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BioAssay: AID 720503

qHTS for Inhibitors of WRN Helicase: BLM Helicase Counterscreen for WRN Inhibitors

Inhibition of DNA repair is proposed as a strategy for combating cancer. Synthetic lethality is an approach that exploits preexisting DNA repair deficiencies in certain tumors to develop inhibitors of DNA repair pathways that compensate for the tumor-associated DNA repair deficiency. Because helicases play critical roles in the DNA damage response and in DNA repair, particularly in actively more ..
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 Tested Compounds
 Tested Compounds
All(611)
 
 
Active(368)
 
 
Inactive(110)
 
 
Inconclusive(133)
 
 
 Tested Substances
 Tested Substances
All(611)
 
 
Active(368)
 
 
Inactive(110)
 
 
Inconclusive(133)
 
 
AID: 720503
Data Source: NCGC (WRN-helicase-BLM-f1)
BioAssay Type: Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Probe Production Network
Deposit Date: 2013-07-09

Data Table ( Complete ):           View Active Data    View All Data
Target
BioActive Compounds: 368
Related Experiments
AIDNameTypeComment
651767qHTS for Inhibitors of WRN Helicase: SummarySummarydepositor-specified cross reference
651768qHTS for Inhibitors of WRN HelicaseConfirmatorydepositor-specified cross reference
720497qHTS for Inhibitors of WRN Helicase: Confirmatory Assay for Cherry-picked Compounds.Confirmatorysame project related to Summary assay
720499qHTS for Inhibitors of WRN Helicase: Thiazole Orange DNA Binding CounterscreenConfirmatorysame project related to Summary assay
Description:
Inhibition of DNA repair is proposed as a strategy for combating cancer. Synthetic lethality is an approach that exploits preexisting DNA repair deficiencies in certain tumors to develop inhibitors of DNA repair pathways that compensate for the tumor-associated DNA repair deficiency. Because helicases play critical roles in the DNA damage response and in DNA repair, particularly in actively dividing and replicating cells, characterization of synthetic lethal relationships of DNA helicases may be of value in developing improved anticancer treatment strategies; moreover, small molecules that specifically target a given DNA helicase may be useful for understanding its role in cellular nucleic acid metabolism. The goal of this project is to identify small molecule, non-covalent chemical inhibitors of the Werner syndrome (WS) helicase (WRN), which plays an important role in cell proliferation, the replication stress response, and DNA repair. Werner syndrome is a premature aging disorder that displays many clinical symptoms of aging at an accelerated rate. The WRN gene product that is defective in the chromosomal instability disorder has DNA helicase and exonuclease activities and interacts with a number of nuclear proteins to maintain genomic stability.

A 1536-well format, fluorescence based, high-throughput screen (AID 651768) has been developed in collaboration with the Brosh Laboratory at the National Institute of Aging (NIA) to screen for inhibitors of the WRN. This counterscreen is used to determine the activity of the validated WRN inhibitors against another related helicase, the Bloom's syndrome (BLM) helicase. Compounds / series that exhibit a 10-fold selectivity will be will be included for further consideration and target characterization of the WRN inhibitors.

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

MLPCN Grant: MH096530
Assay Submitter (PI): Robert Brosh, National Institute of Aging
Protocol
Three uL of reaction buffer (25 mM Tris-HCl pH 8.0, 50 mM NaCl, 2 mM MgCl2, 1 mM DTT, 0.01% Tween-20, and 1 nM BLM) was dispensed into a 1536-well Greiner black assay plate via solenoid-valve based nanoliter dispensers. Compounds (23 nl each in columns 5-48) and control (23 nl each in column 2 as dose-response) were transferred to the assay plate via a Kalypsys pin-tool equipped with a 1536-pin array. The plates were incubated for 15 min at room temperature, followed by the addition of 1 ul substrates (100 nM DNA and 2 mM ATP, final concentrations) to start the reaction. The plates were transferred into a ViewLux CCD imager where the reaction was measured in a discontinuous kinetic mode (two reads spaced 15 minutes apart, with intermediate room temperature incubation of the covered assay plate) using the standard red-fluorescence optics (excitation filter 525 nm, emission filter 598 nm). Enzyme-free and vehicle (DMSO) controls were included for signal normalization.
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
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 0.0009677143 uM (0.000967714μM**)% Activity at given concentration.Float%
16Activity at 0.00290 uM (0.00290314μM**)% Activity at given concentration.Float%
17Activity at 0.00871 uM (0.00870949μM**)% Activity at given concentration.Float%
18Activity at 0.026 uM (0.0261284μM**)% Activity at given concentration.Float%
19Activity at 0.078 uM (0.0783853μM**)% Activity at given concentration.Float%
20Activity at 0.235 uM (0.235156μM**)% Activity at given concentration.Float%
21Activity at 0.705 uM (0.705467μM**)% Activity at given concentration.Float%
22Activity at 2.116 uM (2.1164μM**)% Activity at given concentration.Float%
23Activity at 6.349 uM (6.34921μM**)% Activity at given concentration.Float%
24Activity at 19.05 uM (19.0476μM**)% Activity at given concentration.Float%
25Activity at 57.14 uM (57.1429μM**)% Activity at given concentration.Float%
26Compound QCNCGC designation for data stage: 'qHTS', 'qHTS Verification', 'Secondary Profiling'String

* Activity Concentration. ** Test Concentration.
Additional Information
Grant Number: MH096530

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