Screening Pesticides for Estrogenic and (anti)Androgenic Activity in Support of Endocrine Disruptor Screening Program (EDSP) Revival_SETAC2024
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EPA’s Endocrine Disruptor Screening Program (EDSP) was established in 1998 to identify pesticides and other chemicals that disrupt human endocrine system signaling. EPA plans to use EDSP’s tiered testing framework to support the risk assessment of pesticides under FIFRA, the Federal Insecticide, Fungicide, and Rodenticide Act. Out of 403 pesticides undergoing FIFRA registration review in 2023, 317 lacked adequate reproductive toxicity data, including 30 pesticides with potential estrogenic (human estrogen receptor; ER) and/or anti-androgenic activity (human androgen receptor; AR) identified by ToxCast high-throughput screening (HTS). We propose using sensitive, validated in vitro cell-based bioassays to supplement ToxCast data for these 30 pesticides and compare purported AR antagonist activity with concurrent cytotoxicity assessment. To measure ER agonist activity, we used the T47D-kbluc cell line, which endogenously expresses ER-alpha and was stably transfected with an ER-sensitive luciferase reporter gene. Two out of four suspected ER agonists, fenhexamid (in vitro EC50: 16.84 µM) and flumetralin (in vitro EC50: 2.43 µM), showed significant estrogenic activity in our assay when compared to 17-beta estradiol positive control, while hydramethylnon and pyridaben were ER-inactive and exhibited cytotoxicity at high concentrations. AR antagonist activity was measured using a CV1 cell line virally transduced with an AR gene (chAR) and AR-sensitive luciferase reporter gene. To ensure that any apparent AR antagonist effects were receptor-mediated, rather than cytotoxic in nature, this transcriptional activation assay was run alongside the MTT rapid colorimetric cytotoxicity assay. These Tier 1-type in vitro assays enable us to efficiently and accurately screen for pesticides that pose the greatest risk to human and environmental health and support appropriate resource allocation for Tier 2 in vivo testing. Furthermore, the in vitro screening approach used to assess the ER and anti-AR activity of these pesticides provides a model for validating HTS results and the continued expansion of tiered screening programs like EDSP.
The views expressed in this abstract are those of the author(s) and do not necessarily represent the views or policies of the U.S. Environmental Protection Agency.