High-Throughput Assessment of Metabolism-Induced Toxicity of Compounds on a 384-Pillar Plate
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.
Key wordsMetabolism-induced toxicity Drug-metabolizing enzymes 384-Pillar plate 3D cell culture High-throughput screening (HTS)
This work was partially supported by the US Environmental Protection Agency (US EPA Transform Tox Testing Challenge), Medical & Bio Device (MBD) Korea, the Cleveland State University (Faculty Innovation Fund), and the National Institutes of Health (NIEHS R01ES025779).
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