Neoplastic and Stromal Cells Contribute to an Extracellular Matrix Gene Expression Profile Defining a Breast Cancer Subtype Likely to Progress

Abstract

We recently showed that differential expression of extracellular matrix (ECM) genes delineates four subgroups of breast carcinomas (ECM1, -2, -3- and -4) with different clinical outcome. To further investigate the characteristics of ECM signature and its impact on tumor progression, we conducted unsupervised clustering analyses in 6 additional independent datasets of invasive breast tumors from different platforms for a total of 643 samples. Use of four different clustering algorithms identified ECM3 tumors as an independent group in all datasets tested. ECM3 showed a homogeneous gene pattern, consisting of 58 genes encoding 43 structural ECM proteins. From 26 to 41% of the cases were ECM3-enriched, and analysis of datasets relevant to gene expression in neoplastic or corresponding stromal cells showed that both stromal and breast carcinoma cells can coordinately express ECM3 genes. In in vitro experiments, β-estradiol induced ECM3 gene production in ER-positive breast carcinoma cell lines, whereas TGFβ induced upregulation of the genes leading to ECM3 gene classification, especially in ER-negative breast carcinoma cells and in fibroblasts. Multivariate analysis of distant metastasis-free survival in untreated breast tumor patients revealed a significant interaction between ECM3 and histological grade (p = 0.001). Cox models, estimated separately in grade I–II and grade III tumors, indicated a highly significant association between ECM3 and worse survival probability only in grade III tumors (HR = 3.0, 95% CI = 1.3–7.0, p = 0.0098). Gene Set Enrichment analysis of ECM3 compared to non-ECM3 tumors revealed significant enrichment of epithelial-mesenchymal transition (EMT) genes in both grade I–II and grade III subsets of ECM3 tumors. Thus, ECM3 is a robust cluster that identifies breast carcinomas with EMT features but with accelerated metastatic potential only in the undifferentiated (grade III) phenotype. These findings support the key relevance of neoplastic and stroma interaction in breast cancer progression.

Extracellular matrix proteins as diagnostic markers of breast carcinoma

“…The expression profile of ECM‐related genes was studied in two publicly available datasets of BC and normal tissue samples (GSE5764) (Turashvili et al, ) and (GSE59595) (Huang et al, ). Class comparison analysis was performed using the ECM3 gene list (Triulzi et al, ) and the BrB‐ArrayTool developed by Dr. Richard Simon and the BRB‐ArrayTools Development Team (v4.2.0, http://linus.nci.nih.gov/BRB-ArrayTools.html). A cut‐off was set at p < 0.01 and FDR < 0.1 for identification of differentially expressed genes in normal versus tumor tissue.…”

Activated d16HER2 Homodimers and SRC Kinase Mediate Optimal Efficacy for Trastuzumab

“…Data were normalized to GAPDH (23). Relative abundance of d16HER2 mRNA compared with that of WTHER2 was calculated by the comparative C t method (24), with d16HER2 transcript levels indicated as the ratio 2…”

“…Pathways differentially enriched in activated d16HER2 tumors were evaluated by Gene Set Enrichment Analysis using GSEA v2.0.13 (26) on a 193-cancer-related gene set (24). Permutation type was applied 1,000 times.…”

Stromal Responses among Carcinomas—Letter

“…We found that ECM3, characterized by upmodulation of DTFassociated genes (6), has an independent negative prognostic role only in chemotherapy-untreated grade 3 breast carcinomas, whereas it is slightly associated with better outcome in differentiated tumors (5). Thus, even in absence of a significant association between DTF and specific tumor characteristics, this stromal signature may influence disease progression differently in the same tumor type according to characteristics of the neoplastic cells.…”