Citation:

Stoker, T. High Throughput in vitro chemical screening for thyroid target sodium iodide symporter (NIS): species comparison, prioritization and computational approaches. British Toxicology Society Annual Meeting, Newcastle, Northumberland, UK, April 04 - 06, 2022.

Impact/Purpose:

This talk will provide an overview of the screening of ToxCast chemicals for sodium-iodide symporter (NIS) inhibition, the first step in the biosynthesis of thyroid hormones in humans and wildlife.  The overview provides a general roadmap from the initial development and validation to the implementation of the testing of Ph I, II and e1k chemicals.  The number of potential actives are discussed and secondary assays for species comparisons and extension to in vivo responses for one case study.  

Description:

Thyroid uptake of iodide via the sodium-iodide symporter (NIS) is the first step in the biosynthesis of thyroid hormones that are critical for health and development in humans and wildlife. The NIS has been a well-known target of the endocrine disrupting chemical perchlorate, but until now few environmental chemicals had been screened for NIS inhibition.  We first validated the use of an in vitro radioactive iodide uptake (RAIU) assay using a transduced human NIS-HEK (hNIS) cell line to identify NIS inhibitors.  Once the assay was validated and demonstrated good performance using known positive and negative controls, our laboratory screened over 1800 chemicals from the ToxCast Phase II (ph1v2, ph2 and e1k) libraries for NIS inhibition in the hNIS-HEK RAIU.  An initial concentration of 1×10-4 M is tested in the “single conc” testing with 651 chemicals demonstrating >20% RAIU inhibition were further tested in multiple-concentration, parallel RAIU and cell viability assays.  After filtering out chemicals with significant cytotoxicity, a total of 164 chemicals were identified as active NIS inhibitors. A novel chemical ranking system that incorporates multi-concentration RAIU and cytotoxicity responses was also developed as a standardized method for chemical prioritization.  This cytotoxicity-adjusted potency scoring (with NaClO4 having a reference score of 200) revealed 12 chemicals with moderate to strong RAIU inhibition (scored >100).  Chemotype enrichments were derived for the combined ~1,800 dataset to identify enriched features of NIS inhibitors, as well as chemotypes affiliated with cytotoxicity providing new information to support future data interpretation, structure-activity relationship (SAR), chemical use, and regulation. Top-ranked chemicals from the original hNIS screening were also evaluated in both the Fischer rat follicular cell line (FRTL-5) and hNIS RAIU assay using newly sourced chemicals to further strengthen the testing paradigm and to enable a rat vs. human species comparison. Some of the more potent NIS inhibitors are currently under evaluation in short-term weanling and pubertal rodent models to examine their effects on thyroid hormone synthesis in vivo. This work significantly expands current knowledge of NIS inhibition potential in environmental chemicals and contributes to an initiative to expand coverage of thyroid molecular targets, as well as the development of thyroid adverse outcome pathways (AOPs). This abstract does not necessarily reflect EPA policy. 

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