An Aromatic Diamidine That Targets Kinetoplast DNA, Impairs the Cell Cycle in Trypanosoma cruzi, and Diminishes Trypomastigote Release from Infected Mammalian Host Cells

Abstract

dTrypanosoma cruzi is the etiological agent of Chagas disease, affecting approximately 10 million people in the Americas and with some 40 million people at risk. The objective of this study was to evaluate the anti-T. cruzi activity of three new diamidines that have a 3,4-ethylenedioxy extension of the thiophene core, designated MB17, MB19, and MB38. All three diamidines exhibited dose-dependent inhibition of epimastigote replication. The mechanisms of action of these diamidines were investigated. Unlike MB17 and MB19, MB38 exhibited a significant increase in the number of annexin-propidium iodide double-labeled cells compared to levels in control parasites. As MB17 had shown a lower 50% inhibitory concentration (IC 50 ) against epimastigote growth, the mechanism of action of this drug was studied in more detail. MB17 triggered a decrease in the intracellular ATP levels. As a consequence, MB17 affected the genomic DNA and kinetoplast DNA (kDNA) and impaired the parasite cell cycle. Moreover, MB17 caused DNA fragmentation, with a more severe effect on kDNA than on nuclear DNA, resulting in dyskinetoplastic cells. MB17 was tested for toxicity and effectiveness for the treatment of infected CHO-K 1 cells, exhibiting a 50% cytotoxic concentration (CC 50 ) of 13.47 ؎ 0.37 M and an IC 50 of 0.14 ؎ 0.12 M against trypomastigote release. MB17 also diminished the infection index by 60% at 0.5 M. In conclusion, despite belonging to the same family, these diamidines have different efficiencies. To summarize, MB17 was the most potent of these diamidines against epimastigotes, producing DNA damage preferentially in kDNA, impairing the parasite cell cycle, and decreasing the infection index and trypomastigote release from infected mammalian host cells, with a high selectivity index (SI) (<90). These data suggest that MB17 could be an interesting lead compound against T. cruzi. A merican trypanosomiasis, or Chagas disease, affects approximately 10 million people, with 40 million people at risk of acquiring the infection. Chagas disease is endemic to South and Central America and the southern states of the United States. Currently, it is also spreading in Europe and the rest of North America due to the immigration of infected people, as well as through transmission of the disease by transfusions, transplants, and congenital mechanisms (1, 2). The disease is caused by the protozoan Trypanosoma cruzi, which shares some distinctive features with other trypanosomes, such as the presence of a flagellum and of a kinetoplast, a complex structure bearing the mitochondrial genome. In this case, the mitochondrial genomic DNA is referred to as kinetoplast DNA, or kDNA. It consists of a great number of relaxed circular DNA molecules interlocked with each other to form a catenated DNA network (3). T. cruzi has a complex life cycle, which occurs within invertebrate and vertebrate hosts (4-6). Chagas disease presents two clinical phases, the acute phase, which appears shortly after infection and is characterized by an evident parasitemia and ...

Recruitment kinetics of the homologous recombination pathway in procyclic forms of Trypanosoma brucei after ionizing radiation treatment

“…To determine whether IR treatment led to DNA strand breaks, a terminal deoxynucleotidyl transferase dUTP nick end-labeling (TUNEL) assay was performed according to protocols that were already standardized in trypanosomatids 16 , 32 , 33 (see the material and methods section). After IR treatment, the parasites were TUNEL labeled, and their fluorescence intensity was quantified by flow cytometry.…”

A Fluorinated Phenylbenzothiazole Arrests the Trypanosoma cruzi Cell Cycle and Diminishes the Infection of Mammalian Host Cells

“…An alteration of the cell cycle consisting of the accumulation of parasites in the G/M phase could be a hallmark for DNA damage (31,38). Our results revealed that BT10-and BT3-treated cells (Fig.…”

“…The infection index was calculated as the arithmetical product of the percentage of infected cells and the number of parasites per cell. The effect of BT10 on trypomastigote production was determined by collecting samples of the extracellular medium on the fifth day postinfection and counting the number of trypomastigotes in a Neubauer chamber (38). The results correspond to the means from three independent experiments, each performed in triplicate.…”

Mitigating RNA Toxicity in Myotonic Dystrophy using Small Molecules

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