microRNA-93-5p promotes hepatocellular carcinoma progression via a microRNA-93-5p/MAP3K2/c-Jun positive feedback circuit.

Cumulative evidence suggests that microRNAs (miRNAs) promote gene expression in cancers. However, the pathophysiologic relevance of miRNA-mediated RNA activation in hepatocellular carcinoma (HCC) remains to be established. Our previous miRNA expression profiling in seven-paired HCC specimens revealed miR-93-5p as an HCC-related miRNA. In this study, miR-93-5p expression was assessed in HCC tissues and cell lines by quantitative real-time PCR and fluorescence in situ hybridization. The correlation of miR-93-5p expression with survival and clinicopathological features of HCC was determined by statistical analysis. The function and potential mechanism of miR-93-5p in HCC were further investigated by a series of gain- or loss-of-function experiments in vitro and in vivo. We identified that miR-93-5p, overexpressed in HCC specimens and cell lines, leads to poor outcomes in HCC cases and promotes proliferation, migration, and invasion in HCC cell lines. Mechanistically, rather than decreasing target mRNA levels as expected, miR-93-5p binds to the 3'-untranslated region (UTR) of mitogen-activated protein kinase kinase kinase 2 (MAP3K2) to directly upregulate its expression and downstream p38 and c-Jun N-terminal kinase (JNK) pathway, thereby leading to cell cycle progression in HCC. Notably, we also demonstrated that c-Jun, a downstream effector of the JNK pathway, enhances miR-93-5p transcription by targeting its promoter region. Besides, downregulation of miR-93-5p significantly retarded tumor growth, while overexpression of miR-93-5p accelerated tumor growth in the HCC xenograft mouse model. Altogether, we revealed a miR-93-5p/MAP3K2/c-Jun positive feedback loop to promote HCC progression in vivo and in vitro, representing an RNA-activating role of miR-93-5p in HCC development.

UBE2T promotes proliferation via G2/M checkpoint in hepatocellular carcinoma.

BACKGROUND: Growing evidence suggests that the ubiquitin-proteasome system is involved in the pathogenesis and recurrence of hepatocellular carcinoma (HCC); yet, little is known about the role of ubiquitin-conjugating enzyme E2T (UBE2T) in HCC. MATERIALS AND METHODS: UBE2T levels were detected in HCC tissues and hepatoma cell lines using quantitative reserve transcriptase-polymerase chain reaction and Western blot analysis. Next, the changes of phenotypes after UBE2T knockdown or overexpression were evaluated using in vitro methods. Finally, the mechanism of UBE2T in HCC was tested using ex vivo and in vivo methods. RESULTS: In the present study, we reported that UBE2T mRNA and protein levels were significantly upregulated in HCC tissues compared to adjacent non-tumor tissues. Additionally, suppression of UBE2T expression inhibited proliferation, colony formation, tumorigenesis, migration, and invasion of hepatoma cells, whereas UBE2T overexpression led to the opposite outcomes. Moreover, suppression of UBE2T expression resulted in an increase in G2/M phase and a decrease in the percentage of cells in G1 phase, which indicated a cell cycle arrest at the G2/M phase. In contrast, the percentage of cells in G2/M phase decreased following UBE2T overexpression. Further study indicated that UBE2T regulated the G2/M transition by modulating cyclin B1 and cyclin-dependent kinase 1. CONCLUSION: Taken together, the findings of the present study uncover biological functions of UBE2T in hepatoma cells, and delineate preliminary molecular mechanisms of UBE2T in modulating HCC development and progression.

Comprehensive analysis of long non­coding RNA­messenger RNA­microRNA co­expression network identifies cell cycle­related lncRNA in hepatocellular carcinoma.

Long non­coding RNAs (lncRNAs) have been shown to contribute to progression and prognosis of hepatocellular carcinoma (HCC). However, expression profiling and interaction of lncRNAs with messenger RNAs (mRNAs) and microRNAs (miRNAs) remain largely unknown in HCC. The expression profiling of lncRNAs, mRNA and miRNAs was obtained using microarray. The Gene Ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes pathway analysis were used to characterize potential functions of differentially expressed mRNAs. Cytoscape was applied to construct an lncRNA­miRNA­mRNA co­expression network and candidate lncRNAs were validated via quantitative PCR in 30 pairs of HCC and adjacent tumor­free tissues. In this study, 1,056 upregulated and 1,288 downregulated lncRNAs were identified, while 2,687 mRNAs and 6 miRNAs were aberrantly expressed in HCC compared with adjacent tumor­free tissues. Potential functions of differentially expressed mRNAs were demonstrated to significantly participate in modulating critical genes in the cell cycle, such as cyclin E1 and cyclin B2. After screening, 95 lncRNAs, 5 miRNAs and 36 mRNAs were recruited for construction of lncRNA­mRNA­miRNA co­expression network in the cell cycle pathway. Subsequently, the top 5 lncRNAs that potentially modulate critical genes in the cell cycle were selected as the candidates for further verification. Kaplan­Meier curves using the Cancer Genome Atlas database showed that 13 targeted mRNAs were associated with overall survival of HCC patients. Finally, three lncRNAs, including ENST00000522221, lnc­HACE1­6:1 and lnc­ICOSLG­11:1, are significantly upregulated in HCC tissues compared with adjacent tumor­free tissues. These findings suggest that lncRNAs play essential roles in the pathogenesis of HCC via regulating coding genes and miRNAs, and may be important targets for diagnosis and treatment of this disease.

Overexpressed pepsinogen C is associated with poor prognosis in human hepatocellular carcinoma: a tissue microarray study.

Background: Aberrant expression of pepsinogen C (PGC) has been observed in human cancers. However, its role in hepatocellular carcinoma (HCC) remains to be established. The goal of this study is to illustrate PGC expression and to evaluate its clinical relevance in HCC. Materials and methods: PGC expression was examined in 75 pairs of HCC and adjacent non-tumor tissues using tissue microarray. The correlations between its expression and clinical parameters were also analyzed. Results: PGC overexpression was significantly associated with larger tumor size (≥5 cm; P=0.017) and incomplete encapsulation (P<0.0001). Cox regression model demonstrated that PGC expression and tumor size were independent prognostic factors for overall survival (OS) and disease-free survival (DFS) in HCC. The subgroup analysis by Kaplan-Meier uncovered that OS and DFS were much worse in high PGC level group than in low PGC level group with large tumor size subgroup, while no difference of OS was noted between the two groups with low tumor size subgroup. Conclusion: PGC plays a tumorigenesis role in HCC progression, which may lead to a novel insight to the potential biomarker and novel therapeutic strategies for HCC patients.

microRNA-19a-3p promotes tumor metastasis and chemoresistance through the PTEN/Akt pathway in hepatocellular carcinoma.

microRNA-19a-3p (miR-19a-3p) has been reported to regulate cell proliferation in hepatocellular carcinoma (HCC), but its role in HCC metastasis remains unknown. In this study, miR-19a-3p was noted to be upregulated in HCC specimens and cell lines. Aberrant expression of miR-19a-3p stimulated HCC cell metastasis, and phosphatase and tensin homolog (PTEN) was shown to be a direct target of miR-19a-3p. miR-19a-3p-mediated HCC metastasis was reversed by restoration of PTEN or could be imitated by silencing of PTEN. Modulation of miR-19a-3p also altered expression of phosphorylated Akt, a downstream mediator of PTEN. Moreover, aberrant expression of miR-19a-3p induced sorafenib resistance by regulating the PTEN/Akt pathway. In conclusion, ectopic expression of miR-19a-3p contributes to HCC metastasis and chemoresistance by modulating PTEN expression and the PTEN-dependent pathways.

Microarray Expression Profiling of microRNAs Reveals Potential Biomarkers for Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) remains a major health problem for delayed diagnosis, inefficient surveillance and poor prognosis. Recent studies have indicated that non-coding RNAs contribute to the development of new strategies for diagnosis and treatment of HCC. In the present study, we employed 18 pairs of HCC and matched non-tumor tissues for the identification of differentially expressed microRNAs (miRNAs) in HCC, among which 7 paired specimens were selected randomly for microarray detection. Totally, twenty-three miRNAs were screened out to have statistically significant differences with the threshold of P < 0.01 and fold-change ≥ 2.0 or ≤ 0.5 using miRNA microarray. In the validation stage, two miRNAs exhibited higher expression levels in the HCC tissues compared with those in the matched non-tumor tissues, whereas the expression levels of ten miRNAs were lower in the HCC tissues than those in the matched non-tumor tissues. In further analysis, eight miRNAs, including miR-4270, miR-125b-5p, miR-199a-3p, miR-10a-5p, miR-424-5p, miR-195-5p, miR-106b-5p and miR-3651, were retained, when another constraint about the signal intensity of microarray probes was established. Among these miRNAs, our study was the first to show the higher expression level of miR-3651 and the lower expression level of miR-4270 in HCC. The areas under the receiver-operating-characteristic curve values of miR-3651 and miR-4270 were 0.730 and 0.967, respectively, indicating their potential diagnostic values. Our results may help provide the context for expanded interpretations of miRNA studies involved in the progression of liver disease, potentially serving as a diagnostic tool of HCC.

Circulating miR-338-5p is a potential diagnostic biomarker in colorectal cancer.

OBJECTIVE: The expression of miR-338-5p has been reported to be upregulated in colorectal cancer (CRC) tissues. Clinicopathological features indicate that miR-338-5p overexpression correlates with the metastatic status of CRC. This study was aimed to investigate the diagnostic value of serum miR-338-5p for CRC. METHODS: Peripheral blood samples were collected from 210 participants, including 80 patients with CRC, 50 with colorectal polyps and 80 healthy controls. Serum miR-338-5p was quantified by quantitative reverse-transcription polymerase chain reaction. The area under the receiver operating characteristic curve (AUROC) was used to estimate the diagnostic value of miR-338-5p, carcinoembryonic antigen (CEA) and the combination of these two biomarkers. RESULTS: Serum miR-338-5p levels (fold change) in patients with CRC and colorectal polyps, and controls were 4.94 ± 1.13, 4.12 ± 0.75 and 3.07 ± 0.75, respectively. Significant differences were observed between the groups (P < 0.001). The AUROC of miR-338-5p was 0.923 (95% CI 0.882-0.964) and 0.845 (95% CI 0.792-0.898), respectively, for distinguishing CRC from healthy controls or from those without CRC. The AUROC of the combination of miR-338-5p and CEA was 0.932 (95% CI 0.882-0.964), with a sensitivity of 85%, a specificity of 88.8% at a cut-off value of 8.16. CONCLUSIONS: Circulating miR-338-5p may serve as a potential noninvasive diagnostic biomarker for detecting CRC. The combination of miR-338-5p and CEA exhibits the highest diagnostic value in our study.

Serum microRNA signatures and metabolomics have high diagnostic value in hepatocellular carcinoma.

BACKGROUND: Many new diagnostic biomarkers have been developed for hepatocellular carcinoma (HCC). We selected two methods with high diagnostic value, the detection of serum microRNAs and metabolomics based on gas chromatography/mass spectrometry (GC/MS), and attempted to establish appropriate models. METHODS: We reviewed the diagnostic efficiencies of all microRNAs identified by previous diagnostic tests. Then we chose appropriate microRNAs to validate the diagnostic efficiencies, and determined the optimal combination. We included 66 patients with HCC and 82 healthy controls (HCs) and detected the expression of the microRNAs. GC/MS analysis was performed, and we used three multivariate statistical methods to establish diagnostic models. The concentration of alpha feto-protein (AFP) was determined for comparison with the novel models. RESULTS: 82 published studies and 92 microRNAs were ultimately included in this systematic review. Seven microRNAs were selected for further validation of their diagnostic efficiencies. Among which, miR-21, miR-106b, miR-125b, miR-182 and miR-224 had a significantly different expression in HCC patients. The combination of miR-21, miR-106b and miR-224 had the highest area under the curve (AUC) at 0.950 with a sensitivity of 80.3% and a specificity of 92.7%. The GC/MS analysis exhibited an excellent diagnostic value and the AUC reached 1.0. In comparison, the AUC of the traditional biomarker, AFP, was 0.755. CONCLUSION: MicroRNAs and metabolomics shows promising potential as new diagnostic methods due to their high diagnostic value compared with traditional biomarkers.