Our ultimate aim is the use of autotransplantation in combination with gene transfer for the cure of patients with homozygous Beta thalassemia and sickle cell anemia. The specific goal is to define the optimal conditions for the transfer and expression of human Beta globin genes into bone marrow cells. Studies to date have indicated that retroviral vectors can be used to efficiently transfer exogenous genes into mouse bone marrow cells. In addition, it has been shown that it is possible to express human Beta globin genes in erythroid cells in a tissue-specific manner. While other methods for treating patients with Beta thalassemia and sickle cell anemia exist, the potential use of gene transfer to treat these disorders is particularly exciting since it could provide a cure by appropriate expression of the normal Beta globin gene. In our initial studies, retroviral vectors containing the human Beta globin and the neomycin resistance genes will be used to transfer human Beta globin genes into mouse bone marrow cells, and expression assessed after transplantation into irradiated mice. These experiments will also test whether enhancer sequences derived from cloned DNA segments from within the Beta globin gene complex as well as from other sources can increase Beta globin expression. Amplification of integrated human Beta globin genes will be another method for attempting to increase Beta globin production. A vector containing a dihydrofolate reductase (DHFR) gene will be used for this purpose. A mutant mouse strain with Beta thalassemia as well as normal mice will be used to assess the expression of human Beta globin genes in intact mice. The anemia in these Beta thalassemic mice has already been corrected by a human Beta globin gene inserted transgenically in mouse embryos. In further experiments, when safe and appropriate amphotropic retroviral vectors become available, we plan to introduce the human Beta globin gene into monkey bone marrow cells. These experiments in primates will hopefully be the final step toward demonstrating the feasibility of effective gene therapy of human patients with Beta thalassemia and sickle cell anemia.
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