Summary of Study ST001902

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 PR001197. The data can be accessed directly via it's Project DOI: 10.21228/M8041N 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.

Perform statistical analysis  |  Show all samples  |  Show named metabolites  |  Download named metabolite data  
Download mwTab file (text)   |  Download mwTab file(JSON)   |  Download data files (Contains raw data)
Study IDST001902
Study TitleMetabolomics analysis of AsPC-1 PDAC cells treated with Porcupine inhibitor (LGK974)
Study SummaryWNT signaling promotes pancreatic ductal adenocarcinoma (PDAC) through diverse effects on proliferation, differentiation, survival, and stemness. A subset of PDAC with inactivating mutations in ring finger protein 43 (RNF43) have growth dependency on autocrine WNT ligand signaling, which renders them susceptible to porcupine inhibitors (PORCNi) that block WNT ligand acylation and secretion. For this study, non-targeted metabolomic analyses were performed to explore the therapeutic response of RNF43-mutant PDAC to the PORCNi LGK974. AsPC-1 (RNF43-mutant) PDAC cells were treated with 25 nM LGK974 to explore stable isotope-resolved metabolomics with uniform 1, D-glucose [U13-C6] labeling.
Institute
University of California, Los Angeles
DepartmentPathology & Laboratory Medicine
LaboratoryDawson Lab
Last NameDawson
First NameDavid
Address10833 LeConte Avenue
Emailddawson@mednet.ucla.edu
Phone310-825-0618
Submit Date2021-07-19
Num Groups2
Total Subjects6
Raw Data AvailableYes
Raw Data File Type(s)mzML
Analysis Type DetailAPI-MS
Release Date2022-04-19
Release Version1
David Dawson David Dawson
https://dx.doi.org/10.21228/M8041N
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:


Project:

Project ID:PR001197
Project DOI:doi: 10.21228/M8041N
Project Title:Metabolomics analysis of AsPC-1 PDAC cells treated with Porcupine inhibitor (LGK974)
Project Summary:WNT signaling promotes pancreatic ductal adenocarcinoma (PDAC) through diverse effects on proliferation, differentiation, survival, and stemness. A subset of PDAC with inactivating mutations in ring finger protein 43 (RNF43) have growth dependency on autocrine WNT ligand signaling, which renders them susceptible to porcupine inhibitors (PORCNi) that block WNT ligand acylation and secretion. For this study, non-targeted metabolomic analyses were performed to explore the therapeutic response of RNF43-mutant PDAC to the PORCNi LGK974. AsPC-1 (RNF43-mutant) PDAC cells were treated with 25 nM LGK974 to explore stable isotope-resolved metabolomics with uniform 1, D-glucose [U13-C6] labeling.
Institute:University of California, Los Angeles
Department:Pathology & Laboratory Medicine
Laboratory:Dawson Lab
Last Name:David
First Name:Dawson
Address:10833 Le Conte Avenue, Los Angeles, CA, 90095, USA
Email:ddawson@mednet.ucla.edu
Phone:(310) 267-2799

Subject:

Subject ID:SU001980
Subject Type:Mammal
Subject Species:Homo sapiens
Taxonomy ID:9606

Factors:

Subject type: Mammal; Subject species: Homo sapiens (Factor headings shown in green)

mb_sample_id local_sample_id Treatment
SA176338KA-01625 nM LGK974
SA176339KA-01425 nM LGK974
SA176340KA-01325 nM LGK974
SA176341KA-011Control
SA176342KA-012Control
SA176343KA-010Control
Showing results 1 to 6 of 6

Collection:

Collection ID:CO001973
Collection Summary:AsPC-1 PDAC cells were treated with DMSO vehicle control or 25 nM LGK974 (in triplicate per group). 1 x 10^6 cells were washed with ice-cold 150 mM ammonium acetate twice before adding 1 mL of ice-cold 80% methanol. After vigorous vortexing, samples were centrifuged at maximum speed, the aqueous layer was transferred to a glass vial and the metabolites were dried under vacuum. Metabolites were resuspended in 50 μL 70% acetonitrile (ACN) and 5 μL of this solution used for the mass spectrometer-based analysis. The analysis was performed on a Q Exactive (Thermo Fisher Scientific) in polarity-switching mode with positive voltage 4.0 kV and negative voltage 4.0 kV. The mass spectrometer was coupled to an UltiMate 3000RSLC (Thermo Fisher Scientific) UHPLC system. Mobile phase A was 5 mM ammonium acetate (NH4AcO), pH 9.9, B was acetonitrile and the separation achieved on a Luna 3 mm NH2 100 A column (150 × 2.0 mm, Phenomenex). The flow was 200 μL/min, and the gradient ran from 15% A to 95% A in 18 min, followed by an isocratic step for 9 min and re-equilibration for 7 min. Metabolites were detected and quantified as area under the curve based on retention time and accurate mass (≤ 3 p.p.m.) using the TraceFinder 3.1 (Thermo Fisher Scientific). Relative amounts of metabolites between various conditions, as well as percentage of [13C6] glucose labelling, were calculated and corrected for naturally occurring 13C abundance.
Sample Type:Cultured cells

Treatment:

Treatment ID:TR001992
Treatment Summary:AsPC-1 PDAC cells were plated at 500,000 cells per well in 6 well plates in ultra-low attachment conditions. Cells were pre-treated with 25 nM LGK974 or DMSO vehicle control for 12 hours, maintained in normal media conditions. Media was exchanged with [13C6] glucose labeling media and cells were re-treated with 25 nM LGK974 or DMSO vehicle control for 24 hours.

Sample Preparation:

Sampleprep ID:SP001986
Sampleprep Summary:Cells were washed with ice-cold 150 mM ammonium acetate twice before adding 1 mL of ice-cold 80% methanol. After vigorous vortexing, samples were centrifuged at maximum speed, the aqueous layer was transferred to a glass vial and the metabolites were dried under vacuum. Metabolites were resuspended in 50 μL 70% acetonitrile (ACN) and 5 μL of this solution used for the mass spectrometer-based analysis. The analysis was performed on a Q Exactive (Thermo Fisher Scientific) in polarity-switching mode with positive voltage 4.0 kV and negative voltage 4.0 kV. The mass spectrometer was coupled to an UltiMate 3000RSLC (Thermo Fisher Scientific) UHPLC system. Mobile phase A was 5 mM ammonium acetate (NH4AcO), pH 9.9, B was acetonitrile and the separation achieved on a Luna 3 mm NH2 100 A column (150 × 2.0 mm, Phenomenex). The flow was 200 μL/min, and the gradient ran from 15% A to 95% A in 18 min, followed by an isocratic step for 9 min and re-equilibration for 7 min. Metabolites were detected and quantified as area under the curve based on retention time and accurate mass (≤ 3 p.p.m.) using the TraceFinder 3.1 (Thermo Fisher Scientific). Relative amounts of metabolites between various conditions, as well as percentage of [13C6]glucose labeling, were calculated and corrected for naturally occurring 13C abundance.

Combined analysis:

Analysis ID AN003093 AN003094
Analysis type MS MS
Chromatography type HILIC HILIC
Chromatography system Q Exactive Q Exactive
Column Luna 3 mm NH2 100 A Luna 3 mm NH2 100 A
MS Type API API
MS instrument type Orbitrap Orbitrap
MS instrument name Thermo Q Exactive Orbitrap Thermo Q Exactive Orbitrap
Ion Mode POSITIVE NEGATIVE
Units area area

Chromatography:

Chromatography ID:CH002284
Chromatography Summary:Mobile phase A was 5 mM ammonium acetate (NH4AcO), pH 9.9, B was acetonitrile and the separation achieved on a Luna 3 mm NH2 100 A column (150 × 2.0 mm, Phenomenex). The flow was 200 μL/min, and the gradient ran from 15% A to 95% A in 18 min, followed by an isocratic step for 9 min and re-equilibration for 7 min.
Instrument Name:Q Exactive
Column Name:Luna 3 mm NH2 100 A
Flow Rate:200 uL/min
Solvent A:100% water; 5 mM ammonium acetate, pH 9.9
Solvent B:100% acetonitrile
Chromatography Type:HILIC

MS:

MS ID:MS002875
Analysis ID:AN003093
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:API
MS Comments:Metabolites were detected and quantified as area under the curve based on retention time and accurate mass (≤ 3 p.p.m.) using the TraceFinder 3.1 (Thermo Fisher Scientific). Relative amounts of metabolites between various conditions, as well as percentage of [13C6] glucose labeling, were calculated and corrected for naturally occurring 13C abundance.
Ion Mode:POSITIVE
Ion Spray Voltage:4000
  
MS ID:MS002876
Analysis ID:AN003094
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:API
MS Comments:Metabolites were detected and quantified as area under the curve based on retention time and accurate mass (≤ 3 p.p.m.) using the TraceFinder 3.1 (Thermo Fisher Scientific). Relative amounts of metabolites between various conditions, as well as percentage of [13C6] glucose labeling, were calculated and corrected for naturally occurring 13C abundance.
Ion Mode:NEGATIVE
Ion Spray Voltage:4000
  logo