Nasopharyngeal carcinoma (NPC) is known for its high-metastatic potential. Here we report the identification of the proteoglycan serglycin as a functionally significant regulator of metastasis in this setting. Comparative genomic expression profiling of NPC cell line clones with high-and low-metastatic potential revealed the serglycin gene (SRGN) as one of the most upregulated genes in highly metastatic cells. RNAi-mediated inhibition of serglycin expression blocked serglycin secretion and the invasive motility of highly metastatic cells, reducing metastatic capacity in vivo. Conversely, serglycin overexpression in poorly metastatic cells increased their motile behavior and metastatic capacity in vivo. Growth rate was not influenced by serglycin in either highly or poorly metastatic cells. Secreted but not bacterial recombinant serglycin promoted motile behavior, suggesting a critical role for glycosylation in serglycin activity. Serglycin inhibition was associated with reduced expression of vimentin but not other epithelial-mesenchymal transition proteins. In clinical specimens, serglycin expression was elevated significantly in liver metastases from NPC relative to primary NPC tumors. We evaluated the prognostic value of serglycin by immunohistochemical staining of tissue microarrays from 263 NPC patients followed by multivariate analyses. High serglycin expression in primary NPC was found to be an unfavorable independent indicator of distant metastasis-free and disease-free survival. Our findings establish that glycosylated serglycin regulates NPC metastasis via autocrine and paracrine routes, and that it serves as an independent prognostic indicator of metastasis-free survival and disease-free survival in NPC patients. Cancer Res; 71(8); 3162-72. Ó2011 AACR.

Nasopharyngeal carcinoma (NPC) has the highest metastatic potential among head and neck cancers. Distant metastasis is the major cause of treatment failure. The role of interleukin-8 (IL-8) in NPC progression remains unknown. Our multivariate survival analyses of 255 patients with NPC revealed that higher IL-8 expression in primary NPC tissue was an independent prognostic factor for overall survival, disease-free survival, and distant metastasis-free survival of the patients. In vitro study revealed that IL-8 was highly expressed in the established high-metastasis NPC clone S18 relative to the low-metastasis cells. Suppression of IL-8 by short-hairpin RNA reduced the expression of IL-8 in S18 cells and subsequently inhibited migration, invasion, and hepatic metastasis of the cells without influencing cellular growth. Overexpression of IL-8 in S26 cells resulted in increased migration, invasion, and metastasis capabilities of the cells without affecting cellular growth. Exogenous IL-8 enhanced the migration and invasion of low-metastasis CNE-2 cells in a dose-dependent manner. An epithelial–mesenchymal transition (EMT) could be induced by IL-8 in various NPC cell lines. The high level of phosphorylated AKT in S18 cells could be suppressed by knocking down IL-8 expression. Further, IL-8-promoted migration and invasion could be abolished by either the application of the phosphoinositide-3-kinase inhibitor LY294002 or the knock down of AKT expression by using small-interfering RNA. In summary, IL-8 serves as an independent prognostic indicator of overall survival, disease-free survival, and metastasis-free survival for patients with NPC. IL-8 promotes NPC metastasis via autocrine and paracrine means, involving activation of AKT signaling and inducing EMT in NPC cells.

Nasopharyngeal carcinoma (NPC) has the highest metastasis rate among head and neck cancers with unclear mechanism. WNT5A belongs to the WNT family of cysteine-rich secreted glycoproteins. Our previous high-throughput gene expression profiling revealed that WNT5A was up-regulated in highly metastatic cells. In the present study, we first confirmed the elevated expression of WNT5A in metastatic NPC tissues at both the mRNA and protein levels. We then found that WNT5A promoted epithelial-mesenchymal transition (EMT) in NPC cells, induced the accumulation of CD24-CD44+ cells and side population, which are believed to be cancer stem cell characteristics. Moreover, WNT5A promoted the migration and invasion of NPC cells in vitro, while in vivo treatment with recombinant WNT5A promoted lung metastasis. Knocking down WNT5A diminished NPC tumorigenesis in vivo. When elevated expression of WNT5A coincided with the elevated expression of vimentin in the primary NPC, the patients had a poorer prognosis. Among major signaling pathways, protein kinase C (PKC) signaling was activated by WNT5A in NPC cells. A positive feedback loop between WNT5A and phospho-PKC to promote EMT was also revealed. Taken together, these data suggest that WNT5A is an important molecule in promoting stem cell characteristics in NPC, leading to tumorigenesis and metastasis.

Nasopharyngeal carcinoma (NPC) is one of the most common malignancies in southern China and Southeast Asia, with the highest metastasis rate among head and neck cancers. The mechanisms underlying NPC progression remain poorly understood. Genome-wide expression profiling on 18 NPC vs. 18 noncancerous nasopharyngeal tissues together with GeneGo pathway analysis and expression verification in NPC cells and tissues revealed a potential role of urokinase-type plasminogen activator receptor (uPAR) in NPC progression, which has not been investigated in NPC. We then observed that uPAR expression is increased in poorly differentiated, highly metastatic NPC cells compared with lowly metastatic cells or differentiated NPC cells. In vitro studies demonstrated that uPAR regulates NPC cell growth, colony formation, migration, and invasion and promotes the epithelial-mesenchymal transition (EMT). Additional tumor xenograft and spontaneous metastasis experiments revealed that uPAR promotes NPC cell growth and metastasis in vivo. The JAK-STAT pathway is involved in uPAR-regulated signaling in NPC cells as determined by immunoblotting. Moreover, uPAR-mediated growth and motility is partially abolished upon treatment with the Jak1/Jak2 inhibitor INCB018424. We suppressed uPA expression in uPAR-overexpressing NPC cells and found that uPAR-mediated cellular growth and motility is not exclusively dependent on uPA. In summary, uPAR is a significant regulator of NPC progression and could serve as a promising therapeutic target.

Objectives: To investigate the expression of vimentin and Ki-67 proteins in cervical squamous cell carcinoma (CSCC) and their relationships with patient clinicopathological features. Materials and Methods:Fifty-seven CSCC samples archived in Department of Pathology in the First Affiliated Hospital of Wenzhou Medical University were selected. The expression of vimentin and Ki-67 proteins in CSCC tissue were detected using immunohistochemical SP method, and correlations between them and their relationships with clinicopathological features were analyzed. Results: Among 57 CSCC tissues, there were 43 with positive expression of Vimentin, and the positive rate was 75.4%; there were 57 cases with positive expression of Ki-67, and the positive rate came up to 100.0%. The results of Pearson correlation analysis displayed that the expression of vimentin had a significantly-positive correlation with Ki-67 in CSCC tissue (r=0.984, co0.000). The expression of both Ki-67 and vimentin was intimately associated with the presence or absence of local invasion and lymph node metastasis as well as differentiated degrees of the tumor (P=0.003, 0.017, 0.000; P=0.001, 0.008, 0.003) instead of the age, tumor size and clinical staging (P>0.05). Conclusions: Epithelial-mesenchymal transition (EMT) tends to appear in poorly-differentiated CSCC tissue, and the up-regulation of vimentin expression is accompanied by high expression of Ki-67, suggesting that invasion and metastasis readily occur in these tumor cells.

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