Tumor angiogenesis and destruction of the extracellular matrix (ECM) are two essential factors for tumor invasion and metastasis. Previous studies concentrated on the tumor characteristics but there has been a lack of study of the tumor microenvironment. Genes altered in the tumor microenvironment may play important roles in the destruction process and be further involved in tumor progression.
To contribute to the understanding of tumor stroma roles in breast progression, we compared transcriptome profiles between cancer stroma (stroma surrounding invasive breast primary tumors) and normal breast stroma. A total of 3708 differentially expressed genes (DEGs) were identified in the GSE26910 dataset, whereas 665 DEGs were found in GSE10797. By overlapping the sets, 146 common DEGs were found.
Among the common DEGs, genes encoding matrix components were significantly dysregulated, including MMP1, POSTN, SPARC, COL1A1, COL5A1, COL5A2, COL10A1 and COL11A1. These altered genes were significantly associated with the pathways of “extracellular matrix organization”, “ECM-receptor interaction” and “focal adhesion”. Our findings indicated that these genes and pathways are related to the establishment of the tumor microenvironment.
The degradation of the basal membrane and the ECM results from proteolysis. Most members of the proteolytic system are mediated by AP-1 transcription factors through binding to TPA-responsive elements (TREs) in the promoter or enhancer region of the target genes . The AP-1 factor is a heterodimer consisted of proteins belonging to the Fos, Jun, ATF and JDP families . In our PPI network analysis of these common genes, JUN, FOS and ATF3 were recognized as key nodes. This suggests that they may play a critical role in tumor environment via AP-1 factor.
JUN/c-Jun is a member of the Jun family, FOS/c-Fos is a member of the Fos family and ATF3 belongs to the ATF family. AP-1 regulates several cytological processes, including differentiation, cell death, proliferation, oncogenic transformation, apoptosis and cell migration . Due to its constitutive activation and the diverse responses it induces, AP-1 plays a crucial role in promoting tumor invasion and migration .
In our study, JUN, FOS and ATF3 were involved in 12 of the top 50 significant biological processes. These included “negative regulation of cellular process” and “negative regulation of biological process”.
FOS and ATF3, STAT1, COL1A1 and FN1 were also identified as key nodes. STAT1 mediates both type I (alpha and beta) and type II (gamma) IFNs that are associated with cell growth, regulation, and antiviral and immune defense . Stromal STAT1 expression promotes tumor progression in breast cancer. COL1A1 and FN1 are individual ECM genes and have been reported to be associated with tumor invasion and metastasis. Increased extracellular levels of COL1A1 promote tumor cell invasiveness in culture and metastasis in animal models . Furthermore, a high level of COL1A1 increases the likelihood of clinical metastasis of multiple human solid tumors [42, 43]. The elevated expression of FN1 promotes lymph node metastasis in human oral squamous cell carcinoma by promoting vascular endothelial growth factor-C expression and the epithelial–mesenchymal transition (EMT) . However, no study on COL1A1 and FN1 associations with breast cancer has been reported. Therefore, we performed Oncomine analysis to investigate their expression in breast cancer. High expression levels of COL1A1 and FN1 were associated with high grade or advanced stage of breast cancer and poor prognosis.
In summary, breast cancer stroma-related genes, including JUN, FOS, ATF3, STAT1, COL1A1 and FN1, were all associated with tumor invasion and metastasis. This reveals that cancer stroma-related genes have important roles in tumor progression.
Despite the improvement of tumor detection tools (including imaging techniques like MRI and PET; and analysis of blood biomarkers shed by the tumor, such as proteins and cell free nucleic acids) and tumor therapies, there are still limitations. Clinical detection of tumors is limited to masses 1 cm in diameter. Resistance to therapy has been one of the major challenges in treating cancer .
Tumor microenvironment has been considered as prospective breakthrough point in early tumor detection and tumor treatment recently. Based on the two hallmarks of the tumor microenvironment, acidity and low oxygenation, the macromolecular near-infrared poly (ethylene glycol)-conjugatediridium (iii) complex is designed to successively respond to acidity and hypoxia while amplifying detection sensitivity via signal propagation. Primary tumors and metastasis tumor nodules as small as 1 mm in mice have been successfully detected . In addition, tumor microenvironment biomarkers contribute to tumor detection and treatment. MMP-9, a major regulator inducing ECM component degradation, is highly expressed in many human cancer types compared with normal controls. Therefore, an exogenously administered tumor-penetrating nanosensor with an MMP-9 peptide substrate in its surface has been created . Importantly, this nanosensor is reported to detect nodules with median diameters smaller than 2 mm in an orthotopic model of ovarian cancer. In terms of drug resistance, different components of the microenvironment have been found to participate in the development of chemoresistance and inhibit MMPs, although chemokine signaling is effective when combined with traditional chemotherapies .
In this study, COL1A1 and FN1 were all ECM genes with elevated mRNA levels in tumor stromal cells. Therefore, they might be used as key candidate genes for tumor detection and treatment.
The strength of our study is that we combined gene expression data of stroma surrounding invasive breast primary tumors and normal stroma from two GEO databases and obtained conserved genes. It is known that as breast tumors progress from ductal carcinoma in situ (DCIS) to invasive ductal carcinoma (IDC), the ECM undergoes increased collagen fiber linearization and thickening due to deposition and cross-linking of the collagen. In IDC, the collagen fibers are aligned perpendicularly to the tumor boundary, forming migration tracks for invasive tumor cells to exit the tumor tissue and enter the bloodstream . Thus, the alteration of stroma composition from DCIS to IDC is important for tumor invasion. Further study may focus on the gene expression of cancer stroma in this process.
Our study also has several limitations. First, the analysis of more similar or larger databases may be necessary for identification of the conservative key genes. Second, experimental validation of the expressions of these conserved genes is needed. Third, investigation the roles of these critical genes should be conducted in the future.