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Development of Simplified in Vitro P-Glycoprotein Substrate Assay and in Silico Prediction Models To Evaluate Transport Potential of P-Glycoprotein

2019; American Chemical Society; Volume: 16; Issue: 5 Linguagem: Inglês

10.1021/acs.molpharmaceut.8b01143

1543-8392

Rikiya Ohashi, Reiko Watanabe, Tsuyoshi Esaki, Tomomi Taniguchi, Nao Torimoto‐Katori, Tomoko Watanabe, Yuko Ogasawara, Tsuyoshi Takahashi, M. Tsukimoto, Kenji Mizuguchi,

Pharmacological Effects and Toxicity Studies

For efficient drug discovery and screening, it is necessary to simplify P-glycoprotein (P-gp) substrate assays and to provide in silico models that predict the transport potential of P-gp. In this study, we developed a simplified in vitro screening method to evaluate P-gp substrates by unidirectional membrane transport in P-gp-overexpressing cells. The unidirectional flux ratio positively correlated with parameters of the conventional bidirectional P-gp substrate assay (R2 = 0.941) and in vivo Kp,brain ratio (mdr1a/1b KO/WT) in mice (R2 = 0.800). Our in vitro P-gp substrate assay had high reproducibility and required approximately half the labor of the conventional method. We also constructed regression models to predict the value of P-gp-mediated flux and three-class classification models to predict P-gp substrate potential (low-, medium-, and high-potential) using 2397 data entries with the largest data set collected under the same experimental conditions. Most compounds in the test set fell within two- and three-fold errors in the random forest regression model (71.3 and 88.5%, respectively). Furthermore, the random forest three-class classification model showed a high balanced accuracy of 0.821 and precision of 0.761 for the low-potential classes in the test set. We concluded that the simplified in vitro P-gp substrate assay was suitable for compound screening in the early stages of drug discovery and that the in silico regression model and three-class classification model using only chemical structure information could identify the transport potential of compounds including P-gp-mediated flux ratios. Our proposed method is expected to be a practical tool to optimize effective central nervous system (CNS) drugs, to avoid CNS side effects, and to improve intestinal absorption.