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BioAssay: AID 743132

Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical dose-response gel-based ABPP in vitro in mouse brain

Name: Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical dose-response gel-based ABPP in vitro in mouse brain. ..more
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 Tested Compounds
 Tested Compounds
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Active(1)
 
 
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 Tested Substances
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Active(1)
 
 
AID: 743132
Data Source: The Scripps Research Institute Molecular Screening Center (LYPLA1&2_INH_FLUO_3XIC50VITRO_MUS_GEL_BASED_ABPP)
BioAssay Type: Panel, Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Probe Production Network, Assay Provider
BioAssay Version:
Deposit Date: 2013-11-26
Hold-until Date: 2014-08-09
Modify Date: 2014-08-11

Data Table ( Complete ):           View Active Data    View All Data
BioActive Compound: 1
Related Experiments
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AIDNameTypeProbeComment
2174Counterscreen for PME1 inhibitors: fluorescence polarization-based primary biochemical high throughput screening assay to identify inhibitors of lysophospholipase 1 (LYPLA1).Screening depositor-specified cross reference: Primary screen (LYPLA1 inhibitors in singlicate)
2177Counterscreen for PME1 inhibitors: fluorescence polarization-based primary biochemical high throughput screening assay to identify inhibitors of lysophospholipase 2 (LYPLA2).Screening depositor-specified cross reference: Primary screen (LYPLA2 inhibitors in singlicate)
2202Summary of probe development efforts to identify inhibitors of lysophospholipase 1 (LYPLA1).Summary2 depositor-specified cross reference: Summary (LYPLA1 inhibitors)
2203Summary of probe development efforts to identify inhibitors of lysophospholipase 2 (LYPLA2).Summary1 depositor-specified cross reference: Summary (LYPLA2 inhibitors)
2232Counterscreen for PME1 inhibitors: fluorescence polarization-based biochemical high throughput confirmation assay to identify inhibitors of lysophospholipase 2 (LYPLA2).Screening depositor-specified cross reference: Confirmation screen (LYPLA2 inhibitors in triplicate)
2233Counterscreen for PME1 inhibitors: fluorescence polarization-based biochemical high throughput confirmation assay for inhibitors of lysophospholipase 1 (LYPLA1).Screening depositor-specified cross reference: Confirmation screen (LYPLA1 inhibitors in triplicate)
493105Assay provider results from the probe development effort to identify dual inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical gel-based ABPP inhibition of recombinant and endogenous enzymeOther depositor-specified cross reference: Confirmation screen (LYPLA1 and LYPLA2 inhibitors in singlicate)
493108Late stage assay provider results from the probe development effort to identify dual inhibitors of LYPLA1 and LYPLA2: fluorescence-based cell-based inhibitionOther depositor-specified cross reference: Late stage screen (LYPLA1 and LYPLA2 inhibitors in singlicate)
493109Late stage assay provider results from the probe development effort to identify dual inhibitors of LYPLA1 and LYPLA2: LC-MS/MS assay to assess binding of compounds to active siteOther depositor-specified cross reference: Late stage LCMS assay (LYPLA1)
493110Late stage assay provider results from the probe development effort to identify dual inhibitors of LYPLA1 and LYPLA2: Gel-based Activity-Based Protein Profiling (ABPP) IC50 for LYPLA1 and LYPLA2Confirmatory depositor-specified cross reference: Late stage dose response (LYPLA1 and LYPLA2 inhibitors in triplicate)
493111Late stage assay provider results from the probe development effort to identify dual inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical gel-based ABPP inhibition and selectivityOther depositor-specified cross reference: Late stage screen (LYPLA1 and LYPLA2 inhibitors in singlicate)
493154Late stage assay provider results from the probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Gel-based Activity-Based Protein Profiling (ABPP) IC50 for off-target ABHD11Confirmatory depositor-specified cross reference: Late stage dose response counterscreen (ABHD11 inhibitors in triplicate)
493161Late stage assay provider results from the probe development effort to identify dual inhibitors of LYPLA1 and LYPLA2: absorbance-based cell-based dose response assay to determine cytotoxicity of inhibitor compoundsConfirmatory depositor-specified cross reference: Late stage dose response counterscreen (T-cell cytotoxicity in quadruplicate)
504482Late stage assay provider results from the probe development effort to identify inhibitors of LYPLA1: fluorescence-based cell-based gel-based Activity-Based Protein Profiling (ABPP) IC50 for anti-target ABHD11Confirmatory depositor-specified cross reference: Late stage dose repsonse (ABHD11 inhibitors in triplicate)
504498Late stage assay provider results from the probe development effort to identify inhibitors of LYPLA1: LC-MS/MS assay to assess binding of compounds to active site of anti-target ABHD11Other depositor-specified cross reference: Late stage MOA assay (ABDH11 LCMS)
504505Late stage assay provider results from the probe development effort to identify inhibitors of LYPLA1: fluorescence-based cell-based gel-based Activity-Based Protein Profiling (ABPP) percent inhibition for anti-target ABHD11Other depositor-specified cross reference: Late stage screen (ABHD11 inhibitors in singlicate, in situ)
504507Late stage assay provider results from the probe development effort to identify inhibitors of LYPLA1: fluorescence-based biochemical gel-based Activity-Based Protein Profiling (ABPP) IC50 for anti-target ABHD11 Set 2Confirmatory depositor-specified cross reference: Late stage dose response (ABHD11 inhibitors in triplicate)
504510Late stage assay provider results from the probe development effort to identify inhibitors of LYPLA1: absorbance-based cell-based dose response assay to determine cytotoxicity of inhibitor compounds set 2Confirmatory depositor-specified cross reference: Late stage dose repsonse counterscreen (T-cell cytotoxicity in quadruplicate)
504520Late stage assay provider results from the probe development effort to identify inhibitors of LYPLA1: fluorescence-based biochemical gel-based Activity-Based Protein Profiling (ABPP) inhibition and selectivityOther depositor-specified cross reference: Late stage screen (LYPLA1 and LYPLA2 inhibitors in singlicate)
504522Late stage assay provider results from the probe development effort to identify inhibitors of LYPLA1: LC-MS-based cell-based SILAC Activity-Based Protein Profiling (ABPP) for anti-target ABHD11Other depositor-specified cross reference: Late stage (LC-MS-based cell-based SILAC ABPP for anti-target ABHD11)
504892Late stage assay provider results from the probe development effort to identify inhibitors of ABHD11: Fluorescence-based biochemical gel-based Activity-Based Protein Profiling (ABPP) inhibition of the human isoform of ABHD11Other depositor-specified cross reference: Late stage assay (human isoform of ABHD11 inhibitors)
651978Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA1: LCMS-based Activity-Based Protein Profiling (ABPP) SILAC selectivity analysis in vitroOther depositor-specified cross reference: Late stage ABPP in vitro SILAC assay (LYPLA 1)
651979Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA1: LCMS-based cell-based Activity-Based Protein Profiling (ABPP) SILAC selectivity analysis in situOther depositor-specified cross reference: Late stage ABPP in situ SILAC assay (LYPLA 1)
651980Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA2: LCMS-based cell-based Activity-Based Protein Profiling (ABPP) SILAC selectivity analysis in situOther depositor-specified cross reference: Late stage assay (SILAC selectivity analysis in situ)
651981Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA2: LCMS-based Activity-Based Protein Profiling (ABPP) SILAC selectivity analysis in vitroOther depositor-specified cross reference: Late stage assay (SILAC selectivity analysis in vitro)
651985Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical gel-based ABPP evaluation of activity in vivoOther depositor-specified cross reference: Late stage assay (LYPLA1 and LYPLA2 inhibitors activity in vivo)
651986Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical gel-based ABPP evaluation of activity in situOther depositor-specified cross reference: Late stage assay (LYPLA1 and LYPLA2 inhibitors activity in situ)
651987Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical gel-based ABPP gel filtration assay to assess binding modeOther depositor-specified cross reference: Late stage assay (binding mode)
651988Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical gel-based ABPP inhibition and selectivityOther depositor-specified cross reference: Late stage assay (inhibition and selectivity)
651990Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical dose-response gel-based ABPPConfirmatory depositor-specified cross reference: Late stage dose-response (LYPLA1 and LYPLA2 inhibitors in triplicate)
651991Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA1 and LYPLA2: Absorbance-based cell-based dose response assay to determine cytotoxicity of inhibitor compoundsConfirmatory depositor-specified cross reference: Late stage dose-response (cytotoxicity 6 replicates)
651998Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA1: Substrate-based fluorescence-based biochemical determination of kinetic parametersConfirmatory depositor-specified cross reference: Late stage ABPP biochemical kinetics assay (LYPLA 1)
652001Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA2: Substrate-based fluorescence-based biochemical determination of kinetic parametersConfirmatory depositor-specified cross reference: Late stage assay (LYPLA2 kinetic parameters in quadruplicate)
652003Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA1: fluorescence-based biochemical dose-response assayConfirmatory depositor-specified cross reference: Late stage ABPP biochemical dose response assay (LYPLA 1) (4XIC50)
652004Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA2: fluorescence-based biochemical dose-response assayConfirmatory depositor-specified cross reference: Late stage dose response (LYPLA2 inhibitors in quadruplicate)
652018Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA1: LCMS-based Activity-Based Protein Profiling (ABPP) SILAC selectivity analysis in vitro, Set 2Other depositor-specified cross reference: Late stage assay (LYPLA1 and LYPLA2 inhibition and selectivity in singlicate)
2143Summary of probe development efforts to identify inhibitors of Protein Phosphatase Methylesterase 1 (PME-1).Summary3 same project related to Summary assay
652029Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA2: Fluorescence-based biochemical gel-based ABPP inhibitionOther same project related to Summary assay
652030Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA2: Fluorescence-based biochemical gel-based ABPP inhibition and selectivityOther same project related to Summary assay
743117Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical gel-based ABPP inhibition potency and selectivityOther same project related to Summary assay
743118Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical dose-response gel-based ABPP in vitroConfirmatory same project related to Summary assay
743119Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical dose-response gel-based ABPP in situConfirmatory same project related to Summary assay
743127Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Absorbance-based cell-based dose response assay to determine cytotoxicity of inhibitor compoundsConfirmatory same project related to Summary assay
743133Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical gel-based ABPP gel filtration assay to assess inhibitor binding modeOther same project related to Summary assay
743134Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical gel-based ABPP assay to assess in vivo activityOther same project related to Summary assay
743137Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: LCMS-based cell-based Activity-Based Protein Profiling (ABPP) SILAC selectivity analysisOther same project related to Summary assay
2143Summary of probe development efforts to identify inhibitors of Protein Phosphatase Methylesterase 1 (PME-1).Summary3 same project related to Summary assay
652029Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA2: Fluorescence-based biochemical gel-based ABPP inhibitionOther same project related to Summary assay
652030Late stage assay provider results from the probe development effort to identify selective inhibitors of LYPLA2: Fluorescence-based biochemical gel-based ABPP inhibition and selectivityOther same project related to Summary assay
743117Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical gel-based ABPP inhibition potency and selectivityOther same project related to Summary assay
743118Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical dose-response gel-based ABPP in vitroConfirmatory same project related to Summary assay
743119Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical dose-response gel-based ABPP in situConfirmatory same project related to Summary assay
743127Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Absorbance-based cell-based dose response assay to determine cytotoxicity of inhibitor compoundsConfirmatory same project related to Summary assay
743133Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical gel-based ABPP gel filtration assay to assess inhibitor binding modeOther same project related to Summary assay
743134Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical gel-based ABPP assay to assess in vivo activityOther same project related to Summary assay
743137Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: LCMS-based cell-based Activity-Based Protein Profiling (ABPP) SILAC selectivity analysisOther same project related to Summary assay
Description:
Source (MLPCN Center Name): The Scripps Research Institute Molecular Screening Center (SRIMSC)
Center Affiliation: The Scripps Research Institute (TSRI)
Assay Provider: Benjamin Cravatt, TSRI
Network: Molecular Libraries Probe Production Centers Network (MLPCN)
Grant Proposal Number: 1 R01 CA132630
Grant Proposal PI: Benjamin Cravatt, TSRI
External Assay ID: LYPLA1&2_INH_FLUO_3XIC50VITRO_MUS_GEL_BASED_ABPP

Name: Late stage assay provider results from the extended probe development effort to identify inhibitors of LYPLA1 and LYPLA2: Fluorescence-based biochemical dose-response gel-based ABPP in vitro in mouse brain.

Description:

Protein palmitoylation is an essential post-translational modification necessary for trafficking and localization of regulatory proteins that play key roles in cell growth and signaling. Numerous proteins have been identified as targets of palmitoylation, including cytoskeletal proteins, kinases, receptors, and other proteins involved in various aspects of cellular signaling and homeostasis [1]. Using a global chemo-proteomic method for the metabolic incorporation and identification of palmitoylated proteins, we were able to identify hundreds of palmitoylated proteins, revealing palmitoylation as a widespread post-translational modification (PTM) [2]. Palmitoylation involves an acyl-thioester linkage to specific cysteines [3,4]. Given the labile properties of thioesters, palmitoylation is potentially reversible and may be regulated in a manner analogous to other PTMs (e.g., phosphorylation). As such, identification of proteins responsible for the dynamic modulation of palmitoylation is paramount to understanding its patho/physiological roles. For example, multiple oncogenes, including HRAS and SRC, require palmitoylation for malignant transformation [5], suggesting protein palmitoyl thioesterases may have tumor suppressor activity required to repress aberrant growth signaling. More than a decade ago, the cytosolic serine hydrolase acyl-protein thioesterase 1 (APT1) was identified as an in vitro HRAS palmitoyl thioesterase [6]. Initially classified as lysophospholipase 1 (LYPLA1) [7], the enzyme has since been demonstrated to have several hundred-fold higher activity as a protein thioesterase. While the in vitro data [6,8] provided an intriguing clue to its possible role in vivo, prior to our studies, little was known about the in vivo thioesterase activity of LYPLA1. Upon retroviral shRNA knockdown of LYPLA1, we found that HRAS was robustly hyper-palmitoylated, providing the first evidence that the endogenous enzyme is a functional protein palmitoyl thioesterase capable of regulating HRAS palmitoylation in mammalian cells. However, shRNA resulted in only an 80% reduction in LYPLA1 expression (unpublished). LYPLA2 (a.k.a. APT2) is 65% identical to LYPLA1, and also exhibits lysophospholipase activity in vitro, but its potential role as a thioesterase is unknown [9]. shRNA knockdown studies of LYPLA2 revealed only partial knockdown of the enzyme, making substrate identification inconclusive (unpublished). A principle goal of post-genomic research is the determination of the molecular and cellular role of uncharacterized enzymes like LYPLA1 and LYPLA2. As such, selective inhibitors of LYPLA1 and/or LYPLA2 would greatly aid investigations into the biological function of these enzymes. Several inhibitors of LYPLA1 have been described [10,11], but none of these agents have proven capable of inhibiting LYPLA1 activity in cells. We recently reported optimization of a covalent dual LYPLA1/LYPLA2 inhibitor ML211 (SID 99445338), and identification of reversible compounds ML348 (SID 160654487) and ML349 (SID 160654496) that act as selective LYPLA1 and LYPLA2 inhibitors, respectively (see Probe Reports on the NIH Bookshelf at http://www.ncbi.nlm.nih.gov/books/NBK47352/). These compounds represent an important advance in chemical tools for investigating the biological function(s) of LYPLA1 and LYPLA2. However, ML211 (SID 99445338), owing to its modest solubility, does not have demonstrated in vivo activity. Irreversible chemical probes have distinct advantages vs. reversible compounds for use in living systems, as confirming target engagement is a comparatively straightforward process, and near-complete and sustained target inhibition can be achieved without extensive optimization of physicochemical and pharmacokinetic properties. The goal of this current investigation is to develop an irreversible, in vivo-active dual LYPLA1/LYPLA2 inhibitor to add to the arsenal of available chemical tools for identification of LYPLA1 and LYPLA2 substrates and evaluating the role of these enzymes in dynamic de-palmitoylation and tumorigenesis.

References:

1. Dekker, F.J., et al., Small-molecule inhibition of APT1 affects Ras localization and signaling. Nat. Chem. Biol., 2010. 6(6): p. 449-56.
2. Duncan, J.A. and A.G. Gilman, A cytoplasmic acyl-protein thioesterase that removes palmitate from G protein alpha subunits and p21(RAS). J. Biol. Chem., 1998. 273(25): p. 15830-7.
3. Sugimoto, H., H. Hayashi, and S. Yamashita, Purification, cDNA cloning, and regulation of lysophospholipase from rat liver. J. Biol. Chem., 1996. 271(13): p. 7705-11.
4. Toyoda, T., H. Sugimoto, and S. Yamashita, Sequence, expression in Escherichia coli, and characterization of lysophospholipase II. Biochim. Biophys. Acta, 1999. 1437(2): p. 182-93.
5. Biel, M., et al., Synthesis and evaluation of acyl protein thioesterase 1 (APT1) inhibitors. Chemistry, 2006. 12(15): p. 4121-43.
6. Deck, P., et al., Development and biological evaluation of acyl protein thioesterase 1 (APT1) inhibitors. Angew. Chem. Int. Ed. Engl., 2005. 44(31): p. 4975-80.
7. Jessani, N., et al., Enzyme activity profiles of the secreted and membrane proteome that depict cancer cell invasiveness. Proc. Natl. Acad. Sci. U. S. A., 2002. 99(16): p. 10335-40.
8. Leung, D., et al., Discovering potent and selective reversible inhibitors of enzymes in complex proteomes. Nat. Biotechnol., 2003. 21(6): p. 687-91.
9. Bachovchin, D.A., et al., Identification of selective inhibitors of uncharacterized enzymes by high-throughput screening with fluorescent activity-based probes. Nat. Biotechnol., 2009. 27(4): p. 387-94.
10. Forner, F., et al., Quantitative proteomic comparison of rat mitochondria from muscle, heart, and liver. Mol. Cell. Proteomics, 2006. 5(4): p. 608-19.
11. Schubert, C., The genomic basis of the Williams-Beuren syndrome. Cell. Mol. Life Sci., 2009. 66(7): p. 1178-97.

Keywords:

late stage, late stage AID, assay provider, powders, LYPLA1, lysophospholipase 1, LYPLA2, lysophospholipase 2, APT1, acyl-protein thioesterase 1, APT2, acyl-protein thioesterase 2, serine hydrolase, palmitoylation, counterscreen, inhibitor, inhibition, dose response, IC50, proteome, lysate, activity-based protein profiling, ABPP, gel-based ABPP, fluorophosphonate rhodamine, FP-Rh, Scripps, Scripps Research Institute Molecular Screening Center, SRIMSC, Molecular Libraries Probe Production Centers Network, MLPCN
Panel Information
Panel Assay with Multiple Targets/Outcomes
PID§NameSubstancePanel TargetsDescriptionAdditional Information
ActiveInactive
1LYPLA1/21 [gi:6678760]
[gi:7242156]
Taxonomy id: 10090
Gene id: 18777
Gene id: 26394
2ABHD61 [gi:31560264]
Taxonomy id: 10090
Gene id: 66082
3FAAH1 [gi:123253900]
Taxonomy id: 10090
Gene id: 14073

§ Panel component ID.
Protocol
Assay Overview:
The purpose of this assay is to determine IC50 values of powder samples of test compounds for LYPLA1, LYPLA2, and anti-targets ABHD6 and FAAH in a complex proteomic lysate using an activity-based proteomic profiling (ABPP) assay. In this assay, a complex proteome is incubated with test compound followed by reaction with a serine hydrolase-specific rhodamine-conjugated fluorophosphonate (FP-Rh) activity-based probe. The reaction products are separated by SDS-PAGE and visualized in-gel using a flatbed fluorescence scanner. The percentage activity remaining is determined by measuring the integrated optical density (IOD) of the bands. As designed, test compounds that act as inhibitors will prevent enzyme-probe interactions, thereby decreasing the proportion of bound fluorescent probe, giving lower fluorescence intensity in the band in the gel. Percent inhibition is calculated relative to a DMSO (no compound) control. Compounds are tested in triplicate in a 9-point dilution series starting at a nominal concentration of 100 uM.
Protocol Summary:
Mouse brain membrane proteome (50 uL reaction volume; 1 mg/ml in DPBS) was treated with test compound (1 uL of a 50x stock in DMSO) for 30 minutes at 37 degrees Celsius. FP-Rh (1 uL of 50x stock in DMSO) was added to a final concentration of 2 uM. The reactions was incubated for 30 minutes at 25 degrees Celsius, quenched with 16 uL 4x SDS-PAGE loading buffer (reducing), separated by SDS-PAGE and visualized by in-gel fluorescent scanning. The percentage activity remaining was determined by measuring the integrated optical density of the target and anti-target bands relative to a DMSO-only (no compound) control. Due to the overlapping nature of the LYPLA1 and LYPLA2 bands, the proteins were quantified together. IC50 values were determined from dose-response curves from three replicates at each inhibitor concentration: 0.00001, 0.0001, 0.001, 0.01, 0.1, 1, 10, 50 and 100 uM.
Percent inhibition was calculated as follows:
%_Inhibition = ( 1 - ( IOD_Test_Compound - IOD_Low_Control ) / ( IOD_High_Control - IOD_Low_Control ) ) * 100
Where:
Test_Compound is defined as target treated with test compound.
High_Control is defined as target treated with DMSO only (no compound).
Low_Control is defined as background in a blank region of the gel.
For each test compound, percent inhibition was plotted against the log of the compound concentration. A three parameter equation describing a sigmoidal dose-response curve was then fitted using XLFit (IDBS) normalized from 0 to 100 for each assay. The software-generated IC50 values were reported.
Assay Outcome:
Compounds with an IC50 less than or equal to 1 uM were considered active. Compounds with an IC50 greater than 1 uM were considered inactive.
PubChem Activity Outcome:
Compounds were considered active if they were active in one or more assays. Compounds were considered inactive if they were inactive in all assays.
List of Reagents:
Mouse brain membrane proteome (provided by the Assay Provider)
FP-Rh (Thermo #88318)
DPBS (Cellgro 20-031-CV)
Comment
This assay was performed by the assay provider with powder samples of synthetic test compounds.
Result Definitions
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TIDNameDescriptionPID§Panel TargetsHistogramTypeUnit
OutcomeThe BioAssay activity outcomeOutcome
1LYPLA1/2 OutcomeOne of Active or Inactive for LYPLA1/2.1
Outcome
2LYPLA1/2 IC50*The concentration at which 50 percent of the activity in the inhibition assay is observed; (IC50) shown in micromolar.1FloatμM
3LYPLA1/2 Inhibition at 0.00001 uM [1] (1e-05μM**)Inhibition of endogenous LYPLA1/2 upon 0.00001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.1Float%
4LYPLA1/2 Inhibition at 0.00001 uM [2] (1e-05μM**)Inhibition of endogenous LYPLA1/2 upon 0.00001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.1Float%
5LYPLA1/2 Inhibition at 0.00001 uM [3] (1e-05μM**)Inhibition of endogenous LYPLA1/2 upon 0.00001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.1Float%
6LYPLA1/2 Inhibition at 0.0001 uM [1] (0.0001μM**)Inhibition of endogenous LYPLA1/2 upon 0.0001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.1Float%
7LYPLA1/2 Inhibition at 0.0001 uM [2] (0.0001μM**)Inhibition of endogenous LYPLA1/2 upon 0.0001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.1Float%
8LYPLA1/2 Inhibition at 0.0001 uM [3] (0.0001μM**)Inhibition of endogenous LYPLA1/2 upon 0.0001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.1Float%
9LYPLA1/2 Inhibition at 0.001 uM [1] (0.001μM**)Inhibition of endogenous LYPLA1/2 upon 0.001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.1Float%
10LYPLA1/2 Inhibition at 0.001 uM [2] (0.001μM**)Inhibition of endogenous LYPLA1/2 upon 0.001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.1Float%
11LYPLA1/2 Inhibition at 0.001 uM [3] (0.001μM**)Inhibition of endogenous LYPLA1/2 upon 0.001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.1Float%
12LYPLA1/2 Inhibition at 0.01 uM [1] (0.01μM**)Inhibition of endogenous LYPLA1/2 upon 0.01uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.1Float%
13LYPLA1/2 Inhibition at 0.01 uM [2] (0.01μM**)Inhibition of endogenous LYPLA1/2 upon 0.01uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.1Float%
14LYPLA1/2 Inhibition at 0.01 uM [3] (0.01μM**)Inhibition of endogenous LYPLA1/2 upon 0.01uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.1Float%
15LYPLA1/2 Inhibition at 0.1 uM [1] (0.1μM**)Inhibition of endogenous LYPLA1/2 upon 0.1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.1Float%
16LYPLA1/2 Inhibition at 0.1 uM [2] (0.1μM**)Inhibition of endogenous LYPLA1/2 upon 0.1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.1Float%
17LYPLA1/2 Inhibition at 0.1 uM [3] (0.1μM**)Inhibition of endogenous LYPLA1/2 upon 0.1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.1Float%
18LYPLA1/2 Inhibition at 1 uM [1] (1μM**)Inhibition of endogenous LYPLA1/2 upon 1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.1Float%
19LYPLA1/2 Inhibition at 1 uM [2] (1μM**)Inhibition of endogenous LYPLA1/2 upon 1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.1Float%
20LYPLA1/2 Inhibition at 1 uM [3] (1μM**)Inhibition of endogenous LYPLA1/2 upon 1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.1Float%
21LYPLA1/2 Inhibition at 10 uM [1] (10μM**)Inhibition of endogenous LYPLA1/2 upon 10uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.1Float%
22LYPLA1/2 Inhibition at 10 uM [2] (10μM**)Inhibition of endogenous LYPLA1/2 upon 10uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.1Float%
23LYPLA1/2 Inhibition at 10 uM [3] (10μM**)Inhibition of endogenous LYPLA1/2 upon 10uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.1Float%
24LYPLA1/2 Inhibition at 50 uM [1] (50μM**)Inhibition of endogenous LYPLA1/2 upon 50uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.1Float%
25LYPLA1/2 Inhibition at 50 uM [2] (50μM**)Inhibition of endogenous LYPLA1/2 upon 50uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.1Float%
26LYPLA1/2 Inhibition at 50 uM [3] (50μM**)Inhibition of endogenous LYPLA1/2 upon 50uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.1Float%
27LYPLA1/2 Inhibition at 100 uM [1] (100μM**)Inhibition of endogenous LYPLA1/2 upon 100uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.1Float%
28LYPLA1/2 Inhibition at 100 uM [2] (100μM**)Inhibition of endogenous LYPLA1/2 upon 100uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.1Float%
29LYPLA1/2 Inhibition at 100 uM [3] (100μM**)Inhibition of endogenous LYPLA1/2 upon 100uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.1Float%
30ABHD6 OutcomeOne of Active or Inactive for ABHD6.2Outcome
31ABHD6 IC50*The concentration at which 50 percent of the activity in the inhibition assay is observed; (IC50) shown in micromolar.2FloatμM
32ABHD6 Inhibition at 0.00001 uM [1] (1e-05μM**)Inhibition of endogenous ABHD6 upon 0.00001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.2Float%
33ABHD6 Inhibition at 0.00001 uM [2] (1e-05μM**)Inhibition of endogenous ABHD6 upon 0.00001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.2Float%
34ABHD6 Inhibition at 0.00001 uM [3] (1e-05μM**)Inhibition of endogenous ABHD6 upon 0.00001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.2Float%
35ABHD6 Inhibition at 0.0001 uM [1] (0.0001μM**)Inhibition of endogenous ABHD6 upon 0.0001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.2Float%
36ABHD6 Inhibition at 0.0001 uM [2] (0.0001μM**)Inhibition of endogenous ABHD6 upon 0.0001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.2Float%
37ABHD6 Inhibition at 0.0001 uM [3] (0.0001μM**)Inhibition of endogenous ABHD6 upon 0.0001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.2Float%
38ABHD6 Inhibition at 0.001 uM [1] (0.001μM**)Inhibition of endogenous ABHD6 upon 0.001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.2Float%
39ABHD6 Inhibition at 0.001 uM [2] (0.001μM**)Inhibition of endogenous ABHD6 upon 0.001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.2Float%
40ABHD6 Inhibition at 0.001 uM [3] (0.001μM**)Inhibition of endogenous ABHD6 upon 0.001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.2Float%
41ABHD6 Inhibition at 0.01 uM [1] (0.01μM**)Inhibition of endogenous ABHD6 upon 0.01uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.2Float%
42ABHD6 Inhibition at 0.01 uM [2] (0.01μM**)Inhibition of endogenous ABHD6 upon 0.01uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.2Float%
43ABHD6 Inhibition at 0.01 uM [3] (0.01μM**)Inhibition of endogenous ABHD6 upon 0.01uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.2Float%
44ABHD6 Inhibition at 0.1 uM [1] (0.1μM**)Inhibition of endogenous ABHD6 upon 0.1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.2Float%
45ABHD6 Inhibition at 0.1 uM [2] (0.1μM**)Inhibition of endogenous ABHD6 upon 0.1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.2Float%
46ABHD6 Inhibition at 0.1 uM [3] (0.1μM**)Inhibition of endogenous ABHD6 upon 0.1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.2Float%
47ABHD6 Inhibition at 1 uM [1] (1μM**)Inhibition of endogenous ABHD6 upon 1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.2Float%
48ABHD6 Inhibition at 1 uM [2] (1μM**)Inhibition of endogenous ABHD6 upon 1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.2Float%
49ABHD6 Inhibition at 1 uM [3] (1μM**)Inhibition of endogenous ABHD6 upon 1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.2Float%
50ABHD6 Inhibition at 10 uM [1] (10μM**)Inhibition of endogenous ABHD6 upon 10uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.2Float%
51ABHD6 Inhibition at 10 uM [2] (10μM**)Inhibition of endogenous ABHD6 upon 10uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.2Float%
52ABHD6 Inhibition at 10 uM [3] (10μM**)Inhibition of endogenous ABHD6 upon 10uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.2Float%
53ABHD6 Inhibition at 50 uM [1] (50μM**)Inhibition of endogenous ABHD6 upon 50uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.2Float%
54ABHD6 Inhibition at 50 uM [2] (50μM**)Inhibition of endogenous ABHD6 upon 50uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.2Float%
55ABHD6 Inhibition at 50 uM [3] (50μM**)Inhibition of endogenous ABHD6 upon 50uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.2Float%
56ABHD6 Inhibition at 100 uM [1] (100μM**)Inhibition of endogenous ABHD6 upon 100uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.2Float%
57ABHD6 Inhibition at 100 uM [2] (100μM**)Inhibition of endogenous ABHD6 upon 100uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.2Float%
58ABHD6 Inhibition at 100 uM [3] (100μM**)Inhibition of endogenous ABHD6 upon 100uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.2Float%
59FAAH OutcomeOne of Active or Inactive for FAAH.3Outcome
60FAAH IC50*The concentration at which 50 percent of the activity in the inhibition assay is observed; (IC50) shown in micromolar.3FloatμM
61FAAH Inhibition at 0.00001 uM [1] (1e-05μM**)Inhibition of endogenous FAAH upon 0.00001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.3Float%
62FAAH Inhibition at 0.00001 uM [2] (1e-05μM**)Inhibition of endogenous FAAH upon 0.00001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.3Float%
63FAAH Inhibition at 0.00001 uM [3] (1e-05μM**)Inhibition of endogenous FAAH upon 0.00001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.3Float%
64FAAH Inhibition at 0.0001 uM [1] (0.0001μM**)Inhibition of endogenous FAAH upon 0.0001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.3Float%
65FAAH Inhibition at 0.0001 uM [2] (0.0001μM**)Inhibition of endogenous FAAH upon 0.0001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.3Float%
66FAAH Inhibition at 0.0001 uM [3] (0.0001μM**)Inhibition of endogenous FAAH upon 0.0001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.3Float%
67FAAH Inhibition at 0.001 uM [1] (0.001μM**)Inhibition of endogenous FAAH upon 0.001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.3Float%
68FAAH Inhibition at 0.001 uM [2] (0.001μM**)Inhibition of endogenous FAAH upon 0.001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.3Float%
69FAAH Inhibition at 0.001 uM [3] (0.001μM**)Inhibition of endogenous FAAH upon 0.001uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.3Float%
70FAAH Inhibition at 0.01 uM [1] (0.01μM**)Inhibition of endogenous FAAH upon 0.01uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.3Float%
71FAAH Inhibition at 0.01 uM [2] (0.01μM**)Inhibition of endogenous FAAH upon 0.01uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.3Float%
72FAAH Inhibition at 0.01 uM [3] (0.01μM**)Inhibition of endogenous FAAH upon 0.01uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.3Float%
73FAAH Inhibition at 0.1 uM [1] (0.1μM**)Inhibition of endogenous FAAH upon 0.1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.3Float%
74FAAH Inhibition at 0.1 uM [2] (0.1μM**)Inhibition of endogenous FAAH upon 0.1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.3Float%
75FAAH Inhibition at 0.1 uM [3] (0.1μM**)Inhibition of endogenous FAAH upon 0.1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.3Float%
76FAAH Inhibition at 1 uM [1] (1μM**)Inhibition of endogenous FAAH upon 1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.3Float%
77FAAH Inhibition at 1 uM [2] (1μM**)Inhibition of endogenous FAAH upon 1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.3Float%
78FAAH Inhibition at 1 uM [3] (1μM**)Inhibition of endogenous FAAH upon 1uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.3Float%
79FAAH Inhibition at 10 uM [1] (10μM**)Inhibition of endogenous FAAH upon 10uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.3Float%
80FAAH Inhibition at 10 uM [2] (10μM**)Inhibition of endogenous FAAH upon 10uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.3Float%
81FAAH Inhibition at 10 uM [3] (10μM**)Inhibition of endogenous FAAH upon 10uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.3Float%
82FAAH Inhibition at 50 uM [1] (50μM**)Inhibition of endogenous FAAH upon 50uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.3Float%
83FAAH Inhibition at 50 uM [2] (50μM**)Inhibition of endogenous FAAH upon 50uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.3Float%
84FAAH Inhibition at 50 uM [3] (50μM**)Inhibition of endogenous FAAH upon 50uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.3Float%
85FAAH Inhibition at 100 uM [1] (100μM**)Inhibition of endogenous FAAH upon 100uM compound treatment as assessed by gel-based competitive ABPP. Replicate 1.3Float%
86FAAH Inhibition at 100 uM [2] (100μM**)Inhibition of endogenous FAAH upon 100uM compound treatment as assessed by gel-based competitive ABPP. Replicate 2.3Float%
87FAAH Inhibition at 100 uM [3] (100μM**)Inhibition of endogenous FAAH upon 100uM compound treatment as assessed by gel-based competitive ABPP. Replicate 3.3Float%

* Activity Concentration. ** Test Concentration. § Panel component ID.
Additional Information
Grant Number: 1 R01 CA132630

Classification
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