Summary of Study ST003366

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001941. The data can be accessed directly via it's Project DOI: 10.21228/M8TM76 This work is supported by NIH grant, U2C- DK119886.

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This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

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Study IDST003366
Study TitleMalate dehydrogenase (MDH) inhibition assay: Fasnall does not affect MDH activity in vitro
Study TypeIntracellular metabolomics, medium metabolomics
Study SummaryThe enzymatic activity of porcine malate dehydrogenase (MDH) was reconstructed using NADH and oxaloacetate as substrates. LC-MS/MS was used to monitor the reaction by measuring the accumulation of malic acid and NAD+, as well as the consumption of oxaloacetate and NADH. Fasnall does not affect MDH activity in vitro.
Institute
Wistar Institute
DepartmentMolecular and Cellular Oncogenesis Program, Ellen and Ronald Caplan Cancer Center
LaboratorySchug's Lab
Last NameMukha
First NameDzmitry
Address3601 Spruce St, Philadelphia, PA 19104, USA
Emaildmukha@wistar.org
Phone+12154956903
Submit Date2024-07-17
Num Groups12
Total Subjects48
PublicationsSubmission Pending
Raw Data AvailableYes
Raw Data File Type(s)mzML, wiff
Analysis Type DetailLC-MS
Release Date2024-08-14
Release Version1
Dzmitry Mukha Dzmitry Mukha
https://dx.doi.org/10.21228/M8TM76
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:

Project:

Project ID:PR001941
Project DOI:doi: 10.21228/M8TM76
Project Title:The shutdown of NADH oxidation via Respiratory Complex I mimics fatty acid biosynthesis inhibition
Project Type:LC-MS Quantitative Analysis
Project Summary:Proliferating cancer cells actively utilize anabolic processes for biomass production, including de novo biosynthesis of amino acids, nucleotides, and fatty acids. The key enzyme of the fatty acid biosynthesis pathway, fatty acid synthase (FASN), is widely recognized as a promising therapeutic target in cancer and other health conditions. Here, we establish a metabolic signature of FASN inhibition using a panel of pharmacological inhibitors (GSK2194069, TVB-2640, TVB-3166, C75, cerulenin, and Fasnall). We find that the activity of some commonly used FASN inhibitors is inconsistent with the metabolic signature of FASN inhibition (accumulation of malonate, succinate, malonyl coenzyme A, succinyl coenzyme A, and other metabolic perturbations). Moreover, we show that one of these putative FASN inhibitors, Fasnall, is a respiratory Complex I inhibitor that mimics FASN inhibition through NADH accumulation and consequent depletion of the tricarboxylic acid cycle metabolites. We demonstrate that Fasnall impairs tumor growth in several oxidative phosphorylation-dependent cancer models, including combination therapy-resistant melanoma patient-derived xenografts. Fasnall administration does not reproduce neurological side effects in mice reported for other Complex I inhibitors. Our results have significant implications for understanding the FASN role in human health and disease and provide evidence of therapeutic potential for Complex I inhibitors with fast systemic clearance. Our findings also highlight the continuing need for validation of small molecule inhibitors to distinguish high-quality chemical probes and to expand the understanding of their application.
Institute:Wistar Institute
Department:Molecular and Cellular Oncogenesis Program, Ellen and Ronald Caplan Cancer Center
Laboratory:Schug's Lab
Last Name:Mukha
First Name:Dzmitry
Address:3601 Spruce St., Philadelphia, Pennsylvania 19104, USA
Email:dmukha@wistar.org
Phone:+12154956903
Funding Source:This work was supported by grants from the National Institutes of Health (NIH) National Cancer Institute (NCI) DP2 CA249950-01 (Z.T.S.), NIH NCI P01 CA114046 (Z.T.S.), Melanoma Research Foundation 717173 (Z.T.S.), and Susan G. Komen CCR19608782 (Z.T.S.).
Publications:Submission Pending
Contributors:Dzmitry Mukha, Jena Dessain, Seamus O’Connor, Katherine Pniewski, Fabrizio Bertolazzi, Jeet Patel, Mary Mullins, Zachary T. Schug

Subject:

Subject ID:SU003487
Subject Type:Synthetic sample

Factors:

Subject type: Synthetic sample; Subject species: - (Factor headings shown in green)

mb_sample_id local_sample_id Sample source MDH Fasnall Time
SA36649704 OAA No ENZ No Fasnall 00 minMDH assay FALSE 0 uM 0 min
SA36649806 OAA No ENZ No Fasnall 10 minMDH assay FALSE 0 uM 10 min
SA36649907 OAA No ENZ No Fasnall 20 minMDH assay FALSE 0 uM 20 min
SA36650005 OAA No ENZ No Fasnall 05 minMDH assay FALSE 0 uM 5 min
SA36650512 OAA Porcine 1 to 100 0.5 uM Fasnall 00 minMDH assay TRUE 0.5 uM 0 min
SA36650614 OAA Porcine 1 to 100 0.5 uM Fasnall 10 minMDH assay TRUE 0.5 uM 10 min
SA36650715 OAA Porcine 1 to 100 0.5 uM Fasnall 20 minMDH assay TRUE 0.5 uM 20 min
SA36650813 OAA Porcine 1 to 100 0.5 uM Fasnall 05 minMDH assay TRUE 0.5 uM 5 min
SA36650108 OAA Porcine 1 to 100 No Fasnall 00 minMDH assay TRUE 0 uM 0 min
SA36650210 OAA Porcine 1 to 100 No Fasnall 10 minMDH assay TRUE 0 uM 10 min
SA36650311 OAA Porcine 1 to 100 No Fasnall 20 minMDH assay TRUE 0 uM 20 min
SA36650409 OAA Porcine 1 to 100 No Fasnall 05 minMDH assay TRUE 0 uM 5 min
SA36651340 OAA Porcine 1 to 100 10 uM Fasnall 00 minMDH assay TRUE 10.0 uM 0 min
SA36651442 OAA Porcine 1 to 100 10 uM Fasnall 10 minMDH assay TRUE 10.0 uM 10 min
SA36651543 OAA Porcine 1 to 100 10 uM Fasnall 20 minMDH assay TRUE 10.0 uM 20 min
SA36651641 OAA Porcine 1 to 100 10 uM Fasnall 05 minMDH assay TRUE 10.0 uM 5 min
SA36650916 OAA Porcine 1 to 100 1.0 uM Fasnall 00 minMDH assay TRUE 1.0 uM 0 min
SA36651018 OAA Porcine 1 to 100 1.0 uM Fasnall 10 minMDH assay TRUE 1.0 uM 10 min
SA36651119 OAA Porcine 1 to 100 1.0 uM Fasnall 20 minMDH assay TRUE 1.0 uM 20 min
SA36651217 OAA Porcine 1 to 100 1.0 uM Fasnall 05 minMDH assay TRUE 1.0 uM 5 min
SA36652144 OAA Porcine 1 to 100 20 uM Fasnall 00 minMDH assay TRUE 20.0 uM 0 min
SA36652246 OAA Porcine 1 to 100 20 uM Fasnall 10 minMDH assay TRUE 20.0 uM 10 min
SA36652347 OAA Porcine 1 to 100 20 uM Fasnall 20 minMDH assay TRUE 20.0 uM 20 min
SA36652445 OAA Porcine 1 to 100 20 uM Fasnall 05 minMDH assay TRUE 20.0 uM 5 min
SA36651720 OAA Porcine 1 to 100 2.0 uM Fasnall 00 minMDH assay TRUE 2.0 uM 0 min
SA36651822 OAA Porcine 1 to 100 2.0 uM Fasnall 10 minMDH assay TRUE 2.0 uM 10 min
SA36651923 OAA Porcine 1 to 100 2.0 uM Fasnall 20 minMDH assay TRUE 2.0 uM 20 min
SA36652021 OAA Porcine 1 to 100 2.0 uM Fasnall 05 minMDH assay TRUE 2.0 uM 5 min
SA36652524 OAA Porcine 1 to 100 3.0 uM Fasnall 00 minMDH assay TRUE 3.0 uM 0 min
SA36652626 OAA Porcine 1 to 100 3.0 uM Fasnall 10 minMDH assay TRUE 3.0 uM 10 min
SA36652727 OAA Porcine 1 to 100 3.0 uM Fasnall 20 minMDH assay TRUE 3.0 uM 20 min
SA36652825 OAA Porcine 1 to 100 3.0 uM Fasnall 05 minMDH assay TRUE 3.0 uM 5 min
SA36653348 OAA Porcine 1 to 100 40 uM Fasnall 00 minMDH assay TRUE 40.0 uM 0 min
SA36653450 OAA Porcine 1 to 100 40 uM Fasnall 10 minMDH assay TRUE 40.0 uM 10 min
SA36653551 OAA Porcine 1 to 100 40 uM Fasnall 20 minMDH assay TRUE 40.0 uM 20 min
SA36653649 OAA Porcine 1 to 100 40 uM Fasnall 05 minMDH assay TRUE 40.0 uM 5 min
SA36652928 OAA Porcine 1 to 100 4.0 uM Fasnall 00 minMDH assay TRUE 4.0 uM 0 min
SA36653030 OAA Porcine 1 to 100 4.0 uM Fasnall 10 minMDH assay TRUE 4.0 uM 10 min
SA36653131 OAA Porcine 1 to 100 4.0 uM Fasnall 20 minMDH assay TRUE 4.0 uM 20 min
SA36653229 OAA Porcine 1 to 100 4.0 uM Fasnall 05 minMDH assay TRUE 4.0 uM 5 min
SA36653732 OAA Porcine 1 to 100 5.0 uM Fasnall 00 minMDH assay TRUE 5.0 uM 0 min
SA36653834 OAA Porcine 1 to 100 5.0 uM Fasnall 10 minMDH assay TRUE 5.0 uM 10 min
SA36653935 OAA Porcine 1 to 100 5.0 uM Fasnall 20 minMDH assay TRUE 5.0 uM 20 min
SA36654033 OAA Porcine 1 to 100 5.0 uM Fasnall 05 minMDH assay TRUE 5.0 uM 5 min
SA36654136 OAA Porcine 1 to 100 6.0 uM Fasnall 00 minMDH assay TRUE 6.0 uM 0 min
SA36654238 OAA Porcine 1 to 100 6.0 uM Fasnall 10 minMDH assay TRUE 6.0 uM 10 min
SA36654339 OAA Porcine 1 to 100 6.0 uM Fasnall 20 minMDH assay TRUE 6.0 uM 20 min
SA36654437 OAA Porcine 1 to 100 6.0 uM Fasnall 05 minMDH assay TRUE 6.0 uM 5 min
SA36654502 DMEM/F12 no addsNA NA NA NA
SA36654603 BlankNA NA NA NA
SA36654752 DMEM/F12 no addsNA NA NA NA
SA36654853 BlankNA NA NA NA
SA36654901 BlankNA NA NA NA
Showing results 1 to 53 of 53

Collection:

Collection ID:CO003480
Collection Summary:20 µl of the assay mix was mixed with 80 µl of 100% methanol (80% final).
Collection Protocol Filename:DM_metabolomics_samples.txt
Sample Type:Enzymatic assay
Collection Method:80% methanol extraction
Storage Conditions:-80℃
Collection Vials:1.5 ml plastic centrifuge tubes
Storage Vials:1.5 ml plastic centrifuge tubes

Treatment:

Treatment ID:TR003496
Treatment Summary:The MDH activity reconstruction was performed with different Fasnall concentrations.
Treatment Compound:Fasnall
Treatment Doseduration:20 min
Treatment Vehicle:DMSO

Sample Preparation:

Sampleprep ID:SP003494
Sampleprep Summary:The samples were centrifuged at 18,000 g 4C for 20 min. After transferring the supernatant to new tubes, centrifugation was repeated with the same parameters.
Sampleprep Protocol Filename:DM_metabolomics_samples.txt
Processing Storage Conditions:4℃
Extraction Method:80% methanol
Extract Enrichment:None
Extract Cleanup:None
Extract Storage:-80℃
Sample Resuspension:None
Sample Derivatization:None
Sample Spiking:None
Subcellular Location:Enzymatic assay samples

Combined analysis:

Analysis ID AN005516 AN005517
Analysis type MS MS
Chromatography type HILIC HILIC
Chromatography system Shimadzu 20AD Shimadzu 20AD
Column Merck SeQuant ZIC-pHILIC (150 x 2.1mm,5um) Merck SeQuant ZIC-pHILIC (150 x 2.1mm,5um)
MS Type ESI ESI
MS instrument type Triple quadrupole Triple quadrupole
MS instrument name ABI Sciex 5500 QTrap ABI Sciex 5500 QTrap
Ion Mode NEGATIVE POSITIVE
Units Counts per second (cps) Counts per second (cps)

Chromatography:

Chromatography ID:CH004194
Methods Filename:DM_metabolomics_samples.txt
Instrument Name:Shimadzu 20AD
Column Name:Merck SeQuant ZIC-pHILIC (150 x 2.1mm,5um)
Column Pressure:900-3000 psi
Column Temperature:40
Flow Gradient:0-12.5 min, 80-30% B; 12.5-15 min, 30% B; 15-15.2 min, 30-80% B; 15.2-22.5 min, 80% B
Flow Rate:0-20 min, 0.2 ml/min; 20-21 min 0.2-0.3 ml/min; 21-22 min, 0.3 ml/min; 22-22.1 min, 0.2 ml/min; 22.1-22.5 min, 0.2 ml/min
Injection Temperature:4
Sample Injection:1-5 ul
Solvent A:100% Water; 0.01% ammonium hydroxide; 20 mM ammonium bicarbonate
Solvent B:100% Acetonitrile
Chromatography Type:HILIC

MS:

MS ID:MS005241
Analysis ID:AN005516
Instrument Name:ABI Sciex 5500 QTrap
Instrument Type:Triple quadrupole
MS Type:ESI
MS Comments:SCIEX Analyst 1.7.3
Ion Mode:NEGATIVE
Capillary Temperature:500 °C
Capillary Voltage:-4500
Dry Gas Flow:70
Dry Gas Temp:500 °C
Ion Source Temperature:500 °C
Ion Spray Voltage:-4500
Mass Accuracy:UNIT
Source Temperature:500 °C
Spray Voltage:-4500
Dataformat:.WIFF
Analysis Protocol File:DM_metabolomics_samples.txt
  
MS ID:MS005242
Analysis ID:AN005517
Instrument Name:ABI Sciex 5500 QTrap
Instrument Type:Triple quadrupole
MS Type:ESI
MS Comments:SCIEX Analyst 1.7.3
Ion Mode:POSITIVE
Capillary Temperature:500 °C
Capillary Voltage:4500
Dry Gas Flow:70
Dry Gas Temp:500 °C
Ion Source Temperature:500 °C
Ion Spray Voltage:4500
Mass Accuracy:UNIT
Source Temperature:500 °C
Spray Voltage:4500
Dataformat:.WIFF
Analysis Protocol File:DM_metabolomics_samples.txt
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