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Updated: May 11, 2026
In Vitro Drug Screening Against All Life Cycle Stages of Trypanosoma cruzi Using Parasites Expressing β-galactosidase
Published on: November 5, 2021
An MRC-5 cell based high-throughput, high-content imaging assay to identify hits against Trypanosoma cruzi
Bilal Zulfiqar1, Melissa L Sykes1, Fanny B Escudié2
1Discovery Biology, Centre for Cellular Phenomics, Griffith University, Nathan, Queensland, 4111, Australia; School of Environment and Science, Griffith University, Nathan, Queensland, 4111, Australia.
Abstract:
The discovery of effective therapies for Trypanosoma cruzi (T. cruzi), the causative agent of Chagas disease, remains one of the most pressing challenges in parasitology and global health. Despite the significant burden posed by Chagas disease, especially in Latin America, only two drugs, benznidazole and nifurtimox, are currently available for treatment. These drugs are often limited by side effects and long treatment durations. There is an urgent need for effective new therapies, requiring innovative and physiologically relevant assay platforms for high-throughput identification of compounds active against T. cruzi. Here we discuss the development, optimization, evaluation and validation of a robust and highly reproducible high-throughput, high-content imaging assay in a 384-well microplate format to quantitatively assess the effects of compounds on intracellular T. cruzi amastigotes infecting MRC-5 human lung fibroblasts. The multiplexed assay design enables concurrent evaluation of compound-induced cytotoxicity on host cells within the same well, serving as an early indicator of host cell viability and compound selectivity.
Insights
Developing new Chagas disease treatments is crucial. This study presents a high-throughput imaging assay to efficiently screen compounds against Trypanosoma cruzi (T. cruzi) while assessing host cell toxicity.
Area of Science:
- Parasitology
- Drug Discovery
- Global Health
Background:
- Chagas disease, caused by Trypanosoma cruzi (T. cruzi), significantly impacts global health, particularly in Latin America.
- Current treatments (benznidazole, nifurtimox) have limitations including side effects and long treatment durations.
- There is a critical need for novel, effective therapies against T. cruzi.
Purpose of the Study:
- To develop and validate a high-throughput, high-content imaging assay for T. cruzi drug discovery.
- To quantitatively assess the efficacy of compounds against intracellular T. cruzi amastigotes.
- To concurrently evaluate compound-induced host cell cytotoxicity and selectivity.
Main Methods:
- Development and optimization of a robust, reproducible high-throughput assay in a 384-well microplate format.
- Utilized high-content imaging to quantify intracellular T. cruzi amastigotes in infected MRC-5 human lung fibroblasts.
- Implemented a multiplexed assay design for simultaneous assessment of parasite inhibition and host cell viability.
Main Results:
- The developed assay is highly reproducible and suitable for high-throughput screening.
- The assay enables quantitative assessment of compound effects on intracellular T. cruzi amastigotes.
- Concurrent evaluation of host cell viability provides an early indicator of compound selectivity.
Conclusions:
- This validated high-throughput imaging assay is a valuable platform for identifying novel anti-T. cruzi compounds.
- The assay facilitates efficient screening of potential drug candidates for Chagas disease.
- The multiplexed design aids in the early selection of compounds with favorable safety profiles.
