Objective

Drug induced liver injury is one of the main reasons for safety failure of marketed drugs. Detecting drug candidates which are likely to damage the liver is essential in delivering safe medicines. To this end we have developed a high-content imaging-based screen in monolayer cultures of HepG2 (C3A clone) cells, grown in glucose-free medium. A set of 143 marketed drugs was used to validate the assay. Compounds were acoustically dispensed at 9 half-log spaced concentrations up to 316µM, and cells incubated for 24h. We used Hoecsht 33342, the methyl ester of tetramethylrhodamine (TMRM) and TOTO-3 to detect toxic effects on cell nuclei, mitochondria and cell membrane permeability. Imaging was performed live using the CellInsight platform with on-the-fly analysis. Dose-response curves were fitted to the processed imaging data in GeneData Analyzer and the output analyzed with a custom R script. We were able to classify the set of 143 compounds into DILI positive or negative categories with 64% sensitivity and 78% specificity by using a set of 9 imaging parameters and the maximum plasma concentrations of the drugs.  In the absence of pharmacokinetic information the assay’s sensitivity dropped down to 46%, in order to maintain the same high level of specificity. The high-content assay would deliver the most value when Cmax measurements are available or when reliable Cmax estimates can be calculated for drug candidates.