Development of a bioprinter-based method for adding metabolic competence to high-throughput screening assays
Certain chemicals used in commercial products and present in the environment have the potential to interfere with biological systems. Identifying human health effects using in vitro new approach methodologies (NAMs) is challenged by the absence of metabolism in most test systems, which may lead to the under- or overestimation of potential health hazards. To retrofit existing high-throughput assays with metabolic competence, the EPA developed the alginate immobilization of metabolic enzymes platform (AIME; Deisenroth et al. 2020) and used it to screen the ToxCast library to evaluate chemicals’ estrogenic potential (Hopperstad et al. 2022). The AIME platform improves upon many conventional high-throughput screening assays by incorporating encapsulated hepatic S9-alginate microspheres to allow for consideration of the effects of liver metabolism. The ToxCast library data demonstrated a range of metabolism-dependent effects across a diverse chemical library. These results support the utility of the AIME platform for identifying false positive and false negative target assay effects and reprioritizing hazards based on metabolism-dependent bioactivity. They also highlight the need to evaluate the role of intrinsic xenobiotic metabolism for endocrine and other toxicity-related health effects.
Throughput and accessibility of the lid-based AIME method were improved by incorporating automated bioprinting to deposit S9‐encapsulated microspheres directly into standard microplates with requisite cofactors for phase I and II hepatic metabolism (Hopperstad and Deisenroth 2023). The AIME bioprinting metabolism method will be a useful tool for the tiered testing paradigm outlined in the CompTox Blueprint (Thomas et al. 2019), and is also more amenable to method transfer because it uses commercial hardware rather than custom proprietary lids. Planned activities for 2024 and beyond include an active cooperative research and development agreement with Proctor & Gamble to transfer the AIME method to a commercial contract research organization, application of the AIME method to additional endocrine assays, and application of the method to broad high-throughput phenotypic assays and transcriptomic profiling.