Adeno-associated virus-mediated expression of activated factor V (FVa) for hemophilia phenotypic correction.

Adeno-associated virus (AAV) gene therapy has been successfully applied in hemophilia patients excluding patients with inhibitors. During the coagulation pathway, activated factor V (FVa) functions downstream as a cofactor of activated factor X (FXa) to amplify thrombin generation. We hypothesize that the expression of FVa via gene therapy can improve hemostasis of both factor IX and FVIII deficiencies, regardless of clotting factor inhibitor. A human FVa (hFVa) expression cassette was constructed, and AAV8 vectors encoding hFVa (AAV8/TTR-hFVa) were intravenously administrated into mice with hemophilia A and B with or without FVIII inhibitors. Hemostasis, including hFVa level, activated partial thromboplastin time (aPTT), tail clip, and the saphenous vein bleeding assay (SVBA), was evaluated. In hemophilia B mice, a dose of 4 × 1013 vg/kg AAV8/TTR-hFVa vectors achieved a complete phenotypic correction over 28 weeks. In hemophilia A mice, hemostasis improvement was also achieved, regardless of FVIII inhibitor development. In vivo hemostasis efficacy was confirmed by tail clip and SVBA. Interestingly, while minimal shortening of aPTT was observed at a lower dose of AAV8 vectors, hemostasis improvement was still achieved via in vivo bleeding assays. Collectively, FVa-based AAV gene therapy shows promise for hemostasis correction in hemophilia, regardless of inhibitor development and no potential risk for thrombosis.