INHIBITION OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 REVERSE-TRANSCRIPTASE BY THE 5'-TRIPHOSPHATE-BETA ENANTIOMERS OF CYTIDINE ANALOGS

(-)-beta-L-2',3'-Dideoxycytidine (L-ddC) and (-)-beta-L-2',3'-dideoxy-5-fluorocytidine (L-FddC) have been reported to be potent and selective inhibitors of human immunodeficiency virus type 1 (HIV-1) and type 2 (HIV-2) in vitro. In the present study, the 5'-triphosphates of L-ddC (L-ddCTP) and L-FddC (L-FddCTP) were demonstrated to competitively inhibit HIV-1 reverse transcriptase (RT), with inhibition constants (K(i)s) of 2 and 1.6 mu M, respectively, when a poly(rI) . oligo(dC)(10-15) template primer was used; in comparison K-i values for beta-D-2',3'-dideoxycytidine 5'-triphosphate (D-ddCTP) and beta-D-2',3'-dideoxy-5-fluorocytidine 5'-triphosphate (D-FddCTP) were 1.1 and 1.4 mu M, respectively. Use of the mutant RT at position 184 (substitution of methionine to valine [M184V]), which is associated with resistance to beta-L-2',3'-dideoxy-3'-thiacytidine (3TC) and beta-L-2',3'-dideoxy-5-fluoro-3'-thiacytidine (FTC), resulted in significant increases (50- to 60-fold) in K-i values for L-ddCTP and L-FddCTP, whereas the elevation in K-i values for D-ddCTP and D-FddCTP was moderate (2-fold). L-ddCTP and L-FddCTP did not inhibit human DNA polymerases alpha and beta up to 100 mu M. In contrast, D-ddCTP and D-FddCTP inhibited human DNA polymerase beta, with K-i values of 0.5 and 2.5 mu M, respectively. By using sequencing analysis, L-ddCTP and L-FddCTP exhibited DNA chain terminating activities toward the parental HIV-1 RT, whereas they were not a substrate for the mutant M184V HIV-1 RT. L-ddC and L-FddC did not inhibit the mitochondrial DNA content of human cells up to a concentration of 10 mu M, whereas D-ddC and D-FddC decreased the mitochondrial DNA content by 90% at concentrations of 1 and 10 mu M, respectively. All of these results suggest that further development of L-ddC, and L-FddC in particular, is warranted as a possible anti-HIV candidate.