Hemophilia A (HA) is a common disorder of coagulation caused by deficiency of factor VIII (FVIII). The mainstay of therapy has been replacement therapy with recombinant human FVIII. However, because of high cost, access to therapy, and inhibitory antibody formation, patients continue to suffer from significant long-term morbidity and mortality. The generation of improved versions of FVIII molecules with increased specific activity and/or improved immunological profile would improve hemophilia treatment. The expression of these new variants with liver-directed gene therapy in conjunction with strategies to tolerize patients to the therapeutic protein would constitute important clinical advances. Our development of an optimized helper-dependent adenoviral gene vector (HDV) system has enabled us to achieve long term expression of both secreted and intracellular transgenes with absent chronic toxcity and persistence of vector in small and large animal models. However, acute toxicity remains an obstacle to clinical translation. To overcome this, we have developed a novel balloon catheter based delivery system into the hepatic circulation. We have also developed a novel method for generation of tolerogenic dendritic cells based on calcium phosphate-dependent transduction of HDV expressing TGF2 and IL10. Based on these preliminary data, we will address three important questions in genetic therapy for hemophilia a) Can we increase the therapeutic index by maximizing gene expression of more bioactive FVIII variants? b) Can we increase the maximal toxic dose by overcoming the threshold/nonlinear dose response of Ad gene transfer in canine HA and achieve re-administration of the vector? c) Can we tolerize the recipient of gene transfer or h FVIII by adoptive transfer of autologous tolerogenic DCs? The overall goal of these studies is to address the major obstacle to clinical translation of adenoviral gene therapy in HA: 1) To achieve clinical correction in a large animal model of HA at a dose relevant for human translation and 2) to develop an antigen-specific tolerizing strategy for FVIII expression after gene transfer.

Public Health Relevance

Hemophilia A is a genetic bleeding disorder. Current therapy includes intravenous injections of factor VIII protein. However, patients continue to suffer long term complications in part due to the development of antibodies to this treatment. In this application, we propose to develop a novel form of gene therapy that combines the use of a less toxic helper-dependent adenovirus vector, a more bioactive variant of human factor VIII, and a catheter based delivery system that minimizes the required dose of vector. First, we will determine the best form of human factor VIII to use in a mouse model of hemophilia. Then we will test this variant in the combined approach described above in a dog model of hemophilia. If successful in treating the dog model of hemophilia, we will then apply established methods to enable our repeating this treatment so as to achieve long-term correction. To address the problem of patients rejecting the delivered forms of human factor VIII either from gene therapy or from protein therapy now in use, we will develop a new cell based method to stimulate tolerance to factor VIII. In this approach, we take a special group of immune cells from the hemophilia mouse and treat them so as to make them tolerogenic, i.e., to prevent the rejection phenomenon. These cells can then be injected back into the mouse to prevent the rejection of future factor VIII treatments. In so doing, we will develop methods and approaches that will be broadly applicable to other genetic deficiencies.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Research Project (R01)
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Gene and Drug Delivery Systems Study Section (GDD)
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Link, Rebecca P
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Baylor College of Medicine
Schools of Medicine
United States
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