DNA repair is an essential cellular process mediating response to DNA replication errors, endogenous and exogenous DNA damage, and cell survival. Despite the importance of DNA repair, there has been little analysis of the role of many DNA repair processes in maintenance of the stem cell phenotype as functionally measured in terms of stem cell longevity, long term repopulating capacity after transplantation, and multilineage hematopoietic and lymphoid reconstitution after DNA damage. In this proposal, the investigators will use murine models of DNA repair defects to focus on hematopoietic stem cell repair of methylating agent induced DNA damage. Therapeutic methylating agents cause dose-limiting myelosuppression and are implicated in treatment-related leukemias in humans. Thus, these agents induce DNA damage that is both cytotoxic and mutagenic to hematopoietic stem cells. Two important components of methyl DNA adduct repair are mismatch repair [MMR] and 06-alkylguanine-DNA alkyltransferase [AGT], encoded by the MGMT gene. The investigators have previously shown the importance of MMR and MGMT in preventing the induction of methylating agent induced lymphomas in the mouse. Furthermore, defects in MMR and MGMT have been seen in human leukemias and lymphomas. No studies have examined the impact of DNA repair defects on stem cell function and malignant transformation. Using mice defective in the DNA repair processes of MMR and/or MGMT, the specific aims of this proposal are to: (1) determine whether DNA repair capacity for methylating adducts (i.e., MMR and MGMT) affect key hematopoietic stem cell functions. These characteristics include myeloid and lymphoid proliferation and differentiation, cell cycle distribution and quiescence, long-term repopulating cell survival and longevity, and sequential transplant capacity; (2) determine whether defects in DNA repair affect the competitive long-term repopulation capacity of stem cells exposed to methylating agents; and (3) determine whether the therapeutic methylating agent-induced malignant transformation potential of transplanted stem cells is affected by defects in DNA repair, and whether this mimics treatment-related leukemias in humans. These studies will impact our appreciation of the role of DNA repair in human hematologic malignancies and in the maintenance of human stem cells.
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