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BioAssay: AID 686978

qHTS for Inhibitors of human tyrosyl-DNA phosphodiesterase 1 (TDP1): qHTS in cells in absence of CPT

Human tyrosyl-DNA phosphodiesterase 1 (TDP1) is a novel repair gene, and we propose to use it as a new target for anticancer drug development. TDP1 is not an essential protein, but under treatment with topoisomerase I poison (camptothecin: CPT), TDP1 works as a critical factor for cell survival. To directly identify novel TDP1 inhibitors active in a cellular environment, we have knocked-out the more ..
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 Tested Compounds
 Tested Compounds
All(416837)
 
 
Active(63252)
 
 
Inactive(240194)
 
 
Inconclusive(115703)
 
 
 Tested Substances
 Tested Substances
All(424003)
 
 
Active(64212)
 
 
Inactive(243136)
 
 
Inconclusive(116655)
 
 
AID: 686978
Data Source: NCGC (TDP1100)
BioAssay Type: Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Probe Production Network
Deposit Date: 2013-05-16

Data Table ( Complete ):           Active    All
Target
BioActive Compounds: 63252
Depositor Specified Assays
AIDNameTypeComment
686981qHTS for Inhibitors of human tyrosyl-DNA phosphodiesterase 1 (TDP1): qHTS in cell-based assaysummary
Description:
Human tyrosyl-DNA phosphodiesterase 1 (TDP1) is a novel repair gene, and we propose to use it as a new target for anticancer drug development. TDP1 is not an essential protein, but under treatment with topoisomerase I poison (camptothecin: CPT), TDP1 works as a critical factor for cell survival. To directly identify novel TDP1 inhibitors active in a cellular environment, we have knocked-out the Tdp1 gene in chicken DT40 cells (Tdp1-/-) and generated a complemented counterpart cells that contains a stable transfection of the human TDP1 gene (Tdp1-/-;hTDP1 cells). For the primary screen, Tdp1-/-;hTDP1 cells will be exposed to small molecules in the presence or absence of CPT, and their growth kinetics will be evaluated after 48 hours by measuring ATP activity. If a given compound shows a synergistic effect with CPT, this compound could inhibit the repair pathway of CPT-induced lesions including the TDP1-mediated repair pathway. The hit compounds will then be evaluated in the presence or absence of CPT using Tdp1-/- cells. If a compound shows synergistic effect with CPT in Tdp1-/-;hTDP1 cells, but not with Tdp1-/- cells, such compound could be involved in the TDP1-mediated repair pathway inhibition. In tertiary assays, biochemical gel-based assays will be used to assess whether the hit compounds specifically target TDP1.
To this end, we have screened this cell line against the MLSMR in two primary screens. In one screen we add CPT and in the other we screen in the absence of CPT; this AID represents the screen where CPT was absent.

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

MLPCN Grant: MH095538
Assay Submitter (PI): Yves Pommier, NCI
Protocol
DT40-hTDP1 cells without 20 nM Camptothecin (add 20 microL of DMSO in 1 L of cell culture medium) were dispensed at 400 cells/5microL/well in tissue culture treated 1536-well white wall/solid bottom assay plates (Greiner Bio-One North America, NC, U.S.A.) using a Multidrop Combi 8 channel dispenser (Thermo Fisher, Waltham, MA, USA). 23 nL compounds and controls were transferred using the pin tool (Kalypsys, San Diego, CA, USA) to the assay plates. The assay plates were then incubated at 37 masculineC for a minimum 48 hr. Three microL of Cell Titer Glo solution was added to the plates and incubated at RT in dark for 30 min. Luminescence was read using ViewLux (Perkin Elmer) with 1 sec exposure slow speed, high gain and 2x binning.
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 0.0000295000 uM (2.95e-05μM**)% Activity at given concentration.Float%
16Activity at 0.0000590000 uM (5.9e-05μM**)% Activity at given concentration.Float%
17Activity at 0.0001503265 uM (0.000150327μM**)% Activity at given concentration.Float%
18Activity at 0.0002712146 uM (0.000271215μM**)% Activity at given concentration.Float%
19Activity at 0.0005895491 uM (0.000589549μM**)% Activity at given concentration.Float%
20Activity at 0.00117 uM (0.00117302μM**)% Activity at given concentration.Float%
21Activity at 0.00179 uM (0.0017877μM**)% Activity at given concentration.Float%
22Activity at 0.00299 uM (0.00299115μM**)% Activity at given concentration.Float%
23Activity at 0.00672 uM (0.00672249μM**)% Activity at given concentration.Float%
24Activity at 0.014 uM (0.0144692μM**)% Activity at given concentration.Float%
25Activity at 0.026 uM (0.0255963μM**)% Activity at given concentration.Float%
26Activity at 0.040 uM (0.0395262μM**)% Activity at given concentration.Float%
27Activity at 0.074 uM (0.074417μM**)% Activity at given concentration.Float%
28Activity at 0.167 uM (0.166893μM**)% Activity at given concentration.Float%
29Activity at 0.363 uM (0.362637μM**)% Activity at given concentration.Float%
30Activity at 0.628 uM (0.627936μM**)% Activity at given concentration.Float%
31Activity at 0.975 uM (0.974748μM**)% Activity at given concentration.Float%
32Activity at 1.849 uM (1.84892μM**)% Activity at given concentration.Float%
33Activity at 4.119 uM (4.11858μM**)% Activity at given concentration.Float%
34Activity at 9.037 uM (9.03699μM**)% Activity at given concentration.Float%
35Activity at 15.83 uM (15.8268μM**)% Activity at given concentration.Float%
36Activity at 21.08 uM (21.0829μM**)% Activity at given concentration.Float%
37Activity at 46.23 uM (46.2318μM**)% Activity at given concentration.Float%
38Activity at 92.54 uM (92.542μM**)% Activity at given concentration.Float%
39Activity at 165.6 uM (165.616μM**)% 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: MH095538

Data Table (Concise)
Classification
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