Factor VIII glycoforms
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. An isolated glycosylated protein having factor VIII procoagulant activity comprising the amino acid sequence of SEQ ID NO: 1 and an N-linked oligosaccharide having an alpha-(2,6)-linked sialic acid.
2. A pharmaceutical composition comprising the isolated glycosylated protein of claim 1 and a pharmaceutically acceptable adjuvant.
3. The pharmaceutical composition of claim 2 , wherein the composition does not contain a toxic solubilizing agent.
4. A method of treating hemophilia comprising administering to a human in need thereof a therapeutically effective amount of the pharmaceutical composition of claim 2 .
5. A method of preparing the pharmaceutical composition of claim 2 comprising mixing the isolated glycosylated protein with a pharmaceutically acceptable adjuvant.
6. An isolated glycosylated protein having factor VIII procoagulant activity comprising the amino acid sequence of SEQ ID NO: 1 and a bisecting GlcNAc linked to a core beta-mannose.
7. The isolated glycosylated protein of claim 4 further comprising an alpha-(2,6)-linked sialic acid.
8. A pharmaceutical composition comprising the isolated glycosylated protein of claim 6 and a pharmaceutically acceptable adjuvant.
9. The pharmaceutical composition of claim 8 , wherein the composition does not contain a toxic solubilizing agent.
10. A method of treating hemophilia comprising administering to a human in need thereof a therapeutically effective amount of the pharmaceutical composition of claim 8 .
11. A method of preparing the pharmaceutical composition of claim 8 comprising mixing the isolated glycosylated protein with a pharmaceutically acceptable adjuvant.
12. An isolated protein having factor VIII procoagulant activity, wherein the amino acid sequence of the protein is at least 95% identical to SEQ ID NO: 1 and the protein further comprises a N-linked oligosaccharide comprising alpha-(2,6)-linked sialic acid.
13. A pharmaceutical composition comprising the isolated protein of claim 12 and a pharmaceutically acceptable adjuvant.
14. The pharmaceutical composition of claim 13 , wherein the composition does not contain a toxic solubilizing agent.
15. A method of treating hemophilia comprising administering to a human in need thereof a therapeutically effective amount of the pharmaceutical composition of claim 13 .
16. A method of preparing the pharmaceutical composition of claim 13 comprising mixing the isolated protein with a pharmaceutically acceptable adjuvant.
17. An isolated protein having factor VIII procoagulant activity, wherein the amino acid sequence of the protein is at least 95% identical to SEQ ID NO: 1 and the protein further comprises a N-linked oligosaccharide comprising a bisecting GlcNAc linked to a core beta-mannose.
18. The isolated protein of claim 17 characterized by having the N-linked oligosaccharide further comprising an alpha-(2,6)-linked sialic acid.
19. A pharmaceutical composition comprising the isolated protein of claim 17 and a pharmaceutically acceptable adjuvant.
20. The pharmaceutical composition of claim 19 , wherein the composition does not contain a toxic solubilizing agent.
21. A method of treating hemophilia comprising administering to a human in need thereof a therapeutically effective amount of the pharmaceutical composition of claim 19 .
22. A method of preparing the pharmaceutical composition of claim 19 comprising mixing the isolated protein with a pharmaceutically acceptable adjuvant.
23. An isolated protein having Factor VIII procoagulant activity, wherein the amino acid sequence of the protein and the amino acid sequence of SEQ ID NO: 1 have at least 95% identity and the protein comprises an N-linked oligosaccharide comprising alpha-(2,6)-linked sialic acid, the isolated protein characterized in that at least 75% of the total protein matter present is protein having factor VIII procoagulant activity.
24. The isolated protein of claim 23 characterized in that at least 90% by weight of the total protein matter present is protein having factor VIII procoagulant activity.
25. The isolated protein of claim 8 characterized in that it comprises a heterogeneous population of heterodimers.
26. The isolated protein of claim 17 charaterized in that at least 90% by weight of the total protein matter present is protein having factor VIII procoagulant activity.
27. The isolated protein of claim 17 characterized in that it comprises a heterogeneous population of heterodimers.
28. The isolated protein of claim 12 wherein the protein has the amino acid sequence of SEQ ID NO: 1.
29. The isolated protein of claim 17 wherein the protein has the amino acid sequence of SEQ ID NO: 1.