RETROVIRUS-MEDIATED TRANSFER OF THE HUMAN O-6-METHYLGUANINE-DNA METHYLTRANSFERASE GENE INTO A MURINE HEMATOPOIETIC STEM-CELL LINE AND RESISTANCE TO THE TOXIC EFFECTS OF CERTAIN ALKYLATING-AGENTS

O-6-Methylguanine-DNA methyltransferase (MGMT) is an important DNA rep air protein that plays a key role in cancer chemotherapy by alkylating agents such as carmustine (BCNU) and Dacarbazine (DTIC). Therapy by B CNU and DTIC is reduced by dose-limiting hematological toxicity as a r esult of low MGMT repair activity in bone marrow cells. In this study, we have constructed a Moloney murine leukemia virus retroviral vector containing the human mgmt gene. High-titer retrovirus producer cell l ines have been generated. Retroviral-mediated transfer of the human mg mt gene into murine multi-potent hematopoietic stem cells, FDCP-1, res ulted in the expression of a high level of MGMT activity. In compariso n with the control cells that were transduced with the parent vector, the MGMT-expressing clones were considerably more resistant to the cyt otoxicity of the methylating agents, such as N-methyl-N'-nitro-N-nitro soguanidine, N-nitros-N-methylurea, and temozolomide, as well as the c hloroethylating agents 1-(2-chloroethyl)-1-nitrosourea and BCNU. The p rotection provided by MGMT could be eliminated by the MGMT inactivator O-6-benzylguanine. Thus, the principal lethal lesions produced by the se alkylating agents in the murine hematopoietic stem cells and the MG MT deficiency in these cells can be complemented by retroviral-mediate d gene transduction.