3T3 NIH MURINE FIBROBLASTS AND B78 MURINE MELANOMA-CELLS EXPRESSING THE ESCHERICHIA-COLI N3-METHYLADENINE DNA GLYCOSYLASE-I DO NOT BECOME RESISTANT TO ALKYLATING-AGENTS

The role of alkylation of the N3 position of adenine in the cytotoxici ty of alkylating agents in mammalian cells is still undefined. By co-t ransfecting NIH3T3 murine fibroblast and murine B78 H1 melanoma cells with pSG5tag and pSV2neo, we obtained clones expressing the mRNA of th e bacterial tag gene coding for N3-methyladenine-DNA glycosylase I (Gl y I), which specifically repairs N3-methyladenine. The levels of Gly I were 400 times higher in NIH3T3 pSG5tag (clone 3.9.4.) and 12-33 time s higher in B78 H1 tag clones (2A4, 2A6, 2C3 and 2D1) than in the resp ective control cells. The sensitivity to alkylating agents was evaluat ed in tag-expressing cells in comparison with pSG5, pSV2neo cotransfec ted control cells. As regards the cytotoxic activity of methylating ag ents (N-methylnitrosourea, N-methyl-N'-nitro-N-nitrosoguanidine, dimet hylsulfate and temozolomide) and other alkylators with different struc ture and different interactions with DNA such as CC-1065 and FCE-24517 (minor groove binders known to bind to N3 of adenine), 4-[bis(2-chlor oethyl)amino]L-phenylalanine and cis-diamino-dichloroplatinum II, cyto toxicity was the same for tag-expressing and non-expressing cells. The se results suggest that the increased expression of N3-methyladenine-D NA glycosylase is not necessarily a crucial mechanism for the resistan ce of cells to alkylating agents.