The nucleoside analogs 4'C-methyl thymidine and 4'C-ethyl thymidine block DNA synthesis by wild-type HIV-1 RT and excision proficient NRTI resistant RT variants.

HIV-1 can become resistant to nucleoside analogs by developing an enhanced ability to excise the analogs after they have been incorporated. Excision requires that the analog be located at the 3' terminus of the primer. We have describe nucleoside analogs that do not block DNA synthesis at the point of incorporation, but only after additional dNTPs have been added to the DNA. These nucleoside analogs are called "delayed chain terminators" and are relatively effective inhibitors of drug-resistant HIV-1 reverse transcriptases (RTs) that are excision proficient. However, the first delayed chain terminator that we characterized was poorly phosphorylated in cultured cells. We have examined other nucleoside analogs to determine whether these compounds also act as delayed chain terminators, but were more efficiently converted to the triphosphate form by cellular kinases. These analogs contain substitutions on the deoxyribose sugar ring at the 4' carbon (4'C-methyl dT and 4'C-ethyl dT). Unlike true delayed chain terminators, which terminate DNA synthesis in a spatial sense (DNA synthesis is halted only after additional dNTPs have been incorporated after the analog), 4'C-methyl dTTP causes a pause in DNA synthesis at the point of incorporation. However, HIV-1 RT can eventually extend the primer blocked by the 4' C-Me dTMP analog. 4'C-methyl dTTP blocks DNA synthesis in a temporal sense, rather than in a spatial sense. A primer blocked by 4'C-ethyl dTMP is not extended by HIV-1 RT, and this compound acts like a conventional chain terminator, despite the presence of a 3'-OH group. These compounds effectively block the replication of an HIV-1-based vector that replicates using wild-type HIV-1 RT, but only in the presence of herpes simplex virus thymidine kinase (HSV TK). These compounds are effective against many NRTI drug-resistant RT variants; however, the M184V mutant is relatively resistant.