Clicking 3′-Azidothymidine into Novel Potent Inhibitors of Human Immunodeficiency Virus

3′-Azidothymidine (AZT) was the first approved antiviral for the treatment of human immunodeficiency virus (HIV). Reported efforts in clicking the 3′-azido group of AZT have not yielded 1,2,3-triazoles active against HIV or any other viruses. We report herein the first AZT-derived 1,2,3-triazoles with submicromolar potencies against HIV-1. The observed antiviral activities from the cytopathic effect (CPE) based assay were confirmed through a single replication cycle assay. Structure–activity-relationship (SAR) studies revealed two structural features key to antiviral activity: a bulky aromatic ring and the 1,5-substitution pattern on the triazole. Biochemical analysis of the corresponding triphosphates showed lower ATP-mediated nucleotide excision efficiency compared to AZT, which along with molecular modeling suggests a mechanism of preferred translocation of triazoles into the P-site of HIV reverse transcriptase (RT). This mechanism is corroborated with the observed reduction of fold resistance of the triazole analogue to an AZT-resistant HIV variant (9-fold compared to 56-fold with AZT)

Nucleotide Sugar Pucker Preference Mitigates Excision by HIV‑1 RT

A series of DNA primers containing nucleotides with various sugar pucker conformations at the 3′-terminus were chemically synthesized by solid-phase synthesis. The ability of wild-type (WT) HIV-1 reverse transcriptase (RT) and AZT-resistant (AZTr) RT to excise the 3′-terminal nucleotide was assessed. Nucleosides with a preference for the North conformation were more refractory to excision by both WT-RT and AZTr-RT. We found that DNA primers that contain North puckered-nucleotides at the 3′-terminus can also affect the translocation status of the RT/template/primer complex, which provides an underlying mechanism to avoid being excised. Together, these results point to a correlation between the sugar conformation of the 3′-terminal nucleotide, the precise position of HIV-1 RT on its nucleic acid substrate, and, in turn, its catalytic function. Nucleotide sugar conformation is therefore an important parameter in defining the susceptibility to RT-catalyzed phosphorolytic excision