Summary of Study ST004014

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 PR002501. The data can be accessed directly via it's Project DOI: 10.21228/M8C55X 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 IDST004014
Study TitleCompetitive inhibition of SNAT2-dependent MeAIB uptake by saturating concentrations of L-alanine.
Study SummaryMeAIB uptake in SNAT2-expressing HY15549 cells was inhibited by co-treating with a concentration gradient of L-alanine from 20 mM to 0.04 mM and 0.5 mM of MeAIB.
Institute
University of British Columbia
DepartmentBiochemistry & Molecular Biology
LaboratoryParker laboratory
Last NameParker
First NameSeth
Address950 W 28th Ave, Vancouver, British Columbia, V6H 0B3, Canada
Emailseth.parker@bcchr.ca
Phone6048753121
Submit Date2025-06-19
Raw Data AvailableYes
Raw Data File Type(s)cdf
Analysis Type DetailGC-MS
Release Date2025-07-28
Release Version1
Seth Parker Seth Parker
https://dx.doi.org/10.21228/M8C55X
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR002501
Project DOI:doi: 10.21228/M8C55X
Project Title:Metabolomics analysis of SNAT2-deficient cells: implications for the discovery of selective transporter inhibitors
Project Type:Manuscript
Project Summary:Amino acid uptake by the solute carrier family of transporter proteins is critical to support cell metabolism, and inhibition of transporter activity represents a tractable strategy to restrict nutrient availability to cancer cells. A small molecule inhibitor of the sodium-coupled neutral amino acid transporter 2 (SNAT2), 3-(N-methyl(4-methylphenyl)sulfonamido)-N-(2-trifluoromethylbenzyl)thiophene-2-carboxamide (MMTC/57E), was recently identified and was shown to inhibit cell proliferation when combined with glucose transport inhibitors in breast and pancreatic cancer cell lines. In this study, we use mass spectrometry-based metabolomics and establish cell-based assays for the SNAT2 transporter. We show that SNAT2 knockout cells have significant defects in amino acid availability. Using our established assays, we fail to observe that MMTC/57E inhibits SNAT2 activity likely due to its poor solubility.
Institute:University of British Columbia
Department:Biochemistry & Molecular Biology
Laboratory:Parker laboratory
Last Name:Parker
First Name:Seth
Address:950 W 28th Ave, Vancouver, British Columbia, V6H 0B3, Canada
Email:seth.parker@bcchr.ca
Phone:6048753121

Subject:

Subject ID:SU004152
Subject Type:Mammal
Subject Species:Mus musculus
Taxonomy ID:10090
Gender:Female

Factors:

Subject type: Mammal; Subject species: Mus musculus (Factor headings shown in green)

mb_sample_id local_sample_id Factor Sample source
SA461910B6Treated HY15549
SA461911C10Treated HY15549
SA461912C9Treated HY15549
SA461913C8Treated HY15549
SA461914C7Treated HY15549
SA461915C6Treated HY15549
SA461916C5Treated HY15549
SA461917C4Treated HY15549
SA461918C3Treated HY15549
SA461919C2Treated HY15549
SA461920C1Treated HY15549
SA461921B10Treated HY15549
SA461922B9Treated HY15549
SA461923B8Treated HY15549
SA461924B7Treated HY15549
SA461925B5Treated HY15549
SA461926B4Treated HY15549
SA461927A8Treated HY15549
SA461928A1Treated HY15549
SA461929A2Treated HY15549
SA461930A3Treated HY15549
SA461931A4Treated HY15549
SA461932A5Treated HY15549
SA461933A6Treated HY15549
SA461934A7Treated HY15549
SA461935A9Treated HY15549
SA461936B1Treated HY15549
SA461937A10Treated HY15549
SA461938B2Treated HY15549
SA461939B3Treated HY15549
SA461940A0_r2Untreated HY15549
SA461941C0_r2Untreated HY15549
SA461942C0_r1Untreated HY15549
SA461943B0_r1Untreated HY15549
SA461944B0_r2Untreated HY15549
SA461945A0_r1Untreated HY15549
Showing results 1 to 36 of 36

Collection:

Collection ID:CO004145
Collection Summary:Cells were washed twice with room temperature (20°C) PBS before adding 2 mL of PBS containing 0.5 mM MeAIB and 0-20 mM of L-alanine. After a 30-minute incubation at 20°C, the transport buffer was aspirated and the cells were washed three times with ice cold 0.9% NaCl. To extract metabolites, 1 mL of ice cold 80% methanol containing 2.5 nmol of 13C3-alanine was added to each well. The plates were shaken at ~500 rpm for 15 minutes at room temperature and 900 uL was transferred to a new tube and dried using a SpeedVac for GCMS analysis.
Sample Type:Tumor cells

Treatment:

Treatment ID:TR004161
Treatment Summary:No treatment.

Sample Preparation:

Sampleprep ID:SP004158
Sampleprep Summary:Dried samples were derivatized for GCMS analysis. 25 µL of methoxyamine hydrochloride (20 mg/mL, prepared in pyridine) was added to each tube, which was vortexed and incubated at 37°C for 60 minutes. Following the incubation, 25 µL of MTBSTFA + 1% TBDMSCl was added to each tube, which was vortexed and centrifuged at 21,300 x g for 15-minutes at 20°C. 40 µL of the resulting supernatant was transferred to a plastic sample vial for GCMS analysis.

Chromatography:

Chromatography ID:CH005029
Chromatography Summary:Agilent DB35-MS (30m x 0.25mm, 0.25µm) was interfaced with an inert column (5m x 0.15mm, 0 µm) to the MSD (Mass Selective Detector). Initial oven temperature maintained at 75°C for 3 minutes, ramped to 255°C at 7.5°C/min, held for 10 minutes, ramped to 320°C at 10°C/min, held for 5 minutes. The inlet was maintained at 270°C in splitless mode; an injection volume of 1 µL was used for all samples.
Instrument Name:Agilent 8890
Column Name:Agilent DB-35MS (30m x 0.25mm, 0.25µm) interfaced with an inert column (5m x 0.15mm, 0 µm)
Column Temperature:75°C
Flow Gradient:100% A
Flow Rate:1 mL/min
Injection Temperature:270°C
Solvent A:UHP Helium
Solvent B:None
Oven Temperature:75°C
Chromatography Type:GC

Analysis:

Analysis ID:AN006619
Analysis Type:MS
Chromatography ID:CH005029
Num Factors:2
Num Metabolites:4
Units:ion counts
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