Atividade tripanocida, citotóxica e genotóxica de novos derivados nitro-heterocíclico e os efeitos na expressão de genes do citocromo P450

Abstract

Chagas disease is a tropical, infectious and neglected disease that affects mostly poor regions of developing countries, infecting thousands of people every year. The lack of a safe and effective pharmacological treatment also contributes to the maintenance of the Chagas disease. The Institute of Pharmaceutical Technology, Oswaldo Cruz Foundation, synthesized new bioactive substances in order to develop a safe and effective alternative to the available treatment. They are: PNING 43-14 (ethyl-1-(1-methyl-4-nitro-1H-imidazol-5-yl)-1H-1,2,3-triazole-4-carboxylate), PNING 47-13 (4-cyclopropyl-1-(1-methyl-4-nitro-1H-imidazol-5-yl)-1H-1,2,3-triazole) and PNING 39-14 (1-(1-methyl-4-nitro-1H-imidazol-5-yl)-4-(4-pentylphenyl)-1H-1,2,3-triazole). The aim of this study was to assess the trypanocidal, cytotoxic and genotoxic activities of new nitro-heterocyclic derivatives, and the cytochrome P450 genes expressed, through the treatment of in vitro cell cultures models. The nitro-heterocyclic derivatives showed trypanocidal activity in the treatment (from 5.0 g/mL) of axenic amastigote evolutive form of Trypanosoma cruzi Y strain cell culture, and in the treatment of peritoneal macrophages primary mice cells infected with Trypanosoma cruzi, Y strain cells. In the assessment of cell viability through the colorimetric tests WST-1 and LDH, the nitro-heterocyclic derivatives PNING 43-14 and PNING 47-14 induced damages on high concentrations (500 and 1000 g/mL) and exposure time (48 h), in the treatment of the human hepatocarcinoma cells HepG2, only. The derivative PNING 39-14 induced damage in HepG2 and macrophage RAW 264.7 cells, using low concentrations (5.0 to 100 g/mL) and exposure times (from 3 h), being the most cytotoxic derivative. In the cell death assessment, observed by the annexin V and propidium iodide binding, in the treatment (50 to 1000 g/mL) of RAW 264.7 cells, the derivative PNING 43-14 induced necrosis, while the derivative PNING 39-14 induced apoptosis. In the Salmonella/microsome test, the derivatives induced pair bases substitutions mutations in the treatment of Salmonella enterica TA100 and TA102 strains, and deletions mutations in the TA97 strain, with and without exogenous metabolic activation. In the treatment of strains that overexpress nitroredutases (YG1021) and acetyltransferases (YG1024) enzymes, the derivatives induced cell death, with or without exogenous metabolic activation, mostly by the treatment with the derivative PNING 43-14 (100 to 1000 g/mL). In addition, the derivatives induced (50 to 1000 g/mL; 3 and 24 h) the micronuclei formation in HepG2 and RAW 264.7 cells, in the micronucleus test. In the assessment of cytochrome P450 gene expression in HepG2 cells, the derivatives PNING 43-14 and PNING 47-13 induced the 3A4 gene expression and inhibit the 1A2, 2B6 and 2E1 gene expression, while the derivative PNING 39-14 induced the 1A2, 2B6 and 2E1 gene expression and inhibit the 3A4 gene expression. The gene expression induced by the treatment using the derivative PNING 39-14 is related to cytotoxic and genotoxic positive responses. Despite the toxic positive results, there are lower concentrations of the derivatives that are safe and still hold an important trypanocidal activity.