Sequential inhibitory effects of antitumor agents related to levodopa and dopamine upon DNA synthetic enzymes

Novel antitumor agents related to levodopa and dopamine exhibit a selective and rapid inhibition of DNA synthesis as measured by thymidine incorporation. Our investigations have attempted to determine the biochemical basis of the selective inhibition of tumor cells and in this present study we examined the effects of these agents on thymidylate synthase. The dihydroxybenzene derivatives were found to inhibit thymidylate synthase in situ at concentrations ranging between 100 and 800 microM. The quinols did not inhibit partially purified thymidylate synthase, although the oxidized quinones did cause inhibition at concentrations between 10 and 100 microM. Time course experiments suggested that the inhibition of thymidylate synthase in situ by the dihydroxybenzene derivatives occurs after the inhibition of thymidine incorporation, indicating that an earlier event was critical to the inhibition of DNA synthesis. With the use of a novel in situ assay which measured the release of [3H]water from [5-3H] uridine in intact cells, we were able to show that one of the earliest biochemical events is the inhibition of ribonucleotide reductase and that the inhibition of thymidylate synthase, which is delayed by approximately 30 min, was indirectly mediated possibly through effects on ribonucleotide reductase.