Methods of preparing pharmaceutical compositions for treatment of factor Vlll associated disorders and methods of use

Abstract

The invention concerns glycosylated proteins having human factor VIII activity. In a preferred embodiment, the protein is glycosylated with oligosaccharides that include an alpha-(2,6)-linked sialic acid and a bisecting GlcNAc linked to a core beta-mannose.

1 - 24 . (canceled) 
     
     
         25 . An isolated glycosylated protein having factor VIII procoagulant activity, wherein the amino acid sequence of the protein has at least 62% identity with SEQ ID NO:1 and the protein comprises an N-linked oligosaccharide having an alpha-(2,6)-linked sialic acid. 
     
     
         26 . The isolated protein of  claim 25 , wherein the amino acid sequence has at least 72% identity with SEQ ID NO:1. 
     
     
         27 . The isolated protein of  claim 25 , wherein the amino acid sequence has at least 82% identity with SEQ ID NO:1. 
     
     
         28 . A pharmaceutical composition comprising the isolated protein of  claim 25  and a pharmaceutically acceptable adjuvant. 
     
     
         29 . The pharmaceutical composition of  claim 28  wherein the composition is substantially free of a toxic solubilizing agent. 
     
     
         30 . A method of treating a factor VIII-associated disorder comprising administering to a human in need thereof a therapeutically effective amount of the pharmaceutical composition of  claim 28 . 
     
     
         31 . A method of reducing hemorrhage in a patient due to surgical procedure comprising administering to the patient a therapeutically effective amount of the pharmaceutical composition of  claim 28 . 
     
     
         32 . The method of  claim 31 , wherein the pharmaceutical composition is administered to the patient preoperatively. 
     
     
         33 . The method of  claim 32  further comprising the step of re-administering the pharmaceutical composition 6 to 12 hours after the initial administration. 
     
     
         34 . An isolated glycosylated protein having factor VIII procoagulant activity, wherein the amino acid sequence of the protein has at least 62% identity with SEQ ID NO:1 and the protein comprises an N-linked oligosaccharide having a bisecting GlcNAc linked to a core beta-mannose. 
     
     
         35 . The isolated protein of  claim 34 , wherein the amino acid sequence has at least 72% identity with SEQ ID NO:1. 
     
     
         36 . The isolated protein of  claim 34 , wherein the amino acid sequence has at least 82% identity with SEQ ID NO:1. 
     
     
         37 . A pharmaceutical composition comprising the isolated protein of  claim 34  and a pharmaceutically acceptable adjuvant. 
     
     
         38 . The pharmaceutical composition of  claim 37 , wherein the composition is substantially free of a toxic solubilizing agent. 
     
     
         39 . A method of treating a factor VIII-associated disorder comprising administering to a human in need thereof a therapeutically effective amount of the pharmaceutical composition of  claim 37 . 
     
     
         40 . A method of reducing hemorrhage in a patient due to surgical procedure comprising administering to the patient a therapeutically effective amount of the pharmaceutical composition of  claim 37 . 
     
     
         41 . The method of  claim 40 , wherein the pharmaceutical composition is administered to the patient preoperatively. 
     
     
         42 . The method of  claim 41  further comprising the step of re-administering the pharmaceutical composition 6 to 12 hours after the initial administration.