Novel triazole-based targeted covalent inhibitors of Cruzain as potential antichagasic agents

CERUTTI, JUAN PABLO; DINIZ, LUCAS; FERREIRA, RAFAELA; DEHAEN, WIM; QUEVEDO, MARIO ALFREDO

Rosario

Congreso; 7ma Reunión Internacional de Ciencias Farmacéuticas; 2023

Chagas disease is one of the main neglected tropical diseases worldwide, thus the development of new pharmacotherapeutic agents is of utmost importance. Considering that Cruzain (Cz), the main cysteine protease of T. cruzi, constitutes a chemotherapeutic target with excellent pre-clinical validation, its covalent inhibition has gained high scientific interest. In particular, the design of the so-called targeted covalent inhibitors (TCIs) bearing reactive warhead (WH) groups attracts special attention. In this work, we approached the rational design, synthesis and biological evaluation of novel potential TCIs of Cz, bearing a 1,4-disubstituted 1,2,3-triazole moiety as the main scaffold combined with three different WH. Massive libraries of triazole derivatives were constructed in silico by combining commercially available reagents required for their chemical synthesis. In a second stage, a docking-based high-throughput virtual screening (vHTS) was performed using Cz as target. 47 candidates were identified as presenting promising pharmacodynamic profiles, and were thus selected for chemical synthesis. 24 compounds include an aldehyde as WH, while the remaining 23 triazole derivatives bear a thiol or a thiosulfonate WH. Following Cz inhibition assays, 20 candidates (43%) showed promising inhibitory activity against Cz, with sub-micromolar to low micromolar IC50 values. Inhibition mechanism studies showed that candidates including an aldehyde as WH lead to irreversible covalent inhibitory activity, while derivatives bearing a thiol or a thiosulfonate WH produce a reversible covalent inhibition of Cz. This behaviour, together with the fact that their potency was equivalent or even higher than that of aldehyde derivatives, could be very beneficial in terms of safety. Moreover, molecular dynamics and energetic studies allowed to explain the pattern of interactions and their observed biological activity, providing new insights for the SAR of Cz, mainly for the S1 subsite. Overall, these results show that the design of triazole-based TCIs constitutes a promising strategy towards potent inhibition of Cz, with in silico methods representing a powerful approach towards obtaining high hit rates. In addition, novel promising thiol and thiosulfonate derivatives are envisioned as prototype compounds for future optimisations, paving the way to satisfy the need of new and effective drugs for Chagas disease.