Summary of Study ST003623
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 PR002208. The data can be accessed directly via it's Project DOI: 10.21228/M86Z6P 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 | ST003623 |
| Study Title | NRF2 supports non-small cell lung cancer growth independently of CBP/p300-enhanced glutathione synthesis: Global metabolomics analysis on A549 cells at different NRF2 status (Part 1 of 3) |
| Study Type | Untargeted Metabolomics |
| Study Summary | This study aims at discovering metabolic changes in A549 cancer cells in the presence and absence of NRF2, and a mutant NRF2 genotypes. Metabolites were extracted from cell pellets by using an LLE method with Methanol/Chloroform/water. The aqueous layer was analyzed by HILIC-HRMS. An in-house RT library was used to identify metabolites. Statistical analyses was performed to identify statistically significant changes in the metabolism. 35 metabolites presented differential abundance between NRF2 knockdown and wildtype conditions. In particular, GSH and several glutamate dipeptides were significantly depleted upon NRF2 knockdown, in line with the prevailing role of NRF2 in controlling GSH biosynthesis. Disruption of additional metabolites involved in the PPP (sedoheptulose-7-phosphate, S7P), nucleotide (CMP, IMP) and amino acid (kynurenine, homocitrulline) metabolism were also observed upon NRF2 knockdown. |
| Institute | Genentech Inc. |
| Last Name | Wang |
| First Name | Mike |
| Address | 1 DNA Way, South San Francisco, CA 94080, USA |
| wang.mike@gene.com | |
| Phone | 6502457991 |
| Submit Date | 2024-12-06 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | mzML |
| Analysis Type Detail | LC-MS |
| Release Date | 2025-01-02 |
| Release Version | 1 |
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Collection:
| Collection ID: | CO003746 |
| Collection Summary: | All cell lines were obtained from the Genentech cell bank (Yu et al, 2015) and grown in RPMI or DMEM supplemented with 10% FBS, 1% glutamine, and 1% penicillin-streptomycin at 37°C, 5% CO2. Cell lines were confirmed via STR profiling and were mycoplasma free. A549 and A549shNRF2-BIND rescue cell lines were generated using the Piggybac system. Donor constructs were co-transfected with transposase (Ding et al, 2005) at a 4:1 ratio using Lipofetamine 3000 (Thermo) according to manufacturer’s protocol. 3-days post-transfection, puromycin selection (1µg/mL) was initiated and was performed for ~2 weeks prior to experimentation. A549shNRF2-BIND rescue cells represent polyclonal pools. Cell viability was measured using CellTiter-Glo® Luminescent Cell Viability Assay (Promega) using the Envision 2103 Plate Reader (Promega). Cell growth curves were generated using the IncuCyte System (Sartorius) according to the manufacturer’s protocol. Curve fits were performed in Prism. Clonogenic assays were performed by washing cells in PBS, staining with 0.5% crystal violet solution (Sigma HT90132) for 5 min, and washing 3x with PBS. Four replicates of 20-30 million A549shNRF2-BIND-luc, WT NRF2 or Neh4/5mut cells were treated with 2.5ng/mL dox for 48h. For A485 testing, cells were treated 1µM A-485 or DMSO for 24h before harvesting. After treatment, cells were trypsinized, washed 3x with ice cold PBS and flash frozen in liquid nitrogen for metabolic quenching. |
| Sample Type: | Cultured cells |
