Abstract
Although the extracellular association of Factor VIII (FVIII) and Von Willebrand Factor (VWF) is well established, the intracellular interaction of FVIII and VWF is not well understood. Recently, the importance of intracellular co-localization of FVIII and VWF for in vitro FVIII secretion was demonstrated in endothelial cell lines. Whether intracellular co-localization of FVIII and VWF is required for in vivo FVIII secretion, however, is not known. We previously showed that liver transplantation leads to phenotypic cure of hemophilia A, by virtue of FVIII production in the allograft liver. Because FVIII is synthesized only in the allograft liver but not in endothelial cells of transplant recipients, and VWF is synthesized in extrahepatic tissue, this is an ideal model to study whether co-localization of FVIII and VWF is required for in vivo FVIII secretion. We, therefore, studied FVIII and VWF response after desmopression (DDAVP) infusion, administered at 0.3 mcg/kg by intravenous infusion over 30 minutes, in each of two men with severe hemophilia A (FVIII:C <0.01 U/ml) who had undergone orthotopic liver transplantation for endstage liver disease six months earlier. Both men had HIV and hepatitis C co-infection and were clinically well, with mildly elevated liver function tests, and FVIII:C levels >30% following transplantation. Coagulation studies, drawn before and after DDAVP, revealed that VWF:RCoF and VWF:Ag, but not FVIII:C, increased after DDAVP administration (see Table). The prolonged aPTT and correction in a 1:1 aPTT mix confirmed the absence of an inhibitor in these subjects. The lack of FVIII response to DDAVP supports previous in vitro work, and demonstrates for the first time that intracellular co-localization of FVIII and VWF is essential for in vivo FVIII secretion. These data also suggest that extrahepatic FVIII synthesis is necessary for in vivo response of the DDAVP releasable pool of FVIII. By contrast, co-localization does not appear to be necessary for VWF secretion. Although it is not possible to exclude that a chronic, exhaustive post-transplant increase in VWF may have limited VWF response to DDAVP, it is clear that FVIII did not increase following DDAVP. These findings have important implications for the design of gene therapies for hemophilia A and Von Willebrand Disease.
Subject Demographic Sample aPTT aPTT mix FVIII:C VWF:RCoF VWF:Ag 01-BW 32yoM Hem A Pre-DDAVP 44.4 sec 37.7 sec 0.50 U/ml 2.17 U/ml 2.42 U/ml HIV+/HCV+ Post-DDAVP 44.8 sec 37.4 sec 0.48 U/ml 2.91 U/ml 2.91 U/ml 02-PB 36yoM Hem A Pre-DDAVP 49.5 sec 38.0 sec 0.32 U/ml 1.61 U/ml 2.16 U/ml HIV+/HCV+ Post-DDAVP 50.8 sec 38.5 sec 0.30 U/ml 2.20 U/ml 2.50 U/ml
In Vivo Characterization of Recombinant von Willebrand Factor in Dogs With von Willebrand Disease