title: Quantitative high-throughput screening using a live-cell cAMP assay identifies small-molecule agonists of the TSH receptor.
abstract: 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.
authors.affiliation: National Institutes of Health Chemical Genomics Center, National Human Genome Research Institute, NIH, Bethesda, MD 20892-3370, USA.
authors.forename: Steve
authors.initials: S
authors.lastname: Titus
authors.forename: Susanne
authors.initials: S
authors.lastname: Neumann
authors.forename: Wei
authors.initials: W
authors.lastname: Zheng
authors.forename: Noel
authors.initials: N
authors.lastname: Southall
authors.forename: Sam
authors.initials: S
authors.lastname: Michael
authors.forename: Carleen
authors.initials: C
authors.lastname: Klumpp
authors.forename: Adam
authors.initials: A
authors.lastname: Yasgar
authors.forename: Paul
authors.initials: P
authors.lastname: Shinn
authors.forename: Craig J
authors.initials: CJ
authors.lastname: Thomas
authors.forename: James
authors.initials: J
authors.lastname: Inglese
authors.forename: Marvin C
authors.initials: MC
authors.lastname: Gershengorn
authors.forename: Christopher P
authors.initials: CP
authors.lastname: Austin
mesh_terms: D000465:Algorithms; D002138:Calibration; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000032:analysis*; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D005189:False Positive Reactions; D006801:Humans; D008904:Miniaturization; D008954:Models, Biological; D021281:Quantitative Structure-Activity Relationship; D011989:Receptors, Thyrotropin / Q000819:agonists* / Q000235:genetics; D054852:Small Molecule Libraries / Q000032:analysis*; D014162:Transfection
chemical_list: D011989:Receptors, Thyrotropin; D054852:Small Molecule Libraries; D000242:Cyclic AMP