Summary of Study ST003760

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 PR002343. The data can be accessed directly via it's Project DOI: 10.21228/M8S548 This work is supported by NIH grant, U2C- DK119886. See: https://www.metabolomicsworkbench.org/about/howtocite.php

Study ID	ST003760
Study Title	Untargeted lipidomics of combination gemcitabine/paclitaxel attenuated (CombAT) PDAC cells
Study Summary	In this work produced combination gemcitabine/paclitaxel attenuated (CombAT) Panc1 and MiaPaCa2 cells, and used untargeted lipidomics to profiled changes in their lipidome. This dataset includes CombAT cells and control (CON) cells, alongside internal quality controls (iQC) and blank samples for background subtraction (as described under sample prep). These data revealed extensive increases across 166 lipid species in Panc1 CombAT cell lines compared with CON, including increases in many triacylglycerol (TG) species and total TG. Increases were also seen in several TG species in CombAT MiaPaCa2 cells, with 30 lipid species increased in total when compared with CON. Data were analysed using MS-DIAL v4.9.
Institute	Victor Chang Cardiac Research Institute
Last Name	Hancock
First Name	Sarah
Address	Level 7 Lowy Packer Building, 405 Liverpool Street, Darlinghurst, NSW 2010, Australia
Email	s.hancock@victorchang.edu.au
Phone	+61414537526
Submit Date	2025-02-19
Raw Data Available	Yes
Raw Data File Type(s)	mzML
Analysis Type Detail	LC-MS
Release Date	2025-03-21
Release Version	1

Select appropriate tab below to view additional metadata details:

Project:

Project ID:	PR002343
Project DOI:	doi: 10.21228/M8S548
Project Title:	Untargeted lipidomics of combination gemcitabine/paclitaxel attenuated (CombAT) PDAC cells
Project Summary:	Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive disease with few treatment options and poor survivability. In this work we sought to characterise metabolic adaptations to gemcitabine (GEMC)-based chemotherapy exposure to discover new therapeutic targets for improving treatment efficacy. This project examined the impact of combination gemcitabine/paclitaxel resistance on the lipidomics of Panc1 and MiaPaCa2 PDAC cell lines, finding upregulated de novo lipogenesis in both cell lines.
Institute:	Victor Chang Cardiac Research Institute
Last Name:	Hancock
First Name:	Sarah
Address:	Level 7 Lowy Packer Building, 405 Liverpool Street, Darlinghurst, NSW 2010, Australia
Email:	s.hancock@victorchang.edu.au
Phone:	+61414537526

Subject:

Subject ID:	SU003893
Subject Type:	Cultured cells
Subject Species:	Homo sapiens
Taxonomy ID:	9606

Factors:

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

mb_sample_id	local_sample_id	Sample source	source	group
SA409111	M8C_41_Neg	Pancreas	MiaPaCa2 cells	CombAT
SA409112	M8C_57_Neg	Pancreas	MiaPaCa2 cells	CombAT
SA409113	M8C_57_Pos	Pancreas	MiaPaCa2 cells	CombAT
SA409114	M8C_41_Pos	Pancreas	MiaPaCa2 cells	CombAT
SA409115	M8C_43_Neg	Pancreas	MiaPaCa2 cells	CombAT
SA409116	M8C_43_Pos	Pancreas	MiaPaCa2 cells	CombAT
SA409105	MiaPaCa-2_34_Neg	Pancreas	MiaPaCa2 cells	CON
SA409106	MiaPaCa-2_57_Pos	Pancreas	MiaPaCa2 cells	CON
SA409107	MiaPaCa-2_36_Pos	Pancreas	MiaPaCa2 cells	CON
SA409108	MiaPaCa-2_34_Pos	Pancreas	MiaPaCa2 cells	CON
SA409109	MiaPaCa-2_57_Neg	Pancreas	MiaPaCa2 cells	CON
SA409110	MiaPaCa-2_36_Neg	Pancreas	MiaPaCa2 cells	CON
SA409123	P28C_55_Neg	Pancreas	Panc1 cells	CombAT
SA409124	P28C_55_Pos	Pancreas	Panc1 cells	CombAT
SA409125	P28C_56_Neg	Pancreas	Panc1 cells	CombAT
SA409126	P28C_57_Neg	Pancreas	Panc1 cells	CombAT
SA409127	P28C_57_Pos	Pancreas	Panc1 cells	CombAT
SA409128	P28C_56_Pos	Pancreas	Panc1 cells	CombAT
SA409117	Panc-1_33_Neg	Pancreas	Panc1 cells	CON
SA409118	Panc-1_33_Pos	Pancreas	Panc1 cells	CON
SA409119	Panc-1_34_Neg	Pancreas	Panc1 cells	CON
SA409120	Panc-1_34_Pos	Pancreas	Panc1 cells	CON
SA409121	Panc-1_35_Neg	Pancreas	Panc1 cells	CON
SA409122	Panc-1_35_Pos	Pancreas	Panc1 cells	CON
SA409097	Blank_Neg_20220922100435	-	Blank	Blank
SA409098	Blank_Neg	-	Blank	Blank
SA409099	Blank_Pos	-	Blank	Blank
SA409100	Blank_Pos_20220922093322	-	Blank	Blank
SA409101	IQC_Neg_Start_20220922110658	-	pool	iQC
SA409102	IQC_Pos_Start	-	pool	iQC
SA409103	IQC_Pos_Start_20220922103547	-	pool	iQC
SA409104	IQC_Neg_Start	-	pool	iQC

Showing results 1 to 32 of 32

Showing results 1 to 32 of 32

Collection:

Collection ID:	CO003886
Collection Summary:	Samples were obtained from established immortalised adherent human pancreatic ductal adenocarcinoma cells (Panc1 & MiaPaCa2). Cells were cultured in high glucose DMEM containing 4.5 g/L glucose and 4 mM glutamine and were supplemented with 10% FCS. Cells were cultured under 5% CO2 at 37ºC and routinely passaged every 2-3 days (when ~80-90% confluent). Cell lines were regularly screened for mycoplasma infection.
Sample Type:	Pancreas

Treatment:

Treatment ID:	TR003902
Treatment Summary:	Combination gemcitabine/paclitaxel attenuated (CombAT) cells were produced by serial treatment with escalating doses of gemcitabine/paclitaxel (10:1) for approximately 12 weeks. The final concentration of gemcitabine for Panc1 cells was 28 nM (2.8 nM paclitaxel), and 8 nM (0.8 nM paclitaxel) for MiaPaCa2 cells.

Sample Preparation:

Sampleprep ID:	SP003899
Sampleprep Summary:	Lipids were extracted from cells grown to 80-90% confluency in 6 well plates using a modified methyl tert-butyl ether (MTBE) method. Adherent cells were washed with ice-cold PBS and scraped into methanol containing 0.01% butylated hydroxytoluene and 100 pmol each of internal standards (phosphatidylcholine 19:0/19:0, phosphatidylethanolamine 17:0/17:0, phosphatidylserine 17:0/17:0, lysophosphatidylcholine 17:0, lysophosphatidylethanolamine 14:0, phosphatidylglycerol 17:0/17:0, phosphatidic acid 17:0/17:0, ceramide d18:1/17:0, dihydrosphingomyelin 12:0, cholesteryl ester 22:1, diacylglycerol 17:0/17:0, D5-triacylglycerol 16:0/16:0/16:0 and cardiolipin 14:0/14:0/14:0/14:0). An empty well was also scraped for use in background subtraction. To this, MTBE was added at a ratio of 3:1 v/v, and extracts were rotated overnight at 4ºC. Following overnight rotation, 1 part of ice-cold 150 mM ammonium acetate was added, samples were vortexed vigorously and centrifuged at 2000 x g for 5 min. The upper organic phase containing lipids was transferred into a 1.5 mL autosampler vial and dried under N2 at 37ºC Dried lipids were reconstituted in chloroform:methanol:water (60:30:4.5 v/v/v), transferred to a sleeved vial, and stored at -20ºC until analysis. The aqueous phase containing the protein pellet was dried under N2 and digested with 1M NaOH overnight at 4ºC. Digested protein was diluted 1:2 v/v with water and protein concentration was determined using a Pierce BCA assay kit (ThermoFisher Scientific)

Sampleprep ID:

SP003899

Sampleprep Summary:

Lipids were extracted from cells grown to 80-90% confluency in 6 well plates using a modified methyl tert-butyl ether (MTBE) method. Adherent cells were washed with ice-cold PBS and scraped into methanol containing 0.01% butylated hydroxytoluene and 100 pmol each of internal standards (phosphatidylcholine 19:0/19:0, phosphatidylethanolamine 17:0/17:0, phosphatidylserine 17:0/17:0, lysophosphatidylcholine 17:0, lysophosphatidylethanolamine 14:0, phosphatidylglycerol 17:0/17:0, phosphatidic acid 17:0/17:0, ceramide d18:1/17:0, dihydrosphingomyelin 12:0, cholesteryl ester 22:1, diacylglycerol 17:0/17:0, D5-triacylglycerol 16:0/16:0/16:0 and cardiolipin 14:0/14:0/14:0/14:0). An empty well was also scraped for use in background subtraction. To this, MTBE was added at a ratio of 3:1 v/v, and extracts were rotated overnight at 4ºC. Following overnight rotation, 1 part of ice-cold 150 mM ammonium acetate was added, samples were vortexed vigorously and centrifuged at 2000 x g for 5 min. The upper organic phase containing lipids was transferred into a 1.5 mL autosampler vial and dried under N2 at 37ºC Dried lipids were reconstituted in chloroform:methanol:water (60:30:4.5 v/v/v), transferred to a sleeved vial, and stored at -20ºC until analysis. The aqueous phase containing the protein pellet was dried under N2 and digested with 1M NaOH overnight at 4ºC. Digested protein was diluted 1:2 v/v with water and protein concentration was determined using a Pierce BCA assay kit (ThermoFisher Scientific)

Combined analysis:

Analysis ID	AN006174	AN006175
Chromatography ID	CH004687	CH004687
MS ID	MS005879	MS005880
Analysis type	MS	MS
Chromatography type	Reversed phase	Reversed phase
Chromatography system	Thermo Dionex Ultimate 3000	Thermo Dionex Ultimate 3000
Column	Waters ACQUITY UPLC CSH C18 (100 x 2.1 mm, 1.7 μm)	Waters ACQUITY UPLC CSH C18 (100 x 2.1 mm, 1.7 μm)
MS Type	ESI	ESI
MS instrument type	Orbitrap	Orbitrap
MS instrument name	Thermo Q Exactive HF hybrid Orbitrap	Thermo Q Exactive HF hybrid Orbitrap
Ion Mode	POSITIVE	NEGATIVE
Units	nmol/mg protein	nmol/mg protein

Chromatography:

Chromatography ID:	CH004687
Instrument Name:	Thermo Dionex Ultimate 3000
Column Name:	Waters ACQUITY UPLC CSH C18 (100 x 2.1 mm, 1.7 μm)
Column Temperature:	60°C
Flow Gradient:	0.1-1.5 min 68% A (32% B), 1.4-4 min 55% A (45% B), 4-5 min 48% A (52% B), 5-8 min 42% A (58% B), 8-11 min 34% A (66% B), 11-14 min 30% A (70% B), 14-18 min 25% A (75% B), 18-21 min 3% A (97% B), 21-25 min 3% A (97% B), 25.1-30 min 68% A (32% B).
Flow Rate:	0.26 mL/min
Solvent A:	60% Acetonitrile/40% Water; 10 mM Ammonium Formate; 0.1% Formic acid
Solvent B:	10% Acetonitrile/90% Isopropanol; 10 mM Ammonium Formate; 0.1% Formic acid
Chromatography Type:	Reversed phase

MS:

MS ID:	MS005879
Analysis ID:	AN006174
Instrument Name:	Thermo Q Exactive HF hybrid Orbitrap
Instrument Type:	Orbitrap
MS Type:	ESI
MS Comments:	Source conditions were as follows: a spray voltage 4.0 kV in positive ion mode and 3.5 kV in negative ion mode, capillary temperature 290°C, S lens RF 50, and auxiliary gas heater temperature of 250°C. Nitrogen was used as both source and collision gas, with the sheath gas flow rate set at 20 and the auxiliary gas flow rate at 5 (arbitrary units). Data were acquired in full scan/data-dependent MS2 mode (full-scan resolution 70,000 FWHM, max ion injection time 50 ms, scan range m/z 200–1,500), with the 10 most abundant ions being subjected to collision-induced dissociation using an isolation window of 1.5 Da and a normalized stepped collision energy of 15/27 eV. Product ions were detected at a resolution of 17,500. An exclusion list for background ions was developed using extraction blanks, and mass calibration was performed in both positive and negative ionization modes before analysis to ensure mass accuracy of 5 ppm in full-scan mode.
Ion Mode:	POSITIVE

MS ID:	MS005880
Analysis ID:	AN006175
Instrument Name:	Thermo Q Exactive HF hybrid Orbitrap
Instrument Type:	Orbitrap
MS Type:	ESI
MS Comments:	Source conditions were as follows: a spray voltage 4.0 kV in positive ion mode and 3.5 kV in negative ion mode, capillary temperature 290°C, S lens RF 50, and auxiliary gas heater temperature of 250°C. Nitrogen was used as both source and collision gas, with the sheath gas flow rate set at 20 and the auxiliary gas flow rate at 5 (arbitrary units). Data were acquired in full scan/data-dependent MS2 mode (full-scan resolution 70,000 FWHM, max ion injection time 50 ms, scan range m/z 200–1,500), with the 10 most abundant ions being subjected to collision-induced dissociation using an isolation window of 1.5 Da and a normalized stepped collision energy of 15/27 eV. Product ions were detected at a resolution of 17,500. An exclusion list for background ions was developed using extraction blanks, and mass calibration was performed in both positive and negative ionization modes before analysis to ensure mass accuracy of 5 ppm in full-scan mode.
Ion Mode:	NEGATIVE