Metabolism And Mode Of Selective Inhibition Of Human Immunodeficiency Virus Replication By 3'-Azido-2', 3'-Dideoxy-5-Iodouridine And 3'-Azido-2', 3'-Dideoxy-5-Bromouridine

3'-Azido-2', 3'-dideoxy-5-iodouridine (AzIdUrd) and 3'-azido-2', 3'-dideoxy-5-bromouridine (AzBdUrd), previously shown to be potent and selective inhibitors of human immunodeficiency virus replication in vitro, are minimally toxic to the uninfected human lymphoid cell line H9 (IC50 = 197 and 590 μM, respectively). Both compounds strongly inhibited the incorporation of [³H]thymidine but not [³H]deoxyadenosine into DNA, and we observed no significant inhibition of [³H]uridine incorporation into RNA, or [³H]amino acids into protein. Exposure of H9 cells to AzIdUrd or AzBdUrd (100 μM, 24h) and pulse labeling with [³H]thymidine resulted in approximately a 90% reduction in levels of tritiated dTMP, dTDP, and dTTP relative to control. [¹²⁵I] AzIdUrd was rapidly phosphorylated by H9 cells with the monophosphate accounting for over 90% of total soluble radioactivity. A relatively low but stable level of AzIdUTP was maintained over a 12-hr period. [¹²⁵I] AzIdUrd was phosphorylated by a cell free extract of H9 cells at a rate approximately three times that of thymidine and its phosphorylation was inhibited by excess thymidine. AzIdUrd was found to be a competitive inhibitor of cytosolic thymidine Kinase with a K_i of 2.63 μM and AzIdUMP a weak competitive inhibitor of thymidylate kinase with a K_i of 55.3 μM. Both AzIdUTP and AzBdUTP were potent competitive inhibitors of HIV-1 reverse transcriptase (K_i = 0.028 and 0.043 μM, respectively) and relatively poor inhibitors of H9 cell DNA polymerase a (K_i = 42.0 and 42.7 μM, respectively). Thus, the high therapeutic index of these compounds is due to the sensitivity of the viral reverse transcriptase, coupled with the relative insensitivity of the host cell DNA polymerase a.