Selective Elimination of HIV-1-Infected Macrophages by Targeted Activator of Cell Kill

In the pursuit of an HIV cure, elimination of persistent HIV reservoirs requires strategies that promote immune-mediated or cytopathic clearance of infected cells. While many such approaches are being developed against infected CD4+ T cells, HIV-infected macrophages are comparatively resistant to cell death and to several killing mechanisms being explored for T-cell reservoirs, highlighting the need for strategies that specifically eliminate this reservoir. Targeted Activator of Cell Kill (TACK), developed by Merck Pharmaceuticals, are non-nucleoside reverse transcriptase inhibitor-derived molecules designed to selectively kill HIV-1-infected cells by inducing premature activation of the viral protease. Here, we evaluated the TACK compounds Pyr01 and it?s inactive analog Pyr02, in primary human monocyte-derived macrophages (MDMs). Pyr01 had minimal effects on uninfected MDM viability but potently targeted HIV-1-infected MDMs, with an EC?? of approximately 0.36 ?M. Pyr01 treatment produced a rapid and sustained reduction in extracellular virion-associated HIV-1 RNA and significantly decreased cell-associated viral RNA, whereas Pyr02 exhibited minimal activity. Immunofluorescence microscopy and live-cell imaging further demonstrated a marked loss of HIV-1-infected macrophages following Pyr01 treatment, consistent with selective elimination of infected cells. These findings identify Pyr01 as a specific mediator of HIV-infected macrophage death and establish premature viral protease activation as a strategy for overcoming the intrinsic resistance of macrophage reservoirs to clearance. Targeting this macrophage-specific barrier may complement approaches directed at CD4+ T-cell reservoirs and broaden the cellular reach of HIV cure strategies.