Summary of Study ST002530

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 PR001627. The data can be accessed directly via it's Project DOI: 10.21228/M8D434 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 IDST002530
Study TitleLCMS analysis of amino acid levels in PDAC interstitial fluid samples upon ass1 cancer cell knock out
Study SummaryWe used CRISPR-Cas9 to knockout (KO) Ass1 in mPDAC cells. We tested if inhibiting arginine de novo synthesis would impair PDAC tumor growth. To test this, we generated orthotopic PDAC tumors with mPDAC-Ass1 KO cells and control cells where Ass1 was re-expressed (Ass1KO; mASS1).
Institute
University of Chicago
Last NameApiz Saab
First NameJuan
Address929 E. 57th St.
Emailjapizsaab@uchicago.edu
Phone7738346506
Submit Date2023-03-24
Raw Data AvailableYes
Raw Data File Type(s)raw(Thermo)
Analysis Type DetailLC-MS
Release Date2023-04-13
Release Version1
Juan Apiz Saab Juan Apiz Saab
https://dx.doi.org/10.21228/M8D434
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR001627
Project DOI:doi: 10.21228/M8D434
Project Title:Pancreatic tumors activate arginine biosynthesis to adapt to myeloid-driven amino acid stress
Project Summary:Nutrient stress in the tumor microenvironment requires cancer cells to adopt adaptive metabolic programs to maintain survival and proliferation. Therefore, knowledge of microenvironmental nutrient levels and how cancer cells cope with such nutrition is critical to understand the metabolism underpinning cancer cell biology. Previously, we performed quantitative metabolomics of the interstitial fluid (the local perfusate) of murine pancreatic ductal adenocarcinoma (PDAC) tumors to comprehensively characterize nutrient availability in the microenvironment of these tumors (Sullivan et al., 2019a). Here, we develop Tumor Interstitial Fluid Medium (TIFM), a cell culture medium that contains nutrient levels representative of the PDAC microenvironment, enabling study of PDAC metabolism under physiological nutrition. We show that PDAC cells cultured in TIFM, compared to standard laboratory models, adopt a cellular state more similar to PDAC cells in tumors. Further, using the TIFM model we identified arginine biosynthesis as a metabolic adaptation PDAC cells engage to cope with microenvironmental arginine starvation driven by myeloid cells in PDAC tumors. Altogether, these data show that nutrient availability in tumors is an important determinant of cancer cell metabolism and behavior, and cell culture models that incorporate physiological nutrient availability have improved fidelity and enable the discovery of novel cancer metabolic phenotypes.
Institute:University of Chicago
Last Name:Apiz Saab
First Name:Juan
Address:929 E. 57th St.
Email:japizsaab@uchicago.edu
Phone:7738346506

Subject:

Subject ID:SU002630
Subject Type:Mammal
Subject Species:Mus musculus
Taxonomy ID:10090

Factors:

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

mb_sample_id local_sample_id Sample type
SA25468620210804_17ASS1KO
SA25468720210804_20ASS1KO
SA25468820210804_01ASS1KO
SA25468920210804_13ASS1KO
SA25469020210804_22ASS1KO
SA25469120210804_19ASS1KO
SA25469220210804_11ASS1KO
SA25469320210804_02ASS1KO
SA25469420210804_08ASS1KO
SA25469520210804_05ASS1KO
SA25469620210804_10ASS1KO
SA25469720210804_15ASS1KO;mASS1
SA25469820210804_16ASS1KO;mASS1
SA25469920210804_21ASS1KO;mASS1
SA25470020210804_14ASS1KO;mASS1
SA25470120210804_18ASS1KO;mASS1
SA25470220210804_03ASS1KO;mASS1
SA25470320210804_04ASS1KO;mASS1
SA25470420210804_06ASS1KO;mASS1
SA25470520210804_07ASS1KO;mASS1
SA25470620210804_09ASS1KO;mASS1
SA25470720210804_12ASS1KO;mASS1
Showing results 1 to 22 of 22

Collection:

Collection ID:CO002623
Collection Summary:Tumors were rapidly dissected after euthanizing animals. Tumors were weighed and rinsed in blood bank saline solution (150 mM NaCl) and blotted on filter paper (VWR, Radnor, PA, 28298–020). The process of dissection and tumor preparation took < 3min. Tumors were cut in half and put onto 20µm nylon mesh filters (Spectrum Labs, Waltham, MA, 148134) on top of 50 mL conical tubes, and centrifuged for 10min. at 4°C at 400xg. IF was then collected, snap-frozen in liquid nitrogen and stored at -80°C until further analysis.
Sample Type:Interstitial Fluid

Treatment:

Treatment ID:TR002642
Treatment Summary:Orthotopic tumors were implanted in C57BL6J mice at 8-12 weeks of age. 4 weeks after induction interstitial fluid and tumors were collected.

Sample Preparation:

Sampleprep ID:SP002636
Sampleprep Summary:polar metabolites were extracted from 5µL IF samples using 45µL 75:25:0.1 HPLC grade acetonitrile:methanol:formic acid extraction mix into which a mixture of isotopically labeled amino acids of known concentrations (Cambridge Isotope Laboratories, MSK-A2-1.2) was added. Samples were vortexed for 10 min, and centrifuged at maximum speed for 10 min. 30 μL of each extract was removed and dried under nitrogen gas and stored −80°C.

Combined analysis:

Analysis ID AN004164
Analysis type MS
Chromatography type HILIC
Chromatography system Thermo Dionex Ultimate 3000
Column Waters XBridge Amide (100 x 4.6mm, 3.5um)
MS Type ESI
MS instrument type Orbitrap
MS instrument name Thermo Q Exactive Orbitrap
Ion Mode UNSPECIFIED
Units Peak area (m/z)

Chromatography:

Chromatography ID:CH003083
Chromatography Summary:HPLC
Instrument Name:Thermo Dionex Ultimate 3000
Column Name:Waters XBridge Amide (100 x 4.6mm, 3.5um)
Column Temperature:25
Flow Gradient:0 min, 15% A; 2.5 min, 64% A; 12.4 min, 40% A; 12.5 min, 30% A; 12.5-14 min, 30% A; 14-21 min, 15% A
Flow Rate:150 μL/min
Solvent A:95% water/5% acetonitrile; 10 mM ammonium hydroxide; 10 mM ammonium acetate, pH = 9.0
Solvent B:100% acetonitrile
Chromatography Type:HILIC

MS:

MS ID:MS003911
Analysis ID:AN004164
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:In positive/negative polarity switching mode, an m/z scan range from 60 to 900 was chosen and MS1 data was collected at a resolution of 70,000. The automatic gain control (AGC) target was set at 1 × 106 and the maximum injection time was 200 ms. The targeted ions were subsequently fragmented, using the higher energy collisional dissociation (HCD) cell set to 30% normalized collision energy in MS2 at a resolution power of 17,500. Besides matching m/z, target metabolites are identified by matching either retention time with analytical standards and/or MS2 fragmentation pattern. Data acquisition and analysis were carried out by Xcalibur 4.1 software and Tracefinder 4.1 software, respectively (both from Thermo Fisher Scientific).
Ion Mode:UNSPECIFIED
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