title: Combined Proteomics and Transcriptomics Identifies Carboxypeptidase B1 and Nuclear Factor κB (NF-κB) Associated Proteins as Putative Biomarkers of Metastasis in Low Grade Breast Cancer.
abstract: Current prognostic factors are insufficient for precise risk-discrimination in breast cancer patients with low grade breast tumors, which, in disagreement with theoretical prognosis, occasionally form early lymph node metastasis. To identify markers for this group of patients, we employed iTRAQ-2DLC-MS/MS proteomics to 24 lymph node positive and 24 lymph node negative grade 1 luminal A primary breast tumors. Another group of 48 high-grade tumors (luminal B, triple negative, Her-2 subtypes) was also analyzed to investigate marker specificity for grade 1 luminal A tumors. From the total of 4405 proteins identified (FDR < 5%), the top 65 differentially expressed together with 30 previously identified and control markers were analyzed also at transcript level. Increased levels of carboxypeptidase B1 (CPB1), PDZ and LIM domain protein 2 (PDLIM2), and ring finger protein 25 (RNF25) were associated specifically with lymph node positive grade 1 tumors, whereas stathmin 1 (STMN1) and thymosin beta 10 (TMSB10) associated with aggressive tumor phenotype also in high grade tumors at both protein and transcript level. For CPB1, these differences were also observed by immunohistochemical analysis on tissue microarrays. Up-regulation of putative biomarkers in lymph node positive (versus negative) luminal A tumors was validated by gene expression analysis of an independent published data set (n = 343) for CPB1 (p = 0.00155), PDLIM2 (p = 0.02027) and RELA (p = 0.00015). Moreover, statistically significant connections with patient survival were identified in another public data set (n = 1678). Our findings indicate unique pro-metastatic mechanisms in grade 1 tumors that can include up-regulation of CPB1, activation of NF-κB pathway and changes in cell survival and cytoskeleton. These putative biomarkers have potential to identify the specific minor subpopulation of breast cancer patients with low grade tumors who are at higher than expected risk of recurrence and who would benefit from more intensive follow-up and may require more personalized therapy.
authors.affiliation: From the ‡Masaryk Memorial Cancer Institute, Regional Centre for Applied Molecular Oncology, Brno, Czech Republic; §Masaryk University, Faculty of Science, Department of Biochemistry, Brno, Czech Republic;
authors.forename: Pavel
authors.initials: P
authors.lastname: Bouchal
authors.affiliation: From the ‡Masaryk Memorial Cancer Institute, Regional Centre for Applied Molecular Oncology, Brno, Czech Republic; §Masaryk University, Faculty of Science, Department of Biochemistry, Brno, Czech Republic;
authors.forename: Monika
authors.initials: M
authors.lastname: Dvořáková
authors.affiliation: ¶Proteomics Mass Spectrometry, The Wellcome Trust Sanger Institute, Cambridge CB10 1SA, UK;
authors.forename: Theodoros
authors.initials: T
authors.lastname: Roumeliotis
authors.affiliation: ‖Masaryk University, Faculty of Medicine, Institute of Biostatistics and Analyses, Brno, Czech Republic;
authors.forename: Zbyněk
authors.initials: Z
authors.lastname: Bortlíček
authors.affiliation: ‖Masaryk University, Faculty of Medicine, Institute of Biostatistics and Analyses, Brno, Czech Republic;
authors.forename: Ivana
authors.initials: I
authors.lastname: Ihnatová
authors.affiliation: From the ‡Masaryk Memorial Cancer Institute, Regional Centre for Applied Molecular Oncology, Brno, Czech Republic;
authors.forename: Iva
authors.initials: I
authors.lastname: Procházková
authors.affiliation: **ThermoFisher Scientific, Hemel Hempstead, UK;
authors.forename: Jenny T C
authors.initials: JT
authors.lastname: Ho
authors.affiliation: From the ‡Masaryk Memorial Cancer Institute, Regional Centre for Applied Molecular Oncology, Brno, Czech Republic; §Masaryk University, Faculty of Science, Department of Biochemistry, Brno, Czech Republic;
authors.forename: Josef
authors.initials: J
authors.lastname: Maryáš
authors.affiliation: ‡‡Laboratory of Computational Biology, Center for Human Genetics, University of Leuven, Belgium;
authors.forename: Hana
authors.initials: H
authors.lastname: Imrichová
authors.affiliation: From the ‡Masaryk Memorial Cancer Institute, Regional Centre for Applied Molecular Oncology, Brno, Czech Republic; ‖Masaryk University, Faculty of Medicine, Institute of Biostatistics and Analyses, Brno, Czech Republic;
authors.forename: Eva
authors.initials: E
authors.lastname: Budinská
authors.affiliation: From the ‡Masaryk Memorial Cancer Institute, Regional Centre for Applied Molecular Oncology, Brno, Czech Republic;
authors.forename: Rostislav
authors.initials: R
authors.lastname: Vyzula
authors.affiliation: §§University of Southampton, School of Medicine, Cancer Sciences Division, Institute for Life Sciences-Center for Proteomic Research, Southampton, UK.
authors.forename: Spiros D
authors.initials: SD
authors.lastname: Garbis
authors.affiliation: From the ‡Masaryk Memorial Cancer Institute, Regional Centre for Applied Molecular Oncology, Brno, Czech Republic;
authors.forename: Bořivoj
authors.initials: B
authors.lastname: Vojtěšek
authors.affiliation: From the ‡Masaryk Memorial Cancer Institute, Regional Centre for Applied Molecular Oncology, Brno, Czech Republic; nenutil@mou.cz.
authors.forename: Rudolf
authors.initials: R
authors.lastname: Nenutil
mesh_terms: D014408:Biomarkers, Tumor / Q000235:genetics / Q000378:metabolism*; D001943:Breast Neoplasms / Q000235:genetics* / Q000378:metabolism*; D043424:Carboxypeptidase B / Q000378:metabolism*; D030562:Databases, Protein; D005260:Female; D020869:Gene Expression Profiling / Q000379:methods*; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007150:Immunohistochemistry; D007553:Isotope Labeling; D053208:Kaplan-Meier Estimate; D016328:NF-kappa B / Q000378:metabolism*; D060787:Neoplasm Grading; D009362:Neoplasm Metastasis; D040901:Proteomics / Q000379:methods*; D015203:Reproducibility of Results
chemical_list: D014408:Biomarkers, Tumor; C058173:CPB1 protein, human; D016328:NF-kappa B; D043424:Carboxypeptidase B