Background: Hemophilia A is a congenital bleeding disorder caused by defective or deficient factor VIII (FVIII). The active form of FVIII is the co-factor for the serine protease factor IXa (FIXa) in the membrane-bound intrinsic tenase (FVIIIa-FIXa) complex. The assembly of the FVIIIa-FIXa complex on the activated platelet surface is critical for successful blood clotting.
Objectives: To characterize the role of lipid nanodiscs (ND) for on FVIII function in vivo and test the lipid ND as a delivery system for FVIII. To evaluate the potential of binding recombinant FVIII to ND as improved treatment for Hemophilia A.
Methods: Recombinant porcine FVIII (rpFVIII) was expressed and characterized in solution, and when bound to ND. The rpFVIII, ND and rpFVIII-ND complexes were characterized via transmission electron microscopy. Functional studies were carried out using aPTT tests and time resolved tail snip studies of hemophilic mice.
Results: Functional rpFVIII was successfully assembled on lipid ND. When injected in hemophilic mice, the rpFVIII-ND complexes showed a pronounced pro-coagulant effect, which was stronger than that of rpFVIII alone. While injection of the ND alone showed a pro-coagulant effect this effect was not additive, implying that the rpFVIII-ND complexes have a synergistic effect on the clotting process in hemophilic mice.
Conclusions: Binding of rpFVIII to ND prior to its injection in hemophilic mice significantly improves the therapeutic function of the protein. This represents a meaningful step towards a new approach to modulate blood coagulation at the membrane-bound FVIII level and the assembly of the intrinsic tenase complex.
Keywords: Drug delivery; FVIII deficient mice; Factor VIII; Hemophilia A; Lipid nanodiscs.