Phenotypic Screening In Vitro of Novel Aromatic Amidines against Trypanosoma cruzi

Abstract

The current treatment of Chagas disease (CD), based on nifurtimox and benznidazole (Bz), is unsatisfactory. In this context, we performed the phenotypic in vitro screening of novel mono-and diamidines and drug interaction assays with selected compounds. Ten novel amidines were tested for their activities against bloodstream trypomastigote (BT) and amastigote forms of Trypanosoma cruzi (Y and Tulahuen strains) and their toxicities for mammalian host cells (L929 cells and cardiac cells). Seven of 10 molecules were more active than Bz against BT, with the most active compound being the diamidine DB2267 (50% effective concentration [EC 50 ] ‫؍‬ 0.23 M; selectivity index ‫؍‬ 417), which was 28-fold more active and about 3 times more selective than the standard drug. Five of the six monoamidines were also more active than Bz. The combination of DB2267 and DB2236 in fixed-ratio proportions showed an additive effect (sum of fractional inhibitory concentrations < 4) on BT. Interestingly, when intracellular forms were exposed to DB2267, its activity was dependent on the parasite strain, being effective (EC 50 ‫؍‬ 0.87 ؎ 0.05 M) against a discrete typing unit (DTU) II strain (strain Y) but not against a representative DTU VI strain (strain Tulahuen) even when different vehicles (␤-cyclodextrin and dimethyl sulfoxide) were used. The intrinsic fluorescence of several diamidines allowed their uptake to be studied. Testing of the uptake of DB2236 (inactive) and DB2267 (active) by amastigotes of the Y strain showed that the two compounds were localized intracellularly in different compartments: DB2236 in the cytoplasm and DB2267 in the nucleus. Our present data encourage further studies regarding the activities of amidines and provide information which will help with the identification of novel agents for the treatment of CD.

Imidazole Derivatives as Promising Agents for the Treatment of Chagas Disease

“…CC morphology was evaluated by light microscopy, and the cellular viability of the CCs was determined with a standardized PrestoBlue test. The results were expressed as the difference in reduction between treated and nontreated cells, according to the manufacturer's instructions, and the LC 50 (minimum concentration that reduces cellular viability by 50%) values were determined by nonlinear regression using a sigmoid curve with a variable slope (29).…”

Repurposing Strategy of Atorvastatin against Trypanosoma cruzi: In Vitro Monotherapy and Combined Therapy with Benznidazole Exhibit Synergistic Trypanocidal Activity

“…The top concentrations used were 100 M for BZ and 50 M for AVA in a 24-h assay (against strain Y BT) and 15 M for BZ and 50 M for AVA in a 96-h assay (against strain Tulahuen intracellular forms). The top concentrations were used to prepare AVA and BZ solutions at fixed ratios of 5:0, 4:1, 3:2, 2:3, 1:4, and 0:5, as reported previously (40,41).…”

TrypanocidalActivity of Natural Sesquiterpenoids Involves Mitochondrial Dysfunction, ROS Production and Autophagic Phenotype in Trypanosoma cruzi

“…After 24 h of incubation, the cultures were washed to remove non-internalized parasites, and Pgd, PmTE and Efr were added (1.5 to 6 μg/mL) for 24 h at 37 °C. The results were expressed using the IC 50 /24 h concentration, which corresponded to the value that led to a 50% decrease in the infection index (percentage of infected host cells multiplied by the number of parasites per cell) [ 44 ].…”