Bookmark and Share
BioAssay: AID 2517

qHTS Assay for Inhibitors of the Human Apurinic/apyrimidinic Endonuclease 1 (APE1)

The apurinic/apyrimidinic endonuclease APE1 is the primary mammalian enzyme responsible for the removal of abasic (or AP) sites in DNA and functions centrally in the base excision DNA repair (BER) pathway. Recent studies suggested a link between an overexpression of APE1 in many cancers and resistance of these tumor cells to radio- and chemotherapy. Thus, targeting APE1 could improve the efficacy of current treatment paradigms by promoting selective sensitization or protection of diseased and normal cells, respectively. ..more
_
   
 Tested Compounds
 Tested Compounds
All(345907)
 
 
Active(1173)
 
 
Inactive(334560)
 
 
Inconclusive(10759)
 
 
 Tested Substances
 Tested Substances
All(352185)
 
 
Active(1227)
 
 
Inactive(340097)
 
 
Inconclusive(10861)
 
 
AID: 2517
Data Source: NCGC (APE1917)
BioAssay Type: Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Probe Production Network
BioAssay Version:
Deposit Date: 2010-03-11
Modify Date: 2013-07-16

Data Table ( Complete ):           View Active Data    View All Data
Target
BioActive Compounds: 1173
Related Experiments
Show more
AIDNameTypeComment
1705qHTS Validation Assay for Inhibitors of the Human Apurinic/apyrimidinic Endonuclease 1 (APE1)Confirmatorydepositor-specified cross reference
1707Counterscreen for APE1 Inhibitors: Fluorescent Dye Displacement Validation AssayConfirmatorydepositor-specified cross reference
1708Counterscreen for APE1 Inhibitors: qHTS Validation Assay for Inhibitors of Endonuclease IVConfirmatorydepositor-specified cross reference
2324Probe Development Summary of Inhibitors of the Human Apurinic/apyrimidinic Endonuclease 1 (APE1)Summarydepositor-specified cross reference
2564Counterscreen for APE1 Inhibitors: Fluorescent Dye Displacement AssayConfirmatorydepositor-specified cross reference
2565Counterscreen for APE1 Inhibitors: qHTS Assay for Inhibitors of Endonuclease IVConfirmatorydepositor-specified cross reference
2572Confirmation qHTS Assay for Inhibitors of the Human Apurinic/apyrimidinic Endonuclease 1 (APE1)Confirmatorydepositor-specified cross reference
2573qHTS FP-Based Assay for Inhibitors of the Human Apurinic/apyrimidinic Endonuclease 1 (APE1)Confirmatorydepositor-specified cross reference
2741Counterscreen for APE1 Inhibitors: Confirmatory Fluorescent Dye Displacement AssayConfirmatorydepositor-specified cross reference
488940Radiotracer Incision Assay (RIA) for Inhibitors of Human Apurinic/apyrimidinic Endonuclease 1 (APE1)Confirmatorydepositor-specified cross reference
504595Inhibitors of APE1: Aqueous Solubility ProfilingOthersame project related to Summary assay
504603Inhibitors of APE1: Caco-2 Cell Permeability ProfilingOthersame project related to Summary assay
504618Inhibitors of APE1: Efflux Ratio ProfilingOthersame project related to Summary assay
504624Inhibitors of APE1: Mouse Plasma Stability ProfilingOthersame project related to Summary assay
504643Inhibitors of APE1: Metabolic Stability ProfilingOthersame project related to Summary assay
Description:
The apurinic/apyrimidinic endonuclease APE1 is the primary mammalian enzyme responsible for the removal of abasic (or AP) sites in DNA and functions centrally in the base excision DNA repair (BER) pathway. Recent studies suggested a link between an overexpression of APE1 in many cancers and resistance of these tumor cells to radio- and chemotherapy. Thus, targeting APE1 could improve the efficacy of current treatment paradigms by promoting selective sensitization or protection of diseased and normal cells, respectively.

Inhibition of APE1 activity was screened by utilizing double-stranded short substrate containing tetrahydrofuran (THF) abasic site labeled with rhodamine-type fluorophore (TAMRA) at the 5'-end and with non-fluorescent Black Hole Quencher-2 (BHQ-2) at the opposing 3'-end. An increase in the fluorescence intensity due to incision of the abasic site by APE1 was used to measure the enzyme activity.

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

MLPCN Grant: MH086444
Assay Provider: David M. Wilson, III, National Institute on Aging, NIH
Protocol
Three uL of enzyme solution (0.75 nM APE1 final concentration) was dispensed to 1536-well Greiner black solid bottom plates. Compounds (23 nL of two-fold, 16 pt dilution in duplicate to produce final concentrations in the 5.75 uM - 0.175 nM range) were transferred via Kalypsys pintool. The plates were incubated for 15 min at room temperature, and then 1 uL of substrate solution (50 nM final concentration of TAMRA/BHQ-2) was added to start the reaction. The plates were immediately transferred into ViewLux High-throughput CCD imager (Perkin-Elmer) in order to measure the reaction progress in kinetic mode (three reads every 60 seconds) using 525 nm excitation and 598 nm emission fluorescence protocol. The fluorescence intensity difference between the third and the first time points was used to compute reaction progress. Results were normalized to buffer only columns and NSC13755 positive controls and negative control DMSO treated wells.
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
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 0.0000657143 uM (6.57143e-05μM**)% Activity at given concentration.Float%
16Activity at 0.0001462857 uM (0.000146286μM**)% Activity at given concentration.Float%
17Activity at 0.0003274286 uM (0.000327429μM**)% Activity at given concentration.Float%
18Activity at 0.0007313956 uM (0.000731396μM**)% Activity at given concentration.Float%
19Activity at 0.00163 uM (0.00163429μM**)% Activity at given concentration.Float%
20Activity at 0.00368 uM (0.00367703μM**)% Activity at given concentration.Float%
21Activity at 0.00536 uM (0.00535733μM**)% Activity at given concentration.Float%
22Activity at 0.00876 uM (0.0087617μM**)% Activity at given concentration.Float%
23Activity at 0.020 uM (0.0195625μM**)% Activity at given concentration.Float%
24Activity at 0.031 uM (0.031097μM**)% Activity at given concentration.Float%
25Activity at 0.063 uM (0.063241μM**)% Activity at given concentration.Float%
26Activity at 0.101 uM (0.101146μM**)% Activity at given concentration.Float%
27Activity at 0.178 uM (0.178097μM**)% Activity at given concentration.Float%
28Activity at 0.453 uM (0.453392μM**)% Activity at given concentration.Float%
29Activity at 0.633 uM (0.632726μM**)% Activity at given concentration.Float%
30Activity at 1.022 uM (1.02175μM**)% Activity at given concentration.Float%
31Activity at 2.355 uM (2.35541μM**)% Activity at given concentration.Float%
32Activity at 3.686 uM (3.68628μM**)% Activity at given concentration.Float%
33Activity at 6.190 uM (6.19022μM**)% Activity at given concentration.Float%
34Activity at 12.36 uM (12.3568μM**)% Activity at given concentration.Float%
35Activity at 21.02 uM (21.0177μM**)% Activity at given concentration.Float%
36Activity at 50.43 uM (50.4335μM**)% Activity at given concentration.Float%
37Activity at 65.85 uM (65.8451μM**)% Activity at given concentration.Float%
38Activity at 119.8 uM (119.778μM**)% Activity at given concentration.Float%
39Activity at 186.3 uM (186.286μM**)% 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: MH086444

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