Inhibition of DNA synthesis by nitroheterocycles. II. Mechanisms of cytotoxicity.

Nitroheterocycles have been shown to inhibit the incorporation of 3H-TdR by cultured L-929 cells, and the degree of inhibition is related to their electron-affinity. On the basis of their chemical reactivity and potential clinical utility, nitrofurazone, misonidazole and metronidazole were selected for more detailed studies of the mechanism of inhibition of DNA synthesis. Double-isotope labelling in conjunction with hydroxyapatite chromatography allowed the evaluation of drug effects on initiation of DNA replicons, DNA chain elongation and DNA damage (single-strand breaks), and their correlation with eventual cell viability. Partial inhibition of initiation of DNA synthesis generally preceded other measurable effects, and was not reversed by incubation in the absence of drug. In the absence of DNA strand breaks (at low drug doses or after a repair interval) the rate of elongation was similar in both treated and untreated cell populations. Measurable DNA damage (strand breaks) was predictive for cytotoxicity. At lower drug doses, or under aerobic conditions, DNA synthesis was not always associated with a decrease in plating efficiency (cytotoxicity) but was reflected in decreased colony size (growth rate) of the cells. Thus the aerobic "toxicity" previously reported for chronic exposure to these agents may be better described as a "cytostatic" effect. Under anaerobic conditions (where cell killing is much greater) inhibition of initiation plays a less important role, and the nitroheterocycles are metabolically reduced to intermediates which are truly cytotoxic.