Efeito tripanocida in silico e in vitro de P-aminochalconas sintéticas

Abstract

Chagas disease (CD) is a neglected tropical disease of global health concern, caused by the flagellate protozoan Trypanosoma cruzi, being endemic in Latin America and present in North America and Europe. The treatment against CD recommended by the World Health Organization is benznidazole, however, this therapy is limited and of low safety, requiring new therapeutic options. Chalcones are an important class of natural compounds that have shown antichagasic potential. Thus, the aim of the study was to evaluate the activity of synthetic p-aminochalcones against T. cruzi. The molecules 1-(4-

aminophenyl)-3-phenylprop-2-en-1-one (CPAB) and 1-(4-aminophenyl)-5-phenylpenta- 2,4-dien-1-one (CPAC) were evaluated for their cytotoxicity by the MTT reduction assay

in LLC-MK2 cells. The investigation of the effect on epimastigotes and trypomastigotes was carried out by determining the percentage of viable parasites. In the tests with amastigotes, concentrations of 15.6 and 31.2 μM were used. Then, flow cytometry assays were performed with the epimastigotes forms, in order to evaluate the profile of cell death (7-AAD/AxPE), production of cytoplasmic ROS (DCFH2-DA) and mitochondrial transmembrane potential (Rho123). In addition, molecular docking simulations were performed with the parasite targets, such as Trypanothione reductase, cruzain and TcGAPDH enzymes. In the cytotoxicity analyses, CC50 values estimated at 85.6 ± 9.2 μM were observed for CPAB, while the CPAC molecule proved to be less toxic, with CC50 1115 ± 381.7 μM. The molecules of interest exhibited activity against the epimastigotes forms in the three treatment times for CPAB (IC50 24h = 38.4 ± 6.2 μM; IC50 48h = 20.7 ± 3.3 μM and IC50 72h = 19.2 ± 3 .4 μM) and CPAC (24h IC50 = 74.8 ± 4.2 μM; 48h IC50 = 61.6 ± 5 μM and 72h IC50 = 52.3 ± 1.8 μM). The effect on trypomastigotes showed that CPAB and CPAC chalcones were able to cause parasite death, with estimated LC50 values of 33.3 ± 3.9 μM and 73 ± 15.9 μM, respectively. CPAB and CPAC chalcones were able to reduce the percentage of infected cells and the number of amastigotes per cell, demonstrated by the selectivity index of 2.6 and 15.3, respectively. Cytometry assays demonstrated that the antichagasic effect of the studied molecules was related to a possible necrotic activity with alteration of the mitochondrial potential. In molecular docking studies, targeting the enzymes Trypanothione reductase, cruzain and TcGAPDH, it was possible to observe interactions with the catalytic site and other important regions of these main pharmacological targets of T. cruzi. Thus, it is concluded that the studied chalcones had an antichagasic effect on the evolutionary forms of T. cruzi, with possible induction of necrosis and oxidative stress, with this effect being potentially greater when compared to the reference drug.