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BioAssay: AID 489039

ROR alpha selectivity assay 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-17A, IL-17F, and IL-22. Th17 cells have been implicated in graft versus host disease, autoimmune disease and asthma. RORgamma is induced in naive T helper cells in the presence of more ..
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 Tested Compounds
 Tested Compounds
All(172)
 
 
Active(30)
 
 
Inactive(30)
 
 
Inconclusive(115)
 
 
 Tested Substances
 Tested Substances
All(185)
 
 
Active(31)
 
 
Inactive(30)
 
 
Inconclusive(124)
 
 
AID: 489039
Data Source: NCGC (RORG643)
BioAssay Type: Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Probe Production Network
Deposit Date: 2010-11-22
Hold-until Date: 2011-11-16
Modify Date: 2011-11-16

Data Table ( Complete ):           Active    All
Target
Sequence: RAR-related orphan receptor alpha [Mus musculus]
Description ..   

     More BioActivity Data..
BioActive Compounds: 30
Depositor Specified Assays
AIDNameTypeComment
2551qHTS for inhibitors of ROR gamma transcriptional activityconfirmatoryPrimary qHTS for inhibitors of ROR gamma.
2604Quantitative high throughput screen for inhibitors of ROR gamma transcriptional activity: Summarysummary
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-17A, IL-17F, and IL-22. Th17 cells have been implicated in graft versus host disease, autoimmune disease and asthma. RORgamma 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 nonselective 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 ROR alpha, 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-RORalpha fusion that subsequently induced the UAS-luciferase reporter. Small molecule inhibitors that inhibit RORgamma and RORalpha, or components common to both, will be active in both assays and thus scored as nonselective.
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.
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 0.0001162824 uM (0.000116282μM**)% Activity at given concentration.Float%
15Activity at 0.0002712755 uM (0.000271276μM**)% Activity at given concentration.Float%
16Activity at 0.0005202792 uM (0.000520279μM**)% Activity at given concentration.Float%
17Activity at 0.0008193684 uM (0.000819368μM**)% Activity at given concentration.Float%
18Activity at 0.00156 uM (0.00156084μM**)% Activity at given concentration.Float%
19Activity at 0.00246 uM (0.00245811μM**)% Activity at given concentration.Float%
20Activity at 0.00468 uM (0.00468251μM**)% Activity at given concentration.Float%
21Activity at 0.00737 uM (0.00737432μM**)% Activity at given concentration.Float%
22Activity at 0.014 uM (0.0140475μM**)% Activity at given concentration.Float%
23Activity at 0.022 uM (0.0222268μM**)% Activity at given concentration.Float%
24Activity at 0.042 uM (0.0421586μM**)% Activity at given concentration.Float%
25Activity at 0.072 uM (0.0723714μM**)% Activity at given concentration.Float%
26Activity at 0.126 uM (0.126428μM**)% Activity at given concentration.Float%
27Activity at 0.194 uM (0.193506μM**)% Activity at given concentration.Float%
28Activity at 0.379 uM (0.379169μM**)% Activity at given concentration.Float%
29Activity at 0.630 uM (0.629789μM**)% Activity at given concentration.Float%
30Activity at 1.139 uM (1.1394μM**)% Activity at given concentration.Float%
31Activity at 2.052 uM (2.05216μM**)% Activity at given concentration.Float%
32Activity at 3.414 uM (3.41355μM**)% Activity at given concentration.Float%
33Activity at 5.483 uM (5.48303μM**)% Activity at given concentration.Float%
34Activity at 10.28 uM (10.283μM**)% Activity at given concentration.Float%
35Activity at 19.99 uM (19.9942μM**)% Activity at given concentration.Float%
36Activity at 30.90 uM (30.8997μM**)% Activity at given concentration.Float%
37Activity at 49.16 uM (49.1551μM**)% Activity at given concentration.Float%
38Activity at 92.17 uM (92.1659μM**)% Activity at given concentration.Float%
39Activity at 147.5 uM (147.465μM**)% 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: R03 DA026211-01

Data Table (Concise)
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