Friend or foe: divergent immunomodulatory effects of metabolites derived from the virucidal zinc finger inhibitor SAMT-247.

BACKGROUND: Topical administration of SAMT-247, a mercaptobenzamide thioester zinc finger inhibitor targeting the HIV nucleocapsid Gag zinc finger protein, generates multiple metabolites in mucosal tissues, including Met-A, Met-B, Met-C, and Met-D. Prior studies established that SAMT-247, delivered either as a vaginal gel or via an intravaginal ring (IVR), synergizes with the ΔV1DNA/ALVAC-SIV/ΔV1gp120/alum vaccine regimen to markedly reduce vaginal SIVmac251 acquisition risk. This enhanced protection is associated with augmented protective mucosal immunity and reduced inflammatory responses that promote viral acquisition. METHODS: We performed in vivo characterization of SAMT-247 metabolite distribution in the vaginal compartment of macaques and evaluate the biological activity of individual metabolites using ex vivo rectal mucosal biopsies from vaccinated animals. RESULTS: High concentrations of Met-B, Met-C, and Met-D were detected in vaginal secretions, whereas vaginal tissues contained predominantly Met-D, low levels of Met-A, and no detectable Met-B or Met-C. Functionally, Met-D most closely recapitulated and reinforced the protective immune profile associated with SAMT-247, preserving or expanding IL-17+NKp44+ innate lymphoid cells (ILCs) and CD107a+NKG2A+ natural killer (NK) cells, increasing CD73+ ILCs, NK and dendritic cell as well as IL-10+ dendritic cell and monocyte populations, and reducing inflammatory TNF-α-producing myeloid subsets. Met-B and Met-C partially reproduced this profile, enhancing CD73+ NK/ILC populations and promoting anti-inflammatory myeloid responses, but with more limited effects on antiviral NK/ILC activity and attenuated NKp44+ ILC responses relative to Met-D. In contrast, Met-A, which lacks virucidal activity, failed to induce regulatory CD73+ and IL-10+ responses and diminished both protective IL-17+NKp44+ ILCs and cytolytic CD107a+NKG2A+ NK cells, consistent with inflammatory skewing. In vaccinated macaques receiving SAMT-247-releasing IVRs, plasma Met-A showed limited correlation with rectal mucosal immune responses, partially supporting the ex vivo findings. CONCLUSION: We observed distinct immunomodulatory effects associated with different SAMT-247 metabolites. These findings may guide future delivery strategies that favor tissue-available Met-D while limiting Met-A accumulation. More broadly, this study underscores the importance of metabolite-specific analyses for defining the biological activity and mechanisms of action of therapeutic agents.