MECHANISM OF THE SELECTIVE HYPOXIC CYTOTOXICITY OF 1-METHYL-2-NITROIMIDAZOLE

2-Nitroimidazoles were introduced into radiation therapy to test their ability to radiosensitize hypoxic cells in solid human tumours. In ad dition, they are selectively reduced in hypoxic cells to form reactive metabolites that may be effective cytotoxins. 1-Methyl-2-nitroimidazo le (INO2) was investigated as a model compound to study the mechanism of selective bioreduction in hypoxic cells. Results demonstrated that INO2 was toxic under hypoxic conditions (tested via colony-forming ass ay) at concentrations where no toxicity was observed for aerobic cells . This selective hypoxic toxicity was a function of both concentration and time. The depletion of both glutathione and protein thiols occurr ed under hypoxic conditions and preceded a rise in intracellular calci um levels. Previous work with INO, the nitroso intermediate of INO2 re duction, also showed concentration-dependent cytotoxicity, and glutath ione and protein thiol depletion, which was followed by an increase in intracellular calcium levels. The kinetics of cytotoxicity and cellul ar reactions were slower for the parent compound, INO2 as compared wit h the 2e(-) reductive metabolite, INO, reflecting the limited enzymati c production of the reactive intermediate in the INO2 experiments. Zei osis (membrane blebbing) and chromatin condensation occurred shortly a fter treatment of cells with equitoxic concentrations of both INO2 (un der hypoxic conditions) and INO (under aerobic conditions), suggesting that an apoptotic-like death mechanism may be involved. However, anal ysis of DNA isolated from both INO2- and INO-treated cells, up to 2 hr after treatment, did not reveal any nucleosomal fragmentation, anothe r characteristic feature of cells undergoing apoptosis. The toxicity o f high INO2 concentrations toward CHO cells is consistent with the pro duction of an INO intermediate and has several features characteristic of an apoptotic mechanism of cell death.