In recent clinical trials, gene therapy has had remarkable success in achieving long-term production of therapeutic proteins for the treatment of various genetic disorders, including hemophilia. However, pre-existing immunity to some formulations of gene delivery and the potential for oncogenesis may limit widespread adoption of current and emerging approaches. Chimeric antigen receptor (CAR) therapy has shown that human T cells can be isolated from peripheral blood, modified to express a transgene encoding a CAR targeted to a specific cell surface protein, and then transferred back to patients where they can safely eliminate tumor cells bearing the targeted protein and can persist for long periods of time. Based on the success of CAR T cells, we reasoned that genetically-modified T cells might also serve as a cellular vehicle for protein replacement. Here, we show that T cells can be modified with a lentivirus to secrete functional Factor IX, Factor VIII, and α-galactosidase in vitro. In immune-deficient mice, we also demonstrate that adoptive transfer of Factor IX-secreting human T cells mediates significant levels of Factor IX in vivo (20-40% of normal in NSG mice and 3-8% of normal in NSG-B2m mice) for an extended period. Transfer of Factor IX-secreting mouse T cells to C57BL/6 mice resulted in transient levels of circulating FIX protein that peaked at 2-20% of normal. Additionally, we generated CRISPR-Cas9 guide RNA and a homologous recombination repair template for integration of a Factor IX transgene into the CCR5 locus of human T cells, which can further serve to increase the safety profile of T cell gene therapy. Overall, these studies suggest that T cells can serve as a vehicle for gene therapy.
|Commitee:||Sharp, Andrew, Homann, Dirk, Brody, Joshua, Papapetrou, Eirini|
|School:||Icahn School of Medicine at Mount Sinai|
|School Location:||United States -- New York|
|Source:||DAI-B 81/2(E), Dissertation Abstracts International|
|Subjects:||Genetics, Molecular biology|
|Keywords:||Gene therapy, Hemophilia, T cell therapy|
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