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Unlocking the Proteome of Archival Tissue Collections: Proteomic Analysis of Urinary Bladder Cancer Tissues using Mass Spectrometry-based Proteomics
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. (Lind group)
2017 (English)Independent thesis Advanced level (degree of Master (Two Years)), 20 credits / 30 HE creditsStudent thesis
Abstract [en]

Formalin-fixed and paraffin-embedded (FFPE) and optimal cutting temperature (OCT)-embedded frozen tissue samples have recently gained interest as they exist in vast repositories worldwide and represent a valuable resource for clinical studies. However, unlocking the proteome of these archival tissues has encountered many challenges. Formalin has deleterious effects on protein structure which has been a barrier to use such a tissue material for global discovery and post-translational modification (PTM) analysis using mass spectrometry (MS). Furthermore, a cryopreservation OCT medium composed of polymers, such polyvinyl alcohol and polyethylene glycol, interferes with MS analysis due to ion suppression effects. Recent developments in sample preparation protocols and MS-based proteomics have enabled to investigate these archived samples and opened up the possibility to integrate them into biomarker discovery. In this present study, I demonstrate a streamlined, compatible, reproducible, and high-throughput proteomic workflow for subsequent MS-based ‘shotgun’ and PTM analysis of FFPE and OCT-embedded frozen tissues using nanoLC-MS/MS. For the first time, the comprehensive comparison of protein expression in paired FFPE and OCT-embedded frozen tissues of invasive and non-muscle invasive bladder cancer is presented. Label-free quantification revealed more than 2,000 and 3,000 protein identities in FFPE and OCT-embedded frozen bladder cancer tissues, respectively. In total, 24 and 29 proteins showed differential protein expression in FFPE and OCT/frozen when comparing invasive stage versus non-muscle invasive bladder cancer. Exploring molecular interactions demonstrated that regulated proteins play a crucial role in bladder cancer-associated pathways. These preliminary results, therefore, reveal the potential to apply our optimized proteomic workflow for biomarker and drug target discovery. Furthermore, I tested titanium dioxide phosphopeptide enrichment on paired FFPE and OCT-embedded frozen tissue sample. More than three hundred phosphorylation sites were identified in both types of samples. This proteomic platform could also be a promising tool for PTM analysis in archival tissue samples. 

Place, publisher, year, edition, pages
2017. , p. 77
Series
UPKEM E ; 31
National Category
Analytical Chemistry
Identifiers
URN: urn:nbn:se:uu:diva-351858OAI: oai:DiVA.org:uu-351858DiVA, id: diva2:1211451
Subject / course
Chemistry
Educational program
Master Programme in Chemistry
Supervisors
Examiners
Available from: 2018-05-31 Created: 2018-05-30 Last updated: 2018-05-31Bibliographically approved

Open Access in DiVA

The full text will be freely available from 2019-05-30 10:44
Available from 2019-05-30 10:44

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