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BioAssay: AID 2095

qHTS Assay for Activators of Human Muscle isoform 2 Pyruvate Kinase: Summary

Matthew G. Vander Heiden, M.D., Ph.D. [Harvard Medical School, 77 Avenue Louis Pasteur, Boston, MA 02115] ..more
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 Tested Compounds
 Tested Compounds
All(6)
 
 
Probe(6)
 
 
Active(6)
 
 
 Tested Substances
 Tested Substances
All(6)
 
 
Probe(6)
 
 
Active(6)
 
 
AID: 2095
Data Source: NCGC (PYKH254a)
BioAssay Type: Summary, Candidate Probes/Leads with Supporting Evidence
Depositor Category: NIH Molecular Libraries Probe Production Network
BioAssay Version:
Deposit Date: 2009-11-03
Modify Date: 2012-02-07

Data Table ( Complete ):           View Active Data    View All Data
Target
Sequence: pyruvate kinase, muscle isoform M2 [Homo sapiens]
Description ..   
Protein Family: Pyruvate_Kinase

Gene:PKM     Related Protein 3D Structures     More BioActivity Data..
BioActive Compounds: Chemical Probe: 6    Active: 6
Related Experiments
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AIDNameTypeComment
1540Secondary assay for Activators of Human Pyruvate Kinase M2 isoformConfirmatorydepositor-specified cross reference: human PKM2 activator secondary LDH assay
1631qHTS Assay for Activators of Human Muscle isoform 2 Pyruvate KinaseConfirmatorydepositor-specified cross reference: human PKM2 activator primary luminescent assay
1751Confirmation Concentration-Response Assay for Activators of Human Muscle isoform 2 Pyruvate KinaseConfirmatorydepositor-specified cross reference: human PKM2 activator confirmation luminescent assay
2533Confirmation Concentration-Response Assay for Activators of Human Muscle isoform 2 Pyruvate Kinase: for Probe SARConfirmatorydepositor-specified cross reference: human PKM2 activator confirmation luminescent assay for probe SAR
2534Secondary Concentration-Response Assay for Activators of Human Reticulocyte Pyruvate Kinase: for Probe SARConfirmatorydepositor-specified cross reference: human PKM2 activator confirmation luminescent assay for R isoform for probe SAR
2535Secondary Concentration-Response Assay for Activators of Human Liver Pyruvate Kinase: for Probe SARConfirmatorydepositor-specified cross reference: human PKM2 activator confirmation luminescent assay for L isoform for probe SAR
2536Secondary Concentration-Response Assay for Activators of Human Muscle isoform 1 Pyruvate Kinase: for Probe SARConfirmatorydepositor-specified cross reference: human PKM2 activator confirmation luminescent assay for M1 isoform for probe SAR
2562Secondary LDH Assay for Activators of Human Reticulocyte Pyruvate Kinase: for Probe SARConfirmatorydepositor-specified cross reference: human PKM2 activator secondary LDH assay for R isoform for probe SAR
2576Secondary LDH Assay for Activators of Human Pyruvate Kinase M2 isoform: for Probe SARConfirmatorydepositor-specified cross reference: human PKM2 activator secondary LDH assay for probe SAR
2620Secondary LDH Assay for Activators of Human Pyruvate Kinase M1 Isoform: for Probe SARConfirmatorydepositor-specified cross reference: human PKM2 activator secondary LDH assay for M1 isoform for probe SAR
2625Secondary LDH Assay for Activators of Human Liver Pyruvate Kinase: for Probe SARConfirmatorydepositor-specified cross reference: human PKM2 activator secondary LDH assay for L isoform for probe SAR
2653Secondary Assay for Activators of Human Pyruvate Kinase M2 isoform - Cell Titer Glo Cytotoxicity for Probe SARConfirmatorydepositor-specified cross reference: human PKM2 activator secondary cytotoxicity assay for probe SAR
602359Extended Characterization of Activators of Human Muscle isoform 2 Pyruvate Kinase: SummarySummarydepositor-specified cross reference
602377qHTS for Activators of Human Muscle Isoform Pyruvate Kinase: Extended Characterization Using A Lactate Dehydrogenase (LDH) AssayConfirmatorysame project related to Summary assay
602379qHTS for Activators of Human Muscle Isoform Pyruvate Kinase: Extended Characterization against Liver Pyruvate KinaseConfirmatorysame project related to Summary assay
602381qHTS for Activators of Human Muscle Isoform 2 Pyruvate Kinase (PK): Extended Characterization using Reticulocyte PKConfirmatorysame project related to Summary assay
602383qHTS for Activators of Human Muscle Isoform 2 Pyruvate Kinase: Muscle Isoform 1 AssayConfirmatorysame project related to Summary assay
602384qHTS for Activators of Human Muscle Isoform 2 Pyruvate Kinase: Extended CharacterizationConfirmatorysame project related to Summary assay
624387qHTS for Activators of Human Muscle Isoform 2 Pyruvate Kinase: Extended Characterization using M1 for Probe 2Confirmatorysame project related to Summary assay
624388qHTS for Activators of Human Muscle Isoform 2 Pyruvate Kinase (PK): Extended Characterization using Reticulocyte PK Probe 2Confirmatorysame project related to Summary assay
624389qHTS for Activators of Human Muscle Isoform 2 Pyruvate Kinase: Extended Characterization Using A Lactate Dehydrogenase (LDH) Assay Probe 2Confirmatorysame project related to Summary assay
624390qHTS for Activators of Human Muscle Isoform Pyruvate Kinase: Extended Characterization against Liver Pyruvate Kinase Probe 2Confirmatorysame project related to Summary assay
624391qHTS for Activators of Human Muscle Isoform 2 Pyruvate Kinase: Extended Characterization for Probe 2Confirmatorysame project related to Summary assay
Description:
NIH Chemical Genomics Center [NCGC]
Matthew G. Vander Heiden, M.D., Ph.D. [Harvard Medical School, 77 Avenue Louis Pasteur, Boston, MA 02115]
MLPCN Grant: 1 R03 MH085679-01

The metabolism of cancer cells is altered to support rapid proliferation. Otto Warburg was first to notice that cancer cells produce lactate even in the presence of oxygen [1,2]. This altered metabolism, known as the Warburg effect, is thought to give tumor cells a selective growth advantage relative to normal cells [3]. At least part of this abnormal metabolic phenotype is thought to be due to over expression of specific enzymes such as a tumor specific isozyme of pyruvate kinase - PKM2. Pyruvate kinase operates the final rate determining step of glycolysis where the enzyme converts one molecule of phosphoenol pyruvate and ADP to pyruvate and ATP. Recent work has shown that replacement of PKM2 in cancer cells with an isoform found in typical adult tissue cells (PKM1) can relieve the Warburg effect and promote a state of metabolism characteristic of normal cells [4]. Further, tumors engineered to express only PKM1 showed delayed tumor formation in nude mouse xenografts [4]. It has also been demonstrated that PKM2 has high affinity for binding phosphotyrosine peptides that are characteristic of growth factor signaling pathways present in cancer cells [5]. The binding of phosphopeptides to PKM2 removes the allosteric activator fructose-1,6-bisphosphate which further down-regulates the activity of PKM2 and intensifies the Warburg effect. Therefore, pharmacological activators of PKM2 are thought to be an approach for altering the Warburg effect by shifting the glycolytic flux from a proliferative mode back toward normal energy production.
The primary qHTS data of human PKM2 and confirmatory data are available in PubChem (AIDs: 1631, 1751 and 2533). Follow-up of synthesized analogs was determined using the same luminescent protocol for the PK isoforms M1, L and R (PubChem AIDs for M1, L and R bioluminescent assays are 2536, 2535, and 2534, respectively). A fluorescent secondary assay that determined the activity of PK by coupling the production pyruvate to lactate dehydrogenase oxidation of NADH was also used to determine the activation potency of the synthesized analog. (PubChem AIDs: 1540 and 2576 for M2 isoform; 2625, 2620, 2562 for L, M1 and R isoforms respectively). As well, general cytotoxicity of the compounds was measured in HeLa cells using Cell-TiterGlo (Promega; PubChem AID: 2653).

A bis-sulfonamide compound (CID_650361, ML083) and a heteocycle compound (CID_654376, ML082) were identified as human PKM2 activators.
Protocol
Please refer to other AIDs (1540, 1631, 1751, 2533, 2534, 2535, 2536, 2562, 2576, 2620, 2625 and 2653 for detailed assay protocols.
Comment
This summary is written for the purposes of summarizing the activities of the chemical probe, identified in this project. MLSCN probes are given a score of 100. Less active molecules in the same chemical series as the probe molecules are given a score of 50. Inactive analogues from these series are given a score of 0.
Categorized Comment - additional comments and annotations
From PubChem:
Assay Cell Type: HeLa
From MLP Probe Report:
Probe count: 5
MLP Probe ML# for probe 1: ML083
PubChem Substance ID (SID) for probe 1: 847943
PubChem Compound ID (CID) for probe 1: 650361
Probe type for probe 1: Activator
IC50/EC50 (nM) for probe 1: 0.6
Target for probe 1: hPK- M2 (gi: 33286418)
Anti-target for probe 1: hPK- R
Fold selectivity for probe 1: >30
NCBI Book chapter link for probe 1: http://www.ncbi.nlm.nih.gov/books/NBK47338/ (ID: 2358220)
Grant number for probe 1: MH085679-01
MLP Probe ML# for probe 2: ML170
PubChem Substance ID (SID) for probe 2: 85176568
PubChem Compound ID (CID) for probe 2: 4547230
Probe type for probe 2: Activator
IC50/EC50 (nM) for probe 2: 0.6
Target for probe 2: hPK-M2 (gi: 33286418)
Disease relevance for probe 2: Cancer
Anti-target for probe 2: hPK-R
Fold selectivity for probe 2: 30
NCBI Book chapter link for probe 2: http://www.ncbi.nlm.nih.gov/books/NBK56225/ (ID: 2508544)
Grant number for probe 2: MH085679-01
MLP Probe ML# for probe 3: ML202
PubChem Substance ID (SID) for probe 3: 85267884
PubChem Compound ID (CID) for probe 3: 44246498
Probe type for probe 3: Activator
IC50/EC50 (nM) for probe 3: 73
Target for probe 3: hPyk-M2 (gi: 33286418)
Disease relevance for probe 3: Cancer
Anti-target for probe 3: hPyk-M1, -R, and -L
Fold selectivity for probe 3: >100
NCBI Book chapter link for probe 3: http://www.ncbi.nlm.nih.gov/books/NBK56225/ (ID: 2508544)
Grant number for probe 3: MH085679-01
MLP Probe ML# for probe 4: ML203
PubChem Substance ID (SID) for probe 4: 87225694
PubChem Compound ID (CID) for probe 4: 44543605
Probe type for probe 4: Activator
IC50/EC50 (nM) for probe 4: 129
Target for probe 4: hPyk-M1 (gi: 33286418)
Disease relevance for probe 4: Cancer
Anti-target for probe 4: hPyk-M1, -R, and -L
Fold selectivity for probe 4: >100
NCBI Book chapter link for probe 4: http://www.ncbi.nlm.nih.gov/books/NBK56225/ (ID: 2508544)
Grant number for probe 4: MH085679-01
MLP Probe ML# for probe 5: ML082
PubChem Substance ID (SID) for probe 5: 851783
PubChem Compound ID (CID) for probe 5: 654376
Probe type for probe 5: Activator
IC50/EC50 (nM) for probe 5: 0.2
Target for probe 5: hPK- M2 (gi: 33286418)
Anti-target for probe 5: hPK- L
Fold selectivity for probe 5: >10
PubMed Publication ID (PMID) for probe 1: 20017496
PubMed Publication ID (PMID) for probe 2: 20017496
PubMed Publication ID (PMID) for probe 3: 21735594
PubMed Publication ID (PMID) for probe 4: 21735593
PubMed Publication ID (PMID) for probe 5: 20017496
NCBI Book chapter title for probe 1: Identification of activators for the M2 isoform of human pyruvate kinase
NCBI Book chapter title for probe 2: Identification of activators for the M2 isoform of human pyruvate kinase Version 3
NCBI Book chapter title for probe 3: Identification of activators for the M2 isoform of human pyruvate kinase Version 3
NCBI Book chapter title for probe 4: Identification of activators for the M2 isoform of human pyruvate kinase Version 3
Result Definitions
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TIDNameDescriptionHistogramTypeUnit
OutcomeThe BioAssay activity outcomeOutcome
ScoreThe BioAssay activity ranking scoreInteger
1ML#Molecular library probe numberString
2PhenotypeIndicates type of activity observed based on the compound activity across various assays tested.String
3Potency M2 LuminescenceThe concentration of sample yielding half-maximal inhibition in Kinase-glo assay in detection of ATP depletion.FloatμM
4Potency M1 LuminescenceThe activity outcomes of sample in Kinase-glo assay for M1 isoform.String
5Potency L LuminescenceThe activity outcomes of sample in Kinase-glo assay for L isoform.String
6Potency R LuminescenceThe activity outcomes of sample in Kinase-glo assay for R isoform.String
7Potency M2 LDHThe concentration of sample yielding half-maximal inhibition in LDH assay.FloatμM
8Potency M1 LDHThe activity outcomes of sample in LDH assay for M1 isoform.String
9Potency L LDHThe activity outcomes of sample in LDH assay for L isoform.String
10Potency R LDHThe activity outcomes of sample in LDH assay for R isoform.String
11CytotoxicityThe toxicity outcomes of sample in HeLa cells using Cell-TiterGlo.String
Additional Information
Grant Number: 1 R03 MH085679-01

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