L-ENANTIOMERS AND D-ENANTIOMERS OF 2',3'-DIDEOXYCYTIDINE 5'-TRIPHOSPHATE ANALOGS AS SUBSTRATES FOR HUMAN DNA-POLYMERASES - IMPLICATIONS FOR THE MECHANISM OF TOXICITY

5'-Triphosphates of beta-D and beta-L-enantiomers of 2',3'-dideoxycytidine (ddC), 2',3'-dideoxy-5-fluorocytidine (FddC), 1,3-dioxolane-cytidine (OddC), and 1,3-dioxolane-5-fluorocytidine (FOddC) were evaluated as inhibitors and substrates for human DNA polymerases alpha, beta, gamma, delta, and epsilon. L-ddCTP was not a substrate or inhibitor for any DNA polymerase studied; L-FddCTP was not an inhibitor or substrate for replicative DNA polymerases and was a less potent inhibitor of DNA polymerases gamma and beta than its D-enantiomer by 2 orders of magnitude. In contrast, all L-dioxolane analogs were potent inhibitors and chain terminators for all cellular DNA polymerases studied. The K-i values of their 5'-triphosphates for DNA polymerase gamma were found to be in the following order: D-ddC < n-FddC L-OddC D-FOddC < L-FOddC << L-FddC. The K-i values of L-OddCTP for the reactions catalyzed by DNA polymerases alpha, delta, epsilon, beta and gamma were 6.0, 1.9, 0.4, 3.0, and 0.014 mu M, respectively, and those of L-FOddCTP were 6.5, 1.9, 0.7, 19, and 0.06 mu M, respectively. The K-m values for incorporation of L-OddCTP into the standing points of primer extension were also evaluated and determined to be 1.3, 5.5, 1.5, 2.8, and 0.7 mu M for DNA polymerases alpha, delta, epsilon, beta and gamma, respectively. The incorporation of dioxolane analogs into DNA by replicative DNA polymerases could explain their potent cellular toxicity.