Investigation of resistance to DNA cross-linking agents in 9L cell lines with different sensitivities to chloroethylnitrosoureas

Abstract

The 9L-2, 9L-7, and 9L-8 cell lines, derived from the 9L in vivo rat brain tumor, were treated with nitrosoureas that can alkylate and cross-link DNA and carbamoylate intracellular molecules to various extents. Compared to 9L cells, 9L-2 cells were very resistant to the cytotoxic effects of 1,3-bis(2-chloroethyl)-1-nitrosourea, and to 2-[3-(2-chloroethyl)-3-nitrosoureido]-D-deoxyglucopyranose. The sensitivity of 9L-7 and 9L-8 cell lines to these drugs was intermediate between 9L and 9L-2. Treatment of 9L, 9L-2, 9L-7, and 9L-8 cell lines with 1,3-bis(trans-4-hydroxycyclohexyl)-1-nitrosourea produced approximately the same level of cell kill. Compared to 9L cells, 9L-2 cells are 10-fold more resistant to the cytotoxic effects, 34-fold more resistant to the induction of sister chromatid exchanges, and have 40% fewer DNA interstrand cross-links caused by treatment with 3-(4-amino-2-methyl-5-pyrimidinyl)methyl-1-(2-chloroethyl)-1-nitrosourea . In contrast, treatment of 9L and 9L-2 cells with 1-ethylnitrosourea produced approximately the same level of cell kill and induction of sister chromatid exchanges. Our results suggest that the resistance of 9L-2, 9L-7, and 9L-8 cells is related to DNA cross-linking and not to alkylation or carbamoylation. We studied the effects of other agents that form DNA cross-links with structures different from those formed by treatment with chloroethylnitrosoureas (CENUs) in 9L and 9L-2 cells. In contrast to results obtained with CENUs, 9L-2 cells were 2-fold more sensitive to the cytotoxic effects, 2-fold more sensitive to the induction of sister chromatid exchanges, and had 3-fold more cross-links formed than 9L cells treated with nitrogen mustard. However, the amount of cell kill, number of sister chromatid exchanges induced, and the DNA cross-linking were the same for 9L and 9L-2 cells treated with cis-diamminedichlorplatinum(II). Our results indicate that cellular resistance to CENUs is highly specific and that the mechanism of resistance does not allow cross-resistance with other DNA cross-linking agents. These and other results suggest that when DNA repair processes mediate cellular resistance to CENUs, other cross-linking agents will not be cross-resistant unless they form alkylation products that are affected by repair processes that mediate resistance to CENUs.