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BioAssay: AID 624170

qHTS for Inhibitors of Glutaminase (GLS)

Glutaminase (GLS) catalyzes the hydrolysis of glutamine into glutamate and ammonia. It plays a critical role in glutaminolysis, which is an important energy source for proliferating cells. It was shown that c-Myc, an oncogenic transcription factor known to regulate microRNAs and to stimulate cell proliferation, increases glutamine metabolism by stimulating the expression of GLS via direct more ..
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 Tested Compounds
 Tested Compounds
All(405281)
 
 
Active(844)
 
 
Inactive(397800)
 
 
Inconclusive(6728)
 
 
 Tested Substances
 Tested Substances
All(409400)
 
 
Active(846)
 
 
Inactive(401810)
 
 
Inconclusive(6744)
 
 
AID: 624170
Data Source: NCGC (GLS100)
BioAssay Type: Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Probe Production Network
Deposit Date: 2012-05-22

Data Table ( Complete ):           Active    All
Target
BioActive Compounds: 844
Depositor Specified Assays
AIDNameTypeComment
624154qHTS for Inhibitors of Glutaminase (GLS): Summarysummary
Description:
Glutaminase (GLS) catalyzes the hydrolysis of glutamine into glutamate and ammonia. It plays a critical role in glutaminolysis, which is an important energy source for proliferating cells. It was shown that c-Myc, an oncogenic transcription factor known to regulate microRNAs and to stimulate cell proliferation, increases glutamine metabolism by stimulating the expression of GLS via direct suppression of miR23a and miR23b. Glutamine deprivation, or GLS siRNA treatment in PC3 cancer cells or P-493 B lymphoma cells caused significant decrease in cell proliferation [1]; while a non-potent, non-selective inhibitor of glutaminase BPTES (Ki = 2.9 uM) [2] also showed cancer cell growth inhibition [Slusher lab, unpublished data]. Therefore, GLS has been identified as a target for the treatment of cMyc-expressing cancer, where the upregulated GLS has recently been shown to play a critical metabolic role promoting cancer cell growth.

GLS is also thought to play a pathogenic role on HAD using a human HIV-1 infected macrophage system. Mechanistic studies show that during HIV-1 infection, GLS is tranlocated from mitochondria into the cytosol and extracellular space, where it converts mM levels of glutamine into glutamate, causing an increase level of excitotoxic glutamate [3, 4]. A prototype glutaminase inhibitor is shown to suppress neuronal toxicity caused by HIV-1 infected macrophages [5], suggesting the role of GLS in HIV-induced neurotoxicity. GLS inhibition limits glutamate release and provides neuroprotection against HIV-1 infected macrophages. Interestingly, glutaminase-mediated glutamate release from macrophage has also been shown to occur in a multiple sclerosis model [6], suggesting that GLS inhibition can also be of broad therapeutic interest for neuroinflammatory disorders.
The overall goal of this project is to identify small molecule inhibitors of GLS through quantitative high throughput screening (qHTS) of the MLSMR. A validated potent and selective inhibitor can be used as a tool molecule to further study the pharmacology of its target as well as explore its potential therapeutic application in cancer and inflammatory neurological disorders, such as HIV1-associated dementia (HAD) and multiple sclerosis.


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

MLPCN Grant: DA032470
Assay Submitter (PI): Barbara S. Slusher, Johns Hopkins University
Protocol
Three 3 uL of hKGA is dispensed into black well 1536-well plates in 45 mM KH2PO4 (pH 8.2), 0.02% Tween-20. Then 23 nL of compounds are added through pin tool transfer. To this, 1 uL of 1mM L-Glutamine and 1 uL of a solution of L-Glutamateoxidase, HRP, Amplex red are added. Plate is centrifuged for 25 sec before an initial fluorescence read is obtained on the ViewLux, after 30 min of RT incubation, the final read is taken on the ViewLux.
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 0.074 uM (0.0737μM**)% Activity at given concentration.Float%
16Activity at 0.184 uM (0.184μM**)% Activity at given concentration.Float%
17Activity at 0.369 uM (0.369128μM**)% Activity at given concentration.Float%
18Activity at 0.847 uM (0.847477μM**)% Activity at given concentration.Float%
19Activity at 1.840 uM (1.84μM**)% Activity at given concentration.Float%
20Activity at 2.060 uM (2.06μM**)% Activity at given concentration.Float%
21Activity at 3.360 uM (3.36μM**)% Activity at given concentration.Float%
22Activity at 4.002 uM (4.00201μM**)% Activity at given concentration.Float%
23Activity at 5.886 uM (5.88626μM**)% Activity at given concentration.Float%
24Activity at 9.110 uM (9.11025μM**)% Activity at given concentration.Float%
25Activity at 9.726 uM (9.72592μM**)% Activity at given concentration.Float%
26Activity at 14.48 uM (14.4833μM**)% Activity at given concentration.Float%
27Activity at 19.98 uM (19.9767μM**)% Activity at given concentration.Float%
28Activity at 29.42 uM (29.4153μM**)% Activity at given concentration.Float%
29Activity at 45.54 uM (45.5436μM**)% Activity at given concentration.Float%
30Activity at 48.53 uM (48.5293μM**)% Activity at given concentration.Float%
31Activity at 72.52 uM (72.5184μM**)% Activity at given concentration.Float%
32Activity at 99.36 uM (99.3609μM**)% Activity at given concentration.Float%
33Activity at 147.3 uM (147.251μM**)% Activity at given concentration.Float%
34Activity at 218.9 uM (218.888μM**)% Activity at given concentration.Float%
35Activity at 231.0 uM (231μM**)% Activity at given concentration.Float%
36Compound QCNCGC designation for data stage: 'qHTS', 'qHTS Verification', 'Secondary Profiling'String

* Activity Concentration. ** Test Concentration.
Additional Information
Grant Number: DA032470

Data Table (Concise)
Classification
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