FP34-2 Characterization of a reporter cell line for high throughput screening of endogenous thyroid hormone receptor active compounds

Program: Abstracts - Orals, Featured Poster Presentations, and Posters
Session: FP34-Molecular Mechanisms in Thyroid Hormone Action & Cancer
Basic/Translational
Monday, June 17, 2013: 10:45 AM-11:15 AM
Presentation Start Time: 10:50 AM
Room 103 (Moscone Center)

Poster Board MON-414
J. David Furlow*1, Jaime Freitas2, Lucia N. Dobrawa1, Kyla Walter1, Eric S. Neff1, Monica L. Watson1, Nicole Miller3, Menghang Xia3, Ruili Huang3, Keith A. Houck4 and Albertinka J. Murk2
1UC Davis, Davis, CA, 2Wageningen University, Wageningen, Netherlands, 3NIH, Bethesda, MD, 4US EPA, Research Triangle Park, NC
We have recently reported the development of a stable reporter assay using GH3 cells (GH3.TRE-LUC) for thyroid hormone (TH) receptor (TR) active compounds1 that is amenable to high throughput screening. We confirmed that the parental GH3 cells express endogenous TH signaling components, including TRs, retinoid-X receptors (RXRs), nuclear receptor coactivators and corepressors, Type I and II deiodinases, and TH transporters. Furthermore, we determined that the cells induce a battery of known direct TH target genes including hairless, sonic hedgehog, and type I deiodinase, as well as several novel TH regulated genes. Responses of GH3.TRE-LUC reporter cells to 3,3’,5-triiodothyronine (T3) and TR isotype selective thyromimetics in the reporter gene assay showed excellent concordance with endogenous gene induction. Further, certain compounds of environmental concern such as bisphenol A inhibited T3 mediated induction of the reporter gene in a similar fashion to endogenous genes. To screen for additional TR active compounds, the GH3.TRE-LUC cells were next shown to perform well in initial quantitative high throughput screening assays. Several novel compounds were found in both agonist and antagonist mode screens, but somewhat unexpectedly, several natural and synthetic retinoids were also found to activate the reporter gene. Using RXR and RAR selective compounds, we show that retinoid induction of the reporter gene is likely via RXRs, suggesting that RXRs may be serving as nonsilent partners for the TRs in this context. In summary, the GH3.TRE-LUC cell line provides a new platform for quantitative high throughput assays for TR active compounds, but may also be sensitive to RXR interacting modulators of the RXR/TR heterodimeric complex in these cells.

(1) Freitas J, Cano P, Craig-Veit C, Goodson ML, Furlow JD, Murk AJ. Detection of thyroid hormone receptor disruptors by a novel stable in vitro reporter gene assay.  Toxicol In Vitro. 2011 25(1):257-266

Nothing to Disclose: JDF, JF, LND, KW, ESN, MLW, NM, MX, RH, KAH, AJM

*Please take note of The Endocrine Society's News Embargo Policy at http://www.endo-society.org/endo2013/media.cfm

Sources of Research Support: EPA STAR grant 83516401; Netherlands Organization for Health Research and Development (ZonMW) – NWO Grant 11.400.0075; these views represent the opinions of the authors and do not reflect the policies of the EPA.