The Role Of SUMO-1 In The Diagnosis And Therapy Of Hepatocellular Carcinoma

Background:Small ubiquitin-like modifier (SUMO) is a member of ubiquitin-like gene family, which can covalently modify a large number of proteins. Unlike the function of ubiquitin which degrades the substrates, SUMO-1 regulates the functions of substrate proteins, including transcription, DNA repair, nucleocytoplasmic trafficking, chromosome segregation and chromosome metabolism. More and more cancer-related proteins, such as proliferating cell nuclear antigen, p53, estrogen receptor, NF-kappa B signal regulation, the maintenance of telomerase activation, et al, have been found to be involved in SUMO-1 modification. For example, SUMO-1 modification can inhibit the activity of p53, which will promote the occurrence, development and metastasis of cancers. Tumor suppressor gene p53 has an important role in the occurrence, differentiation, apoptosis, etc, of cells. If the activity of p53 was inhibited, the cells would be prone to become cancers. Deletion of p53 gene in cells can cause the occurrence of many types of cancer. But once the inactivation of the p53 gene was re-activated, which can lead to tumor growth inhibition or even disappearance. SUMO-1 plays an important role in the occurrence and development of cancer.Objectives:Owing to the special function in the occurrence and development of cancer, the aims of the research are mainly to investigate the expression level of SUMO-1 in HCC, to explore the varies of HCC cell in proliferation, apoptosis and cell cycle after silencing the expression of SUMO-1, and to demonstrate the mechanism inducing the changes in HCC. According to the results of the research, the role of SUMO-1 was evaluated in diagnosis and therapy of HCC.Methods:The expression levels of SUMO-1 mRNA and protein in SMMC-7721, HepG2, Hep3B, HCCs and liver tissue surrounding HCCs were examined by means of RT-PCR and western blot. The SUMO-1 mRNAs were examined by means of RT-PCR in primary carcinomas of liver, liver tissues surrounding liver cancers, hemangiomas of liver and focal nodulahyperplasia of liver. The differences of expression level of SUMO-1 mRNA were compared between primary carcinomas of liver and liver tissues surrounding liver cancers, positive serum AFP and negative serum AFP hepatocellular carcinomas, primary carcinomas of liver and benign tumors of liver. The SUMO-1 siRNA was transfected into SMMC-7721 by means of LipofectamineTM 2000. The silencing efficiency of SUMO-1 was examined by RT-PCR and western blot. The cell growth and cell cycle were examined by MTT and flow cytometry (FCM). The cell apoptosis were detected by DeadEndTM Colorimetric TUNEL System. The expression changes of Bcl-2 and c-myc in SMMC-7721 were detected by means of RT-PCR and western blot after being transfected SUMO-1 siRNA.Results:The expression level of SUMO-1 mRNA and protein elevated significantly in SMMC-7721, HepG2, Hep3B and hepatocellular carcinoma tissues. Although the quantities of SUMO-1 mRNA and protein varied in liver tissues surrounding HCC, there was a significant difference between hepatocellular carcinoma tissues and their surrounding liver tissues (P< 0.001). SUMO-1 mRNA all expressed in clinical primary carcinomas of liver, liver tissues surrounding liver cancers, hemangiomas of liver and focal nodulahyperplasia of liver. There were significant differences between primary carcinomas of liver and liver tissues surrounding liver cancers, primary carcinomas of liver and benign tumors of liver. No matter positive or negative of serum AFP in the patients with hepatocellular carcinoma, SUMO-1 mRNA all had over expression levels. The siRNA could significantly silence the expression of SUMO-1 in SMMC-7721.The maximal silencing rate was ulmost 73.43% at 48 hours after being transfected SUMO-1 siRNA. MTT assay revealed that the cell line grew more slowly. FCM showed that the number of G2 stage cells was increased significantly. But apoptosis cells were not found by TUNNEL assay. Following the down-regulation of SUMO-1 in SMMC-7721, the expressions level of Bcl-2, c-myc and a-tubulin were all down-regulated significantly.Conclusion:The expression of SUMO-1 in HCCs showed marked contrast enhancement relative to the liver tissues surrounding HCC. The results implied that SUMO-1 may be an interesting target in the diagnosis and treatment of HCC. Owing to the overexpression of SUMO-1, SUMO-1 mRNA may be a good marker in diagnosing primary carcinoma of liver. SiRNA is a good manner to silence the expression of SUMO-1 in SMMC-7721 in vitro. Owing to the growth inhibition and down-regulation of Bcl-2 and c-myc induced by SUMO-1 siRNA, SUMO-1 plays an important role in development of SMMC-7721. SUMO-1 may be a potential target in treating HCC.