Structure-Based Discovery of Non-Covalent Triazole-Based Cruzipain Inhibitors with Improved Bioactivity Translation.

Chagas disease is a neglected tropical condition with limited treatment options, emphasizing the need for new antichagasic agents. Cruzipain (CZP), the main cysteine protease of Trypanosoma cruzi, is a validated target; however, achieving strong trypanocidal activity through CZP inhibition remains challenging. In this work, we report a series of novel triazole-based noncovalent CZP inhibitors designed by applying state-of-the-art molecular modeling techniques, showing good potency against isolated CZP combined with effective trypanocidal activity. Structure-activity relationships that define key pharmacodynamic features were elucidated, enabling analogues to translate enzymatic inhibition into cellular effects. The computer-aided design workflow combining molecular docking, molecular dynamics simulations, and binding free-energy calculations was key to reveal conserved interaction patterns within the oxyanion hole and the S1/S1' subsites (Gln19, His162, Trp184) of CZP for the most effective inhibitors. Overall, our findings introduce new 1,2,3-triazole-based antichagasic candidates and clarify molecular determinants influencing the translational gap between enzymatic and cellular activity, contributing with a predictive framework for the rational design of CZP-targeted inhibitors for Chagas disease therapy.