Comprehensive micro-scaled proteome and phosphoproteome characterization of archived retrospective cancer repositories

Corinna Friedrich, Simon Schallenberg, Marieluise Kirchner, Matthias Ziehm, Sylvia Niquet, Mohamed Haji, Christin Beier, Jens Neudecker, Frederick Klauschen () and Philipp Mertins ()
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Corinna Friedrich: German Cancer Consortium (DKTK), partner site Berlin
Simon Schallenberg: Institute of Pathology, Charité - Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health
Marieluise Kirchner: Max-Delbrück-Center for Molecular Medicine in the Helmholtz Association (MDC), Proteomics Platform
Matthias Ziehm: Max-Delbrück-Center for Molecular Medicine in the Helmholtz Association (MDC), Proteomics Platform
Sylvia Niquet: Max-Delbrück-Center for Molecular Medicine in the Helmholtz Association (MDC), Proteomics Platform
Mohamed Haji: Max-Delbrück-Center for Molecular Medicine in the Helmholtz Association (MDC), Proteomics Platform
Christin Beier: Max-Delbrück-Center for Molecular Medicine in the Helmholtz Association (MDC), Proteomics Platform
Jens Neudecker: Charité - Universitätsmedizin Berlin
Frederick Klauschen: German Cancer Consortium (DKTK), partner site Berlin
Philipp Mertins: German Cancer Consortium (DKTK), partner site Berlin

Nature Communications, 2021, vol. 12, issue 1, 1-15

Abstract: Abstract Formalin-fixed paraffin-embedded (FFPE) tissues are a valuable resource for retrospective clinical studies. Here, we evaluate the feasibility of (phospho-)proteomics on FFPE lung tissue regarding protein extraction, quantification, pre-analytics, and sample size. After comparing protein extraction protocols, we use the best-performing protocol for the acquisition of deep (phospho-)proteomes from lung squamous cell and adenocarcinoma with >8,000 quantified proteins and >14,000 phosphosites with a tandem mass tag (TMT) approach. With a microscaled approach, we quantify 7,000 phosphosites, enabling the analysis of FFPE biopsies with limited tissue amounts. We also investigate the influence of pre-analytical variables including fixation time and heat-assisted de-crosslinking on protein extraction efficiency and proteome coverage. Our improved workflows provide quantitative information on protein abundance and phosphosite regulation for the most relevant oncogenes, tumor suppressors, and signaling pathways in lung cancer. Finally, we present general guidelines to which methods are best suited for different applications, highlighting TMT methods for comprehensive (phospho-)proteome profiling for focused clinical studies and label-free methods for large cohorts.

Date: 2021
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Persistent link: https://EconPapers.repec.org/RePEc:nat:natcom:v:12:y:2021:i:1:d:10.1038_s41467-021-23855-w

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DOI: 10.1038/s41467-021-23855-w

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