Bookmark and Share
BioAssay: AID 624495

HTS Assay for Allosteric Antagonists of the Human D2 Dopamine Receptor: SAR in D2 Beta Arrestin Assay

There is great interest in identifying small molecule ligands for G-protein coupled receptors (GPCRs) as nearly 50% of all FDA-approved drugs target these important receptor proteins. Unfortunately, many of the ligands that are used as drugs, or as pharmacological tools, are not selective and exhibit problematic or limiting side effects due to inappropriate signaling activities. Amongst the more ..
_
   
 Tested Compounds
 Tested Compounds
All(14)
 
 
Active(13)
 
 
Inconclusive(1)
 
 
 Tested Substances
 Tested Substances
All(14)
 
 
Active(13)
 
 
Inconclusive(1)
 
 
AID: 624495
Data Source: NCGC (DOP2701)
BioAssay Type: Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Probe Production Network
Deposit Date: 2012-08-17
Hold-until Date: 2013-08-14
Modify Date: 2013-08-14

Data Table ( Complete ):           View Active Data    View All Data
Target
BioActive Compounds: 13
Related Experiments
Show more
AIDNameTypeComment
485359HTS Assay for Allosteric Antagonists of the Human D2 Dopamine Receptor: Antagonists SummarySummarydepositor-specified cross reference
485344HTS Assay for Allosteric Antagonists of the Human D2 Dopamine Receptor: Primary Screen for AntagonistsScreeningsame project related to Summary assay
485347HTS Assay for Positive Allosteric Modulators of the Human D2 Dopamine Receptor: Primary Screen for PotentiatorsScreeningsame project related to Summary assay
624453HTS Assay for Allosteric Antagonists of the Human D2 Dopamine Receptor: Hit Validation in primary assayConfirmatorysame project related to Summary assay
624455HTS Assay for Allosteric Antagonists of the Human D2 Dopamine Receptor: Hit Validation in HTRFConfirmatorysame project related to Summary assay
624459HTS Assay for Allosteric Antagonists of the Human D2 Dopamine Receptor: Hit Validation in Parental Cell LineConfirmatorysame project related to Summary assay
624461HTS Assay for Allosteric Antagonists of the Human D2 Dopamine Receptor: Hit Validation in Radioligand AssayOthersame project related to Summary assay
624494HTS Assay for Allosteric Antagonists of the Human D2 Dopamine Receptor: SAR in D2 Binding AssayOthersame project related to Summary assay
624496HTS Assay for Allosteric Antagonists of the Human D2 Dopamine Receptor: SAR in Primary Assay.Confirmatorysame project related to Summary assay
624500HTS Assay for Allosteric Antagonists of the Human D2 Dopamine Receptor: SAR in D3 Beta Arrestin AssayConfirmatorysame project related to Summary assay
624502HTS Assay for Allosteric Antagonists of the Human D2 Dopamine Receptor: SAR in D3 Binding AssayOthersame project related to Summary assay
Description:
There is great interest in identifying small molecule ligands for G-protein coupled receptors (GPCRs) as nearly 50% of all FDA-approved drugs target these important receptor proteins. Unfortunately, many of the ligands that are used as drugs, or as pharmacological tools, are not selective and exhibit problematic or limiting side effects due to inappropriate signaling activities. Amongst the dopamine receptors (DARs), the D2 is arguably one of the most validated drug targets in neurology and psychiatry. For instance, all receptor-based antiparkinsonian drugs primarily work via stimulating the D2 DAR, whereas all FDA-approved antipsychotic agents are antagonists of this receptor. The D2 DAR is also therapeutically targeted in other disorders such as restless legs syndrome, tardive dyskinesia, Tourette's syndrome, and hyperprolactinemia. However, most drugs targeting the D2 DAR are problematic, either being less efficacious than desired or possessing adverse side effects due to the activation or blockade of inappropriate signaling pathways. A novel approach for attaining greater selectivity of therapeutic agents targeting the D2 DAR is needed to identify and develop ligands that exhibit functionally-selective properties.

This is an orthogonal assay to test the lead compounds against the D2 receptor, using a beta arrestin assay.

NIH Molecular Libraries Probe Production Network [MLPCN]
NIH Chemical Genomics Center [NCGC]

Grant: NS0676421
PI Name: David Sibley, National Institutes of Neurological Disorders and Stroke
Protocol
DiscoveRx D2 cells were thawed from frozen stock and seeded at 2,100 cells/ 3 uL/well media (DiscoveRx Plating Reagent 2) with MultiDrop Combi dispenser (Thermo Scientific, Logan, UT) onto white tissue culture treated 1536-well Aurora plates (Brooks Automation, Chelmsford, MA) and allowed to attach overnight at 37 deg C, 5% CO2. Next, 23 nL/well of compound solutions in DMSO were added with a pintool transfer (Kalypsis, San Diego, CA). As the positive control, 1 uL/well dopamine (EC100, 57 uM final concentration) diluted in buffer (HBSS + 10 mM HEPES + SMB + 0.1% Tween-20) was dispensed to 1 column of each plate. The rest of the plate was dispensed with buffer only, 1 uL/well as well. The cells were stimulated by compounds or dopamine control for 90 minutes at 37 deg C, 5% CO2, after which 1.5 ul/well of DiscoveRx detection reagent was added with BioRAPTR FRD dispenser. The detection reagent was prepared by mixing of Galacton Star Substrate, Emerald II solution and PathHunter buffer (supplied by the assay kit) together at 1:5:19 proportion accordingly just prior to dispensing. The plates were incubated at ambient temperature for 1 hour and then the luminescent signal was read on ViewLux plate reader (PerkinElmer, Waltham, MA).
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
Show more
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
5Activity_ScoreActivity score.Integer
6Curve_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
7Fit_LogAC50The logarithm of the AC50 from a fit of the data to the Hill equation (calculated based on Molar Units).Float
8Fit_HillSlopeThe Hill slope from a fit of the data to the Hill equation.Float
9Fit_R2R^2 fit value of the curve. Closer to 1.0 equates to better Hill equation fit.Float
10Fit_InfiniteActivityThe asymptotic efficacy from a fit of the data to the Hill equation.Float%
11Fit_ZeroActivityEfficacy at zero concentration of compound from a fit of the data to the Hill equation.Float%
12Fit_CurveClassNumerical encoding of curve description for the fitted Hill equation.Float
13Excluded_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
14Max_ResponseMaximum activity observed for compound (usually at highest concentration tested).Float%
15Activity at 0.0003225714 uM (0.000322571μM**)% Activity at given concentration.Float%
16Activity at 0.0009677143 uM (0.000967714μM**)% Activity at given concentration.Float%
17Activity at 0.00290 uM (0.00290314μM**)% Activity at given concentration.Float%
18Activity at 0.00871 uM (0.00870949μM**)% Activity at given concentration.Float%
19Activity at 0.026 uM (0.0261284μM**)% Activity at given concentration.Float%
20Activity at 0.078 uM (0.0783853μM**)% Activity at given concentration.Float%
21Activity at 0.235 uM (0.235156μM**)% Activity at given concentration.Float%
22Activity at 0.705 uM (0.705467μM**)% Activity at given concentration.Float%
23Activity at 2.116 uM (2.1164μM**)% Activity at given concentration.Float%
24Activity at 6.349 uM (6.34921μM**)% Activity at given concentration.Float%
25Activity at 19.05 uM (19.0476μM**)% Activity at given concentration.Float%
26Activity at 57.14 uM (57.1429μM**)% Activity at given concentration.Float%
27Compound QCNCGC designation for data stage: 'qHTS', 'qHTS Verification', 'Secondary Profiling'String

* Activity Concentration. ** Test Concentration.
Additional Information
Grant Number: NS0676421

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