Contribution of O6‐methylguanine‐DNA methyltransferase to resistance to 1,3‐(2‐chloroethyl)‐1‐nitrosourea in human brain tumor‐derived cell lines

Abstract

To assess the possible role of the DNA repair protein O⁶‐methylguanine‐DNA methyltransferase (MGMT) in resistance of brain neoplasms to the clinically important chloroethylating agent 1,3‐bis(2‐chloroethyl)‐1‐nitrosourea (BCNU), we quantitated MGMT activity, BCNU survival, and the effect of ablating MGMT activity on the sensitivity of 14 human medulloblastoma‐ and glioma‐derived cell lines. BCNU resistance, measured as 10% survival dose (LD10), differed eightfold among the lines. Elimination of measurable MGMT activity with the substrate analogue inhibitor O⁶‐benzylguanine (O⁶‐BG) revealed a variable but limited contribution of MGMT to survival. In no case did O⁶‐BG reduce LD₁₀ by more than 3.4‐fold. In contrast, 06‐BG reduced the LD10 for N‐methyl‐N′‐nitro‐N‐nitrosoguanidine up to 31‐fold in the same cell lines (Bobola MS, Blank A, Berger MS, Silber JR, Mol Carcinog 13:70–80, 1995). Variability in BCNU survival, manifested as a sevenfold range of LD₁₀, persists after measurable MGMT was eliminated, indicating that another mechanism or mechanisms is operating to limit cytotoxicity. Cells alkylated while suspended in growth medium are more resistant to BCNU and display less dependence on MGMT than cells treated while proliferating on a plastic substratum. When alkylated in suspension, most of the lines are either unresponsive to O⁶‐BG or contain a subpopulation that did not respond to O6‐BG. Our results demonstrate that BCNU resistance is multifactorial and that MGMT makes a modest contribution to resistance in our lines.