Summary of Study ST002712
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 PR001681. The data can be accessed directly via it's Project DOI: 10.21228/M8DT5S 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 | ST002712 |
| Study Title | Ranolazine induced metabolic rewiring improves melanoma responses to targeted therapy and immunotherapy - metabolomics |
| Study Summary | Metabolomics analysis was performed on A375 melanoma cells resistant to BRAF inhibitor vemurafenib (VR) and cells resistant to VR and treated with fatty acid oxidation inhibitor ranolazine. |
| Institute | University of Colorado Denver |
| Last Name | Haines |
| First Name | Julie |
| Address | 12801 E 17th Ave, Room 1303, Aurora, Colorado, 80045, USA |
| julie.haines@cuanschutz.edu | |
| Phone | 3037243339 |
| Submit Date | 2023-05-09 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | raw(Thermo) |
| Analysis Type Detail | LC-MS |
| Release Date | 2023-06-21 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR001681 |
| Project DOI: | doi: 10.21228/M8DT5S |
| Project Title: | Ranolazine induced metabolic rewiring improves melanoma responses to targeted therapy and immunotherapy |
| Project Summary: | Metabolic rewiring affects resistance of melanoma to targeted- and immuno-therapy. We have found that increased fatty acid oxidation (FAO) during late stages of BRAF inhibitor (BRAFi) treatment enables the establishment of acquired resistance. Targeting FAO with ranolazine in vivo once acquired BRAFi-resistance emerges delays tumour recurrence. Single cell RNAseq analysis revealed that ranolazine diminishes the transcriptional NGFRhigh neural crest stem cell subpopulation, which is refractory against BRAFi and immunotherapy. Moreover, by rewiring the methionine salvage pathway, ranolazine enhanced melanoma immunogenicity through increased antigen presentation and interferon signalling. Combination of ranolazine with anti-PD-L1 antibodies strongly improved survival in mice, where it increased lymphocyte infiltration and enhanced anti-tumour responses. Altogether, we show that ranolazine increases the efficacy of targeted melanoma therapy through fatty acid and methionine salvage metabolic rewiring. Importantly, our study suggests that ranolazine could sensitize BRAFi-resistant tumours to immunotherapy, by modulating melanoma cell recognition and immune infiltration. Ranolazine is an FDA and EMA-approved anti-anginal drug with very mild side effects, and our preclinical data encourage its use as a therapeutic option to improve the two main therapeutic strategies currently used to treat metastatic melanoma. |
| Institute: | University of Colorado Denver |
| Laboratory: | Lab of Angelo D'Alessandro in collaboration with lab of Imanol Arozarena |
| Last Name: | Haines |
| First Name: | Julie |
| Address: | 12801 E 17th Ave, Room 1303, Aurora, Colorado, 80045, USA |
| Email: | julie.haines@cuanschutz.edu |
| Phone: | 3037243339 |
Subject:
| Subject ID: | SU002817 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Gender: | Female |
| Cell Biosource Or Supplier: | ATCC |
| Cell Strain Details: | A375 |
Factors:
Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)
| mb_sample_id | local_sample_id | Treatment |
|---|---|---|
| SA273518 | A375P 8 | VEHICLE |
| SA273519 | A375P 7 | VEHICLE |
| SA273520 | A375P 5 | VEHICLE |
| SA273521 | A375P 6 | VEHICLE |
| SA273522 | A375 VR 8 | VEMURA |
| SA273523 | A375 VR 7 | VEMURA |
| SA273524 | A375 VR 6 | VEMURA |
| SA273525 | A375 VR 5 | VEMURA |
| SA273526 | A375 VR-Rano 8 | VEMURA+RANO |
| SA273527 | A375 VR-Rano 6 | VEMURA+RANO |
| SA273528 | A375 VR-Rano 5 | VEMURA+RANO |
| SA273529 | A375 VR-Rano 7 | VEMURA+RANO |
| Showing results 1 to 12 of 12 |
Collection:
| Collection ID: | CO002810 |
| Collection Summary: | All cell lines had been authenticated in 2021. Cells were expanded to generate enough vials from a single batch before the start of the study. Cell lines were cultured in Dulbecco’s Eagle’s Medium (DMEM) (cat#61965026, Gibco) supplemented with 10% fetal bovine serum (cat#10500064, Gibco) plus 1% penicillin/streptomycin (cat#15140122, Gibco). Cells were grown at 37°C in a 5% CO2 environment. |
| Sample Type: | Melanoma cells |
Treatment:
| Treatment ID: | TR002826 |
| Treatment Summary: | Melanoma cells seeded in 6-well plates were treated for 7 days with a high dose (5 or 10μM as indicated) of vemurafenib before switching to a lower concentration of the drug (0,5 or 1μM). Fresh medium and vemurafenib was added once weekly for further 3-4 weeks until arising colonies grew to confluence. Fatty Acid oxidation inhibitors ranolazine, etomoxir or thiroridazine were added once a week. |
Sample Preparation:
| Sampleprep ID: | SP002823 |
| Sampleprep Summary: | Metabolites from frozen cell pellets were extracted with cold 5:3:2 MeOH:acetonitrile:water at a concentration of 2 million cells/mL. Samples were vortexed 30 min at 4 degrees C then supernatants clarified by centrifugation (10 min, 10,000 g, 4 degrees C) and transferred to autosampler vials. |
| Processing Storage Conditions: | 4℃ |
| Extract Storage: | -80℃ |
Combined analysis:
| Analysis ID | AN004395 | AN004396 |
|---|---|---|
| Analysis type | MS | MS |
| Chromatography type | Reversed phase | Reversed phase |
| Chromatography system | Thermo Vanquish | Thermo Vanquish |
| Column | Phenomenex Kinetex C18 (150 x 2.1mm,1.7um) | Phenomenex Kinetex C18 (150 x 2.1mm,1.7um) |
| MS Type | ESI | ESI |
| MS instrument type | Orbitrap | Orbitrap |
| MS instrument name | Thermo Orbitrap Exploris 480 | Thermo Orbitrap Exploris 480 |
| Ion Mode | NEGATIVE | POSITIVE |
| Units | peak area | peak area |
Chromatography:
| Chromatography ID: | CH003297 |
| Chromatography Summary: | Negative C18 |
| Instrument Name: | Thermo Vanquish |
| Column Name: | Phenomenex Kinetex C18 (150 x 2.1mm,1.7um) |
| Column Temperature: | 45 |
| Flow Gradient: | 0-0.5 min 0% B, 0.5-1.1 min 0-100% B, 1.1-2.75 min hold at 100% B, 2.75-3 min 100-0% B, 3-5 min hold at 0% B |
| Flow Rate: | 450 uL/min |
| Sample Injection: | 10 uL |
| Solvent A: | 95% water 5% acetonitrile 1 mM ammonium acetate |
| Solvent B: | 95% acetonitrile 5% water 1 mM ammonium acetate |
| Chromatography Type: | Reversed phase |
| Chromatography ID: | CH003298 |
| Chromatography Summary: | Positive C18 |
| Instrument Name: | Thermo Vanquish |
| Column Name: | Phenomenex Kinetex C18 (150 x 2.1mm,1.7um) |
| Column Temperature: | 45 |
| Flow Gradient: | 0-0.5 min 5% B, 0.5-1.1 min 5-95% B, 1.1-2.75 min hold at 95% B, 2.75-3 min 95-5% B, 3-5 min hold at 5% B |
| Flow Rate: | 450 uL/min |
| Sample Injection: | 10 uL |
| Solvent A: | 100% water 0.1% formic acid |
| Solvent B: | 100% acetonitrile 0.1% formic acid |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS004144 |
| Analysis ID: | AN004395 |
| Instrument Name: | Thermo Orbitrap Exploris 480 |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | We use a Thermo Orbitrap Exploris 120 (not available in the drop down box). Resolution 120,000, scan range 65-975 m/z, maximum injection time 100 ms, microscans 1, automatic gain control (AGC) detection duration 20 msec, source voltage 2.0 kV, capillary temperature 320 C, vaporizer temp 200 C, and sheath gas 50, auxiliary gas 10, and sweep gas 1 (all nitrogen). Data converted to mzXML using RawConverter. Metabolites were annotated and integrated using Maven in conjunction with the KEGG database. |
| Ion Mode: | NEGATIVE |
| MS ID: | MS004145 |
| Analysis ID: | AN004396 |
| Instrument Name: | Thermo Orbitrap Exploris 480 |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | We use a Thermo Orbitrap Exploris 120 (not available in the drop down box). Resolution 120,000, scan range 65-975 m/z, maximum injection time 100 ms, microscans 1, automatic gain control (AGC) detection duration 20 msec, source voltage 3.5 kV, capillary temperature 320 C, vaporizer temp 200 C, and sheath gas 50, auxiliary gas 10, and sweep gas 1 (all nitrogen). Data converted to mzXML using RawConverter. Metabolites were annotated and integrated using Maven in conjunction with the KEGG database. |
| Ion Mode: | POSITIVE |
