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Cytokinetics of a novel 1,2,3-triazene-containing heterocycle, 8-nitro-3-methyl-benzo-1,2,3,5-tetrazepin-4(3H)-one (NIME), in the human epithelial ovarian cancer cell line OVCAR-3

1999; Elsevier BV; Volume: 57; Issue: 7 Linguagem: Inglês

10.1016/s0006-2952(98)00345-1

1873-2968

BertrandJacques Jean-Claude, Amir Mustafa, Zoe Damian, Josie De Marte, DanielaEliza Vasilescu, Rose Yen, ThakHang Chan, Brian Leyland‐Jones,

Chemical Reactions and Isotopes

The mechanism of action of the novel tetrazepinone 8-nitro-3-methyl-benzo-1,2,3,5-tetrazepin-4(3H)-one (NIME), structurally related to the antitumour drug temozolomide, was studied in the human ovarian tumour cell line OVCAR-3. NIME preferentially inhibited DNA synthesis over protein and RNA syntheses at 3 and 24 hr post-treatment. A Maxam-Gilbert sequencing assay showed that NIME induced barely detectable levels of guanine N7 alkylation in an isolated DNA strand, in contrast to temozolomide, a strong alkylating agent containing, like NIME, a cyclic 3-methyl-1,2,3-triazene moiety. Alkaline sucrose density-gradient sedimentation, at concentrations 2- to 10-fold lower than the ones used in the DNA sequencing assay, showed significant DNA damage in OVCAR-3 cells 24 hr after treatment with NIME. This was accompanied by a significant accumulation of cells in late S and G2M. Cell cycle arrest was transient and was reversed after 2–3 days following drug treatment. This was in agreement with bivariate bromodeoxyuridine/propidium iodide analysis, which showed that at 100 μM, a concentration at which the majority of the cells arrested in late S and G2M, a significant fraction of bromodeoxyuridine positive (S-phase) cells escaped the block. In an attempt to elucidate the mechanism underlying these effects, the degradation of NIME in cell culture medium was analyzed by GC–MS (gas chromatography coupled with mass spectrometry). The results showed that, in contrast to temozolomide, NIME did not convert to an open-chain alkyltriazene in cell culture medium, but to a major benzimidazole product, which exerted a minor effect on the cell cycle. This suggests that NIME, despite containing a 3-(alkyl)-1,2,3-triazene moiety, does not act by DNA alkylation but probably by generating a short-lived genotoxic species during its degradation to 6,5-benzofused derivatives.