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BioAssay: AID 624031

S16 Schwann cell viability assay (CellTiter-Glo assay)

Charcot-Marie-Tooth (CMT) disease was first characterized by Jean-Martin Charcot and Pierre Marie in France and, independently, by Howard Henry Tooth in England in 1886. CMT is one of the most common inherited neurological diseases, affecting approximately 1 in 2,500 Americans. CMT patients typically exhibit muscle atrophy in the extremities and sensory loss. The most prevalent type of CMT is more ..
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 Tested Compounds
 Tested Compounds
All(2791)
 
 
Active(105)
 
 
Inactive(2280)
 
 
Inconclusive(410)
 
 
 Tested Substances
 Tested Substances
All(2814)
 
 
Active(106)
 
 
Inactive(2298)
 
 
Inconclusive(410)
 
 
 Related BioAssays
 Related BioAssays
AID: 624031
Data Source: NCGC (cmt-p4-fda-celltiter_regid)
BioAssay Type: Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Probe Production Network
BioAssay Version:
Deposit Date: 2012-04-07
Hold-until Date: 2012-04-30
Modify Date: 2012-04-30

Data Table ( Complete ):           View Active Data    View All Data
BioActive Compounds: 105
Related Experiments
AIDNameTypeComment
624036Inhibitors for the S16 Schwann cell PMP22 intronic elementSummarydepositor-specified cross reference
624030Biochemical firefly luciferase enzyme assay for NPCConfirmatorysame project related to Summary assay
624032S16 Schwann cell PMP22 intronic element firefly luciferase assayConfirmatorysame project related to Summary assay
624044S16 Schwann cell PMP22 intronic element beta-lactamase assayConfirmatorysame project related to Summary assay
Description:
Charcot-Marie-Tooth-Association [CMTA]
National Center for Advancing Translational Sciences [NCATS]

Charcot-Marie-Tooth (CMT) disease was first characterized by Jean-Martin Charcot and Pierre Marie in France and, independently, by Howard Henry Tooth in England in 1886. CMT is one of the most common inherited neurological diseases, affecting approximately 1 in 2,500 Americans. CMT patients typically exhibit muscle atrophy in the extremities and sensory loss. The most prevalent type of CMT is known as CMT1A and caused by a duplication of the PMP22 gene which leads to over-expression of this protein. PMP22 is a glycosylated intrinsic membrane protein accounting for 2-5% of the myelin protein content, and its over-expression results in demyelination and subsequently axonal loss.

A pair of cellular transcription-based assays was developed in the S16 rat Schwann cell lines (Hai, M., et al., 2002) stably transfected with orthogonal reporters - firefly luciferase (Fluc) and beta-lactamase (beta-Lac) - whose expression is driven by an intronic regulatory element of the PMP22 gene (Jones, E.A., et al., 2011) to recapitulate endogenous expression of PMP22. A comprehensive library of approved drugs, the NCGC Pharmaceutical Collection (Huang, R., et al., 2011), was screened in the cross-validating platform of the orthogonal reporter assays to identify drug modulators inhibitory towards PMP22 expression.

A set of counter-screens was devised to address non-specific activity of the library. An ATP-dependent CellTiter-Glo assay (developed by Promega Corp) was utilized to control cell viability. In addition, biochemical Fluc enzyme assay was performed to assess direct inhibition of the Fluc enzymatic activity.
References
Hai, M., Muja, N., DeVries, G.H., Quarles, R.H. and Patel, P.I. Comparative analysis of Schwann cell lines as model systems for myelin gene transcription studies. J Neurosci Res, 2002. 69(4): pp 497-508
Huang, R., Southall, N., Wang, Y., Yasgar A., Shinn P., Jadhav, A. , Nguyen, D.-T. and Austin, C.P., The NCGC pharmaceutical collection: a comprehensive resource of clinically approved drugs enabling repurposing and chemical genomics. Sci Transl Med, 2011. 3(80): p. 80ps16.
Protocol
Native S16 cells were seeded in white solid-bottom 1536 well plates at between 500-750 cells/6uL in DMEM medium containing 10% FBS, w/o phenol red. After overnight incubation at 37 degree Celsius/5% C02, 23 nL of compounds or DMSO were delivered to each well using a pin tool, followed by 24h-incubation at 37 degree Celsius/5% C02. Then one volume of CellTiter-Glo assay reagent (Promega) was added to each well for cell viability measured by a ViewLux plate reader (PerkinElmer). The % activity was determined by normalizing to the average readings from wells containing no cells relative to DMSO-treated cells (0% Activity). The concentration-effect curves were then classified based on curve quality (r2), response magnitude and degree of measured activity.
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, toxic compounds are ranked higher than non-toxic compounds.
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. Toxic 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 PubChem:
Assay Cell Type: S16
From ChEMBL:
Assay Type: Functional
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 0.0001640000 uM (0.000164μM**)% Activity at given concentration.Float%
16Activity at 0.0007350000 uM (0.000735μM**)% Activity at given concentration.Float%
17Activity at 0.0008220000 uM (0.000822μM**)% Activity at given concentration.Float%
18Activity at 0.00164 uM (0.00164μM**)% Activity at given concentration.Float%
19Activity at 0.00382 uM (0.00381809μM**)% Activity at given concentration.Float%
20Activity at 0.00822 uM (0.00822μM**)% Activity at given concentration.Float%
21Activity at 0.019 uM (0.0191059μM**)% Activity at given concentration.Float%
22Activity at 0.041 uM (0.0411μM**)% Activity at given concentration.Float%
23Activity at 0.095 uM (0.0954603μM**)% Activity at given concentration.Float%
24Activity at 0.206 uM (0.206μM**)% Activity at given concentration.Float%
25Activity at 0.477 uM (0.477338μM**)% Activity at given concentration.Float%
26Activity at 1.030 uM (1.03μM**)% Activity at given concentration.Float%
27Activity at 2.387 uM (2.38669μM**)% Activity at given concentration.Float%
28Activity at 5.336 uM (5.33578μM**)% Activity at given concentration.Float%
29Activity at 11.50 uM (11.5μM**)% Activity at given concentration.Float%
30Activity at 25.70 uM (25.7μM**)% Activity at given concentration.Float%
31Activity at 57.50 uM (57.5μM**)% Activity at given concentration.Float%
32Compound QCNCGC designation for data stage: 'qHTS', 'qHTS Verification', 'Secondary Profiling'String

* Activity Concentration. ** Test Concentration.

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