprising administering a therapeutically effective dose of a preparation of  claim 14 .  
     
     
         26 . A method of treating thrombocytopenia comprising administering a therapeutically effective dose of a preparation of  claim 19.