Abstract

Human brain and colon tumor cell lines SF-188 (Mer+) and WiDR (Mer+), which express the DNA repair proteinO⁶-methylguanine-DNA methyl transferase (MGMT), were 3- to 30-fold less sensitive to temozolomide, mitozolomide, andN,N′-bis(2-chloroethyl)-N-nitrosourea (BCNU) than the MGMT-deficient tumor cells SF-126 (Mer−) and BE (Mer−). This differential sensitivity was not observed when these cells were exposed to the novel tetrazepinones PYRZ, NIME, QUINCL, and PYRCL, which contain, like temozolomide and mitozolomide, a ureido-triazene moiety. Flow cytometric studies revealed that temozolomide induced G₂-M arrest in the Mer− cells, but exerted a minor effect on the cycle of the Mer+ cells. Similarly, mitozolomide (25–100 μM) induced a stronger S-phase arrest in the SF-126 cells than in the SF-188 cells. In the same dose range (25–100) BCNU induced a significant cell cycle accumulation in G₂2-M in the SF-126 cells but little in the SF-188 cell line. In contrast, the cell cycle effects of the tetrazepinones were independent of the cell phenotypes. When O⁶-benzylguanine (O⁶-BG) was used to deplete MGMT activity in the SF brain tumor cell lines, significant potentiation of temozolomide (67-fold), mitozolomide (7-fold), and BCNU (3-fold) was observed in the SF-188 cell line. By contrast, O⁶-BG did not potentiate PYRZ, PYRCL, QUINCL, and NIME. Moreover, an MGMT inhibitory assay showed that all the tetrazepinones were capable of inactivating MGMT in the SF-188 cell line, the strongest inhibitor being PYRCL. The results suggest that, unlike temozolomide, mitozolomide, and BCNU, the cytotoxicity of the tetrazepinones does not correlate with the alkylation of the O⁶ position of guanine and that the mechanism of MGMT inactivation by tetrazepinones may differ from that of hitherto known inhibitors.