Anti-Trypanosoma cruzi Potential of New Pyrazole-Imidazoline Derivatives.

Chagas disease, caused by Trypanosoma cruzi, poses a significant public health challenge due to its widespread prevalence, limited therapeutic options, and adverse effects associated with available medications. In this study, we developed 13 novel pyrazole-imidazoline derivatives, inspired by a previously identified cysteine protease inhibitor, and evaluated their antiparasitic activity. Our in silico analyses predicted favorable physicochemical profiles and promising oral bioavailability for these derivatives. Upon phenotypic screening, we observed that these new derivatives exhibited low cytotoxicity (CC50 > 100 µM) and marked efficacy against intracellular amastigotes. Derivative 1k showed high activity (IC50 = 3.3 ± 0.2 µM), selectivity (SI = 73.9), and potency (pIC50 = 5.4). In a 3D cardiac microtissue model, 1k significantly reduced parasite load, matching the efficacy of benznidazole (Bz) even at lower concentrations. Both 1k and Bz effectively prevented parasite recrudescence; however, neither resulted in parasite sterility under the experimental conditions employed. The combination of 1k-Bz yielded an additive interaction, highlighting its potential for in vivo combination therapy. While structural changes abolished cysteine protease inhibition, incorporating a CF3 substituent at the para position and excluding the amino group enhanced antiparasitic activity. These findings reinforce the promise of the pyrazole-imidazoline scaffold and support further structural optimizations to develop innovative candidates for treating Chagas disease.