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BioAssay: AID 2444

Protein Interaction Secondary Assay for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2, Alpha Screen

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 confirm that inhibitors specifically prevent the interaction of TR with the more ..
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 Tested Compounds
 Tested Compounds
All(30)
 
 
Active(17)
 
 
Inactive(12)
 
 
Inconclusive(1)
 
 
 Tested Substances
 Tested Substances
All(30)
 
 
Active(17)
 
 
Inactive(12)
 
 
Inconclusive(1)
 
 
AID: 2444
Data Source: NCGC (TR802)
BioAssay Type: Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Probe Production Network
BioAssay Version:
Deposit Date: 2010-03-01
Hold-until Date: 2010-06-15
Modify Date: 2010-06-15

Data Table ( Complete ):           Active    All
Target
BioActive Compounds: 17
Depositor Specified Assays
AIDNameTypeProbeComment
1469qHTS for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2confirmatory
1479Total Fluorescence Counterscreen for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2confirmatory
1570Concentration Response Confirmation Assay for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2, Fluorescein Fluoroprobeconfirmatory
1571Total Fluorescence Confirmation Counterscreen for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2, Fluorescein Fluoroprobeconfirmatory
1572Total Fluorescence Confirmation Counterscreen for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2confirmatory
1573Concentration Response Confirmation Assay for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2confirmatory
1485Quantitative High-Throughput Screen for Inhibitors of the Interaction of Thyroid Hormone Receptor and Steroid Receptor Coregulator 2: Summarysummary
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 confirm that inhibitors specifically prevent the interaction of TR with the steroid receptor coregulator 2 (SRC2), an orthogonal TR-SRC2 interaction assay using Alpha Screen (PerkinElmer) was employed. This assay detects the interaction of the ligand-binding domain of human TR# with a SRC2 peptide, corresponding to a 20 amino acid region of the nuclear receptor interaction domain. Small molecule inhibitors that block the interaction of TR and SRC2 are detected by a decrease in chemiluminescence.
Protocol
NCGC Assay Protocol Summary:

Fifteen uL/well of 100 nM TRb and 100 nM SRC2-2-peg2-biotin in buffer (25 mM HEPES, 100 mM NaCl, 1 mM DTT, 0.1% BSA, 0.01% NP-40) was dispensed into white 384 well Optiplates (PerkinElmer). Following transfer of 135 nL compound (ranging from 90 nM to 200 uM), plates were incubated 1 hr, and 5 uL acceptor beads (6.3 ug/ml TRb antibody [Santa Cruz #sc-32754] and 40 ug/ml protein A beads in buffer) were added. After 30 min incubation, 5 uL of streptavidin donor beads were added and after 90 min incubation, the plates were read by an Envision (Perkin Elmer) to detect luminescence of the donor and acceptor bead interaction (680 nm excitation and 520-620 nm emission).
Comment
Compound Ranking:

Active compounds were assigned a score between 50 and 100 based on compound potency. Inactive compounds were assigned a score of 0 and inconclusive compounds were assigned a score of 25.
Result Definitions
TIDNameDescriptionHistogramTypeUnit
OutcomeThe BioAssay activity outcomeOutcome
ScoreThe BioAssay activity ranking scoreInteger
1ActivityIndicates type of activity observed: active or inactive.String
2Potency*The concentration of sample yielding half-maximal inhibition.FloatμM
3Compound QCNCGC designation for data stage: 'qHTS', 'qHTS Verification', 'Secondary Profiling'String

* Activity Concentration.
Additional Information
Grant Number: DK058080

Data Table (Concise)
Classification
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