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BioAssay: AID 1573

Concentration Response Confirmation Assay for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2

Thyroid receptor (TR) regulates many homeostatic processes including basal metabolism, cardiovascular function, body weight, and lipid trafficking. TR modulators are potential therapeutics for obesity and hyperlipidemias but current thyroid analogs have undesirable side effects, particularly cardiac stimulation. To identify inhibitors that specifically prevent the interaction of TR with the more ..
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 Tested Compounds
 Tested Compounds
All(109)
 
 
Active(27)
 
 
Inactive(20)
 
 
Inconclusive(64)
 
 
 Tested Substances
 Tested Substances
All(112)
 
 
Active(27)
 
 
Inactive(20)
 
 
Inconclusive(65)
 
 
AID: 1573
Data Source: NCGC (TR757)
BioAssay Type: Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Probe Production Network
Deposit Date: 2009-03-20

Data Table ( Complete ):           Active    All
Target
BioActive Compounds: 27
Depositor Specified Assays
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AIDNameTypeProbeComment
1469qHTS for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2confirmatory
2552Irreversible Binding Assay for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2other
2444Protein Interaction Secondary Assay for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2, Alpha Screenconfirmatory
2448Androgen Receptor Selectivity Assay for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2confirmatory
2447Cell Viability Secondary Assay for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2confirmatory
2479Gene Reporter Secondary Assay for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2 in a Thyroid Receptor Assayconfirmatory
2449PPAR gamma Selectivity Assay For Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2confirmatory
2455Vitamin D Receptor Selectivity Assay for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2confirmatory
2487Secondary Confirmation Assay for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2, Fluorescein Fluoroprobeconfirmatory
2490Secondary Confirmation Assay for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2, Texas Red Fluoroprobeconfirmatory
2512qHTS Assay for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2: Probe Summarysummary1
Description:
NIH Molecular Libraries Probe Production Network [MLPCN]
NIH Chemical Genomics Center [NCGC]

MLPCN Grant: DK058080
Assay Provider: R. Kip Guy, St. Jude Children's Research Hospital

NCGC Assay Overview:
Thyroid receptor (TR) regulates many homeostatic processes including basal metabolism, cardiovascular function, body weight, and lipid trafficking. TR modulators are potential therapeutics for obesity and hyperlipidemias but current thyroid analogs have undesirable side effects, particularly cardiac stimulation. To identify inhibitors that specifically prevent the interaction of TR with the steroid receptor coregulator 2 (SRC2), a fluorescence polarization assay was screened. This assay detects interaction of the ligand-binding domain of human TRb with a Texas Red labeled SRC2 peptide, corresponding to a 20 amino acid region of the nuclear receptor interaction domain (Arnold et al., 2006). Small molecule inhibitors that block the interaction of TR and SRC2 are detected by a decrease in fluorescence polarization.

This assay is a confirmation assay for AID 1469.

Arnold, L. A.; Estebanez-Perpina, E.; Togashi, M.; Shelat, A.; Ocasio, C. A.; McReynolds, A. C.; Nguyen, P.; Baxter, J. D.; Fletterick, R. J.; Webb, P.; Guy, R. K. A high-throughput screening method to identify small molecule inhibitors of thyroid hormone receptor coactivator binding. Sci STKE 2006, p 13.
Protocol
NCGC Assay Protocol Summary:
For screening, 5 uL/well 0.6 uM TRb and 20 nM SRC2 Texas Red in protein buffer (20mM Tris hydrochloride, 100mM NaCl, 10% glycerol, 1mM EDTA, 0.01% NP-40, 1mM DTT, 1uM T3 and 5% DMSO) was dispensed into black solid 1536-well plates (Grenier) using a solenoid-based dispenser. Following transfer of 23 nL compound or DMSO vehicle by a pin tool, the plates were centrifuged 15 s at 1000 RPM and incubated 5 hr at ambient temperature. The plates were read by an Envision (Perkin Elmer) to detect fluorescence polarization of SRC2 Texas Red (555 nm excitation and 632 nm emission). Data were normalized to unbound (all components except TRb) and bound SRC2 Texas Red controls. In this assay, we provide % activity data for each concentration as well as potencies for the active compounds.

Keywords: NIH Roadmap, MLPCN, MLI, MLSMR, qHTS, NCGC, qHTS, thyroid hormone receptor, steroid receptor coactivator, fluorescence polarization
Comment
Compound Ranking:

1. Compounds are first classified as having full titration curves, partial modulation, partial curve (weaker actives), single point activity (at highest concentration only), or inactive. See data field "Curve Description". For this assay, apparent inhibitors are ranked higher than compounds that showed apparent activation.

2. For all inactive compounds, PUBCHEM_ACTIVITY_SCORE is 0. For all active compounds, a score range was given for each curve class type given above. Active compounds have PUBCHEM_ACTIVITY_SCORE between 40 and 100. Inconclusive compounds have PUBCHEM_ACTIVITY_SCORE between 1 and 39. Fit_LogAC50 was used for determining relative score and was scaled to each curve class' score range.
Result Definitions
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TIDNameDescriptionHistogramTypeUnit
OutcomeThe BioAssay activity outcomeOutcome
ScoreThe BioAssay activity ranking scoreInteger
1PhenotypeIndicates type of activity observed: inhibitor, activator, fluorescent, cytotoxic, inactive, or inconclusive.String
2Potency*Concentration at which compound exhibits half-maximal efficacy, AC50. Extrapolated AC50s also include the highest efficacy observed and the concentration of compound at which it was observed.FloatμM
3EfficacyMaximal efficacy of compound, reported as a percentage of control. These values are estimated based on fits of the Hill equation to the dose-response curves.Float%
4Analysis CommentAnnotation/notes on a particular compound's data or its analysis.String
5Curve_DescriptionA description of dose-response curve quality. A complete curve has two observed asymptotes; a partial curve may not have attained its second asymptote at the highest concentration tested. High efficacy curves exhibit efficacy greater than 80% of control. Partial efficacies are statistically significant, but below 80% of control.String
6Fit_LogAC50The logarithm of the AC50 from a fit of the data to the Hill equation (calculated based on Molar Units).Float
7Fit_HillSlopeThe Hill slope from a fit of the data to the Hill equation.Float
8Fit_R2R^2 fit value of the curve. Closer to 1.0 equates to better Hill equation fit.Float
9Fit_InfiniteActivityThe asymptotic efficacy from a fit of the data to the Hill equation.Float%
10Fit_ZeroActivityEfficacy at zero concentration of compound from a fit of the data to the Hill equation.Float%
11Fit_CurveClassNumerical encoding of curve description for the fitted Hill equation.Float
12Excluded_PointsWhich dose-response titration points were excluded from analysis based on outlier analysis. Each number represents whether a titration point was (1) or was not (0) excluded, for the titration series going from smallest to highest compound concentrations.String
13Max_ResponseMaximum activity observed for compound (usually at highest concentration tested).Float%
14Activity at 0.0000110132 uM (1.10132e-05μM**)% Activity at given concentration.Float%
15Activity at 0.0000220824 uM (2.20824e-05μM**)% Activity at given concentration.Float%
16Activity at 0.0000441640 uM (4.4164e-05μM**)% Activity at given concentration.Float%
17Activity at 0.0000884179 uM (8.84179e-05μM**)% Activity at given concentration.Float%
18Activity at 0.0001763765 uM (0.000176376μM**)% Activity at given concentration.Float%
19Activity at 0.0003532042 uM (0.000353204μM**)% Activity at given concentration.Float%
20Activity at 0.0007082659 uM (0.000708266μM**)% Activity at given concentration.Float%
21Activity at 0.00141 uM (0.0014117μM**)% Activity at given concentration.Float%
22Activity at 0.00282 uM (0.00282336μM**)% Activity at given concentration.Float%
23Activity at 0.00565 uM (0.00564671μM**)% Activity at given concentration.Float%
24Activity at 0.011 uM (0.0112936μM**)% Activity at given concentration.Float%
25Activity at 0.023 uM (0.022587μM**)% Activity at given concentration.Float%
26Activity at 0.045 uM (0.0452208μM**)% Activity at given concentration.Float%
27Activity at 0.090 uM (0.0902559μM**)% Activity at given concentration.Float%
28Activity at 0.181 uM (0.180971μM**)% Activity at given concentration.Float%
29Activity at 0.361 uM (0.361491μM**)% Activity at given concentration.Float%
30Activity at 0.722 uM (0.722043μM**)% Activity at given concentration.Float%
31Activity at 1.449 uM (1.4487μM**)% Activity at given concentration.Float%
32Activity at 2.893 uM (2.89284μM**)% Activity at given concentration.Float%
33Activity at 5.786 uM (5.78569μM**)% Activity at given concentration.Float%
34Activity at 11.57 uM (11.5712μM**)% Activity at given concentration.Float%
35Activity at 23.14 uM (23.1424μM**)% Activity at given concentration.Float%
36Activity at 46.29 uM (46.2851μM**)% Activity at given concentration.Float%
37Activity at 92.17 uM (92.1659μM**)% Activity at given concentration.Float%
38Activity at 184.3 uM (184.332μM**)% Activity at given concentration.Float%
39Activity at 368.7 uM (368.664μM**)% Activity at given concentration.Float%
40Compound QCNCGC designation for data stage: 'qHTS', 'qHTS Verification', 'Secondary Profiling'String

* Activity Concentration. ** Test Concentration.
Additional Information
Grant Number: DK058080

Data Table (Concise)
Classification
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