High content analysis assay for prediction of human hepatotoxicity in HepaRG and HepG2 cells

•HCA assay successfully detected clinical DILI potential in HepaRG and HepG2 cells.•HepaRG and HepG2 cells had different responses to clinical DILI compounds.•The known biological changes related to clinical DILI were detected in HCA assay.•HepaRG and HepG2 cells should be used in HCA assays for hepatotoxic evaluation.

Drug-induced liver injury (DILI) results in the termination of drug development or withdrawal of a drug from the market. The establishment of a predictive, high-throughput preclinical test system to evaluate potential clinical DILI is therefore required. Here, we established a high content analysis (HCA) assay in human hepatocyte cell lines such as the HepaRG with normal expression levels of CYP enzymes and HepG2 with extremely low expression levels of CYP enzymes. Clinical DILI or non-DILI compounds were evaluated for reactive oxygen species (ROS) production, glutathione (GSH) consumption, and mitochondrial membrane potential (MMP) attenuation. A proportion of DILI compounds induced ROS generation, GSH depletion, and MMP dysfunction, which was consistent with reported mechanisms of DILI of these compounds. In particular, DILI compounds that deplete GSH via reactive metabolites exhibited a more marked decrease in intracellular GSH or increase in ROS production in HepaRG cells than in HepG2 cells. Comparison of the two cell lines with different levels of CYP expression might help clarify the contribution of metabolism to hepatocyte toxicity. These results suggest that the HCA assay in HepaRG and HepG2 cells might help improve the accuracy of evaluating clinical DILI potential during drug screening.