Expression Of NANOG And SOX2in HCC Patients And Their Mechanism Of Regulating HCC Cell Invasion

Hepatocellular carcinoma (HCC) is the third leading cause of cancer related death in the world. Although surgical resection improves survival, the prognosis of HCC remains unsatisfactory mainly because of frequent intrahepatic spread and extrahepatic metastasis. Consequently, elucidation of the molecular mechanism underlying tumor invasion is crucial to improve the outcome of HCC therapy. Recent publications showed that NANOG and SOX2participated in oncogenesis and tumor progression of various kinds of cancers. In this study, NANOG and SOX2expression of HCC cell lines and tumor tissues was analyzed and further analysis of the roles of NANOG and SOX2in HCC metastasis was carried out through in vivo and in vitro experiments.Part OneThe mechanism of NANOG regulating HCC cell invasion NANOG is a major transcription factors essential to the stem cell self-renewal and are associated with various kinds of cancers. In this part of study, we determined the expression of NANOG in HCC tumor tissues by immunohistochemistry and investigated its correlation with clinicopathologic characteristics of these patients. The results showed that NANOG expression was elevated in patients with metastasis. Kaplan-Meier survival analysis also indicated that NANOG expression was correlated with survival time of HCC patients. However, the NANOG signaling in HCC cell invasion was still elusive. In this part, we investigated its underlying mechanism in the metastasis of HCC. Our data showed that HCC cells overexpressing NANOG were characterized by active epithelial-mesenchymal transition (EMT), and exhibited increased ability of matrigel invasion, soft agar colonization, sphere formation and drug resistance, whereas SB-431542, an antagonist of activin receptor-like kinase (ALK) receptors, attenuated EMT and invasion of HCC cells. NANOG activated NODAL and CRIPTO-1to promote SMAD3phosphorylation and SNAIL expression. The transcriptional activity of NODAL gene was dependent on two NANOG binding motifs in its promoter region. This part also showed a significant correlation between the NANOG expression and the expression of NODAL, P-SMAD3or SNAIL, and the combination of NANOG and P-SMAD3was a potential predictor of poor prognosis of HCC. Additionally, cells in the tumor edge area displayed higher NANOG expression than cells in the tumor center. These results presented novel mechanistic insight into an important role of NANOG in HCC metastasis, and suggest a potential application of NANOG in HCC prognosis and treatment.Part TwoThe mechanism of SOX2regulating HCC cell invasion SOX2is a major transcription factors essential to the stem cell self-renewal and are associated with various kinds of cancers. In this part of study, we determined the expression of SOX2in HCC tumor tissues by immunohistochemistry and investigated their correlation with clinicopathologic characteristics of these patients. The results showed that SOX2expression was elevated in patients with metastasis. Kaplan-Meier survival analysis also indicated that SOX2expression was correlated with survival time of HCC patients. However, the SOX2signaling in HCC cell invasion was still elusive. In this part, we investigated its underlying mechanism in the metastasis of HCC. In this part, our data showed that HCC cells overexpressing Sox2were characterized by active epithelial-mesenchymal transition (EMT), and exhibited increased ability of matrigel invasion, soft agar colonization and sphere formation. We also found Sox2expression was correlated with the transcription activity of SLUG promoter region. These results presented novel mechanistic insight into an important role of Sox2in HCC, and suggest a potential application of Sox2in HCC prognosis and treatment.Conclusions1. NANOG and SOX2expression is elevated in metastatic HCC cell lines and invasive HCC specimens, and is correlated with survival time of HCC patients, which indicates that NANOG and SOX2have potential prognostic value for HCC.2. NANOG and SOX2enhance stem-like property and invasive potential of HCC cells in vitro. Overexpressed NANOG promotes in vivo tumorigenic and metastatic abilities of HCC cells. NANOG and Sox2regulated HCC cell invasion may be through activation of epithelial-mesenchymal transtion.3. Transcription of NODAL and SLUG gene is regulated by NANOG and SOX2respectively and maybe involved in NANOG and SOX2regulated acquisition of invasive ability and EMT phenotypes of HCC cells.The potential value of his workThis study indicated a potential application of NANOG and SOX2in HCC prognosis and treatment as diagnostic biomarkers and drug targets.Novelty of the projectThis study presented novel mechanistic insight into an important role of NANOG and SOX2in HCC metastasis. A signaling feedback loop between the NANOG pathway and EMT through transcriptional activation of NODAL gene was shown to be important in HCC cell invasion. The NANOG and SOX2might regulate HCC cell invasion through transcriptional activation of NODAL and SLUG gene, respectively.