Summary of Study ST003433

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.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

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 IDST003433
Study TitleIntracellular and medium metabolomics of BT-474 cells treated with dimethylmalonate, Fasnall, and GSK2194069
Study SummaryBT-474 human breast cancer cells were treated with dimethylmalonate, Fasnall, and GSK2194069 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 Date2024-08-21
Num Groups11
Total Subjects33
PublicationsSubmission Pending
Raw Data AvailableYes
Raw Data File Type(s)mzML, wiff
Analysis Type DetailLC-MS
Release Date2024-09-12
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:SU003560
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 Sample Type Drug Treatment
SA37801626 Fasnall 1 uM DMM 0 mM cellsBT-474 breast cancer cells Intracellular 1 uM Fasnall
SA37801718 Fasnall 1 uM DMM 0 mM cellsBT-474 breast cancer cells Intracellular 1 uM Fasnall
SA37801810 Fasnall 1 uM DMM 0 mM cellsBT-474 breast cancer cells Intracellular 1 uM Fasnall
SA37801939 Fasnall 1 uM GSK 1 uM cellsBT-474 breast cancer cells Intracellular 1 uM Fasnall + 1 uM GSK2194069
SA37802036 Fasnall 1 uM GSK 1 uM cellsBT-474 breast cancer cells Intracellular 1 uM Fasnall + 1 uM GSK2194069
SA37802133 Fasnall 1 uM GSK 1 uM cellsBT-474 breast cancer cells Intracellular 1 uM Fasnall + 1 uM GSK2194069
SA37802214 Fasnall 1 uM DMM 5 mM cellsBT-474 breast cancer cells Intracellular 1 uM Fasnall + 5 mM DMM
SA37802330 Fasnall 1 uM DMM 5 mM cellsBT-474 breast cancer cells Intracellular 1 uM Fasnall + 5 mM DMM
SA37802422 Fasnall 1 uM DMM 5 mM cellsBT-474 breast cancer cells Intracellular 1 uM Fasnall + 5 mM DMM
SA37802528 Fasnall 40 uM DMM 0 mM cellsBT-474 breast cancer cells Intracellular 40 uM Fasnall
SA37802620 Fasnall 40 uM DMM 0 mM cellsBT-474 breast cancer cells Intracellular 40 uM Fasnall
SA37802712 Fasnall 40 uM DMM 0 mM cellsBT-474 breast cancer cells Intracellular 40 uM Fasnall
SA37802838 Fasnall 40 uM GSK 1 uM cellsBT-474 breast cancer cells Intracellular 40 uM Fasnall + 1 uM GSK2194069
SA37802935 Fasnall 40 uM GSK 1 uM cellsBT-474 breast cancer cells Intracellular 40 uM Fasnall + 1 uM GSK2194069
SA37803041 Fasnall 40 uM GSK 1 uM cellsBT-474 breast cancer cells Intracellular 40 uM Fasnall + 1 uM GSK2194069
SA37803124 Fasnall 40 uM DMM 5 mM cellsBT-474 breast cancer cells Intracellular 40 uM Fasnall + 5 mM DMM
SA37803232 Fasnall 40 uM DMM 5 mM cellsBT-474 breast cancer cells Intracellular 40 uM Fasnall + 5 mM DMM
SA37803316 Fasnall 40 uM DMM 5 mM cellsBT-474 breast cancer cells Intracellular 40 uM Fasnall + 5 mM DMM
SA37803429 Fasnall 0 uM DMM 5 mM cellsBT-474 breast cancer cells Intracellular 5 mM DMM
SA37803521 Fasnall 0 uM DMM 5 mM cellsBT-474 breast cancer cells Intracellular 5 mM DMM
SA37803613 Fasnall 0 uM DMM 5 mM cellsBT-474 breast cancer cells Intracellular 5 mM DMM
SA37803719 Fasnall 5 uM DMM 0 mM cellsBT-474 breast cancer cells Intracellular 5 uM Fasnall
SA37803811 Fasnall 5 uM DMM 0 mM cellsBT-474 breast cancer cells Intracellular 5 uM Fasnall
SA37803927 Fasnall 5 uM DMM 0 mM cellsBT-474 breast cancer cells Intracellular 5 uM Fasnall
SA37804037 Fasnall 5 uM GSK 1 uM cellsBT-474 breast cancer cells Intracellular 5 uM Fasnall + 1 uM GSK2194069
SA37804140 Fasnall 5 uM GSK 1 uM cellsBT-474 breast cancer cells Intracellular 5 uM Fasnall + 1 uM GSK2194069
SA37804234 Fasnall 5 uM GSK 1 uM cellsBT-474 breast cancer cells Intracellular 5 uM Fasnall + 1 uM GSK2194069
SA37804331 Fasnall 5 uM DMM 5 mM cellsBT-474 breast cancer cells Intracellular 5 uM Fasnall + 5 mM DMM
SA37804423 Fasnall 5 uM DMM 5 mM cellsBT-474 breast cancer cells Intracellular 5 uM Fasnall + 5 mM DMM
SA37804515 Fasnall 5 uM DMM 5 mM cellsBT-474 breast cancer cells Intracellular 5 uM Fasnall + 5 mM DMM
SA37804625 Fasnall 0 uM DMM 0 mM cellsBT-474 breast cancer cells Intracellular Control
SA37804717 Fasnall 0 uM DMM 0 mM cellsBT-474 breast cancer cells Intracellular Control
SA37804809 Fasnall 0 uM DMM 0 mM cellsBT-474 breast cancer cells Intracellular Control
SA37804943 BlankNA Blank NA
SA37805001 BlankNA Blank NA
SA37805108 RPMI bottle D2ONA Medium NA
SA37805207 RPMI bottleNA Medium NA
SA37805306 RPMI bottle D2ONA Medium NA
SA37805405 RPMI bottleNA Medium NA
SA37805504 RPMI bottle D2ONA Medium NA
SA37805603 RPMI bottleNA Medium NA
SA37805702 DMEM/F12NA QC NA
SA37805842 DMEM/F12NA QC NA
Showing results 1 to 43 of 43

Collection:

Collection ID:CO003553
Collection Summary: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:TR003569
Treatment Summary:Cells were grown in RPMI-1640 supplemented with 10% dialyzed FBS and treated with various concentrations of dimethylmalonate (DMM) for 24 h.
Treatment Compound:Dimethylmalonate (DMM), Fasnall, and GSK2194069
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:SP003567
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 AN005639 AN005640
Analysis type MS MS
Chromatography type HILIC HILIC
Chromatography system Shimadzu 20AD Shimadzu 20AD
Column Merck SeQuant ZIC-HILIC (150 x 2.1mm,5um) Merck SeQuant ZIC-HILIC (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:CH004282
Instrument Name:Shimadzu 20AD
Column Name:Merck SeQuant ZIC-HILIC (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:MS005363
Analysis ID:AN005639
Instrument Name:ABI Sciex 5500 QTrap
Instrument Type:Triple quadrupole
MS Type:ESI
MS Comments:Data were analyzed with SCIEX Multiquant 3.0.3.
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
Desolvation Gas Flow:70
Desolvation Temperature:500 °C
  
MS ID:MS005364
Analysis ID:AN005640
Instrument Name:ABI Sciex 5500 QTrap
Instrument Type:Triple quadrupole
MS Type:ESI
MS Comments:Data were analyzed with SCIEX Multiquant 3.0.3.
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
Desolvation Gas Flow:70
Desolvation Temperature:500 °C
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