Gene expression profiling of tumour epithelial and stromal compartments during breast cancer progression.
ABSTRACT The progression of ductal carcinoma in situ (DCIS) to invasive ductal carcinoma (IDC) marks a critical step in the evolution of breast cancer. There is some evidence to suggest that dynamic interactions between the neoplastic cells and the tumour microenvironment play an important role. Using the whole-genome cDNA-mediated annealing, selection, extension and ligation assay (WG-DASL, Illumina), we performed gene expression profiling on 87 formalin-fixed paraffin-embedded (FFPE) samples from 17 patients consisting of matched IDC, DCIS and three types of stroma: IDC-S (<3 mm from IDC), DCIS-S (<3 mm from DCIS) and breast cancer associated-normal stroma (BC-NS; >10 mm from IDC or DCIS). Differential gene expression analysis was validated by quantitative real time-PCR, immunohistochemistry and immunofluorescence. The expression of several genes was down-regulated in stroma from cancer patients relative to normal stroma from reduction mammoplasties. In contrast, neoplastic epithelium underwent more gene expression changes during progression, including down regulation of SFRP1. In particular, we observed that molecules related to extracellular matrix (ECM) remodelling (e.g. COL11A1, COL5A2 and MMP13) were differentially expressed between DCIS and IDC. COL11A1 was overexpressed in IDC relative to DCIS and was expressed by both the epithelial and stromal compartments but was enriched in invading neoplastic epithelial cells. The contributions of both the epithelial and stromal compartments to the clinically important scenario of progression from DCIS to IDC. Gene expression profiles, we identified differential expression of genes related to ECM remodelling, and specifically the elevated expression of genes such as COL11A1, COL5A2 and MMP13 in epithelial cells of IDC. We propose that these expression changes could be involved in facilitating the transition from in situ disease to invasive cancer and may thus mark a critical point in disease development.
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ABSTRACT: Although the search for disease biomarkers continues, the clinical return has thus far been disappointing. The complexity of the body's response to disease makes it difficult to represent this response with only a few biomarkers, particularly when many are present at low levels. An alternative to the typical reductionist biomarker paradigm is an assay we call an "immunosignature." This approach leverages the response of antibodies to disease-related changes, as well as the inherent signal amplification associated with antigen-stimulated B-cell proliferation. To perform an immunosignature assay, the antibodies in diluted blood are incubated with a microarray of thousands of random sequence peptides. The pattern of binding to these peptides is the immunosignature. Because the peptide sequences are completely random, the assay is effectively disease-agnostic, potentially providing a comprehensive diagnostic on multiple diseases simultaneously. To explore the ability of an immunosignature to detect and identify multiple diseases simultaneously, 20 samples from each of five cancer cohorts collected from multiple sites and 20 noncancer samples (120 total) were used as a training set to develop a reference immunosignature. A blinded evaluation of 120 blinded samples covering the same diseases gave 95% classification accuracy. To investigate the breadth of the approach and test sensitivity to biological diversity further, immunosignatures of >1,500 historical samples comprising 14 different diseases were examined by training with 75% of the samples and testing the remaining 25%. The average accuracy was >98%. These results demonstrate the potential power of the immunosignature approach in the accurate, simultaneous classification of disease.Proceedings of the National Academy of Sciences of the United States of America. 07/2014;
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ABSTRACT: Background: The human COL11A1 gene has been shown to be up-regulated in stromal cells of colorectal tumours, but, so far, the immunodetection of procollagen 11A1, the primary protein product of COL11A1, has not been studied in detail in human colon adenocarcinomas. Some cancer-associated stromal cells seem to be derived from bone marrow mesenchymal cells; the expression of the COL11A1 gene and the parallel immunodetection of procollagen 11A1 have not been evaluated in these latter cells, either. Methods: We used quantitative RT-PCR and/or immunocytochemistry to study the expression of DES/desmin, VIM/vimentin, ACTA2/alphaSMA (alpha smooth muscle actin) and COL11A1/procollagen 11A1 in HCT 116 human colorectal adenocarcinoma cells, in immortalised human bone marrow mesenchymal cells and in human colon adenocarcinoma-derived cultured stromal cells. The immunodetection of procollagen 11A1 was performed with the new recently described DMTX1/1E8.33 mouse monoclonal antibody. Human colon adenocarcinomas and non-malignant colon tissues were evaluated by immunohistochemistry as well. Statistical associations were sought between anti-procollagen 11A1 immunoscoring and patient clinicopathological features. Results: Procollagen 11A1 was immunodetected in human bone marrow mesenchymal cells and in human colon adenocarcinoma-associated spindle-shaped stromal cells but not in colon epithelial or stromal cells of the normal colon. This immunodetection paralleled, in both kinds of cells, that of the other mesenchymal-related biomarkers studied: vimentin and alpha smooth muscle actin, but not desmin. Thus, procollagen 11A1+ adenocarcinoma-associated stromal cells are similar to "activated myofibroblasts". In the series of human colon adenocarcinomas here studied, a high procollagen 11A1 expression was associated with nodal involvement (p = 0.05), the development of distant metastases (p = 0.017), and advanced Dukes stages (p = 0.047). Conclusion: The immunodetection of procollagen 11A1 in cancer-associated stromal cells could be a useful biomarker for human colon adenocarcinoma characterisation.BMC Cancer 11/2014; 14(1):867. · 3.33 Impact Factor
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ABSTRACT: Accurate diagnosis of invasive breast lesions, when analyzed by Core Needle Biopsy, may suppose a major challenge for the pathologist. Various markers of invasiveness such as laminin, S-100 protein, P63 or calponin have been described, however none of them is completely reliable. The use of a specific marker of the infiltrating tumor microenvironment seems vital to support the diagnosis of invasive against in situ lesions. At this point, Collagen, type XI, alpha 1 (COL11A1), might be helpful since it has been described associated to cancer associated fibroblasts in other tumors such as lung, pancreas or colorectal. This paper aims to analyze the role of COL11A1 as a marker of invasiveness in breast tumor lesions. Two hundred and one breast Core Needle Biopsy samples were analyzed by immunohistochemistry against pro-COL11A1. The results show a significant difference (p <0.0001) when comparing the expression in infiltrative tumors (93%) versus immunostaining of non-invasive lesions (4%). Forty cases of underestimated DCIS were also stained for COL11A1 presenting a sensitivity of 90% when compared with p63 and calponin which not tagged invasion. In conclusion pro-COL11A1 expression is a promising marker of invasive breast lesions, and may be included in immunohistochemical panels aiming at identifying infiltration in problematic breast lesions.Pathology - Research and Practice 01/2014; · 1.21 Impact Factor