DNA/RNA-based classification of Bladder Cancer (BC) supports the existence of multiple molecular subtypes, while investigations at the protein level are scarce. Here, we aimed to investigate if Non-Muscle Invasive Bladder Cancer (NMIBC) can be stratified to biologically meaningful groups based on the proteome. Tissue specimens from 117 patients at primary diagnosis (98 with NMIBC and 19 with MIBC), were processed for high resolution proteomics analysis by liquid chromatography tandem mass spectrometry (LC-MS/MS). The proteomics output was subjected to unsupervised consensus clustering, principal component analysis (PCA), and investigation of subtype-specific features, pathways, and gene sets. NMIBC patients were optimally stratified to three NMIBC Proteomic Subtypes (NPS), differing in size, clinico-pathological and molecular backgrounds: NPS1 (mostly high stage/grade/risk samples) was the smallest in size (17/98) and overexpressed proteins reflective of an immune/inflammatory phenotype, involved in cell proliferation, unfolded protein response and DNA damage response, whereas NPS2 (mixed stage/grade/risk composition) presented with an infiltrated/mesenchymal profile. NPS3 was rich in luminal/differentiation markers, in line with its pathological composition (mostly low stage/grade/risk samples). PCA revealed a close proximity of NPS1 and conversely, remoteness of NPS3 to the proteome of MIBC. Proteins distinguishing these two extreme subtypes were also found to consistently differ at the mRNA levels between high and low risk subtypes of the UROMOL and LUND cohorts. Collectively, this study identifies three proteomic NMIBC subtypes and following a cross-omics validation in two independent cohorts, shortlists molecular features meriting further investigation for their biomarker or potentially therapeutic value. This article is protected by copyright. All rights reserved.

International journal of cancer. 2019 Jul 08 [Epub ahead of print]

Rafael Stroggilos, Marika Mokou, Agnieszka Latosinska, Manousos Makridakis, Vasiliki Lygirou, Emmanouil Mavrogeorgis, Dimitrios Drekolias, Maria Frantzi, William Mullen, Charalampos Fragkoulis, Konstantinos Stasinopoulos, Georgios Papadopoulos, Georgios Stathouros, Andreas C Lazaris, Periklis Makrythanasis, Konstantinos Ntoumas, Harald Mischak, Jerome Zoidakis, Antonia Vlahou

Biotechnology Division, Biomedical Research Foundation of the Academy of Athens, Athens, Greece., Mosaiques Diagnostics GmbH, Hannover, Germany., Department of Biotechnology, Agricultural University of Athens, Athens, Greece., Department of Pathology, School of Medicine, National and Kapodistrian University of Athens, Athens, Greece., British Heart Foundation Glasgow Cardiovascular Research Centre, University of Glasgow, Glasgow, United Kingdom., Department of Urology, General Hospital of Athens ‘Georgios Gennimatas’, Athens, Greece.