The goal of the proposed research is to apply techniques of gene transfer to the treatment of diseases affecting the hematopoietic system in humans. While gene transfer rates in the mouse hematopoietic system can approach 100%, gene transfer rates in large animals and humans have been disappointingly low. We will explore methods to improve gene transduction rates in long-term repopulating hematopoietic cells (stem cells) in the canine model. We will study the optimal exposure time of hematopoietic stem cells to retrovirus vectors, the optimal source of stem cells, and less toxic conditioning regimens or no conditioning with repeated infusions of transduced hematopoietic stem cells. New vectors and retrovirus packaging cell lines will be generated and tested for improved gene transfer and expression. The canine alpha-L-iduronidase-deficient genetic disease model will be used to test our ability to correct a disease that affects hematopoietic cells in the dog. We will continue our current clinical protocol to treat Gaucher disease by transduction of G-CSF- mobilized peripheral blood hematopoietic cells. The focus of this experiment is to examine the persistence and expression of vector sequences following transplantation. Techniques developed in the canine model for improved gene transfer and expression will provide the basis for improvements in human gene therapy experiments. Finally, we have discovered a new retrovirus during testing of human hematopoietic cells for the absence of helper virus following transduction with retroviral vectors. The new virus appears to be of human origin, and we will perform experiments to definitively establish its origin. If indeed this is a human virus, we will explore implications of the virus for ongoing gene therapy involving retrovirus vectors, and its possible role in human disease.
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