Identification of Potential Sodium Iodide Symporter (NIS) Inhibitors in ToxCast Phase1_v2 Chemical Library Using in vitro Radioactive Iodide Uptake (RAIU) Assay
Citation:
Wang, J., D. Hallinger, A. Murr, A. Buckalew, T. Stoker, AND S. Laws. Identification of Potential Sodium Iodide Symporter (NIS) Inhibitors in ToxCast Phase1_v2 Chemical Library Using in vitro Radioactive Iodide Uptake (RAIU) Assay. 56th Annual Meeting, Society of Toxicology, Baltimore, MD, March 12 - 16, 2017.
Impact/Purpose:
This work addresses an urgent need and request from the Office of Coordination and Science Policy to develop in vitro assays to detect potential thyroid toxicants, and now provides a rapid screening approach that targets a key molecular initiating event, the uptake of iodide as mediated by the thyroid sodium iodide symporter (NIS), within the thyroid hormone pathway. Previously, we developed an assay to identify chemicals that can inhibit NIS using a novel cell line (hNIS-HEK293T-EPA) and demonstrated its high level of reproducibility and accuracy using a set of 50 reference and test chemicals. This abstract reports the use of this assay to screen the ToxCast Phase 1_v2 chemical library. It is expected that findings for the Phase1-v2 chemicals will be reported within the upcoming year, and that these data will be utilized during a 2017 FIFRA Science Advisory Panel review to obtain recommendations and feedback for the Agency’s Endocrine Disrupting Screening Program regarding the appropriate use of high throughput in vitro data for the thyroid hormone pathway. This work is of international interest because of the novel cell line created for this RAIU assay and its potential use for the development of an OECD test guideline.
Description:
Identification of Potential Sodium Iodide Symporter (NIS) Inhibitors in ToxCast Phase1_v2 Chemical Library Using in vitro Radioactive Iodide Uptake (RAIU) Assay Jun Wang1,2, Daniel R. Hallinger2, Ashley S. Murr2, Angela R. Buckalew1, Tammy E. Stoker2, Susan C. Laws21Oak Ridge Institute for Science and Education, U.S. Department of Energy, Oak Ridge, TN 37831, USA; 2Endocrine Toxicology Branch, TAD, NHEERL, ORD, U.S. Environmental Protection Agency, RTP, NC, 27711, USA.The NIS is a transmembrane glycoprotein that mediates active uptake of iodide into the thyroid, the critical first step of thyroid hormone synthesis. Inhibition of NIS function by xenobiotic chemicals has been demonstrated to suppress thyroid hormone synthesis and related physiological functions. However, current knowledge regarding NIS inhibition is limited for chemicals under the regulatory purview of the U.S. EPA’s Endocrine Disruptor Screening Program. Previously, we developed a RAIU screening assay to identify NIS inhibitors using a novel cell line (hNIS-HEK293T-EPA) and demonstrated its high reproducibility and accuracy. Here, we tested the ToxCast Phase 1_v2 chemical library with the RAIU assay. A total of 310 blind samples (293 unique chemicals) were initially screened in single-concentration format at 100 µM. Known NIS inhibitors, NaClO4, NaNO3 and NaSCN, were included as RAIU positive controls, while 2,4-dichlorophenoxyacetic acid was included as a negative control for both RAIU and cytotoxicity. Using 20% inhibition as the hit-call threshold, a total of 180 chemicals were further tested in multi-concentration (0.001 - 100 µM) format in the RAIU and cell viability assays. Toxicity-adjusted activity area (TAA, the area between the RAIU and cytotoxicity dose-response curves) was calculated to rank the tested chemicals. Using the TAA metric, 35 chemicals were ranked as higher priority for further evaluation as potential thyroid toxicants. A review of the literature showed that several of these putative NIS inhibitors (captan, fipronil, PFOS, 2-(thiocyanomethylthio) benzothiazole, and triphenyltin hydroxide) have been reported to disrupt thyroid hormone levels in vivo. Additional secondary in vitro and in vivo assays will further support the prioritization of top-ranked chemicals in the RAIU assay. This abstract does not reflect U.S.EPA policy.