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Updated: Oct 22, 2025
High-Throughput Cardiotoxicity Screening Using Mature Human Induced Pluripotent Stem Cell-Derived Cardiomyocyte Monolayers
Published on: March 24, 2023
A Novel High-Content Screening-Based Method for Anti-Trypanosoma cruzi Drug Discovery Using Human-Induced Pluripotent
Diogo Crispim Nascimento Portella1, Erik Aranha Rossi1,2,3, Bruno Diaz Paredes2,3
1Gonçalo Moniz Institute, Oswaldo Cruz Foundation (FIOCRUZ), Salvador, Brazil.
Insights
A new high-content screening method using human stem cell-derived cardiomyocytes (hiPSC-CMs) can rapidly assess anti-Trypanosoma cruzi drug activity and cardiotoxicity. This approach aids in discovering new Chagas disease treatments by evaluating compound efficacy and safety in human heart cells.
Area of Science:
- Parasitology and Tropical Diseases
- Stem Cell Biology
- Drug Discovery and Development
Background:
- Chagas disease, caused by Trypanosoma cruzi, is a major cause of heart failure in Latin America.
- Current treatments for Chagas disease are limited and ineffective during the chronic phase.
- Human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) offer a promising model for preclinical drug screening.
Purpose of the Study:
- To establish a novel high-content screening (HCS) assay using hiPSC-CMs.
- To simultaneously evaluate anti-Trypanosoma cruzi activity and cardiotoxicity of chemical compounds.
- To validate the HCS assay using known anti-Chagas disease compounds.
Main Methods:
- hiPSC-CMs were infected with Trypanosoma cruzi and treated with serial dilutions of compounds.
- Automated multiparametric analysis using high-content imaging assessed anti-parasitic activity (EC50) and cytotoxicity (CC50).
- Sublethal toxicity was evaluated via cytoskeletal integrity, nuclear morphology, mitochondrial function, and NT-pro-BNP release.
Main Results:
- The HCS assay successfully determined EC50 values for anti-T. cruzi activity, comparable to existing data.
- The thiazolidinone compound GT5B demonstrated potent trypanocidal activity in hiPSC-CMs.
- Sublethal toxicity markers correlated with cardiotoxicity, with mitochondrial changes indicating increased cytotoxicity.
Conclusions:
- The developed HCS assay is feasible for rapid assessment of anti-T. cruzi compounds and their cardiotoxicity.
- This hiPSC-CM-based platform can accelerate drug discovery for Chagas disease by identifying effective and safe novel compounds.
- The assay provides valuable insights into compound mechanisms and potential adverse effects on human cardiomyocytes.
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.
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