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This version was published on February
1, 2008
Journal of Biomolecular Screening, Vol. 13, No. 2,
120-127 (2008)
DOI: 10.1177/1087057107313786
Quantitative High-Throughput Screening Using a Live-Cell cAMP Assay Identifies Small-Molecule Agonists of the TSH Receptor
Steve Titus
National Institutes of Health Chemical Genomics Center, National Human Genome Research Institute, Bethesda, Maryland
Susanne Neumann
Clinical Endocrinology Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland
Wei Zheng
National Institutes of Health Chemical Genomics Center, National Human Genome Research Institute, Bethesda, Maryland
Noel Southall
National Institutes of Health Chemical Genomics Center, National Human Genome Research Institute, Bethesda, Maryland
Sam Michael
National Institutes of Health Chemical Genomics Center, National Human Genome Research Institute, Bethesda, Maryland
Carleen Klumpp
National Institutes of Health Chemical Genomics Center, National Human Genome Research Institute, Bethesda, Maryland
Adam Yasgar
National Institutes of Health Chemical Genomics Center, National Human Genome Research Institute, Bethesda, Maryland
Paul Shinn
National Institutes of Health Chemical Genomics Center, National Human Genome Research Institute, Bethesda, Maryland
Craig J. Thomas
National Institutes of Health Chemical Genomics Center, National Human Genome Research Institute, Bethesda, Maryland
James Inglese
National Institutes of Health Chemical Genomics Center, National Human Genome Research Institute, Bethesda, Maryland
Marvin C. Gershengorn
Clinical Endocrinology Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland
Christopher P. Austin
National Institutes of Health Chemical Genomics Center, National Human Genome Research Institute, Bethesda, Maryland, austinc{at}mail.nih.gov
The thyroid-stimulating hormone (TSH; thyrotropin) receptor belongs to the glycoprotein hormone receptor subfamily of 7-transmembrane spanning receptors. TSH receptor (TSHR) is expressed mainly in thyroid follicular cells and is activated by TSH, which regulates the growth and function of thyroid follicular cells. Recombinant TSH is used in diagnostic screens for thyroid cancer, especially in patients after thyroid cancer surgery. Currently, no selective small-molecule agonists of the TSHR are available. To screen for novel TSHR agonists, the authors miniaturized a commercially available cell-based cyclic adenosine 3',5' monophosphate (cAMP) assay into a 1536-well plate format. This assay uses an HEK293 cell line stably transfected with the TSHR coupled to a cyclic nucleotide gated ion channel as a biosensor. From a quantitative high-throughput screen of 73,180 compounds in parallel with a parental cell line (without the TSHR), 276 primary active compounds were identified. The activities of the selected active compounds were further confirmed in an orthogonal homogeneous time-resolved fluorescence cAMP-based assay. Forty-nine compounds in several structural classes have been confirmed as the small-molecule TSHR agonists that will serve as a starting point for chemical optimization and studies of thyroid physiology in health and disease. (Journal of Biomolecular Screening 2008:120-127)
Key Words: thyroid-stimulating hormone • TSH • TSHR • TSHR agonist • quantitative high-throughput screening • qHTS • HTS • probe identification • CNG • PubChem
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