Extracellular matrix signature in grade III breast carcinomas is associated with a distinct proteomic profile and biomechanical characteristics consistent with their poor prognosis

Our recent analysis of extracellular matrix (ECM) gene expression profiles in breast carcinomas identified an expression pattern (ECM3) in ~40% of breast carcinomas that defines an independent group of tumors. The ECM3 pattern has prognostic significance in relation to tumor differentiation status, stratifying a subgroup with poor prognosis only within the most undifferentiated grade III tumors. Conversely, distant metastasis-free survival of non-ECM3-grade III patients was similar to that of the most differentiated grade I-II patients regardless of ECM3 features (Triulzi et al. PlosOne 2013). Because gene expression analyses cannot be assumed to precisely reflect protein expression, to investigate mechanisms underlying the interaction between ECM3 and tumor differentiation status, we performed an in-gel-based proteomic analysis and a mechanical characterization of the elastic modulus of ECM3 and non-ECM3 breast cancers according to tumor grade (III vs. I-II). In-gel proteomic analysis of frozen tumor samples showed 198 spots differentially expressed (p<0.01 one-way Anova) among the four groups of breast carcinoma. Principal component analysis (PCA) revealed significant differences between ECM3 and non-ECM3 within grade III tumors, whereas grade I-II tumors did not show major variations according to ECM classification. Hierarchical Pearson correlation analysis of grade III tumor samples showed that ECM3 tumors overexpressed ECM, cytoskeletal and secreted proteins, whereas chaperone and stress response proteins, cell signaling molecules, and metabolic proteins involved in the glycolytic pathway and TCA cycle were down-modulated. Concerning the mechanical properties of tumor samples, compression tests of the elastic modulus revealed a major stiffness of ECM3-grade III tumors compared to all other groups, with an average (± SD) elastic modulus of 761 ± 113 kPa in ECM3-grade III vs. 378 ± 34.9, 345 ± 34.8 and 480 ± 47.6 in non-ECM3-grade III, ECM3-grades I-II and non-ECM3-grades I-II, respectively. Our results indicate that ECM3-grade III tumors have a peculiar proteomic profile and mechanical characteristics compared to all other breast cancers, underscoring the biological role of the interaction between tumor cell phenotype and ECM features in guiding tumor progression. Molecules and mechanisms involved in this interaction await further elucidation to provide potential therapeutic targets of cancer progression.