Treatment of canine hemophilia A via intraosseous delivery of a platelet-specific factor VIII-lentiviral vector.

Hemophilia A (HemA) is a genetic disease resulting from a factor VIII (FVIII) deficiency. Traditional protein infusions to treat HemA are costly and inconvenient and require repeated dosing. We demonstrated previously that intraosseous (IO) gene therapy via delivery of lentiviral vectors (LVs) into bone marrow targeting FVIII expression in platelets successfully treated HemA mice. To facilitate the translation of this novel strategy to clinical application, we investigated the treatment of HemA dogs using IO gene therapy. The vesicular stomatitis virus-pseudotyped G-cF8-LV incorporating a platelet-specific promoter, Gp1bα, and canine F8 gene was injected into the tibia or iliac bones of 4 HemA dogs. All dogs recovered well from the procedure and had blood chemistry values within normal ranges. Canine FVIII can be detected in platelets with the highest expression ∼40 to 50 days after the procedure, and the expression persisted for the experimental duration in all treated dogs. Copy numbers of the transgene were persistently detected from the genomic DNA isolated from peripheral mononuclear blood cells. The shortened whole-blood clotting time and improved parameters evaluated through thromboelastography testing at multiple time points indicated improved hemostasis after gene therapy. Furthermore, the IO gene therapy was well tolerated and did not produce any toxicity in the treated dogs. Most significantly, the treated dogs experienced fewer bleeding events per year after gene therapy than before. Our study demonstrated a potentially safe and effective in vivo gene therapy strategy for treating people with HemA.