Gar-Yang Chau

MicroRNA‐122, a tumor suppressor microRNA that regulates intrahepatic metastasis of hepatocellular carcinoma

MicroRNAs (miRNAs), which are inhibitors of gene expression, participate in diverse biological functions and in carcinogenesis. In this study, we show that liver‐specific microRNA‐122 (miR‐122) is significantly down‐regulated in liver cancers with intrahepatic metastastasis and negatively regulates tumorigenesis. Restoration of miR‐122 in metastatic Mahlavu and SK‐HEP‐1 cells significantly reduced in vitro migration, invasion, and anchorage‐independent growth as well as in vivo tumorigenesis, angiogenesis, and intrahepatic metastasis in an orthotopic liver cancer model. Because an inverse expression pattern is often present between an miRNA and its target genes, we used a computational approach and identified multiple miR‐122 candidate target genes from two independent expression microarray datasets. Thirty‐two target genes were empirically verified, and this group of genes was enriched with genes regulating cell movement, cell morphology, cell‐cell signaling, and transcription. We further showed that one of the miR‐122 targets, ADAM17 (a disintegrin and metalloprotease 17) is involved in metastasis. Silencing of ADAM17 resulted in a dramatic reduction of in vitro migration, invasion, in vivo tumorigenesis, angiogenesis, and local invasion in the livers of nude mice, which is similar to that which occurs with the restoration of miR‐122. Conclusion: Our study suggests that miR‐122, a tumor suppressor microRNA affecting hepatocellular carcinoma intrahepatic metastasis by angiogenesis suppression, exerts some of its action via regulation of ADAM17. Restoration of miR‐122 has a far‐reaching effect on the cell. Using the concomitant down‐regulation of its targets, including ADAM17, a rational therapeutic strategy based on miR‐122 may prove to be beneficial for patients with hepatocellular carcinoma. (HEPATOLOGY 2009.)

Comprehensive analysis of the independent effect of twist and snail in promoting metastasis of hepatocellular carcinoma

The epithelial‐mesenchymal transition (EMT) is critical for induction of invasiveness and metastasis of human cancers. In this study we investigated the expression profiles of the EMT markers, the relationship between EMT markers and patient/tumor/viral factors, and the interplay between major EMT regulators in human hepatocellular carcinoma (HCC). Reduced E‐cadherin and nonmembranous β‐catenin expression, the hallmarks of EMT, were shown in 60.2% and 51.5% of primary HCC samples, respectively. Overexpression of Snail, Twist, or Slug, the major regulators of EMT, was identified in 56.9%, 43.1%, and 51.4% of primary HCCs, respectively. Statistical analysis determined that Snail and Twist, but not Slug, are major EMT inducers in HCC: overexpression of Snail and/or Twist correlated with down‐regulation of E‐cadherin, nonmembranous expression of β‐catenin, and a worse prognosis. In contrast, there were no such significant differences in samples that overexpressed Slug. Coexpression of Snail and Twist correlated with the worst prognosis of HCC. Hepatitis C‐associated HCC was significantly correlated with Twist overexpression. HCC cell lines with increased Snail and Twist expression (e.g., Mahlavu) exhibited a greater capacity for invasiveness/metastasis than cells with low endogenous Twist/Snail expression (e.g., Huh‐7). Overexpression of Snail or/and Twist in Huh‐7 induced EMT and invasiveness/metastasis, whereas knockdown of Twist or Snail in Mahlavu reversed EMT and inhibited invasiveness/metastasis. Twist and Snail were independently regulated, but exerted an additive inhibitory effect to suppress E‐cadherin transcription. Conclusion: Our study provides a comprehensive profile of EMT markers in HCC, and the independent and collaborative effects of Snail and Twist on HCC metastasis were confirmed through different assays. (HEPATOLOGY 2009.)

Risk factors for early and late recurrence in hepatitis B-related hepatocellular carcinoma ☆

BACKGROUND/AIMS Hepatitis B virus (HBV) levels correlate with the development of hepatocellular carcinoma (HCC), but the role of viral load in HCC recurrence after tumor resection remains unclear. Herein we aimed to investigate the role of viral load in HCC recurrence following tumor resection. METHODS From 1990 to 2002, 193 HBV-related HCC patients who underwent tumor resection in Taipei Veterans General Hospital were enrolled. Serum HBV DNA level and mutations were analyzed for association with early and late recurrence, together with other clinical variables. RESULTS During a follow-up of 58.2+/-44 months, 134 patients had HCC recurrence. Multivariate analysis showed that multinodularity (Hazard ratio [HR], 95% confidence interval [CI]; 2.232, 1.021-4.878), macroscopic venous invasion (4.693, 1.645-13.391), AFP >20 ng/ml (3.891, 1.795-8.475), and cut margin <or= 1cm (3.333, 1.487-7.470) were correlated with early recurrence (within two years of operation) of HCC. In addition, multivariate analysis determined that Ishak hepatic inflammatory activity >6 (4.658, 1.970-11.017), multinodularity (3.266, 1.417-7.526), ICG-15 >10% (2.487, 1.095-5.650) and HBV DNA level >10(6) copies/ml (2.548, 1.040-6.240) were significantly associated with late recurrence (>two years after resection). Patients with high viral loads tended to have higher Ishak inflammatory (7.00+/-3.07 vs. 5.33+/-2.96, p=0.001) and fibrosis scores (4.17+/-2.01 vs. 3.20+/-2.41, p=0.007) than those with lower loads. CONCLUSIONS Tumor factors were associated with early HCC recurrence while high viral loads and hepatic inflammatory activity were associated with late recurrence. Pre- and post-operative antiviral and anti-inflammatory therapies may be crucial in reducing late recurrence.

Prognostic Significance of Vascular Endothelial Growth Factor, Basic Fibroblast Growth Factor, and Angiogenin in Patients With Resectable Hepatocellular Carcinoma After Surgery

Background: Hepatocellular carcinoma (HCC) is a hypervascular malignancy. Vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and angiogenin (ANG) are important angiogenic factors of neoangiogenesis. This study investigated the predictive value of serum VEGF, bFGF, and ANG in tumor recurrence, disease-free survival (DFS), and overall survival (OS) in HCC patients.Methods: Preoperative serum VEGF, bFGF, and ANG were measured in 98 patients with resectable HCC and in 15 healthy controls. The median follow-up time was 43 months.Results: Preoperative serum VEGF was increased in patients with resectable HCC compared with healthy controls (P < .05). Increased serum VEGF was correlated with tumor recurrence (P = .001). Univariate analysis showed that serum VEGF, tumor-node-metastasis stage, tumor size and number, macroscopic portal vein invasion, and microscopic vascular invasion were correlated with OS and DFS. Serum bFGF and ANG were not associated with survival. Multivariate analysis showed that serum VEGF was the most significant predictor of DFS (relative risk, 2.35; 95% confidence interval, 1.26–4.39; P = .007) and OS (relative risk, 3.44; 95% confidence interval, 1.81–6.57; P < .001) in HCC patients after surgical resection.Conclusions:Preoperative serum VEGF is a significant independent predictor of tumor recurrence, DFS, and OS in patients with resectable HCC.

Alteration of the copy number and deletion of mitochondrial DNA in human hepatocellular carcinoma

Somatic mutations in mitochondrial DNA (mtDNA) have been detected in hepatocellular carcinoma (HCC). However, it remains unclear whether mtDNA copy number and mitochondrial biogenesis are altered in HCC. In this study, we found that mtDNA copy number and the content of mitochondrial respiratory proteins were reduced in HCCs as compared with the corresponding non-tumorous livers. MtDNA copy number was significantly reduced in female HCC but not in male HCC. Expression of the peroxisome proliferator-activated receptor γ coactivator-1 was significantly repressed in HCCs (P<0.005), while the expression of the mitochondrial single-strand DNA-binding protein was upregulated, indicating that the regulation of mitochondria biogenesis is disturbed in HCC. Moreover, 22% of HCCs carried a somatic mutation in the mtDNA D-loop region. The non-tumorous liver of the HCC patients with a long-term alcohol-drinking history contained reduced mtDNA copy number (P<0.05) and higher level of the 4977 bp-deleted mtDNA (P<0.05) as compared with non-alcohol patients. Our results suggest that reduced mtDNA copy number, impaired mitochondrial biogenesis and somatic mutations in mtDNA are important events during carcinogenesis of HCC, and the differential alterations in mtDNA of male and female HCC may contribute to the differences in the clinical manifestation between female and male HCC patients.

Identification of SOX4 target genes using phylogenetic footprinting-based prediction from expression microarrays suggests that overexpression of SOX4 potentiates metastasis in hepatocellular carcinoma

A comprehensive microarray analysis of hepatocellular carcinoma (HCC) revealed distinct synexpression patterns during intrahepatic metastasis. Recent evidence has demonstrated that synexpression group member genes are likely to be regulated by master control gene(s). Here we investigate the functions and gene regulation of the transcription factor SOX4 in intrahepatic metastatic HCC. SOX4 is important in tumor metastasis as RNAi knockdown reduces tumor cell migration, invasion, in vivo tumorigenesis and metastasis. A multifaceted approach integrating gene profiling, binding site computation and empirical verification by chromatin immunoprecipitation and gene ablation refined the consensus SOX4 binding motif and identified 32 binding loci in 31 genes with high confidence. RNAi knockdown of two SOX4 target genes, neuropilin 1 and semaphorin 3C, drastically reduced cell migration activity in HCC cell lines suggesting that SOX4 exerts some of its action via regulation of these two downstream targets. The discovery of 31 previously unidentified targets expands our knowledge of how SOX4 modulates HCC progression and implies a range of novel SOX4 functions. This integrated approach sets a paradigm whereby a subset of member genes from a synexpression group can be regulated by one master control gene and this is exemplified by SOX4 and advanced HCC.

Serum Interleukin-10 But Not Interleukin-6 Is Related to Clinical Outcome in Patients With Resectable Hepatocellular Carcinoma

OBJECTIVE To evaluate the clinical significance of preoperative serum levels of interleukin-10 (IL-10) and interleukin-6 (IL-6) in patients with resectable hepatocellular carcinoma (HCC). SUMMARY BACKGROUND DATA IL-10 is an immunosuppressive factor and IL-6 is a multifunctional cytokine that plays a role in host defense mechanisms. Both have been reported to be related to the disease prognosis in some human solid tumors. Their role in human HCC has not been investigated. METHODS Preoperative serum samples of 67 patients with HCC who underwent potentially curative resection and 27 normal healthy donors were assayed. Levels of IL-10 and IL-6 were determined by enzyme-linked immunosorbent assay. The clinical significance of serum IL-10 and IL-6 was evaluated and compared with conventional clinicopathologic factors. RESULTS Levels of IL-10 and IL-6 were significantly higher in patients with HCC than in healthy subjects. There was no correlation between IL-10 and IL-6 levels. Tumor resection resulted in a decrease in IL-10 and IL-6 levels. On univariate analysis, patients with high IL-10 levels had a worse disease-free survival, but IL-6 levels had no correlation with the disease-free survival. Multivariate analysis identified IL-10 levels as a predictor of postresectional outcome, in addition to the well-established clinical risk factors. CONCLUSIONS In patients with HCC, the preoperative serum IL-10 level is related to the clinical outcome. IL-10 may play an important role in the progression of HCC.

Prognostic significance of surgical margin in hepatocellular carcinoma resection: an analysis of 165 Childs' A patients.

The clinical significance of the width of the surgical margin in the resection of hepatocellular carcinoma (HCC) has yet to be clarified.

Comparison of recurrence after hepatic resection in patients with hepatitis B vs. hepatitis C-related small hepatocellular carcinoma in hepatitis B virus endemic area.

Abstract: Purpose: Hepatitis B virus (HBV) and hepatitis C virus (HCV) infection are two important factors in the development of hepatocellular carcinoma (HCC). The carcinogenic mechanism of HBV and HCV is considered to be different. It is interesting to compare the recurrence after hepatic resection in patients with small HCC who were infected with HBV or HCV.

Diabetes mellitus is a risk factor for hepatic decompensation in patients with hepatocellular carcinoma undergoing resection: a longitudinal study.

OBJECTIVES:Patients with hepatocellular carcinoma (HCC) frequently have diabetes mellitus (DM) due to coexisting liver cirrhosis. The aim of this study was to assess the long-term impact of DM on the hepatic regenerative ability of HCC patients undergoing surgical resection.METHODS:We retrospectively studied 245 HCC patients (210 male; age, 61 ± 13 yr) with well-preserved liver functions undergoing resection. Forty (16%) of them were diabetic and were controlled with hypoglycemic agents. The Child-Pugh scoring system was used to evaluate the postoperative liver regeneration ability. The endpoint was the occurrence of hepatic decompensation, defined as a sustained increase in the Child-Pugh score of 2 or more points or the development of tumor recurrence.RESULTS:Seventy-five patients (31%) developed hepatic decompensation during a follow-up period of 27 ± 18 months (range, 3–75). DM (p= 0.001), large (>3 cm) tumor size (p= 0.044), and age > 65 yr (p= 0.058) were the factors associated with hepatic decompensation in univariate analysis. Multivariate Cox regression model analysis confirmed that DM (relative risk [RR] = 2.3, 95% CI = 1.4–3.7, p= 0.001) and tumor size > 3 cm (RR = 1.7, 95% CI = 1.1–2.7, p= 0.046) were independent prognostic predictors associated with the occurrence of hepatic decompensation; the respective 3- and 5-yr cumulative rates were 53% and 64% versus 27% and 50% for diabetic and nondiabetic patients, and 24% and 41% versus 38% and 60% for patients with small (≤3 cm) versus large (>3 cm) tumors.CONCLUSION:HCC patients with DM or large tumor size are at a cumulative increased risk for postoperative hepatic decompensation.