Summary of Study ST002226
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 PR001418. The data can be accessed directly via it's Project DOI: 10.21228/M8F409 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 | ST002226 |
| Study Title | Exometabolomics of HK2, 786-O cells cultured in Plasmax media |
| Study Summary | The objective of this study is to analyse the exometabolomics of human epithelial renal cell line HK2 and clear cell renal cell carcinoma (ccRCC) cell lines 786-O, 786-M1A, 786-M2A, OS-RC-2, OS-LM1 and RFX-631 that are cultured with the Plasmax media. This is part 3 of the study, and the experimental number is MS51. |
| Institute | CECAD Research Center |
| Last Name | Yang |
| First Name | Ming |
| Address | Joseph-Stelzmann-Straße 26, Köln, Koeln, 50931, Germany |
| ming.yang@uni-koeln.de | |
| Phone | 4922147884306 |
| Submit Date | 2022-07-15 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | raw(Thermo) |
| Analysis Type Detail | LC-MS |
| Release Date | 2022-08-03 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR001418 |
| Project DOI: | doi: 10.21228/M8F409 |
| Project Title: | Dynamic partitioning of branched-chain amino acids-derived nitrogen supports renal cancer progression |
| Project Summary: | Metabolic reprogramming is critical for tumor initiation and progression. However, the exact impact of specific metabolic changes on cancer progression is poorly understood. Here, we integrate multimodal analyses of primary and metastatic clonally related clear cell renal cancer cells (ccRCC) grown in physiological media to identify key stage-specific metabolic vulnerabilities. We show that a VHL loss-dependent reprogramming of branched-chain amino acid catabolism sustains the de novo biosynthesis of aspartate and arginine enabling tumor cells with the flexibility of partitioning the nitrogen of the amino acids depending on their needs. Importantly, we identify the epigenetic reactivation of argininosuccinate synthase (ASS1), a urea cycle enzyme suppressed in primary ccRCC, as a crucial event for metastatic renal cancer cells to acquire the capability to generate arginine, invade in vitro and metastasize in vivo. Overall, our study uncovers a novel mechanism of metabolic flexibility occurring during ccRCC progression, paving the way for the development of novel stage-specific therapies. |
| Institute: | CECAD Research Center, University Hospital Cologne |
| Last Name: | Yang |
| First Name: | Ming |
| Address: | Joseph-Stelzmann-Straße 26, CECAD Research Center, Köln, Koeln, 50931, Germany |
| Email: | ming.yang@uni-koeln.de |
| Phone: | +4922147884306 |
Subject:
| Subject ID: | SU002312 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
Factors:
Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)
| mb_sample_id | local_sample_id | Cell line | Sample type |
|---|---|---|---|
| SA212296 | MS51-17 | 786-M1A | conditioned media |
| SA212297 | MS51-18 | 786-M1A | conditioned media |
| SA212298 | MS51-19 | 786-M1A | conditioned media |
| SA212299 | MS51-16 | 786-M1A | conditioned media |
| SA212300 | MS51-20 | 786-M1A | conditioned media |
| SA212301 | MS51-23 | 786-M2A | conditioned media |
| SA212302 | MS51-25 | 786-M2A | conditioned media |
| SA212303 | MS51-21 | 786-M2A | conditioned media |
| SA212304 | MS51-24 | 786-M2A | conditioned media |
| SA212305 | MS51-11 | 786-O | conditioned media |
| SA212306 | MS51-15 | 786-O | conditioned media |
| SA212307 | MS51-14 | 786-O | conditioned media |
| SA212308 | MS51-12 | 786-O | conditioned media |
| SA212309 | MS51-13 | 786-O | conditioned media |
| SA212310 | MS51-07 | HK2 | conditioned media |
| SA212311 | MS51-10 | HK2 | conditioned media |
| SA212312 | MS51-08 | HK2 | conditioned media |
| SA212313 | MS51-06 | HK2 | conditioned media |
| SA212314 | MS51-09 | HK2 | conditioned media |
| SA212315 | MS51-02 | No cell | control media |
| SA212316 | MS51-01 | No cell | control media |
| SA212317 | MS51-04 | No cell | control media |
| SA212318 | MS51-05 | No cell | control media |
| SA212319 | MS51-03 | No cell | control media |
| SA212320 | MS51-31 | OSLM1B | conditioned media |
| SA212321 | MS51-33 | OSLM1B | conditioned media |
| SA212322 | MS51-35 | OSLM1B | conditioned media |
| SA212323 | MS51-32 | OSLM1B | conditioned media |
| SA212324 | MS51-34 | OSLM1B | conditioned media |
| SA212325 | MS51-26 | OSRC2 | conditioned media |
| SA212326 | MS51-30 | OSRC2 | conditioned media |
| SA212327 | MS51-27 | OSRC2 | conditioned media |
| SA212328 | MS51-28 | OSRC2 | conditioned media |
| SA212329 | MS51-29 | OSRC2 | conditioned media |
| SA212330 | MS51-40 | RFX631 | conditioned media |
| SA212331 | MS51-39 | RFX631 | conditioned media |
| SA212332 | MS51-37 | RFX631 | conditioned media |
| SA212333 | MS51-36 | RFX631 | conditioned media |
| SA212334 | MS51-38 | RFX631 | conditioned media |
| Showing results 1 to 39 of 39 |
Collection:
| Collection ID: | CO002305 |
| Collection Summary: | 1.5x105 cells were seeded onto a 6-well plate and cultured in the Plasmax media for 24h. The conditioned medium at 24 hours was analysed for exometabolomics, using media without cells as controls. Quantification of protein content and cell counting was carried using additional plates seeded in parallel. |
| Sample Type: | Cultured cells |
Treatment:
| Treatment ID: | TR002324 |
| Treatment Summary: | Cells were cultured in the Plasmax media supplemented with 2.5% FBS for 24 hours. |
Sample Preparation:
| Sampleprep ID: | SP002318 |
| Sampleprep Summary: | Media samples collected at t=0 and t=24hrs were centrifuged at 4 °C for 10 min. 50 µl of supernatants was further extracted with 350 µl of metabolite extraction solution (50% methanol, 30% acetonitrile, 20% ultrapure water, 5 µM final concentration valine-d8). The solution was then centrifuged at 4 °C for 10 min at max speed and the supernatant collected onto LC-MS vials. |
Combined analysis:
| Analysis ID | AN003635 |
|---|---|
| Analysis type | MS |
| Chromatography type | HILIC |
| Chromatography system | Thermo Dionex Ultimate 3000 |
| Column | SeQuant ZIC-pHILIC |
| MS Type | ESI |
| MS instrument type | Orbitrap |
| MS instrument name | Thermo Q Exactive Orbitrap |
| Ion Mode | UNSPECIFIED |
| Units | peak area |
Chromatography:
| Chromatography ID: | CH002690 |
| Chromatography Summary: | Chromatographic separation of metabolites was achieved using a Millipore Sequant ZIC-pHILIC analytical column (5 µm, 2.1 × 150 mm) equipped with a 2.1 × 20 mm guard column (both 5 mm particle size) with a binary solvent system. Solvent A was 20 mM ammonium carbonate, 0.05% ammonium hydroxide; Solvent B was acetonitrile. The column oven and autosampler tray were held at 40 °C and 4 °C, respectively. The chromatographic gradient was run at a flow rate of 0.200 mL/min as follows: 0–2 min: 80% B; 2-17 min: linear gradient from 80% B to 20% B; 17-17.1 min: linear gradient from 20% B to 80% B; 17.1-23 min: hold at 80% B. Samples were randomized and the injection volume was 5 µl. A pooled quality control (QC) sample was generated from an equal mixture of all individual samples and analysed interspersed at regular intervals. |
| Instrument Name: | Thermo Dionex Ultimate 3000 |
| Column Name: | SeQuant ZIC-pHILIC |
| Column Temperature: | 40 |
| Flow Gradient: | 0-2 min: 80% B; 2-17 min: linear gradient from 80% B to 20% B; 17-17.1 min: linear gradient from 20% B to 80% B; 17.1-23 min: hold at 80% B |
| Flow Rate: | 0.200 mL/min |
| Solvent A: | 100% water; 20 mM ammonium carbonate; 0.05% ammonium hydroxide |
| Solvent B: | 100% acetonitrile |
| Chromatography Type: | HILIC |
MS:
| MS ID: | MS003386 |
| Analysis ID: | AN003635 |
| Instrument Name: | Thermo Q Exactive Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | Metabolites were measured with a Thermo Scientific Q Exactive Hybrid Quadrupole-Orbitrap Mass spectrometer (HRMS) coupled to a Dionex Ultimate 3000 UHPLC. The mass spectrometer was operated in full-scan, polarity-switching mode, with the spray voltage set to +4.5 kV/-3.5 kV, the heated capillary held at 320 °C, and the auxiliary gas heater held at 280 °C. The sheath gas flow was set to 55 units, the auxiliary gas flow was set to 15 units, and the sweep gas flow was set to 0 unit. HRMS data acquisition was performed in a range of m/z = 70–900, with the resolution set at 70,000, the AGC target at 1 × 106, and the maximum injection time (Max IT) at 120 ms. Metabolite identities were confirmed using two parameters: (1) precursor ion m/z was matched within 5 ppm of theoretical mass predicted by the chemical formula; (2) the retention time of metabolites was within 5% of the retention time of a purified standard run with the same chromatographic method. Chromatogram review and peak area integration were performed using the Thermo Fisher software Tracefinder 5.0 and the peak area for each detected metabolite was normalized against the total ion count (TIC) of that sample to correct any variations introduced from sample handling through instrument analysis. The normalized areas were used as variables for further statistical data analysis. |
| Ion Mode: | UNSPECIFIED |
