Summary of Study ST002855
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 PR001782. The data can be accessed directly via it's Project DOI: 10.21228/M8CB14 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 | ST002855 |
| Study Title | Targeting Pancreatic Cancer Metabolic Dependencies through Glutamine Antagonism (Lipidomics mus musculus) |
| Study Summary | Pancreatic ductal adenocarcinoma (PDAC) cells utilize glutamine (Gln) to support proliferation and redox balance. Earlier attempts to inhibit Gln metabolism using glutaminase inhibitors resulted in rapid metabolic reprogramming and therapeutic resistance. Here, we demonstrated that treating PDAC cells with a Gln antagonist, 6-Diazo-5-oxo-L-norleucine (DON), led to a metabolic crisis in vitro. In addition, we observed a profound decrease in tumor growth in various in vivo models using DRP-104 (sirpiglenastat), a pro-drug version of DON that was designed to circumvent DON associated toxicity. We found that ERK signaling is increased as a compensatory mechanism. Combinatorial treatment of DRP-104 and Trametinib led to a significant increase in survival in a syngeneic model PDAC. These proof-of-concept studies suggested that broadly targeting Gln metabolism could provide a therapeutic avenue for PDAC. The combination with an ERK signaling pathway inhibitor could further improve the therapeutic outcome. |
| Institute | New York University |
| Last Name | Encarnacion Rosado |
| First Name | Joel |
| Address | Smilow Research Building Room 907G New York, NY 10016 |
| joel.encarnacion-rosado@nyulangone.org | |
| Phone | 646-501-8984 |
| Submit Date | 2023-09-06 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | mzXML |
| Analysis Type Detail | LC-MS |
| Release Date | 2024-09-12 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR001782 |
| Project DOI: | doi: 10.21228/M8CB14 |
| Project Title: | Targeting Pancreatic Cancer Metabolic Dependencies through Glutamine Antagonism. |
| Project Type: | Manuscript |
| Project Summary: | Pancreatic ductal adenocarcinoma (PDAC) cells utilize glutamine (Gln) to support proliferation and redox balance. Earlier attempts to inhibit Gln metabolism using glutaminase inhibitors resulted in rapid metabolic reprogramming and therapeutic resistance. We demonstrated that treating PDAC cells with a Gln antagonist, 6-Diazo-5-oxo-L-norleucine (DON), led to a metabolic crisis in vitro. In addition, we observed a profound decrease in tumor growth in various in vivo models using DRP-104 (sirpiglenastat), a pro-drug version of DON designed to circumvent DON-associated toxicity. |
| Institute: | New York University |
| Department: | Radiation Oncology |
| Laboratory: | Alec C Kimmelman |
| Last Name: | Encarnacion Rosado |
| First Name: | Joel |
| Address: | Smilow Research Building Room 907G New York, NY 10016 |
| Email: | jencarnacionrosado@salk.edu, Alec.Kimmelman@nyulangone.org |
| Phone: | 646-501-8984 |
Subject:
| Subject ID: | SU002967 |
| Subject Type: | Cultured cells |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
| Gender: | Not applicable |
Factors:
Subject type: Cultured cells; Subject species: Mus musculus (Factor headings shown in green)
| mb_sample_id | local_sample_id | Vehicle |
|---|---|---|
| SA308784 | 15_DON_3 | DON |
| SA308785 | 15_DON_2 | DON |
| SA308786 | 15_DON_1 | DON |
| SA308787 | 15_CNT_2 | Vehicle |
| SA308788 | 15_CNT_3 | Vehicle |
| SA308789 | 15_CNT_1 | Vehicle |
| Showing results 1 to 6 of 6 |
Collection:
| Collection ID: | CO002960 |
| Collection Summary: | Samples were subjected to an LCMS analysis to detect and identify phospholipid molecules and quantify the relative levels of identified lipids. A lipid extraction was carried out on each sample based on the method by Vorkas et. al., cite#1and#2. The dried samples were resolubilized in 10 μL of a 4:3:1 mixture (isopropanol:acetonitrile:water) and analyzed by UPLC-MS/MS with a polarity switching method modified from Vorkas et. al., cite #1. The LC column was a WatersTM CSH-C18 (2.1 x100 mm, 1.7 μm) coupled to a Dionex Ultimate 3000TM system and the column oven temperature was set to 55oC for the gradient elution. The flow rate of 0.3 mL/min was used with the following buffers; A) 60:40 acetonitrile:water, 10 mM ammonium formate, 0.1% formic acid and B) 90:10 isopropanol:acetonitrile, 10 mM ammonium formate, 0.1% formic acid. The gradient profile was as follows; 40-43%B (0-1.25 min), 43-50%B (1.25-2 min), 50-54%B (2-11 min), 54-70%B (11-12 min), 70-99%B (12-18 min), 70-99%B (18-32min), 99-40%B (23-24 min), hold 40%B (1 min). Injection volume was set to 1 μL for all analyses (25 min total run time per injection). MS analyses were carried out by coupling the LC system to a Thermo Q Exactive HFTM mass spectrometer operating in heated electrospray ionization mode (HESI). Method duration was 20 min with a polarity switching data-dependent Top 10 method for both positive and negative modes. Spray voltage for both positive and negative modes was 3.5kV and capillary temperature was set to 320oC with a sheath gas rate of 35, aux gas of 10, and max spray current of 100 μA. The full MS scan for both polarities utilized 120,000 resolution with an AGC target of 3e6 and a maximum IT of 100 ms, and the scan range was from 350-2000 m/z. Tandem MS spectra for both positive and negative mode used a resolution of 15,000, AGC target of 1e5, maximum IT of 50 ms, isolation window of 0.4 m/z, isolation offset of 0.1 m/z, fixed first mass of 50 m/z, and 3-way multiplexed normalized collision energies (nCE) of 10, 35, 80. The minimum AGC target was 5e4 with an intensity threshold of 1e6. All data were acquired in profile mode. The resulting lipids were identified by searching the LipidBlast tandem mass spectral library of lipids cite #3. The top scoring structure match for each data-dependent spectrum was returned using an in-house script for MSPepSearch_x64. Putative lipids were sorted from high to low by their reverse dot scores, and duplicate structures were discarded, retaining only the top-scoring MS2 spectrum and the neutral chemical formula, detected m/z, and detected polarity (+ or -) of the putative lipid was recorded. References 1. Vorkas, P. A. et al. Untargeted UPLC-MS Profiling Pipeline to Expand Tissue Metabolome Coverage: Application to Cardiovascular Disease. Anal. Chem. 87, 4184–4193 (2015). 2. Vorkas, P. A. et al. Metabolic phenotyping of atherosclerotic plaques reveals latent associations between free cholesterol and ceramide metabolism in atherogenesis. J. Proteome Res. 14, 1389–1399 (2015). 3. https://www.ncbi.nlm.nih.gov/pubmed/23817071 |
| Sample Type: | Cultured cells |
Treatment:
| Treatment ID: | TR002976 |
| Treatment Summary: | HY19636 cells were plated in a six-well plate at 1.5x10^6 cells/well and allowed to attach overnight in DMEM. Next, cells were washed with PBS twice and cultured for 24 hours in DMEM supplemented with 10% dialyzed serum was added. Cells were pre-treated with DON (25µM) overnight, media was removed and washed with PBS. Then, cells were frozen in -80C and until metabolite extraction. |
Sample Preparation:
| Sampleprep ID: | SP002973 |
| Sampleprep Summary: | Samples were analyzed with the global and untargeted lipidomics LCMS assays after scaling the lipid extraction to a measured aliquot (~5e6/mL) for each of the 6 samples. SPLASH® LIPIDOMIX® Mass Spec Standard was used as an extraction standard. |
Combined analysis:
| Analysis ID | AN004677 |
|---|---|
| Analysis type | MS |
| Chromatography type | HILIC |
| Chromatography system | Thermo Dionex Ultimate 3000 |
| Column | WatersTM CSH-C18 (2.1 x100 mm, 1.7 μm) |
| MS Type | ESI |
| MS instrument type | Orbitrap |
| MS instrument name | Thermo Q Exactive HF hybrid Orbitrap |
| Ion Mode | UNSPECIFIED |
| Units | ion counts |
Chromatography:
| Chromatography ID: | CH003519 |
| Instrument Name: | Thermo Dionex Ultimate 3000 |
| Column Name: | WatersTM CSH-C18 (2.1 x100 mm, 1.7 μm) |
| Column Temperature: | 55C |
| Flow Gradient: | The gradient profile was as follows; 80-20%B (0-30 min), 20-80%B (30-31 min), 80-80%B (31-42 min) |
| Flow Rate: | 1 μL for all analyses (25 min total run time per injection) |
| Solvent A: | The flow rate of 0.3 mL/min was used with the following buffers; A) 60:40 acetonitrile:water, 10 mM ammonium formate, 0.1% formic acid and B) 90:10 isopropanol:acetonitrile, 10 mM ammonium formate, 0.1% formic acid. The gradient profile was as follows; 40-43%B (0-1.25 min), 43-50%B (1.25-2 min), 50-54%B (2-11 min), 54-70%B (11-12 min), 70-99%B (12-18 min), 70-99%B (18-32min), 99-40%B (23-24 min), hold 40%B (1 min) |
| Solvent B: | acetonitrile |
| Chromatography Type: | HILIC |
MS:
| MS ID: | MS004424 |
| Analysis ID: | AN004677 |
| Instrument Name: | Thermo Q Exactive HF hybrid Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | MS analyses were carried out by coupling the LC system to a Thermo Q Exactive HFTM mass spectrometer operating in heated electrospray ionization mode (HESI). Method duration was 20 min with a polarity switching data-dependent Top 10 method for both positive and negative modes. Spray voltage for both positive and negative modes was 3.5kV and capillary temperature was set to 320C with a sheath gas rate of 35, aux gas of 10, and max spray current of 100 μA. The full MS scan for both polarities utilized 120,000 resolution with an AGC target of 3e6 and a maximum IT of 100 ms, and the scan range was from 350-2000 m/z. Tandem MS spectra for both positive and negative mode used a resolution of 15,000, AGC target of 1e5, maximum IT of 50 ms, isolation window of 0.4 m/z, isolation offset of 0.1 m/z, fixed first mass of 50 m/z, and 3-way multiplexed normalized collision energies (nCE) of 10, 35, 80. The minimum AGC target was 5e4 with an intensity threshold of 1e6. All data were acquired in profile mode. The resulting lipids were identified by searching the LipidBlast tandem mass spectral library of lipids. The top scoring structure match for each data-dependent spectrum was returned using an in-house script for MSPepSearch_x64. Putative lipids were sorted from high to low by their reverse dot scores, and duplicate structures were discarded, retaining only the top-scoring MS2 spectrum and the neutral chemical formula, detected m/z, and detected polarity (+ or -) of the putative lipid was recorded. The “-” is for not-detected peaks. After the correction, zero values were input for non-detected values instead of the blank threshold to avoid false positive. |
| Ion Mode: | UNSPECIFIED |
