A new recombinant procoagulant protein derived from the cDNA encoding human factor VIII

We have constructed new B domain deletion derivatives of human factor Vm (FVm) by manipulating the cDNA using recombinant DNA techniques. One of these new derivatives, FVIII ΔII, in which amino acids 771(pro)–1666(asp) have been deleted, no longer contains the protease cleavage site at amino acid position 1648(arg) –1649(glu) known to be involved in the initial step of FVin processing. We have expressed this molecule in both baby hamster kidney (BHK) 21 cells using the vaccinia virus (VV) expression system and have established Chinese hamster ovary (CHO) derived permanent cell lines expressing either recombinant (r)FVIII or FVIII ΔII AD. The characteristics of FVIII ΔII AD have been compared to those of rFVIII and/or plasma derived (pd) FVIII. FVIII ΔII All has the following properties: (i) it exhibits FVDI procoagulant activity; (ii) it is expressed at 5-fold higher levels than is the complete molecule in comparable systems; (iii) it migrates for the most part as a single major band on SDS-PAGE, hi contrast to the complete molecule; (iv) it is activated to a greater extent by thrombin than is either rFVm or pdFVIII; and (v) it retains the ability to bind von Willebrand factor (vWf).