159. Vector Genome Loss Does Not Account for Decline of Plasma cFVIII Protein in Hemophilic Dogs That Demonstrate Long Term Clinical Correction Following Helper-Dependent Adenoviral Gene Therapy

Helper-dependent adenoviral vectors (HDV) offer the advantage of a large cloning capacity, reduced toxicity and extremely efficient transduction of target tissues. These vectors remain episomal in host target cells limiting the possibility of insertional mutagenesis but potentially reducing the duration of transgene expression due to vector genome loss. In previous studies using these vectors in various animal models of disease, long-term expression of both intracellular and secreted proteins has been achieved. However, in canine models of both hemophilia A and B, this has not been the case. The level of canine factor VIII (cFVIII) transgene product in the plasma of hemophilic dogs treated with HDV gene therapy declines at a greater rate than would be expected based on data from other models of disease including hemophilic mice. In these experiments we treated two hemophilia A dogs from the colony at the University of North Carolina with a HDV encoding cFVIII under the control of a liver tissue-restricted phosphoenolpyruvate carboxykinase (PEPCK) promoter. One animal received 1 × 10 12 vp/kg and a second received 3 × 1012 vp/kg. The whole blood clotting time (WBCT) in both of the animals was significantly improved for up to 10 months. However, the plasma level of cFVIII declined from peaks at two weeks post-treatment of 1.3% normal in the low dose animal and 70% normal in the high dose animal to less than 1% at 2 months in both. To determine the reason for this a third animal was injected with 3 × 10 12 vp/kg and underwent survival liver biopsy at days 18 and 79 after vector administration correlating with the high (35% normal) and low (