Abstract
Chagas disease is caused by Trypanosoma cruzi infection and remains a relevant cause of chronic heart failure in Latin America. The pharmacological arsenal for Chagas disease is limited, and the available anti-T. cruzi drugs are not effective when administered during the chronic phase. Cardiomyocytes derived from human-induced pluripotent stem cells (hiPSC-CMs) have the potential to accelerate the process of drug discovery for Chagas disease, through predictive preclinical assays in target human cells. Here, we aimed to establish a novel high-content screening- (HCS-) based method using hiPSC-CMs to simultaneously evaluate anti-T. cruzi activity and cardiotoxicity of chemical compounds. To provide proof-of-concept data, the reference drug benznidazole and three compounds with known anti-T. cruzi activity (a betulinic acid derivative named BA5 and two thiazolidinone compounds named GT5A and GT5B) were evaluated in the assay. hiPSC-CMs were infected with T. cruzi and incubated for 48 h with serial dilutions of the compounds for determination of EC50 and CC50 values. Automated multiparametric analyses were performed using an automated high-content imaging system. Sublethal toxicity measurements were evaluated through morphological measurements related to the integrity of the cytoskeleton by phalloidin staining, nuclear score by Hoechst 33342 staining, mitochondria score following MitoTracker staining, and quantification of NT-pro-BNP, a peptide released upon mechanical myocardial stress. The compounds showed EC50 values for anti-T. cruzi activity similar to those previously described for other cell types, and GT5B showed a pronounced trypanocidal activity in hiPSC-CMs. Sublethal changes in cytoskeletal and nucleus scores correlated with NT-pro-BNP levels in the culture supernatant. Mitochondrial score changes were associated with increased cytotoxicity. The assay was feasible and allowed rapid assessment of anti-T. cruzi action of the compounds, in addition to cardiotoxicity parameters. The utilization of hiPSC-CMs in the drug development workflow for Chagas disease may help in the identification of novel compounds.
Highlights
Chagas disease, caused by the hemoflagellate protozoan Trypanosoma cruzi, affects approximately 6 to 7 million people worldwide [1, 2]
There is an urgent need to increase the therapeutic arsenal for chronic Chagas disease through drug discovery or repurposing [2, 6]
To evaluate the susceptibility of human-induced pluripotent stem cells (hiPSCs)-CMs to T. cruzi infection and to define the optimal MOI to be used in the assay, a preliminary test was performed using MOIs 5 and 10. hiPSCCMs were highly permissive to T. cruzi infection, leading to similar infection rates (54% and 44% for MOIs 5 and 10, respectively)
Summary
Chagas disease, caused by the hemoflagellate protozoan Trypanosoma cruzi, affects approximately 6 to 7 million people worldwide [1, 2]. Cardiac involvement occurs in up to 30% of the cases, leading to chronic Chagas cardiomyopathy (CCC), a disease that continues to be ranked among the most frequent etiologies of chronic heart failure in Latin American countries [3, 4]. There is an urgent need to increase the therapeutic arsenal for chronic Chagas disease through drug discovery or repurposing [2, 6]. In this context, the incorporation of innovative approaches in the preclinical in vitro screening process of anti-T. cruzi may contribute to accelerate the drug discovery process
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