6280

S100A9

S100 calcium binding protein A9;S100A9;S100 calcium binding protein A9

MRP-14|S100 calcium-binding protein A9 (calgranulin B)|calgranulin B|calgranulin-B|calprotectin L1H subunit|leukocyte L1 complex heavy chain|migration inhibitory factor-related protein 14|protein S100-A9

1q21

protein-coding

The goal of the present study is to unveil the gene expression profile specific to the biological processes of human breast epithelial cell invasion and migration using an MCF10A model genetically engineered to constitutively activate the H-ras or N-ras signaling pathway. We previously showed that H-Ras, but not N-Ras, induces MCF10A cell invasion/migration, whereas both H-Ras and N-Ras induce cell proliferation and phenotypic transformation. Thus, these cell lines provide an experimental system to separate the gene expression profile associated with cell invasion apart from cell proliferation/transformation. Analysis of whole human genome microarray revealed that 412 genes were differentially expressed among MCF10A, N-Ras MCF10A, and H-Ras MCF10A cells and hierarchical clustering separated 412 genes into four clusters. We then tested whether S100A8 and S100A9, two of the genes which are most highly up-regulated in an H-Ras-specific manner, play a causative role for H-Ras-mediated MCF10A cell invasion and migration. Importantly, small interfering RNA-mediated knockdown of S100A8/A9 expression significantly reduced H-Ras-induced invasion/migration. Conversely, the induction of S100A8/A9 expression conferred the invasive/migratory phenotype to parental MCF10A cells. Furthermore, we provided evidence of signaling cross-talk between S100A8/A9 and the mitogen-activated protein kinase signaling pathways essential for H-Ras-mediated cell invasion and migration. Taken together, this study revealed S100A8/A9 genes as candidate markers for metastatic potential of breast epithelial cells. Our gene profile data provide useful information which may lead to the identification of additional potential targets for the prognosis and/or therapy of metastatic breast cancer.

S100A8/A9 proteins are members of EF-hand calcium-binding proteins secreted by neutrophils and activated monocytes. S100A8/A9 has cell growth-promoting activity at low concentrations by binding to the receptor for advanced glycation end products (RAGE). In this study, we report for the first time that S100A8/A9 promoted the invasion of breast cancer cells depending on RAGE. In addition, RAGE binding to S100A8/A9 promoted the phosphorylation of LIN-11, Isl1, and MEC-3 protein domain kinase, as well as cofilin. This phosphorylation is a critical step in cofilin recycling and actin polymerization. Interestingly, RAGE binding to S100A8/A9 enhanced cell mesenchymal properties and induced epithelial-mesenchymal transition. Mechanistically, RAGE binding to S100A8/A9 stabilized Snail through the NF-kappaB signaling pathway. Based on these observations, RAGE expression in breast cancer cells was associated with lymph node and distant metastases in patients with invasive ductal carcinoma. Moreover, RAGE binding to S100A8/A9 promoted lung metastasis in vivo. In summary, our in vitro and in vivo results indicated that RAGE binding to S100A8/A9 played an important role in breast cancer invasion/metastasis. This study identified both RAGE and S100A8/A9 as potential anti-invasion targets for therapeutic intervention in breast cancer.

To better understand the mechanism underlying colorectal carcinoma (CRC) genesis or metastasis, and to search for potential markers for CRC prognosis, a comparative proteomic analysis was performed on CRC tissue. Proteins were extracted from normal colorectal mucosa, non-metastatic CRC (nmCRC) and metastatic CRC (mCRC) tissue samples. Protein profiling of each sample was performed by two-dimensional electrophoresis coupled with MALDI-TOF MS, followed by confirmation by Western blotting. Thirty-one proteins were found to be differentially expressed between normal mucosa, nmCRC and mCRC tissue. In 126 paraffin-embedded CRC samples, three differentially expressed proteins, identified as LASP-1, S100A9 and RhoGDI by proteomic analysis, were detected by immunohistochemical staining to determine the clinicopathological characteristics of these proteins in CRC. Increased expression levels of these proteins were found in CRC, especially mCRC, compared with normal mucosa. The results provide the basis for searching for potential markers for CRC genesis and metastasis, and also provide clues for elucidating the mechanism of CRC progression. The pattern changes identified have the potential to be used for the design of marker panels for assistance in diagnostic and therapeutic strategies in CRC.

The tumor microenvironment plays an important role in modulating tumor progression. Earlier, we showed that S100A8/A9 proteins secreted by myeloid-derived suppressor cells (MDSC) present within tumors and metastatic sites promote an autocrine pathway for accumulation of MDSC. In a mouse model of colitis-associated colon cancer, we also showed that S100A8/A9-positive cells accumulate in all regions of dysplasia and adenoma. Here we present evidence that S100A8/A9 interact with RAGE and carboxylated glycans on colon tumor cells and promote activation of MAPK and NF-kappaB signaling pathways. Comparison of gene expression profiles of S100A8/A9-activated colon tumor cells versus unactivated cells led us to identify a small cohort of genes upregulated in activated cells, including Cxcl1, Ccl5 and Ccl7, Slc39a10, Lcn2, Zc3h12a, Enpp2, and other genes, whose products promote leukocyte recruitment, angiogenesis, tumor migration, wound healing, and formation of premetastatic niches in distal metastatic organs. Consistent with this observation, in murine colon tumor models we found that chemokines were upregulated in tumors, and elevated in sera of tumor-bearing wild-type mice. Mice lacking S100A9 showed significantly reduced tumor incidence, growth and metastasis, reduced chemokine levels, and reduced infiltration of CD11b(+)Gr1(+) cells within tumors and premetastatic organs. Studies using bone marrow chimeric mice revealed that S100A8/A9 expression on myeloid cells is essential for development of colon tumors. Our results thus reveal a novel role for myeloid-derived S100A8/A9 in activating specific downstream genes associated with tumorigenesis and in promoting tumor growth and metastasis.

With increasing therapeutic use of minimally invasive therapy for treatment of early gastric cancer, prediction of lymph node metastasis is important. In search of tissue biomarkers for prediction of lymph node metastasis of gastric adenocarcinoma, we analyzed gastric adenocarcinoma tissue using proteomic methods. We have done 2D-PAGE and MALDI-TOF MS analysis in matched normal and gastric cancer tissues. We also evaluated the clinicopathological significance of expression of S100A8 and S100A9 in gastric adenocarcinoma using a tissue microarray of 218 gastric adenocarcinoma specimens. Cell invasion and migration assay were performed to confirm functional role of S100A8 and S100A9 using small hairpin RNA lentivirus. We identified 8 up-regulated and 5 down-regulated proteins in gastric cancer tissues compared to matched normal mucosa. Of these, expression of S100A8 and S100A9 occurred mainly in stromal cells and inflammatory cells between tumor cells. Correlation was observed between small lesion size, decreased depth of invasion, a tendency to absence of lymphovascular tumor emboli, a decrease in perineural invasion and lymph node metastasis, and expression of stromal S100A8. In addition, increased expression of stromal S100A9 in gastric adenocarcinoma was associated with small lesion size and a decrease in lymph node metastasis. Functional analysis confirmed that down-regulation of S100A8 and S100A9 by small hairpin RNA lentivirus induced an increase of migration and invasion in gastric cancer cell lines. Taken together, these findings suggest that S100A8 and S100A9 are negative regulators of lymph node metastasis of gastric adenocarcinoma and can be used as biomarkers for prediction of lymph node metastasis in gastric adenocarcinoma.