Hemophilia B is an X-linked bleeding disorder that results from a defect in the gene encoding coagulation factor IX (FIX), a serine protease that is critical for blood clotting. Persons with severe hemophilia B have functional FIX levels that are less than 1% of normal values and have frequent bleeding episodes, which are associated with crippling arthropathy and early death.[1,2] Current treatment involves frequent intravenous injections of FIX protein concentrate (i.e., two to three times a week). However, this treatment is prophylactic rather than curative, is extremely expensive, and is associated with inhibitor formation. Somatic gene therapy for hemophilia B offers the potential for a cure through continuous endogenous production of FIX after a single administration of vector, especially since a small rise in circulating FIX to at least 1% of normal levels can substantially ameliorate the bleeding phenotype.
At present, gene transfer mediated by an adenovirus-associated virus (AAV) vector shows the greatest promise for long-term correction of hemophilia B in the preclinical setting.[3-6] However, a combined phase 1 and 2 study that involved serotype 2-based AAV vectors (AAV2) showed only transient expression of FIX and suggested that stable expression of therapeutic levels of FIX may be limited by a capsid-specific cytotoxic T-cell response against the transduced hepatocytes.
Patients who met the entry criteria and did not have neutralizing antibodies to AAV8, as determined by an in vivo transduction-inhibition assay (see Table 1 and the Methods section in the Supplementary Appendix, available with the full text of this article at NEJM.org), were enrolled after providing written informed consent. Participants 1 through 5 were recruited in 2010 and Participant 6 was recruited early in 2011. All were admitted to the Royal Free Hospital for vector administration and subsequent observation overnight. Participants were enrolled sequentially into one of three cohorts according to dose level: low (2^1011 vector genomes [vg] per kilogram of body weight), intermediate (6^1011 vg per kilogram), and high (2*1012 vg per kilogram).
The study was sponsored by St. Jude Children's Research Hospital and was conducted independently by the authors in accordance with the protocol, which is available at NEJM.org, and with the provisions of the Good Clinical Practice guidelines.
Characteristics of the Study Participants
Six men with severe hemophilia B (FIX activity, <1% of normal values) were enrolled under a protocol approved by the relevant ethics boards and regulatory agencies, after providing written informed consent. All except one participant received regular prophylaxis with FIX concentrates (two or three times per week) before gene transfer. Participant 4 was receiving targeted prophylaxis (about once weekly), which was tailored to his sports activities and the associated risk of traumatic injury. Participant 2 had a null mutation in the FIX gene, and Participant 6 had a promoter mutation; consequently, neither had FIX protein expression. The other four participants had missense mutations, which resulted in normal plasma levels of FIX antigen but less than 1% clotting activity. All participants had modest levels (>5 relative units) of anti-AAV2 IgG antibodies before gene transfer.[2-4]
In summary, we have found that a single peripheral-vein infusion of our scAAV2/8-LP1-hFIXco vector consistently leads to long-term expression of the FIX transgene at therapeutic levels, without acute or long-lasting toxicity in patients with severe hemophilia B. Immune-mediated, AAV-capsid-induced elevations in aminotransferase levels remain a concern, but our data suggest that this process may be controlled by a short course of glucocorticoids, without loss of transgene expression. Follow-up of larger numbers of patients for longer periods of time is necessary to fully define the benefits and risks and to optimize dosing. However, this gene-therapy approach, even with the associated risk of transient hepatic dysfunction, has the potential to convert the severe bleeding phenotype into a mild form of the disease or to reverse it entirely.[1-6]
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- Macfarlane R.G, Barnett B: The haemostatic possibilities of snake-venom. Lancet 1934;2:985-987.