Retrovirus-mediated transfer of the human O6-methylguanine-DNA methyltransferase gene into a murine hematopoietic stem cell line and resistance to the toxic effects of certain alkylating agents.
O6-Methylguanine-DNA methyltransferase (MGMT) is an important DNA repair protein that plays a key role in cancer chemotherapy by alkylating agents such as carmustine (BCNU) and Dacarbazine (DTIC). Therapy by BCNU and DTIC is reduced by dose-limiting hematological toxicity as a result 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 cells lines have been generated. Retroviral-mediated transfer of the human mgmt gene into murine multi-potent hematopoietic stem cells, FDCP-1, resulted in the expression of a high level of MGMT activity. In comparison with the control cells that were transduced with the parent vector, the MGMT-expressing clones were considerably more resistant to the cytotoxicity of the methylating agents, such as N-methyl-N'-nitro-N-nitrosoguanidine, N-nitroso-N-methyl-urea, and temozolomide, as well as the chloroethylating agents 1-(2-chloroethyl)-1-nitrosourea and BCNU. The protection provided by MGMT could be eliminated by the MGMT inactivator O6-benzylguanine. Thus, the principal lethal lesions produced by these alkylating agents in the murine hematopoietic stem cells and the MGMT deficiency in these cells can be complemented by retroviral-mediated gene transduction.
Wang
G
G
Department of Pharmacology and Toxicology, Dartmouth Medical School, Hanover, NH, USA.
Weiss
C
C
Sheng
P
P
Bresnick
E
E
D016475:3T3 Cells; D000818:Animals; D018906:Antineoplastic Agents, Alkylating / Q000633:toxicity*; D002330:Carmustine / Q000633:toxicity; D002460:Cell Line; D003606:Dacarbazine / Q000633:toxicity; D004351:Drug Resistance / Q000235:genetics; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000187:drug effects*; D006801:Humans; D008780:Methyltransferases / Q000235:genetics*; D051379:Mice; D008979:Moloney murine leukemia virus / Q000235:genetics*; D019853:O(6)-Methylguanine-DNA Methyltransferase
D018906:Antineoplastic Agents, Alkylating; D003606:Dacarbazine; D008780:Methyltransferases; D019853:O(6)-Methylguanine-DNA Methyltransferase; D002330:Carmustine