Novel Synthetic Chalcones Display Potent Anti- Leishmania ( Viannia ) braziliensis Profile and Inhibit Proteolytic Activities

Abstract American Tegumentary Leishmaniasis, primarily caused by Leishmania (Viannia) braziliensis, remains a major public health challenge in Latin America. The variable efficacy and toxicity of pentavalent antimonials underscore the urgent need for new therapeutic candidates. Among the compounds currently explored, chalcones have emerged as promising bioactive scaffolds, exhibiting antiparasitic activity and the potential to inhibit key proteases, which are essential enzymes to virulence and pathogenesis of the parasite. In this study, chalcone derivatives obtained via a novel synthetic approach were evaluated for their antileishmanial activity against L.(V.)braziliensis. Their drug-likeness properties were initially assessed in silico, and the inhibitory activity against proteases was subsequently evaluated using promastigote protein extracts. In Tris–HCl buffer, compound 3j exhibited the most pronounced inhibitory effect, 36% residual activity, followed by compounds 3d and 2′,4′,6′-trimethoxy-chalcone (TMC), which showed 47% residual activity. In acetate buffer, the most effective inhibitors were 3b and 3j (34%), and 3d (35%). Cytotoxicity assays were performed in murine macrophages, and the leishmanicidal effects were evaluated against promastigotes to determine compound selectivity index (SI). TMC, 3b, 3d, 3h, and 3i exhibited SI ≥ 16.74 and were tested against intracellular amastigotes. They decreased ≥34% macrophage infection by intracellular amastigotes, and demonstrated a predictive profile for high metabolic stability with low organ toxicity risk. The findings suggest that these chalcones may act through a multitarget mechanism involving the inhibition of parasite proteases, supporting their progression to advanced stages of therapeutic validation.