Related Experiment Video
Updated: May 25, 2026
Small Molecule Screening and Toxicity Testing in Early-stage Zebrafish Larvae
Published on: March 7, 2025
Comparisons between in vitro whole cell imaging and in vivo zebrafish-based approaches for identifying potential
Adrian Hill1, Natalie Mesens, Margino Steemans
1Evotec Ltd., Abingdon, Oxford, United Kingdom.
Abstract:
Drug-induced liver injury (DILI) is a major cause of attrition during both the early and later stages of the drug development and marketing process. Reducing or eliminating drug-induced severe liver injury, especially those that lead to liver transplants or death, would be tremendously beneficial for patients. Therefore, developing new pharmaceuticals that have the highest margins and attributes of hepatic safety would be a great accomplishment. Given the current low productivity of pharmaceutical companies and the high costs of bringing new medicines to market, any early screening assay(s) to identify and eliminate pharmaceuticals with the potential to cause severe liver injury in humans would be of economic value as well. The present review discusses the background, proof-of-concept, and validation studies associated with high-content screening (HCS) by two major pharmaceutical companies (Pfizer Inc and Jansen Pharmaceutical Companies of Johnson & Johnson) for detecting compounds with the potential to cause human DILI. These HCS assays use fluorescent-based markers of cell injury in either human hepatocytes or HepG2 cells. In collaboration with Evotec, an independent contract lab, these two companies also independently evaluated larval zebrafish as an early-stage in vivo screen for hepatotoxicity in independently conducted, blinded assessments. Details about this model species, the need for bioanalysis, and, specifically, the outcome of the phenotypic-based zebrafish screens are presented. Comparing outcomes in zebrafish against both HCS assays suggests an enhanced detection for hepatotoxicants of most DILI concern when used in combination with each other, based on the U.S. Food and Drug Administration DILI classification list.
Insights
Early screening assays can identify drug-induced liver injury (DILI). Combining high-content screening (HCS) with zebrafish models enhances detection of potential hepatotoxicants, improving drug safety.
Area of Science:
- Pharmacology and Toxicology
- Drug Development
- Hepatotoxicity Research
Background:
- Drug-induced liver injury (DILI) is a significant challenge in drug development, leading to high attrition rates and patient harm.
- Developing safer pharmaceuticals with improved hepatic safety profiles is crucial for patient well-being and reducing healthcare costs.
- Early identification of compounds with DILI potential is economically valuable due to the high cost and low productivity in pharmaceutical research.
Purpose of the Study:
- To review the validation of high-content screening (HCS) assays for detecting DILI potential.
- To evaluate the utility of larval zebrafish as an in vivo model for early hepatotoxicity screening.
- To compare the effectiveness of HCS and zebrafish models in identifying DILI-causing compounds.
Main Methods:
- High-content screening (HCS) assays utilizing fluorescent markers of cell injury in human hepatocytes or HepG2 cells.
- In vivo screening of larval zebrafish for hepatotoxicity, conducted in blinded assessments.
- Independent evaluation of HCS and zebrafish models by major pharmaceutical companies (Pfizer, Johnson & Johnson) in collaboration with Evotec.
Main Results:
- HCS assays effectively identify compounds with DILI potential using human cells.
- Larval zebrafish screens provide valuable in vivo data for hepatotoxicity assessment.
- Combining HCS assays with zebrafish screens demonstrated enhanced detection of critical hepatotoxicants compared to individual methods.
Conclusions:
- Integrated screening strategies employing both HCS and zebrafish models offer improved prediction of DILI.
- These combined approaches can significantly aid in the early elimination of potentially harmful drug candidates.
- The findings support the use of these assays to enhance the safety and efficiency of drug development pipelines.
