Stable Expression of Recombinant Factor VIII Molecules Using a Bovine Papillomavirus Vector

Abstract

The bleeding disorder in hemophilia A results from a deficiency or abnormality of Factor VIII (FVIII), a member of the coagulation cascade. FVIII is a large glycoprotein (∼350,000 daltons) that is activated by a series of proteolytic cleavages. During activation, a large internal domain (B domain) is removed, resulting in an active complex comprised of the amino and carboxyl subunits of the parental molecule. Using a bovine papillomavirus expression vector system, we have established stable, genetically engineered cell lines harboring either full-length FVIII cDNA or variant FVIII cDNA (ΔFVIII), the latter containing an extensive deletion in the region encoding the B domain. We demonstrate that the two recombinant FVIII molecules manifest the biological attributes of native FVIII. Relative to full-length FVIII transformants, cells harboring ΔFVIII cDNA are five to eight times more efficient in expressing coagulant activity. This difference is due to a post-transcriptional event.