We propose an open label, dose-escalation, phase I/II study in which a single dose of the novel self- complementary AAV vector, scAAV2/8 LP1 hFIXco, will be administered into a peripheral vein of adult subjects with severe hemophilia B. Hemophilia B (HB) is an X-linked recessive bleeding disorder that results from a defect in the factor IX (FIX) gene which encodes a serine protease critical for appropriate fibrin clot formation. Clinically the disease is characterized by frequent spontaneous bleeding, most commonly into such sites as joints and soft tissues, but which can also occur in the brain and potentially be life threatening. Our extensive studies in murine models and rhesus macaques have shown that scAAV2/8 LP1 hFIXco vector establishes therapeutic levels of FIX at relatively low doses in these animals, compared to vectors evaluated in previous clinical trials. This study differs from previous HB clinical trials with AAV vectors in three important aspects. Firstly, an AAV8 pseudotyped vector will be used instead of AAV2, primarily because of the substantially lower prevalence of pre-existing immunity to this AAV serotype in humans. The second difference relates to the use of a vector containing a self-complementary genome which, because of its ability to rapidly form stable, transcriptionally active, double stranded linear molecules in target tissues, offers the unique opportunity to mediate efficient therapeutic gene transfer potentially at lower doses of vector. Finally, because the biodistribution of vector is predominantly to the liver regardless of the route of administration, scAAV particles will be administered via a peripheral vein. We propose to test three dose levels: 2x1010, 6x1010 and 2x1011 vector genomes per kilogram body weight. 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 hFIX at or above 3% of normal;2) to describe the immune responses to the hFIX transgene product and AAV capsid proteins and 3) to access viral shedding into various body fluids. Recruitment will be limited to adults (greater than or equal to 18 years of age) with a confirmed diagnosis of severe HB resulting from a missense mutation in the hFIX gene. Patients will be observed for at least 42 days following vector administration before enrollment of the next patient. Dose escalation will proceed based on safety (primary) and efficacy (secondary) criteria. Immunosupression will not be given routinely;rather, only if a patient develops acute, significant transaminitis (ALT or bilirubin >10X normal) or persistent (>16 weeks) ALT or bilirubin elevation. The occurrence of any of the following will result in immediate suspension of the trial: 1) The occurrence of Grade IV toxicity in one patient or Grade III toxicity in two patients at a given dose level. 2) The development of neutralizing antibodies to FIX following gene transfer in one subject. 5) Death of a patient at any time point after gene transfer that is possibly, probably, or definitely related to the study agent. The inadequacies of current therapy for hemophilia B have fuelled interest in alternative treatment approaches, including gene transfer. Successful AAV-mediated gene transfer for hemophilia B 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 disorders affecting the liver, such as lysosomal storage and urea cycle disorders, are targeted.
