Long non-coding RNAs (lncRNAs) have previously been implicated in human disease states, especially cancer. Although the aberrant expression of lncRNAs has been observed in cancer, the biological functions and molecular mechanisms underlying aberrantly expressed lncRNAs in hepatocellular carcinoma (HCC) have not been widely established. In the present study, we investigated a novel lncRNA, termed URHC (up-regulated in hepatocellular carcinoma), and evaluated its role in the progression of HCC. Expression profiling using a lncRNA microarray revealed that URHC was highly expressed in 3 HCC cell lines compared to normal hepatocytes. Quantitative real-time polymerase chain reaction (qRT-PCR) analyses confirmed that URHC expression was increased in hepatoma cells and HCC tissues. Moreover, using qRT-PCR, we confirmed that URHC expression was up-regulated in 30 HCC cases (57.7%) and that its higher expression was correlated with poor overall survival. We further demonstrated that URHC inhibition reduced cell proliferation and promoted apoptosis. We hypothesize that URHC may function by regulating the sterile alpha motif and leucine zipper containing kinase AZK (ZAK) gene, which is located near URHC on the same chromosome. We found that ZAK mRNA levels were down-regulated in HCC tissues and the expression levels of ZAK were negatively correlated with those of URHC in the above HCC tissues. Next, we confirmed that URHC down-regulated ZAK, which is involved in URHC-mediated cell proliferation and apoptosis. Furthermore, ERK/MAPK pathway inactivation partially accounted for URHC-ZAK-induced cell growth and apoptosis. Thus, we concluded that high URHC expression can promote cell proliferation and inhibit apoptosis by repressing ZAK expression through inactivation of the ERK/MAPK pathway. These findings may provide a novel mechanism and therapeutic targets for the treatment of HCC.

B‐cell‐specific Moloney murine leukemia virus insertion site 1 (BMI1) is a member of the polycomb group of transcriptional repressors. Until now, its expression and functional significance in pancreatic carcinogenesis was unknown. In the present study, we demonstrated that expression of BMI1 was markedly up‐regulated in pancreatic cancer cell lines and surgically resected cancer specimens. In addition, BMI1 expression levels correlated positively with the presence of lymph node metastases and negatively with patient survival rates, suggesting a role for BMI1 in the progression of pancreatic cancer. Furthermore, stable down‐regulation of BMI1 suppressed cell growth, delayed the G1/S transition, and enhanced the susceptibility of different pancreatic cell lines to apoptosis following expression of a lentiviral‐mediated shRNA targeted for BMI1. Expression of the short‐hairpin RNA also correlated with the up‐regulation of p21 and Bax and the down‐regulation of cyclin D1, cyclin‐dependent kinase (CDK)‐2 and ‐4, Bcl‐2, and phospho‐Akt. Finally, growth suppression following BMI1 depletion was confirmed in a nude mouse model. In conclusion, our findings indicate that BMI1 plays an important role in the late progression of pancreatic cancer and may represent a novel therapeutic target for the treatment of pancreatic cancer. (Cancer Sci 2010)

Hepatocellular carcinoma (HCC) can be derived from malignant transformed adult hepatic progenitor cells. However, the regulatory factors and molecular mechanisms underlying the process are not well defined. Our previous microRNA (miRNA) microarray analysis revealed a significant decrease of miR-200a level in F344 rat HCC side population (SP) fraction cells versus their normal counterparts. In the present study, we further investigated the effect of miR-200a on hepatic oval cell (HOC) phenotypes. We first confirmed downregulated miR-200a levels in rat hepatoma cells compared with WB-F344 cells. Next, by lentivirus-mediated loss-of-function studies, we showed that stable knockdown of miR-200a confers a mesenchymal phenotype to WB-F344 cells, including an elongated cell morphology, enhanced cell migration ability and expression of epithelial mesenchymal transition (EMT)-representative markers. Concomitantly, several cancer stem cell (CSC)-like traits appeared in these cells, which exhibit enhanced spheroid-forming capacity, express putative hepatic CSC markers and display superior resistance to chemotherapeutic drugs in vitro. Furthermore, bioinformatics analysis, luciferase assays and western blot analysis identified β-catenin (CTNNB1) as a direct and functional target of miR-200a. Knockdown of miR-200a partially activated Wnt/β-catenin signaling, and silencing of β-catenin functionally attenuated anti-miR-200a effects in vitro in WB-F344 cells. At length, in vivo xenograft assay demonstrated the acquisition of tumorigenicity of WB-F344 cells after miR-200a siliencing. Collectively, our findings indicate that miR-200a may function as an important regulatory factor in neoplastic transition of HOCs by targeting the β-catenin pathway.

Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal solid tumor due to the lack of reliable early detection markers and effective therapies. MicroRNAs (miRNAs), noncoding RNAs that regulate gene expression, are involved in tumorigenesis and have a remarkable potential for the diagnosis and treatment of malignancy. In this study, we investigated aberrantly expressed miRNAs involved in PDAC by comparing miRNA expression profiles in PDAC cell lines with a normal pancreas cell line and found that miR-135a was significantly down-regulated in the PDAC cell lines. The microarray results were validated by qRT-PCR in PDAC tissues, paired adjacent normal pancreatic tissues, PDAC cell lines, and a normal pancreas cell line. We then defined the tumor-suppressing significance and function of miR-135a by constructing a lentiviral vector to express miR-135a. The overexpression of miR-135a in PDAC cells decreased cell proliferation and clonogenicity and also induced G1 arrest and apoptosis. We predicted Bmi1 may be a target of miR-135a using bioinformatics tools and found that Bmi1 expression was markedly up-regulated in PDAC. Its expression was inversely correlated with miR-135a expression in PDAC. Furthermore, a luciferase activity assay revealed that miR-135a could directly target the 3'-untranslated region (3'-UTR) of Bmi1. Taken together, these results demonstrate that miR-135a targets Bmi1 in PDAC and functions as a tumor suppressor. miR-135a may offer a new perspective for the development of effective miRNA-based therapy for PDAC.