Dataset Information

Glycoprotein Signatures for Aggressive Prostate Cancer via SWATH mass spectrometry

ABSTRACT: The identification of biomarkers indicating the level of aggressiveness of prostate cancer (PCa) addresses an urgent clinical need to minimize the general over-treatment of patients with non-aggressive PCa, which account for the majority of PCa cases. In this study we combined N-glycopeptide isolation and SWATH-MS towards the molecular characterization of tumor malignancy and aggressiveness. Here, we isolated formerly N-linked glycopeptides from normal prostate (n=10), non-aggressive (n=22), aggressive (n=16) and metastatic PCa (n=25) tumor tissues and analyzed the samples by SWATH-MS, an emerging data independent mass spectrometric acquisition method that generates a single MS file containing fragment ion spectra of all ionized species of a sample. The resulting datasets were searched using a targeted data analysis strategy where a priori spectral reference library representing known N-glycosites of the human proteome was used to identify groups of signals in the SWATH-MS data. On-average we identified 1430 N-glycosites from each SWATH map of which 1057 were quantified across all samples. The 220 glycoproteins that showed significant quantitative changes associated diverse biological processes with the level of PCa aggressiveness and metastasis and indicated functional relationships with common PCa genomic mutations.

INSTRUMENT(S): TripleTOF 5600

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Prostate Gland

DISEASE(S): Prostate Cancer

SUBMITTER: Yansheng Liu

LAB HEAD: Ruedi Aebersold

PROVIDER: PXD000704 | Pride | 2014-04-23

REPOSITORIES: Pride

ACCESS DATA

Dataset's files

Source:

			Action	DRS
	5600_interact.pep.xml	Pepxml
	5600_interact.prot.xml	Xml
	Orbitrap_interact.prot.xml	Xml
	Oribitrap_interact.pep.xml	Pepxml
	feature_alignment_openSWATHresultPX_yanliu_20140122.csv	Csv

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Publications

Glycoproteomic analysis of prostate cancer tissues by SWATH mass spectrometry discovers N-acylethanolamine acid amidase and protein tyrosine kinase 7 as signatures for tumor aggressiveness.

Liu Yansheng Y Chen Jing J Sethi Atul A Li Qing K QK Chen Lijun L Collins Ben B Gillet Ludovic C J LC Wollscheid Bernd B Zhang Hui H Aebersold Ruedi R

Molecular & cellular proteomics : MCP 20140416 7

The identification of biomarkers indicating the level of aggressiveness of prostate cancer (PCa) will address the urgent clinical need to minimize the general overtreatment of patients with non-aggressive PCa, who account for the majority of PCa cases. Here, we isolated formerly N-linked glycopeptides from normal prostate (n = 10) and from non-aggressive (n = 24), aggressive (n = 16), and metastatic (n = 25) PCa tumor tissues and analyzed the samples using SWATH mass spectrometry, an emerging da ...[more]

PMID: 24741114

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Project description:Glycobiology plays a central role in prostate cancer (PCa), as evidenced by the aberrant glycosylation in PCa-derived glycoproteins. Understanding the protein glycosylation changes underling PCa onset and progression may open opportunities for the discovery of novel biomarkers for PCa detection and stratification as well as targets for PCa metastasis. Advances in mass spectrometry-based glycomics and glycoproteomics have opened up exciting avenues for deep characterisation of the immensely, yet poorly understood complex glycoproteome. In this study, we have used integrated glycomics and glycoproteomics to explore the protein, cell and tumour-grade specific N- and O- glycosylation signatures in surgically-removed fresh PCa tissues grouped into five PCa disease grades and tissues from benign prostatic hyperplasia patients (BPH). Porous graphitised carbon–liquid chromatography (PGC–LC)-MS/MS analysis was used to profile the N-and O-glycome, revealing PCa grade specific N and O glycosylation changes, including oligomannosidic and paucimannosidic, bisecting-GlcNAc and highly branched tri- and tetra-antennary fucosylated and sialylated N-glycans as well as sialylated core 1 and 2 type O-glycans. High resolution LC-MS/MS was then employed to capture the micro and macroheterogeneity of 1,085 N-glycosites (>7,400 unique N-glycopeptides) from 540 N-glycoproteins and 360 O-glycosites (>500 unique O-glycopeptides) from 178 O-glycoproteins and reveal protein and cell specific N- and O- glycosylation changes associated with PCa progression. We also showed PCa grade-specific increase in the expression of oligosaccharyltransferase (OST) subunits that correlate with changes in the site occupancy of N-glycoproteins. In conclusion, this study shows that integrated glycomics and glycoproteomics can provide unprecedented insight into key molecular features and site-specific glycan heterogeneity contributing to the highly diverse tumour-microenvironment and allow us to deconstruct the regulation of the protein, cell and tumour-grade-specific glycosylation associated with PCa onset and development.

Project description:Background: Aldo-keto reductase (AKR) 1C family member 3 (AKR1C3), one of four identified human AKR enzymes, catalyzes steroid, prostaglandin, and xenobiotic metabolism. In the prostate, AKR1C3 is up-regulated in localized and advanced prostate adenocarcinoma, and is associated with prostate cancer (PCa) aggressiveness. Here we provide initial evidence for potential roles of AKR1C3 in PCa progression. Methods: Spatial distribution of AKR1C3 was analyzed using immunohistochemical staining in prostate adenocarcinoma tissue array. Human PCa PC-3 cells were stably transfected with AKR1C3 cDNA to establish PC3-AKR1C3 transfectants. Microarray and bioinformatics analyses were performed to identify pathways that are activated by elevated AKR1C3 expression in PCa cells. Functional confirmation of microarray and bioinformatics results was performed by immunoblot analysis and an in vitro Matrigel angiogenesis assay. Results: Elevated AKR1C3 expression was specifically limited to human prostate adenocarcinoma. Microarray and bioinformatics analysis suggested that elevated AKR1C3 expression in PC-3 cells modulates estradiol and androgen metabolism and activates insulin growth factor (IGF)-1 and Akt signaling pathways. Immunoblots confirmed that phosphorylated levels of IGF-1 receptor (IGF-1R) and Akt are significantly up-regulated in PC3-AKR1C3 as compared to mock transfectants. PC3-AKR1C3 transfectants promoted endothelial cell tube formation in Matrigel as compared to parental PC-3 cells and mock transfectants. Conclusion: Microarray and bioinformatics data followed by biological analyses suggest that elevated AKR1C3 expression in PC-3 cells promotes PCa angiogenesis and aggressiveness. These results suggest AKR1C3 can promote the aggressiveness of PCa through modulating estrogen and androgen metabolism with subsequent activation of growth factor IGF-1 and cytoplasmic Akt signaling pathways. Total RNA from mock- and ACR1C3 transfected PC-3 cells was isolated, with 2 or 3 biological replicates each. Gene expression data from AKR1C3 transfected PC-3 cells were compared with mock-transfected data.