Correction of Purine Nucleoside Phosphorylase Deficiency by Retroviral-Mediated Gene Transfer in Mouse S49 T Cell Lymphoma: A Model for Gene Therapy of T Cell Immunodeficiency

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Abstract
To determine the effectiveness of retroviral-mediated purine nucleoside phosphorylase (PNP) gene transfer and expression for metabolic correction of PNP deficiency, we used as a gene transfer target the NSU-1 subline of murine S49 T lymphoma cells, an in vitro genetic model of PNP deficiency. NSU-1 cells were transduced with recombinant retroviruses that express either the murine or human PNP coding sequences under transcriptional regulation of the Moloney murine leukemia virus (Mo-MLV) long terminal repeat (LTR), resulting in expression of substantial levels of PNP activity. Untransduced or control virus-transduced NSU-1 cells were extremely sensitive to deoxyguanosine, a PNP substrate that is toxic for lymphoid cells. However, PNP-virus transduction of NSU-1 cells metabolically corrected the sensitivity of these cells to deoxyguanosine, resulting in near wild-type levels of growth inhibition. These results demonstrate that retroviral-mediated PNP gene transfer and expression corrects the metabolic defect observed in PNP-deficient murine lymphoid cells, suggesting that PNP gene transfer and expression in human lymphoid cells might similarly correct substrate-mediated toxicity and provide an effective genetic therapy. Absence of purine nucleoside phosphorylase (PNP) in humans is associated with T cell immunodeficiency, a candidate condition for human gene therapy due to symptomatology restricted to the lymphohematopoietic system. Immune toxicity in PNP deficiency is likely caused by systemic accumulation of deoxyguanosine, one of the PNP substrates. In this article, metabolic correction of deoxyguanosine toxicity is demonstrated by retroviral-mediated PNP gene transfer in PNP-deficient S49 T lymphoma cells as an in vitro genetic model for PNP deficiency. These in vitro results suggest that retroviral-mediated PNP gene transfer and expression may also be effective for in vivo metabolic correction of lymphoid cells in PNP-deficient patients.

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