M184V: The Major Determinant of Cooperative ISL Resistance When in Combination with Multiple TAM2 Mutations in HIV-1 Subtype C.

Islatravir is a novel nucleoside reverse transcriptase translocation inhibitor recently approved for the treatment of HIV-1 infection in virologically suppressed PLWH. While we previously documented high-level phenotypic resistance to islatravir (ISL) in HIV-1 subtype C variants harboring combinations of type 2 thymidine analogue mutations (TAM2) along with the M184V resistance-associated mutation, the primary contributor to the observed phenotype remains unclear. This report dissects the relative contribution of M184V to ISL resistance in the context of TAM2-containing variants through systematic reversion analysis. We show that reverting mutant 184V to wildtype M184 resulted in substantial sensitization to ISL, with fold-change reductions ranging from 8.1 to 13.4-fold across three TAM2 genetic backgrounds. Contrary to the well-documented antagonism between M184V and TAMs in zidovudine resistance, these two resistance pathways exhibit cooperative rather than antagonistic interactions in the context of ISL resistance. These findings provide critical evidence for refinement of genotypic resistance interpretation algorithms for ISL and have direct implications for the future clinical management of ISL-based antiretroviral therapy in people living with HIV. Importantly, elucidating the mechanistic basis of M184V-TAM2 cooperation would establish a more predictive framework for understanding how complex resistance mutation patterns impact ISL susceptibility and guide treatment decisions in virologically experienced individuals. Considering the diminished use of TAM2-seletive antiretroviral drugs in contemporary ART regimens, islatravir is likely to be an effective treatment option.