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BioAssay: AID 2763

Concentration response confirmation VP16 counterscreen for inhibitors of ROR gamma transcriptional activity

The retinoic acid-related orphan receptor (ROR) gamma is a transcription factor that has a central role in the differentiation of Th17 cells, a subset of T helper cells that secrete the inflammatory cytokines IL-17, IL-17F, and IL-22. Th17 cells have been implicated in graft versus host disease, autoimmune disease and asthma. ROR gamma is induced in naive T helper cells in the presence of more ..
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 Tested Compounds
 Tested Compounds
All(250)
 
 
Active(97)
 
 
Inactive(96)
 
 
Inconclusive(57)
 
 
 Tested Substances
 Tested Substances
All(250)
 
 
Active(97)
 
 
Inactive(96)
 
 
Inconclusive(57)
 
 
AID: 2763
Data Source: NCGC (RORG486)
BioAssay Type: Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Probe Production Network
Deposit Date: 2010-04-14

Data Table ( Complete ):           View Active Data    View All Data
Target
BioActive Compounds: 97
Related Experiments
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AIDNameTypeComment
2546VP16 counterscreen qHTS for inhibitors of ROR gamma transcriptional activityConfirmatorydepositor-specified cross reference: Primary counterscreen tested in the VP16 cell line at 6 concentrations from 46 uM to 3 nM
2604Quantitative high throughput screen for inhibitors of ROR gamma transcriptional activity: SummarySummarydepositor-specified cross reference
2551qHTS for inhibitors of ROR gamma transcriptional activityConfirmatorysame project related to Summary assay
2762Concentration response confirmation assay for inhibitors of ROR gamma transcriptional activityConfirmatorysame project related to Summary assay
489036VP16 selectivity assay for inhibitors of ROR gamma transcriptional activityConfirmatorysame project related to Summary assay
489037Secondary confirmation assay for inhibitors of ROR gamma transcriptional activityConfirmatorysame project related to Summary assay
489038DHR3 selectivity assay for inhibitors of ROR gamma transcriptional activityConfirmatorysame project related to Summary assay
489039ROR alpha selectivity assay for inhibitors of ROR gamma transcriptional activityConfirmatorysame project related to Summary assay
492954Mouse Th17 T cell differentiation assay for inhibitors of ROR gamma transcriptional activityConfirmatorysame project related to Summary assay
492962Mouse Th1 T cell differentiation assay for inhibitors of ROR gamma transcriptional activityOthersame project related to Summary assay
Description:
NIH Molecular Libraries Probe Production Network [MLPCN]
NIH Chemical Genomics Center [NCGC]

MLPCN Grant: R03 DA026211-01
Assay Provider: Dan Littman, New York University

Assay Overview:

The retinoic acid-related orphan receptor (ROR) gamma is a transcription factor that has a central role in the differentiation of Th17 cells, a subset of T helper cells that secrete the inflammatory cytokines IL-17, IL-17F, and IL-22. Th17 cells have been implicated in graft versus host disease, autoimmune disease and asthma. ROR gamma is induced in naive T helper cells in the presence of TGF-beta combined with IL-6, IL-21, or IL-23, and thereafter directs the expression of the Th17 lineage cytokines. To identify nonspecific inhibitors, a Drosophila Schneider cell line was stably transfected with two vectors: a gene expressing a fusion of the Gal4 DNA binding domain and the transactivation domain of the viral transcription factor VP16, under the control of the metallothionine promoter and a Photinus luciferase reporter regulated by the Gal4 binding site enhancer, UAS. The addition of copper to the medium induced expression of the Gal4-VP16 gamma fusion that subsequently induced the UAS-luciferase reporter. Small molecule inhibitors that inhibit components common to the RORgt and VP16 assays, such as the Gal4 DNA binding domain or the UAS luciferase reporter, will be active in both assays and thus scored as nonspecific.
Protocol
Assay Protocol Summary:

For screening, 600 cells in 4 uL/well were dispensed into white 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 incubated 1 hr at ambient temperature, and 1 uL/well copper sulfate (700 uM final concentration) was added. The plates were centrifuged 15 s at 1000 RPM and incubated 20 hr at ambient temperature. After addition of 1.5 uL Photinus luciferase detection reagent, the plates were incubated 10 min at ambient temperature and then read by an ViewLux (Perkin Elmer) to detect luminescence.
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.
Categorized Comment - additional comments and annotations
From ChEMBL:
Assay Type: Functional
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 0.0000520279 uM (5.20279e-05μM**)% Activity at given concentration.Float%
15Activity at 0.0002586940 uM (0.000258694μM**)% Activity at given concentration.Float%
16Activity at 0.0007760819 uM (0.000776082μM**)% Activity at given concentration.Float%
17Activity at 0.00233 uM (0.00232825μM**)% Activity at given concentration.Float%
18Activity at 0.00698 uM (0.00698474μM**)% Activity at given concentration.Float%
19Activity at 0.021 uM (0.0209542μM**)% Activity at given concentration.Float%
20Activity at 0.023 uM (0.0225014μM**)% Activity at given concentration.Float%
21Activity at 0.063 uM (0.0626363μM**)% Activity at given concentration.Float%
22Activity at 0.090 uM (0.0900058μM**)% Activity at given concentration.Float%
23Activity at 0.188 uM (0.188493μM**)% Activity at given concentration.Float%
24Activity at 0.563 uM (0.563011μM**)% Activity at given concentration.Float%
25Activity at 0.720 uM (0.720046μM**)% Activity at given concentration.Float%
26Activity at 1.694 uM (1.69428μM**)% Activity at given concentration.Float%
27Activity at 2.880 uM (2.88018μM**)% Activity at given concentration.Float%
28Activity at 5.092 uM (5.09187μM**)% Activity at given concentration.Float%
29Activity at 5.760 uM (5.76037μM**)% Activity at given concentration.Float%
30Activity at 15.23 uM (15.2292μM**)% Activity at given concentration.Float%
31Activity at 23.04 uM (23.0415μM**)% Activity at given concentration.Float%
32Activity at 46.08 uM (46.0829μM**)% Activity at given concentration.Float%
33Compound QCNCGC designation for data stage: 'qHTS', 'qHTS Verification', 'Secondary Profiling'String

* Activity Concentration. ** Test Concentration.
Additional Information
Grant Number: R03 DA026211-01

Data Table (Concise)
Data Table ( Complete ):     View Active Data    View All Data
Classification
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