Structure-enabled enhancement of potency in a metronidazole–benznidazole hybrid: design, synthesis, and evaluation of antitrypanosomal activity of a benzylamide-linked 5-nitroimidazole

Benznidazole ( 1 ) remains the primary antiparasitic drug used clinically for Chagas disease, underscoring the need to discover novel treatments targeting Trypanosoma cruzi ( T. cruzi ) infection. In this sense, the present work reports the in silico design of a novel hybrid ( 3 ) containing the 5-nitroimidazole core from the commercial antiparasitic drug metronidazole ( 2 ) and the N -benzylacetamide moiety from 1 . These structural motifs were designed to target T. cruzi nitroreductase type I (TcNTR) for activation and to boost intracellular drug levels (lipophilicity). The in silico predictions suggested that the presence of the N -benzylacetamide moiety might enhance the anti-T . cruzi activity of 2 and improve the drug-likeness. To validate the predictions, hybrid 3 was synthesized with a yield of 53% and structurally characterized (melting point, 1 H and 13 C NMR, and HRMS). The in vitro assays of hybrid 3 against T. cruzi amastigotes (Tulahuen strain C2C4 LacZ ) supported the in silico predictions, showing a lower IC 50 for 3 (67.73 ± 8.98 µM) than for 2 (100 µM). Despite hybrid 3 having an activity 45-fold lower than that of 1 , the results provide insights for future hybrid optimization.