Summary of Study ST003365

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 IDST003365
Study TitleIntracellular and medium metabolomics for BT-474 breast cancer cells treated with a range of Fasnall and GSK2194069 concentrations for 24 h
Study TypeIntracellular metabolomics, medium metabolomics
Study SummaryIntracellular and medium metabolomics for BT-474 breast cancer cells treated with a range of Fasnall concentrations for 24 h
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 Groups15
Total Subjects45
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:SU003486
Subject Type:Cultured cells
Subject Species:Homo sapiens
Taxonomy ID:9606
Age Or Age Range:60
Gender:Female
Cell Strain Details:BT-474, breast cancer cell line

Factors:

Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)

mb_sample_id local_sample_id Sample source Drug Treatment
SA36644946 BT474 med GSK 1000BT-474 Breast cancer cells 1000 nM GSK2194069
SA36645006 BT474 cells GSK 1000BT-474 Breast cancer cells 1000 nM GSK2194069
SA36645138 BT474 med GSK 1000BT-474 Breast cancer cells 1000 nM GSK2194069
SA36645230 BT474 med GSK 1000BT-474 Breast cancer cells 1000 nM GSK2194069
SA36645313 BT474 cells GSK 1000BT-474 Breast cancer cells 1000 nM GSK2194069
SA36645420 BT474 cells GSK 1000BT-474 Breast cancer cells 1000 nM GSK2194069
SA36644349 BT474 med Fasnall 1BT-474 Breast cancer cells 1 uM Fasnall
SA36644441 BT474 med Fasnall 1BT-474 Breast cancer cells 1 uM Fasnall
SA36644508 BT474 cells Fasnall 1BT-474 Breast cancer cells 1 uM Fasnall
SA36644633 BT474 med Fasnall 1BT-474 Breast cancer cells 1 uM Fasnall
SA36644722 BT474 cells Fasnall 1BT-474 Breast cancer cells 1 uM Fasnall
SA36644815 BT474 cells Fasnall 1BT-474 Breast cancer cells 1 uM Fasnall
SA36645518 BT474 cells GSK 200BT-474 Breast cancer cells 200 nM GSK2194069
SA36645644 BT474 med GSK 200BT-474 Breast cancer cells 200 nM GSK2194069
SA36645736 BT474 med GSK 200BT-474 Breast cancer cells 200 nM GSK2194069
SA36645828 BT474 med GSK 200BT-474 Breast cancer cells 200 nM GSK2194069
SA36645904 BT474 cells GSK 200BT-474 Breast cancer cells 200 nM GSK2194069
SA36646011 BT474 cells GSK 200BT-474 Breast cancer cells 200 nM GSK2194069
SA36646117 BT474 cells Fasnall 40BT-474 Breast cancer cells 40 uM Fasnall
SA36646224 BT474 cells Fasnall 40BT-474 Breast cancer cells 40 uM Fasnall
SA36646343 BT474 med Fasnall 40BT-474 Breast cancer cells 40 uM Fasnall
SA36646410 BT474 cells Fasnall 40BT-474 Breast cancer cells 40 uM Fasnall
SA36646535 BT474 med Fasnall 40BT-474 Breast cancer cells 40 uM Fasnall
SA36647245 BT474 med GSK 500BT-474 Breast cancer cells 500 nM GSK2194069
SA36647305 BT474 cells GSK 500BT-474 Breast cancer cells 500 nM GSK2194069
SA36647419 BT474 cells GSK 500BT-474 Breast cancer cells 500 nM GSK2194069
SA36647537 BT474 med GSK 500BT-474 Breast cancer cells 500 nM GSK2194069
SA36647612 BT474 cells GSK 500BT-474 Breast cancer cells 500 nM GSK2194069
SA36647729 BT474 med GSK 500BT-474 Breast cancer cells 500 nM GSK2194069
SA36646634 BT474 med Fasnall 5BT-474 Breast cancer cells 5 uM Fasnall
SA36646716 BT474 cells Fasnall 5BT-474 Breast cancer cells 5 uM Fasnall
SA36646850 BT474 med Fasnall 5BT-474 Breast cancer cells 5 uM Fasnall
SA36646923 BT474 cells Fasnall 5BT-474 Breast cancer cells 5 uM Fasnall
SA36647042 BT474 med Fasnall 5BT-474 Breast cancer cells 5 uM Fasnall
SA36647109 BT474 cells Fasnall 5BT-474 Breast cancer cells 5 uM Fasnall
SA36647821 BT474 cells controlBT-474 Breast cancer cells Control
SA36647939 BT474 med controlBT-474 Breast cancer cells Control
SA36648007 BT474 cells controlBT-474 Breast cancer cells Control
SA36648131 BT474 med controlBT-474 Breast cancer cells Control
SA36648214 BT474 cells controlBT-474 Breast cancer cells Control
SA36648347 BT474 med controlBT-474 Breast cancer cells Control
SA36648451 BT474 med Fasnall 40BT-474 40 uM Fasnall
SA36648532 BT474 med no cellsNA Control
SA36648640 BT474 med no cellsNA Control
SA36648748 BT474 med no cellsNA Control
SA36648854 BlankNA NA
SA36648953 DMEM/F12 no addsNA NA
SA36649052 BlankNA NA
SA36649125 BlankNA NA
SA36649203 DMEM/F12 no addsNA NA
SA36649326 DMEM/F12 no addsNA NA
SA36649427 BlankNA NA
SA36649502 BlankNA NA
SA36649601 BlankNA NA
Showing results 1 to 54 of 54

Collection:

Collection ID:CO003479
Collection Summary:For medium metabolites, 100 µl of medium was mixed with 400 µl of 100% methanol (80% final). For intracellular metabolite samples, the medium was aspirated, and cells were washed with PBS volume matching the volume of the medium. Metabolites were extracted with ice-cold 80% methanol. The volume of the solvent was 500 µl per 6-cm Petri dish (scaled according to the ratio of surface areas for other cell containers). After adding the methanol solution, cells were scraped from the plates, and all the content was transferred to Eppendorf tubes.
Collection Protocol Filename:DM_metabolomics_samples.txt
Sample Type:Breast cancer cells
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:TR003495
Treatment Summary:Cells were grown in RPMI-1640 supplemented with 10% dialyzed FBS and treated with various drug concentrations.
Treatment Compound:Fasnall, GSK2194069
Treatment Doseduration:24 h
Treatment Vehicle:DMSO
Cell Growth Container:6-cm Petri dishes
Cell Media:RPMI-1640
Cell Envir Cond:37C, 5% CO2
Cell Pct Confluence:~70%
Cell Media Lastchanged:24 h before collection

Sample Preparation:

Sampleprep ID:SP003493
Sampleprep Summary:Intracellular metabolites were extracted with ice-cold 80% methanol, and medium samples were extracted with 100% methanol. 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:Intracellular metabolites and medium metabolites

Combined analysis:

Analysis ID AN005514 AN005515
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:CH004193
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:MS005239
Analysis ID:AN005514
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:MS005240
Analysis ID:AN005515
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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