Long-Term Genetic Modification of Rhesus Monkey Hematopoietic Cells Following Transplantation of Adenoassociated Virus Vector-Transduced CD34+Cells

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Abstract
We have explored the potential of recombinant adenoassociated virus (AAV) vectors for gene transfer of the human β-globin gene and the genetic modification of primate pluripotent hematopoietic stem cells (P-PHSCs). Transduction of P-PHSCs was tested in a preclinical bone marrow transplantation model in rhesus monkeys. CD34+ cells were transduced ex vivo and autologously transplanted without prior selection into irradiated rhesus monkeys. Vector-transduced peripheral blood mononuclear cells and granulocytes were present in the circulation for more than 15 months after transplantation. Approximately 1 in 105 cells in the circulation was vector modified. The vector was detected in the bone marrow, in granulocytes, and in purified populations of B and T cells, thus demonstrating multilineage repopulation by vector-transduced stem cells. Comparison of transduction protocols suggested that short-term culture of P-PHSCs enhances transduction and subsequent repopulation by rAAV-transduced cells. These results demonstrate that rAAV vectors can be used for the permanent genetic modification of a rhesus monkey hematopoietic system in the absence of selective pressure. Recombinant adenoassociated virus (rAAV) vectors are being evaluated for gene transfer into hemopoietic stem cells. In vitro data suggest that rAAV vectors efficiently transduce hematopoietic progenitor cells. In this study, rAAV-transduced CD34+ cells were grafted into irradiated autologous rhesus monkeys. Recombinant AAV vector-containing hematopoietic cells were shown to persist in the circulation of the rhesus monkey for more than 15 months following transplantation. These results show that a strategy that employs rAAV-mediated gene transfer of hematopoietic stem cells holds promise for the permanent genetic modification of the hematopoietic system of primates.