Orthogonal assay for validation of Tox21 PPARγ data and applicability to in silico prediction model.

High-throughput screening data from the Tox21 database is used for prioritizing hazardous chemicals and building in silico-based toxicity prediction models. One of the Tox21 dataset, peroxisome proliferator-activated receptor-gamma (PPARγ), a nuclear receptor superfamily, identified various endpoints in HEK293 cells. PPARγ mediates various toxic effects when its receptors are activated or inhibited by ligands such as thiazolidinedione and GW9662. In this study, an orthogonal assay was constructed to verify the effectiveness of the Tox21 PPARγ data, and the effect of highly reliable data on in silico model construction was investigated. The orthogonal assay was a reporter gene assay based on the PPARγ ligand binding domain in CV-1 cells. Only 39% of agonists and 55% of antagonists had similar responses in CV-1 and HEK293 cells. Thus, the effectiveness of Tox21 data on PPARγ may vary depending on the cell line. However, in silico PLS-DA analysis with only high-reliability data (i.e., the same response in both cell lines), yielded more accurate prediction of the activity of potential chemical ligands, despite the small number of samples. Thus, obtaining reliable chemical screening data for PPARγ through orthogonal analysis, even for only limited chemicals, supports the construction of highly predictive in silico models with improved screening efficiency.