High-Throughput Assessment of Metabolism-Induced Toxicity of Compounds on a 384-Pillar Plate

Abstract

A variety of oxidative and conjugative enzymes are involved in the metabolism of compounds including drugs, which can be converted into toxic metabolites by Phase I drug-metabolizing enzymes (DMEs), such as the cytochromes P450 (CYP450s), and/or detoxified by Phase II DMEs, such as UDP-glucuronosyltransferases (UGTs), sulfotransferases (SULTs), and glutathione S-transferases (GSTs). Traditionally, primary hepatocytes containing a complete set of DMEs have been widely used as a gold standard to assess metabolism-induced compound toxicity. However, primary hepatocytes are expensive, have high donor variability in expression levels of DMEs, and rapidly lose liver-specific functions when the cells are maintained under standard in vitro cell culture conditions over time. To address this issue and rapidly profile metabolism-induced drug toxicity, we have developed a 384-pillar plate, which is complementary to conventional 384-well plates. In this chapter, we provide step-by-step procedures for three-dimensional (3D) cell printing on the 384-pillar plate coupled with DMEs and compounds in the 384-well plate for high-throughput assessment of metabolism-induced toxicity.