We propose an open label, dose-escalation, phase I/II study in which a single dose of the novel recombinant AAV vector, AAV2/8 HLP FVIII-V3co, will be administered into a peripheral vein of adult subjects with severe hemophilia A . Our premise is that this will be a safe and efficacious approach for limiting the bleeding propensity of these patients . Hemophilia A (HA) is an X-linked recessive bleeding disorder that results from a defect in the factor VIII (FVIII) gene. FVIII encodes a glycoprotein procofactor which, when activated by thrombin, interacts with factor IXa in the coagulation cascade, leading to clot formation. Clinically, the disease is characterized by frequent spontaneous bleeding, which can be life-threatening. There are several novel aspects to our approach to liver-targeted AAV-mediated FVIII gene transfer, including 1) a unique FVIII cDNA that meets the size constraints of AAV and mediates synthesis of fully functional FVIII protein, 2) an expression cassette that improves the efficiency of FVIII expression and secretion and 3) an improved method of recombinant AAV vector production. Our extensive studies in murine models and rhesus macaques have shown that AAV2/8 HLP FVIII-V3co vector safely establishes therapeutic levels of FVIII in these animals. In addition, we now have the unique experience of having conducted a successful phase I/II clinical trial of liver-targeted, AAV-mediated FIX gene transfer for hemophilia B that has provided new insights to the design of our AAV-mediated FVIII gene transfer trial. We propose to test three dose levels: 6x1011, 2x1012 and 6x1012 vg/kg. The primary objective of the study is to assess the safety of systemic administration of this vector while the secondary objectives are: 1) to determine the dose of vector particles required to achieve stable expression of FVIII ?5% of normal; 2) to explore the impact on quality of life, emotional health and neurocognitive function; 3) to describe the immune responses to the FVIII transgene product and AAV capsid proteins and 4) to assess viral shedding into various body fluids. Recruitment will be limited to adults (?18 years of age) with a confirmed diagnosis of severe HA caused by a mutation in the FVIII gene not associated with a high incidence of inhibitor formation. In addition, participants must have had ?50 exposure days of treatment with FVIII protein concentrate without having developed an inhibitor. Dose escalation will proceed based on safety (primary) and efficacy (secondary) criteria. Immunosuppression will not be given routinely to patients receiving the lowest dose of vector; rather, only for those low dose patients who develop ALT elevation >1.5-times baseline. However, based on our AAV2/8 FIX experience, all patients in the intermediate and high dose cohorts will receive prophylactic steroids from weeks 6-13. Stopping rules are in place which include: 1) the occurrence of Grade IV or V toxicity in one patient or Grade III toxicity in two patients at a given dose level or 2) the development of neutralizing antibodies to FVIII in one patient.
The inadequacies of current therapy for hemophilia A have fuelled interest in alternative treatment approaches, including gene transfer. Successful AAV-mediated gene transfer for hemophilia A offers the potential of effective lifetime prevention of bleeding and its complications for patients with this disorder. In addition, the information gained from this study will be important in planning additional future strategies with AAV vectors in which other diseases affecting the liver, such as lysosomal storage and urea cycle disorders, are targeted.!