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BioAssay: AID 652105

qHTS for Inhibitors of phosphatidylinositol 5-phosphate 4-kinase (PI5P4K)

Phosphatidylinositol (PI) signaling has been shown to impact a large and diverse set of cellular processes including proliferation, survival, and growth; their dysregulation is common in cancer and other diseases. Recently, PI5P has been shown to regulate the tumor suppressor ING2. In addition to finding PI5P, we discovered the type II phosphatidylinositol 5-phosphate 4-kinase (PI5P4K) family more ..
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 Tested Compounds
 Tested Compounds
All(328848)
 
 
Active(4076)
 
 
Inactive(318371)
 
 
Inconclusive(6420)
 
 
 Tested Substances
 Tested Substances
All(329140)
 
 
Active(4081)
 
 
Inactive(318634)
 
 
Inconclusive(6425)
 
 
AID: 652105
Data Source: NCGC (PI5P4K100)
BioAssay Type: Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Probe Production Network
Deposit Date: 2013-03-14

Data Table ( Complete ):           View Active Data    View All Data
Target
BioActive Compounds: 4076
Related Experiments
AIDNameTypeComment
652103qHTS for Inhibitors of phosphatidylinositol 5-phosphate 4-kinase (PI5P4K): SummarySummarydepositor-specified cross reference
743285qHTS for Inhibitors of PI5P4K: ADP-Glo counter assayConfirmatorydepositor-specified cross reference
743379On Hold
743380On Hold
743381On Hold
743382On Hold
743383On Hold
743286qHTS for Inhibitors of PI5P4K: Confirmation in Primary AssayConfirmatorysame project related to Summary assay
Description:
Phosphatidylinositol (PI) signaling has been shown to impact a large and diverse set of cellular processes including proliferation, survival, and growth; their dysregulation is common in cancer and other diseases. Recently, PI5P has been shown to regulate the tumor suppressor ING2. In addition to finding PI5P, we discovered the type II phosphatidylinositol 5-phosphate 4-kinase (PI5P4K) family (alpha, beta, gamma isoforms) that catalyze the conversion of PI5P to PI(4,5)P2. Alternatively, PI(4,5)P2 can also be synthesized through phosphatidylinositol-4-phosphate (PI4P) by the type I phosphatidylinositol 4-phosphate 5-kinases (PI4P5K). These PI5P4K enzymes therefore represent one way by which cells can regulate endogenous PI5P levels and play important roles in insulin signaling and in stress responses. Furthermore, recent findings have demonstrated a critical role for PI5P4K in tumor cell growth and support a potential role in oncogenesis for PI5P4K.

This proposal aims to identify and optimize specific modulators of human PI5P4K, phosphatidylinositol-5-phosphate 4-kinase (Type II PIPK), a family of kinases that regulate AKT signaling and tumor cell growth. In this project we aim to identify inhibitors of human PI5P4K using a quantitative high-throughput screening approach against the MLSMR containing ~400,000 small molecules.

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

MLPCN Grant: MH096575
Assay Submitter (PI): Atsuo Sasaki, Beth Israel Deaconess Medical Center
Protocol
2 microL of PI5P4Kalpha/substrate buffer (columns 1-2, 5-48) are dispensed into Greiner white solid bottom 1536-well assay plates; column 3 receives only PI5P4Kalpha buffer as 0x substrate controls; column 4 receives only substrate buffer as 0x PI5P4Kalpha controls. Final assay buffer consists of 0.038 M HEPES pH 7.4, 0.28 mM EGTA, 0.1% CHAPS, and 5% DMSO (final concentration). Compounds are then transferred via Kalypsys pin tool equipped with 1536-pin array (10 nL slotted pins, V&P Scientific, San Diego, CA). Following addition of compound, 1 microL of ATP (5 microM final concentration, all columns) is added to initiate the reaction. ATP is diluted in an assay buffer consisting of 0.02 M HEPES pH 7.4, 60 mM MgCl2, 15 microM ATP, and 0.1% CHAPS. The plates are then incubated at room temperature for 60 minutes. ADP-Glo reagent 1 is added (2 microL) and plates incubated for 45 minutes, before ADP-Glo reagent 2 (4 microL) is added and plates incubated for 30 minutes. They are then transferred to a ViewLux high-throughput CCD imager (PerkinElmer) wherein single end-point measurements of luminescence are acquired using a clear filter.
Comment
Compounds whose "max activity" is <= -50 are considered "active" and assigned a score of 90; compounds where the "max activity" is > -50% and < -30% are consider "inconclusive" and are assigned a score of 50; for compounds with a "max activity" >= -30% they are considered "inactive" and are assigned a score of 10.
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 0.00130 uM (0.00130269μM**)% Activity at given concentration.Float%
16Activity at 0.00391 uM (0.00390808μM**)% Activity at given concentration.Float%
17Activity at 0.012 uM (0.0117243μM**)% Activity at given concentration.Float%
18Activity at 0.035 uM (0.0351728μM**)% Activity at given concentration.Float%
19Activity at 0.106 uM (0.105519μM**)% Activity at given concentration.Float%
20Activity at 0.317 uM (0.316556μM**)% Activity at given concentration.Float%
21Activity at 0.950 uM (0.949668μM**)% Activity at given concentration.Float%
22Activity at 2.849 uM (2.849μM**)% Activity at given concentration.Float%
23Activity at 8.547 uM (8.54701μM**)% Activity at given concentration.Float%
24Activity at 15.40 uM (15.4μM**)% Activity at given concentration.Float%
25Activity at 25.64 uM (25.641μM**)% Activity at given concentration.Float%
26Activity at 76.90 uM (76.9μM**)% Activity at given concentration.Float%
27Activity at 76.92 uM (76.9231μM**)% Activity at given concentration.Float%
28Compound QCNCGC designation for data stage: 'qHTS', 'qHTS Verification', 'Secondary Profiling'String

* Activity Concentration. ** Test Concentration.
Additional Information
Grant Number: MH096575

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