Fostemsavir analog BMS-818251 has enhanced viral neutralization potency and similar escape mutation profile

BMS-818251, a fostemsavir analog, is a next-generation HIV-1 attachment inhibitor with enhanced potency and a similar resistance profile. By using <i>ex vivo</i> viral outgrowth assays with HIV+ donor samples, we demonstrate here that BMS-818251 exhibits superior viral suppression compared to temsavir, the active form of fostemsavir. To map potential resistance pathways, we employed deep mutational scanning and pseudotyped virus neutralization assays to identify escape mutations within the HIV-1 envelope glycoprotein (Env). These mutations were largely clustered around the BMS-818251 binding site, with key resistance mutations reducing drug-binding affinity. Several of the enriched mutations, such as S375I/N, M426L, and M475I, have been previously observed in fostemsavir-treated patients, highlighting their clinical relevance. Isothermal titration calorimetry revealed reduced binding affinity as the primary mechanism of resistance, though with notable exceptions, such as R429G, suggesting additional factors to influence viral escape. <i>Ex vivo</i> Env sequencing confirmed selection of resistance mutations under BMS-818251 pressure, reinforcing the predictive value of deep mutational scanning for <i>in vivo</i> resistance monitoring. Compared to fostemsavir, BMS-818251 achieved more effective viral suppression at lower concentrations, even in donor samples harboring preexisting resistance mutations. These findings support the continued development of BMS-818251 as a promising alternative to fostemsavir, with potential benefits for patients with multidrug-resistant HIV-1.