Summary of Study ST003486
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 PR002140. The data can be accessed directly via it's Project DOI: 10.21228/M80R74 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 | ST003486 |
| Study Title | Tryptophan metabolite profiling of cell supernatants (culture media) from TNBC cell line BT549 grown in suspension with TDO inhibition. |
| Study Summary | Indole-focused metabolomics analysis of cell supernatants (i.e. cell culture media) from TNBC cell lines with pharmacological inhibition of TDO2 by AT-0174 or 680c91. |
| Institute | University of Colorado Anschutz Medical Campus |
| 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 | 2024-09-17 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | raw(Thermo) |
| Analysis Type Detail | LC-MS |
| Release Date | 2024-10-03 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR002140 |
| Project DOI: | doi: 10.21228/M80R74 |
| Project Title: | Blocking tryptophan catabolism reduces triple-negative breast cancer invasive capacity |
| Project Summary: | Anchorage-independent triple-negative breast cancer (TNBC) cells exhibit elevated levels of the tryptophan (TRP) catabolizing enzyme tryptophan 2,3-dioxygenase 2 (TDO2) compared to the same cells grown in two-dimensional culture. Tracing of 13C11-TRP demonstrated that anchorage-independent culture and/or inflammatory cytokines that activate nuclear factor kappa-light-chain-enhancer of activated B (NFκB) increase TRP catabolism and production of downstream catabolites such as kynurenine (KYN), which activate the aryl hydrocarbon receptor (AhR). TDO2 expression is heterogeneous within TNBC cell lines. To determine the function of TDO2, both pharmacologic inhibition and genetic manipulation were conducted. TDO2 knockdown revealed a compensatory increase in indoleamine 2,3-dioxygenase 1 (IDO1), a non-homologous TRP catabolizing enzyme, indicating that dual inhibition of these two enzymes is necessary to reliably block TRP catabolism. Thus, we tested a newly developed TDO2/IDO1 dual inhibitor, AT-0174, and found that it effectively inhibits TNBC TRP catabolism. Furthermore, AT-0174 treatment or AhR inhibitor significantly decreased TNBC anchorage-independent survival, invasive capacity, and expression of mesenchymal genes and protein, while exogenous KYN increased invasion through AhR-mediated ZEB1 expression. Thus, dual inhibition of TDO2/IDO1 may prove efficacious against TNBC progression. |
| Institute: | University of Colorado Anschutz Medical Campus |
| Laboratory: | Lab of Angelo D'Alessandro in collaboration with lab of Jennifer Richer |
| 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: | SU003614 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
| Gender: | Female |
Factors:
Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)
| mb_sample_id | local_sample_id | treatment | Sample source |
|---|---|---|---|
| SA383988 | BTS-68048-1M | 10 uM 680c91 | culture media |
| SA383989 | BTS-68048-2M | 10 uM 680c91 | culture media |
| SA383990 | BTS-68048-3M | 10 uM 680c91 | culture media |
| SA383991 | BTS-10A48-1M | 10 uM AT0174 | culture media |
| SA383992 | BTS-10A48-2M | 10 uM AT0174 | culture media |
| SA383993 | BTS-10A48-3M | 10 uM AT0174 | culture media |
| SA383985 | BTS-1A48-1M | 1 uM AT0174 | culture media |
| SA383986 | BTS-1A48-2M | 1 uM AT0174 | culture media |
| SA383987 | BTS-1A48-3M | 1 uM AT0174 | culture media |
| SA383994 | BTS-D48-1M | DMSO vehicle | culture media |
| SA383995 | BTS-D48-2M | DMSO vehicle | culture media |
| SA383996 | BTS-D48-3M | DMSO vehicle | culture media |
| Showing results 1 to 12 of 12 |
Collection:
| Collection ID: | CO003607 |
| Collection Summary: | At the end of the cell culturing timecourse, the media were harvested and centrifuged at 1500 rpm, 4℃. The resulting supernatants were collected and frozen at -80 ℃. |
| Sample Type: | Culture Media |
Treatment:
| Treatment ID: | TR003623 |
| Treatment Summary: | TNBC cell line BT549 was cultured for 24 hours then treated with DMSO (control), 1 µM AT-0174, 10 µM AT-0174, or 10 uM 680c91 for 48 hours. Culture media was not changed during the duration of the experiments. |
Sample Preparation:
| Sampleprep ID: | SP003621 |
| Sampleprep Summary: | Culture media aliquots were thawed on ice then 20 uL was treated with 480 uL of ice cold 5:3:2 methanol:acetonitrile:water (v/v/v). Samples were vortexed 30 min at 4 degrees C followed by centrifugation for 10 min at 18,000 g. Aliquots of supernatant (50 uL) were dried using a speedvac then reconstituted in an equivalent volume of 0.1% formic acid. Samples were maintained at 4°C until analysis that same day. |
| Processing Storage Conditions: | 4℃ |
| Extract Storage: | -80℃ |
Combined analysis:
| Analysis ID | AN005724 |
|---|---|
| Analysis type | MS |
| Chromatography type | Reversed phase |
| Chromatography system | Thermo Vanquish |
| Column | Phenomenex Kinetex C18 (150 x 2.1mm,1.7um) |
| MS Type | ESI |
| MS instrument type | Orbitrap |
| MS instrument name | Thermo Q Exactive Orbitrap |
| Ion Mode | POSITIVE |
| Units | peak area |
Chromatography:
| Chromatography ID: | CH004343 |
| 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: | 6 uL |
| Solvent A: | 100% water; 0.1% formic acid |
| Solvent B: | 100% acetonitrile; 0.1% formic acid |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS005447 |
| Analysis ID: | AN005724 |
| Instrument Name: | Thermo Q Exactive Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | Resolution 70,000, scan range 65-900 m/z, maximum injection time 200 ms, microscans 2, automatic gain control (AGC) 3 x 10^6 ions, source voltage 4.0 kV, capillary temperature 320 C, and sheath gas 45, auxiliary gas 15, and sweep gas 0 (all nitrogen). Data converted to mzXML using RawConverter. Metabolites were annotated and integrated using Maven in conjunction with the KEGG database. |
| Ion Mode: | POSITIVE |
