Helper‐dependent adenoviral gene therapy mediates long‐term correction of the clotting defect in the canine hemophilia A model

Background: Adenoviral vector‐mediated gene therapy might have potential for long‐term correction of the monogenic disease hemophilia A. Objective: In this study, we tested the efficacy of administering a helper‐dependent adenoviral vector (HDV) designed for maximal liver‐restricted canine factor VIII (cFVIII) expression on three out‐bred hemophilia A dogs. Methods: Three FVIII‐deficient animals from the University of North Carolina colony were injected with 1 × 1012 (Dog A), and 3 × 1012 (Dog B and C) vp kg−1 helper‐dependent adenoviral vector, and we performed systematic analysis of toxicity, persistence of therapeutic gene expression, and molecular analysis of gene transfer. Results: We observed acute dose‐dependent elevation in liver enzymes and thrombocytopenia after injection, although both were transient and resolved within 2 weeks. The whole blood clotting time (WBCT), plasma FVIII concentration, FVIII activity, and activated partial thromboplastin time in all animals improved significantly after treatment, and two animals receiving a higher dose reached near normal WBCT with low‐level FVIII activity until terminal sacrifice at 3 months, and 2 years. Importantly, the treated dogs suffered no bleeding events after injection. Moreover, we observed persistent vector‐specific DNA and RNA in liver tissue collected from one high‐dose animal at days 18 and 79, and could not detect the formation of inhibitory antibodies. Conclusion: Although vector‐associated toxicity remains an obstacle, a single injection of HDV led to long‐term transgene expression and vector persistence in two FVIII‐deficient animals with conversion of their severe phenotype to a moderate one.