Hong Tu

Gain of miR-151 on chromosome 8q24.3 facilitates tumour cell migration and spreading through downregulating RhoGDIA

Recurrent chromosomal aberrations are often observed in hepatocellular carcinoma (HCC), but little is known about the functional non-coding sequences, particularly microRNAs (miRNAs), at the chromosomal breakpoints in HCC. Here we show that 22 miRNAs are often amplified or deleted in HCC. MicroRNA-151 (miR-151), a frequently amplified miRNA on 8q24.3, is correlated with intrahepatic metastasis of HCC. We further show that miR-151, which is often expressed together with its host gene FAK, encoding focal adhesion kinase, significantly increases HCC cell migration and invasion in vitro and in vivo, mainly through miR-151-5p, but not through miR-151-3p. Moreover, miR-151 exerts this function by directly targeting RhoGDIA, a putative metastasis suppressor in HCC, thus leading to the activation of Rac1, Cdc42 and Rho GTPases. In addition, miR-151 can function synergistically with FAK to enhance HCC cell motility and spreading. Thus, our findings indicate that chromosome gain of miR-151 is a crucial stimulus for tumour invasion and metastasis of HCC.

Hypoxia‐inducible factor 1 alpha–activated angiopoietin‐like protein 4 contributes to tumor metastasis via vascular cell adhesion molecule‐1/integrin β1 signaling in human hepatocellular carcinoma

Angiopoietin‐like protein 4 (ANGPTL4) plays complex and often contradictory roles in vascular biology and tumor metastasis, but little is known about its function in hepatocellular carcinoma (HCC) metastasis. In the present study, we showed that hypoxia‐inducible factor 1α (HIF‐1α) directly up‐regulates ANGPTL4, and its stableness positively correlates with ANGPTL4 expression in HCC tissue. Overexpression of ANGPTL4 significantly increased HCC cell transendothelial migration in vitro and intrahepatic and distal pulmonary metastasis in vivo, whereas silencing ANGPTL4 expression or treatment with a neutralizing antibody specific for ANGPTL4 protein resulted in a reduced transendothelial migration. We also found that serum ANGPTL4 is higher in HCC patients, compared to healthy control, and correlates with intrahepatic metastasis and histological grade. Further, secreted ANGPTL4 promotes transendothelial migration and metastasis of HCC cells in vitro and in vivo through the up‐regulation of vascular cell adhesion molecule‐1 (VCAM‐1) of human umbilical vein endothelial cells and the activation of the VCAM‐1/integrin β1 axis. Conclusion: ANGPTL4 is a target gene of HIF‐1α and acts as an important regulator in the metastasis of HCC. Serum ANGPTL4 correlates with tumor progression and metastasis and might be used to indicate prognosis in HCC patients. (HEPATOLOGY 2011 54:910–919;)

Sequential accumulation of the mutations in core promoter of hepatitis B virus is associated with the development of hepatocellular carcinoma in Qidong, China

BACKGROUND/AIMS To investigate the mutations in hepatitis B virus (HBV) that might be related to hepatocellular carcinoma (HCC) in the high-risk area Qidong, China. METHODS DNA sequences of HBV basal core promoter (BCP) and the overlapping X gene were determined in 58 HCC and 71 chronic hepatitis (CH) patients. In addition, a consecutive series of plasma samples from 15 HCC cases were employed to compare the CP/X sequences before and after the occurrence of HCC. RESULTS T1762/A1764 double mutation was frequently found in Qidong patients, regardless of clinical status (65.5% in HCC and 73.2% in CH, P>0.05). Unexpectedly, the adjacent T1766/A1768 mutation significantly increased the risk of HCC (P<0.05). Moreover, the prevalence of triple mutations in BCP was significantly higher in patients with HCC than those with CH (P<0.05). The longitudinal study demonstrated that the mutations in BCP were gradually accumulated during the development of HCC. Colony formation assay showed while A1764 mutation alone did not alter the colony-inhibitory activity of HBx, double or triple mutations largely abrogated this effect. CONCLUSIONS The complex mutation involving T1766/A1768 was closely related to HCC. The enhanced risk of HCC caused by BCP variants could be attributable partially to the aberrant activity of HBx.

K-ras mutational status predicts poor prognosis in unresectable pancreatic cancer

OBJECTIVE To determine the prognostic value of K-ras mutations in plasma DNA of unresectable pancreatic cancer patients. METHODS Blood samples were collected from 91 patients with unresectable pancreatic cancer prior to treatment. K-ras gene was amplified from the circulating plasma DNA. Mutations were detected by direct sequencing. The relationship between the types of K-ras gene and prognosis of unresectable pancreatic cancer was evaluated. RESULTS K-Ras codon 12 mutations were found in 30 of 91(33%) plasma DNA samples, 17mutations were c.35G>A (p.G12D), 11 were c.35G>T (p.G12V) and only 2 were c.34G>C (p.G12R)). K-ras codon 12 mutations could significantly reflect the clinical parameters, including TNM tumor staging (P=0.033) and liver metastasis (P=0.014). The median survival time of patients with K-ras mutations was shorter than that of patients with wild-type K-ras gene (3.9 months vs. 10.2 months, P<0.001). K-ras codon 12 mutation from plasma DNA was an independent negative prognostic factor for survival (hazard ratio, 7.39; 95% confidence interval, 3.69-14.89). CONCLUSION K-ras mutation in plasma DNA is a predictive biomarker for a poor prognosis of unresectable pancreatic cancer patients.

Development of T Cells Redirected to Glypican-3 for the Treatment of Hepatocellular Carcinoma

Purpose: The aim of our study is to elucidate whether T cells expressing GPC3-targeted chimeric antigen receptor (CAR) can efficiently eliminate GPC3-positive HCC cells and their potential in the treatment of HCC. Experimental Design: T cells expressing a first-generation and third-generation GPC3-targeted CAR were prepared using lentiviral vector transduction. The in vitro and in vivo cytotoxic activities of the genetically engineered CAR T cells were evaluated against various HCC cell lines. Results: GPC3-targeted CAR T cells could efficiently kill GPC3-positive HCC cells but not GPC3-negative cells in vitro. These cytotoxic activities seemed to be positively correlated with GPC3 expression levels in the target cells. In addition, T cells expressing the third-generation GPC3-targeted CAR could eradicate HCC xenografts with high level of GPC3 expression and efficiently suppress the growth of HCC xenografts with low GPC3 expression level in vivo. The survival of the mice bearing established orthotopic Huh-7 xenografts was significantly prolonged by the treatment with the third-generation GPC3-targeted CAR T cells. Conclusions: GPC3-targeted CAR T cells could potently eliminate GPC3-positive HCC cells, thereby providing a promising therapeutic intervention for GPC3-positive HCC. Clin Cancer Res; 20(24); 6418–28. ©2014 AACR.

Sphingosine kinase 1 promotes tumour cell migration and invasion via the S1P/EDG1 axis in hepatocellular carcinoma

Sphingosine kinase 1 (SphK1), which phosphorylates sphingosine to sphingosine‐1‐phosphate (S1P), is overexpressed in various types of cancers, and may act as an oncogene in tumorigenesis. However, little is known about the precise role of the SphK1/S1P pathway in human liver cancer, especially regarding the metastasis of hepatocellular carcinoma (HCC).

MiRNA-10a is upregulated in NSCLC and may promote cancer by targeting PTEN

MicroRNAs (miRNAs) are involved in human cancer including non-small cell lung cancer (NSCLC). In this study, we compared miRNA expression microarray of SPC-A-1sci (high metastatic) and SPC-A-1 (weakly metastatic) cells. We found that miRNA-10a was up-regulated in NSCLC compared with corresponding normal tissues. High expression of miR-10a was associated with tumor node metastasis and lymph node metastasis. Furthermore, overexpression of miR-10a promoted NSCLC cell proliferation, migration and invasion in vitro. We found that PTEN was a direct target of miR-10a in NSCLC. Also miR-10a activated the PTEN/AKT/ERK pathway. We suggest that miR-10a contributes to NSCLC by targeting PTEN.

Comparison Study on the Complete Sequence of Hepatitis B Virus Identifies New Mutations in Core Gene Associated with Hepatocellular Carcinoma

Background: Mutations in the hepatitis B virus (HBV) genome may influence the activity of liver disease. The aim of this study was to identify new viral variations associated with hepatocellular carcinoma (HCC). Methods: We carried out a comparison study on the complete sequence of HBV isolated from 20 HCC and 35 non-HCC patients in Qidong, China, an area with a high incidence of HCC. We compared the HBV sequences in a consecutive series of plasma samples from four HCC cases before and after the occurrence of HCC. In addition, we selected four mutations in the HBV core (C) gene to verify their relationships to HCC in an independent set of 103 HCC cases and 103 sex- and age-matched non-HCC controls. Results: The pre-S deletion and 12 point mutations, namely, the pre-S2 start codon mutation, T53C in the pre-S2 gene, T766A in the S gene, G1613A, C1653T, A1762T, G1764A in the X gene, and G1899A, C2002T, A2159G, A2189C, and G2203W (A or T) in the pre-C/C gene, showed close associations with HCC. In the validation study, A2159G, A2189C, and G2203W showed consistent associations with HCC by univariate analysis. Multivariate analysis showed that A2189C and G2203W were independent risk factors for HCC. The odds ratios (95% confidence interval) were 3.99 (1.61-9.92) and 9.70 (1.17-80.58), respectively, for A2189C and G2203W. Conclusions: These results implicate A2189C and G2203W as new predictive markers for HCC. Impact: The complete genome analysis of HBV provided pilot data for the identification of novel mutations that could serve as markers for HCC. Cancer Epidemiol Biomarkers Prev; 19(10); 2623–30. ©2010 AACR.

Reevaluation of glypican-3 as a serological marker for hepatocellular carcinoma

BACKGROUND Glypican-3 (GPC3) is a novel histochemical marker of hepatocellular carcinoma (HCC). However, its utility as a serologic marker for HCC is not conclusive. METHODS A total of 1037 subjects, including 155 patients with HCC, 180 with chronic hepatitis, 124 with liver cirrhosis, 442 with non-HCC cancer and 136 healthy controls, were analyzed for serum GPC3 (sGPC3) by an ELISA constructed with 2 monoclonal antibodies. RESULTS The average level of sGPC3 in HCC patients was 99.94±267.2ng/ml, which was significantly higher than in patients with chronic hepatitis (10.45±46.02ng/ml, P<0.0001), liver cirrhosis (19.44±50.88ng/ml, P=0.0013), non-HCC cancer (20.50±98.33ng/ml, P<0.0001) and healthy controls (4.14±31.65ng/ml, P<0.0001). The sensitivity of sGPC3 in HCC diagnosis was 40.0%, whereas the specificity was 98.5%, 94.4% and 87.1% in healthy controls, chronic hepatitis patients and liver cirrhosis patients, respectively. In addition, 13.5% (28/207) of lung cancer patients and 13.2% (9/68) of thyroid cancer patients had positive results with sGPC3. CONCLUSION Serum GPC3 is a potential marker for HCC. However, the presence of sGPC3 in patients with lung cancer and thyroid cancer might limit its application as a single marker in the diagnosis of HCC.

Temporal acquisition of sequential mutations in the enhancer II and basal core promoter of HBV in individuals at high-risk for hepatocellular carcinoma

To investigate the roles of mutations in enhancer II (Enh II) and basal core promoter (BCP) of hepatitis B virus (HBV) in hepatocellular carcinoma (HCC), we determined the sequence of Enh II/BCP in 152 HCC and 136 non-HCC patients from a high-incidence area of East China. A longitudinal study was conducted on 21 cases in which serial plasma samples were available before HCC. In total, six point mutations, including T1653, V1753, T1762, A1764, T1766 and A1768, were found to occur more frequently in HCC patients. Multivariate analysis showed that the T1653 [odds ratio (OR), 2.07; 95% confidence interval (CI), 1.114-3.845] and V1753 (OR, 3.099; 95% CI, 1.520-6.317) were independent factors that were associated with HCC. Although a T1762/A1764 double mutation was found in 73.0% of the HCC patients and 66.9% of the non-HCC patients, if the combined pattern with other adjacent mutations was not taken into account, it alone showed a lower frequency in HCC patients compared with non-HCC patients (19.7 versus 34.6%, P = 0.005). Interestingly, while the OR of HCC patients with a double mutation was only 0.393 (95% CI, 0.234-0.660), it increased to 1.861 (95% CI, 1.161-2.984) with a triple mutation and to 4.434 (95% CI, 1.630-12.063) with a quadruple mutation. The longitudinal study demonstrated that the mutations in Enh II/BCP accumulated during the development of HCC. In conclusion, the T1653 and V1753 mutations were independent risk factors for HCC in East China. The T1762/A1764 double mutation was necessary but not sufficient to produce an association between Enh II/BCP mutations and HCC.