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Evaluating the DevTox-Germ Layer Reporter Platform for Screening Chemicals under the Toxic Substances Control Act
Citation:
Gamble, J. AND C. Deisenroth. Evaluating the DevTox-Germ Layer Reporter Platform for Screening Chemicals under the Toxic Substances Control Act. Presented at SOT Conference 2024: A New Approach Method (NAM) to Screen for the Impact of Endogenous Stress on Chemical Toxicity, Salt Lake City, UT, March 10 - 14, 2024. https://doi.org/10.23645/epacomptox.25533316
Impact/Purpose:
Presentation/Poster presented to SOT Conference 2024: A New Approach Method (NAM) to Screen for the Impact of Endogenous Stress on Chemical Toxicity
Description:
Background and Purpose The US EPA is committed to reviewing new chemicals for risks to human health and the environment as required by the Toxic Substances Control Act (TSCA). With limited exposure and hazard information available for new chemical submissions, the US EPA created the New Chemicals Collaborative Research Program (NCCRP) with the goal of modernizing human health and ecological risk assessments by incorporating innovative New Approach Methods (NAMs) into chemical reviews (EPA-HQ-OPPT-2022-0218-0001). One aim of the NCCRP is to apply in vitro NAMs to evaluate key areas of potential hazard including developmental toxicity. The stem cell-based DevTox-GLR (germ layer reporter) model platform enables rapid and human relevant screening for potential developmental toxicants (doi.org/10.3390/toxics10070392). The purpose of this study was to evaluate the overall predictivity and suitability of the model platform assays for screening data-poor, TSCA-relevant chemicals selected for the NCCRP. Methods This model platform is comprised of multiple assays (DevTox GLR-Endo, DevTox GLR-Meso, DevTox GLR-Ecto, DevTox GLR-Pluri), each of which assess different differentiation pathways for lineages associated with gastrulation. The RUES2-GLR transgenic pluripotent cell line expresses fluorescent reporter fusion protein biomarkers SOX17 (endoderm marker), Brachyury (mesoderm marker), and SOX2 (ectoderm and pluripotency marker). Through directed differentiation to each of the three germ layers, or maintenance in a pluripotent state, the assays were used to measure chemical perturbations via high-content image analysis of fluorescent biomarker expression to flag hazard potential for developmental toxicity. A training set of twelve known developmental toxicants (13-cis retinoic acid, 5,5-diphenylhydantoin, 5-fluouracil, all-trans retinoic acid, bisphenol A, busulfan, diethylstilbestrol, methotrexate, pomalidomide, sunitinib, thalidomide and valproic acid) and four non-developmental toxicants (acetaminophen, folic acid, penicillin G and saccharin) were tested on each assay to determine assay performance and predictivity. Qualification and selection of the most robust method, DevTox GLR-Endo, was then applied to screen 237 TSCA-relevant chemicals and reference compounds. Results Robust Z’-factor scores of 0.8, 0.4, 0.9, and 0.9 were observed for DevTox GLR-Endo, DevTox GLR-Meso, DevTox GLR-Ecto, and DevTox GLR-Pluri assays, respectively. Balanced accuracies for the chemical training set were 92% (DevTox GLR-Endo), 58% (DevTox GLR-Meso), 71% (DevTox GLR-Ecto), and 92% (DevTox GLR-Pluri). Additional testing between DevTox GLR-Endo and DevTox GLR-Pluri with 63 developmental and non-developmental toxicants revealed predictivity of 75% and 68%, respectively. Preliminary results for NCCRP screening display bioactivity for approximately 10% of tested chemicals (including controls) in the DevTox GLR-Endo assay when removing effects of cytotoxicity. Conclusions In conclusion, DevTox GLR-Endo provides the most robust performance and highest predictivity among the DevTox GLR platform assays and was selected for screening TSCA relevant chemicals for the NCCRP. These results will aid in modernizing the chemicals review process through incorporation of in vitro NAMs to inform risk assessment and decision making for human health effects. This abstract does not necessarily reflect EPA policy, nor endorse or recommend any products mentioned.
URLs/Downloads:
DOI: Evaluating the DevTox-Germ Layer Reporter Platform for Screening Chemicals under the Toxic Substances Control Act
JOHNGAMBLE_SOT2024_POSTERPRESENTATION_V3.PDF (PDF, NA pp, 1474.009 KB, about PDF)