Summary of Study ST002527
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.
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.
| Study ID | ST002527 |
| Study Title | HPLC-MS-MS analysis amino acid levels in PDAC IF samples upon arginase inhibition |
| Study Summary | To test the hypothesis that myeloid arginase-1 activity could be responsible for pancreatic ductal adenocarninoma microenvironmental arginine starvation (PMID: 30990168), we generated orthotopic allograft mPDAC tumors in a mouse model with myeloid specific Arg1 knockout (LysM-Cre+/+-; Arg1fl/fl) and control animals (Arg1fl/fl). We also tested this with pharmacological inhibition of arginase-1 with the small-molecule inhibitor CB-1158. We then isolated IF from these tumors at end-stage and measured the levels of amino acids including arginine and ornithine in these samples. |
| Institute | University of Chicago |
| Last Name | Apiz Saab |
| First Name | Juan |
| Address | 929 E. 57th St. |
| japizsaab@uchicago.edu | |
| Phone | 7738346506 |
| Submit Date | 2022-08-05 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | raw(Thermo) |
| Analysis Type Detail | LC-MS |
| Release Date | 2023-04-13 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
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: | SU002627 |
| 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 | Genotype | Treatment |
|---|---|---|---|
| SA254603 | T202 | Arg1fl/fl | NA |
| SA254604 | T237 | Arg1fl/fl | NA |
| SA254605 | T111 | Arg1fl/fl | NA |
| SA254606 | T217 | Arg1fl/fl | NA |
| SA254607 | T236 | LysM-Cre+/+; Arg1fl/fl | NA |
| SA254608 | T203 | LysM-Cre+/+; Arg1fl/fl | NA |
| SA254609 | T215 | LysM-Cre+/+; Arg1fl/fl | NA |
| SA254610 | T298 | LysM-Cre+/+; Arg1fl/fl | NA |
| SA254611 | T214 | LysM-Cre+/+; Arg1fl/fl | NA |
| SA254612 | T233 | LysM-Cre+/+; Arg1fl/fl | NA |
| SA254613 | T234 | LysM-Cre+/+; Arg1fl/fl | NA |
| SA254614 | T245 | NA | CB1158 |
| SA254615 | T242 | NA | CB1158 |
| SA254616 | T241 | NA | CB1158 |
| SA254617 | T244 | NA | CB1158 |
| SA254618 | T231 | NA | CB1158 |
| SA254619 | T238 | NA | Vehicle |
| SA254620 | T239 | NA | Vehicle |
| SA254621 | T400 | NA | Vehicle |
| SA254622 | T282 | NA | Vehicle |
| SA254623 | T278 | NA | Vehicle |
| Showing results 1 to 21 of 21 |
Collection:
| Collection ID: | CO002620 |
| 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: | TR002639 |
| Treatment Summary: | For genetically modified mice, LysM-Cre and Arg1fl/fl C57BL6J mice were bred to generate LysM-Cre +/+; Arg1fl/fl and litter mate control Arg1fl/fl mice. Orthotopic pancreatic cancer tumors were implanted in C57BL6J mice at 8-12 weeks of age. At endpoint, mice were euthanized by cervical dislocation, and tumors were harvested for TIF extraction. For Pharmacological inhibition, orthotopic tumors were implanted in C57BL6J mice at 8-12 weeks of age. 4 weeks after induction, once tumors were close end stage disease, CB-1158 (MedChem Express) dissolved in sterile water was administered by oral gavage at 100mg/kg. The acidity caused by the HCl in the drug solution was neutralized by adding equivalent amount of NaOH and an equivalent NaCl in sterile water solution was prepared as vehicle. 2hrs after treatment with CB-1158 or vehicle, mice were euthanized by cervical dislocation, and tumors were harvested for TIF extraction. |
Sample Preparation:
| Sampleprep ID: | SP002633 |
| 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 | AN004161 |
|---|---|
| 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: | CH003080 |
| 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% (vol/vol) water, 5% (vol/vol) acetonitrile, 10 mM ammonium hydroxide, 10 mM ammonium acetate, pH = 9.0 |
| Solvent B: | 100% Acetonitrile |
| Chromatography Type: | HILIC |
MS:
| MS ID: | MS003908 |
| Analysis ID: | AN004161 |
| 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 |
