227. A Dose-Escalating Preclinical Study to Determine the Efficacy, MED, and Safety of a Clinical Candidate Vector in a Mouse Model of Hemophilia B

Abstract

Despite the availability of safe and effective recombinant FIX therapeutics, patients with Hemophilia B could obtain additional clinical benefit by having longer-acting therapeutic options. Although longer-acting recombinant protein FIX therapies are becoming available, the ultimate long-acting FIX therapy could be provided by a gene therapy approach, for example with rAAV vectors encoding FIX. Promising clinical data has been generated over the last few years in an academic setting, but these results must be replicated and extended in a commercial setting for these new therapies to become broadly available to patients.Dose-dependent inflammatory responses that impact the transgene expression have been reported in clinical trials with rAAV FIX vectors, highlighting the need for clinical rAAV vectors to have the highest possible potency such that the overall vector dose can be reduced. We have developed a potent AAV serotype rh10 vector containing a wild-type FIX gene under the control of a highly active liver specific promoter with the goal of achieving approximately 10% of normal FIX levels in patients.In preparation for a clinical trial, we tested this rAAVrh10FIX vector for aspects of efficacy and safety in the factor IX knockout (FIX-KO) mouse model of Hemophilia B. A dose-dependent increase in FIX protein and activity was observed following intravenous administration at vector doses between 1.6×1010 and 5.0×1013 GC/kg. FIX-KO mice that received AAVrh10-hFIXco at 1.6 × 1010 GC/kg achieved between 5% and 8% of normal hFIX levels at 2 weeks post-dose and maintained this level for the 90-day study period. FIX-KO mice that received rAAVrh10FIX vector at 5.0 × 1010 GC/kg achieved between 30% and 42% of normal hFIX levels at 14 days post-dose and maintained this level for the 90-day study period. From these results we can estimate that the vector dose required to achieve the stated goal of 10% of normal FIX levels of 10% would be between 1.6 × 1010 GC/kg and 5.0 × 1010 GC/kg. Expression of hFIX antigen correlated well with measured activity of hFIX antigen, with doses between 1.6 × 1010 GC/kg and 5.0 × 1010 GC/kg providing between 8% and 35% activity of normal hFIX levels.FIX-KO mice provide an excellent model to examine the effects of AAVrh10-hFIXco on hemophilia B-related complications. FIX KO mice are prone to spontaneous bleeds and often suffer from fatal hemorrhages after handling or trauma. In this study, 4 out of 7 animals in the Day 90 control group succumbed to hemophilia-related complications. Single dose intravenous administration of AAVrh10-hFIXco at doses of 1.6 × 1010, 5.0 × 1010, 1.6 × 1011, 5.0 × 1011, 5.0 × 1012, and 5.0 × 1013 GC/kg to FIX-KO mice did not result in test article-related mortality, clinical pathology, gross pathology, or histopathologic findings.