Gene Therapy for Blood Protein Deficiencies

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Hemophilia B occurs in 1:30,000 males and is associated with a life-long bleeding diathesis. Although IV injection of Factor IX can prevent or stop bleeding, this treatment is inconvenient, expensive, and can transmit infections. Hepatic gene therapy could permanently correct the clinical manifestations of hemophilia. Retroviral vectors (RV) can result in long-term and therapeutic levels of expression of coagulation factors from the liver in rodents, and are currently being used in a clinical trial for Hemophilia A in humans. However, there are two major problems that must be solved before RV-mediated hepatic gene therapy will be used routinely: 1) identify ways to achieve a higher efficiency of stable gene transfer without major toxicity; and 2) identify methods for blocking an immune response to the therapeutic gene in the context of RV-mediated hepatic gene therapy. This project will address both of these issues. The first aim is to determine if delivery of an RV expressing the canine Factor IX (cFIX) cDNA into the liver can reduce the bleeding manifestations of Hemophilia B dogs obtained from a colony that rarely makes antibodies to the canine protein. This should allow us to quantify gene expression without the confounding issue of an immune response. Initial studies will use neonatal dogs, as their high baseline level of hepatocyte replication allows transduction of 9 percent of liver cells. Subsequent studies will use hepatocyte growth factor to induce replication in young adult dogs. Animals will be evaluated for cFIX levels, development of antibodies, bleeding, and for other adverse effects. The second aim will address the second major problem of RV-mediated hepatic gene therapy, that of immune responses to the therapeutic gene product. In this aim, we will try to block immune responses to the de novo expression of a transgene from an RV in mice by either performing neonatal gene transfer, or by injecting immunoinhibitory agents at the time of gene therapy in young adults. Although mice are optimal for initial studies due to cost considerations, approaches that function in inbred mice sometimes fail in outbred larger animals. We will therefore test any immunomodulatory approaches that function in mice for their efficacy in normal and Hemophilia B dogs in Aim III. Success in this project might lead to a safe, effective, and permanent therapy for Hemophilia B.