Summary of Study ST003488

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.

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Study IDST003488
Study TitleTryptophan metabolite profiling of cell supernatants (culture media) from TNBC cell line BT549 grown in suspension with TDO2 knockdown and IDO1 compensation.
Study SummaryCulture media harvested from TNBC cell line BT549 with stable TDO2 knockdown (shTDO2) but harbor IDO1 compensation. The cells were treated with TDO2/IDO1 dual inhibitor AT-0174, IDO1 inhibitor epacadostat, or TDO2 inhibitor 680C91 for 48 hours.
Institute
University of Colorado Anschutz Medical Campus
Last NameHaines
First NameJulie
Address12801 E 17th Ave, Room 1303, Aurora, Colorado, 80045, USA
Emailjulie.haines@cuanschutz.edu
Phone3037243339
Submit Date2024-09-17
Raw Data AvailableYes
Raw Data File Type(s)raw(Thermo)
Analysis Type DetailLC-MS
Release Date2024-10-03
Release Version1
Julie Haines Julie Haines
https://dx.doi.org/10.21228/M80R74
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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:SU003616
Subject Type:Cultured cells
Subject Species:Homo sapiens
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
SA384009AA22-2M-710uM 680c91 culture media
SA384010AA22-2M-810uM 680c91 culture media
SA384011AA22-2M-910uM 680c91 culture media
SA384012AA22-2M-410uM AT0174 culture media
SA384013AA22-2M-510uM AT0174 culture media
SA384014AA22-2M-610uM AT0174 culture media
SA384015AA22-2M-1210uM Epacadotstat culture media
SA384016AA22-2M-1010uM Epacadotstat culture media
SA384017AA22-2M-1110uM Epacadotstat culture media
SA384018AA22-2M-2DMSO culture media
SA384019AA22-2M-1DMSO culture media
SA384020AA22-2M-3DMSO culture media
SA384021AA22-4M-3EV culture media
SA384022AA22-4M-2EV culture media
SA384023AA22-4M-1EV culture media
SA384027AA22-3M-1shSCR culture media
SA384028AA22-3M-3shSCR culture media
SA384029AA22-3M-2shSCR culture media
SA384030AA22-3M-12shTDO2-054 culture media
SA384031AA22-3M-10shTDO2-054 culture media
SA384032AA22-3M-11shTDO2-054 culture media
SA384033AA22-3M-6shTDO2-082 culture media
SA384034AA22-3M-5shTDO2-082 culture media
SA384035AA22-3M-4shTDO2-082 culture media
SA384036AA22-3M-9shTDO2-098 culture media
SA384037AA22-3M-8shTDO2-098 culture media
SA384038AA22-3M-7shTDO2-098 culture media
SA384024AA22-4M-6TDO2 OE culture media
SA384025AA22-4M-5TDO2 OE culture media
SA384026AA22-4M-4TDO2 OE culture media
Showing results 1 to 30 of 30

Collection:

Collection ID:CO003609
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:TR003625
Treatment Summary:TNBC cell line BT549 with stable TDO2 knockdown (shTDO2) was first cultured for 24 hours and treated with DMSO (control), 10 µM AT-0174 and 10 µM 680c91 or 10 µM Epacadostat for 48 hours. For comparison, BT549 with stable TDO2 overexpression (TDO2 OE) were also cultured relative to empty vector (EV). Culture media was not changed during the duration of the experiments.

Sample Preparation:

Sampleprep ID:SP003623
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 AN005726
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:CH004345
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:MS005449
Analysis ID:AN005726
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
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