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Hepatitis B virus (HBV), hepatitis C virus (HCV), human papilloma virus (HPV), Epstein-Barr virus (EBV), human T-cell lymphotropic virus type 1 (HTLV-1), Kaposi's sarcoma-associated herpesvirus (KSHV), and Merkel cell polyomavirus (MCV) contribute to about 10-15% global burden of human cancers. Conventional chemotherapy or molecular target therapies have been used to treat virus-associated cancers. However, a more proactive approach would be the use of antiviral treatment to suppress or eliminate viral infections to prevent the occurrence of cancer in the first place. Antiviral treatments against chronic HBV and HCV infection have achieved this goal, with significant reduction in the incidence of hepatocellular carcinoma in treated patients. Antiviral treatments for EBV, KSHV, and HTLV-1 had limited success in treating refractory EBV-associated lymphoma and post-transplant lymphoproliferative disorder, KSHV-associated Kaposi's sarcoma in AIDS patients, and HTLV-1-associated acute, chronic, and smoldering subtypes of adult T-cell lymphoma, respectively. Therapeutic HPV vaccine and RNA interference-based therapies for treating HPV-associated infection or cervical cancers also showed some encouraging results. Taken together, antiviral therapies have yielded promising results in cancer prevention and treatment. More large-scale studies in a real-world setting are necessary to confirm the efficacy of antiviral therapy. Further investigation for more effective and convenient antiviral regimens warrants more attention.

Background: Tumor-infiltrating leukocytes (TILs) are immune cells surrounding tumor cells, and several studies have shown that TILs are potential survival predictors in different cancers. However, few studies have dissected the differences between hepatitis B- and hepatitis C-related hepatocellular carcinoma (HBV-HCC and HCV-HCC). Therefore, we aimed to determine whether the abundance and composition of TILs are potential predictors for survival outcomes in HCC and which TILs are the most significant predictors., Methods: Two bioinformatics algorithms, ESTIMATE and CIBERSORT, were utilized to analyze the gene expression profiles from 6 datasets, from which the abundance of corresponding TILs was inferred. The ESTIMATE algorithm examined the overall abundance of TILs, whereas the CIBERSORT algorithm reported the relative abundance of 22 different TILs. Both HBV-HCC and HCV-HCC were analyzed., Results: The results indicated that the total abundance of TILs was higher in non-tumor tissue regardless of the HCC type. Alternatively, the specific TILs associated with overall survival (OS) and recurrence-free survival (RFS) varied between subtypes. For example, in HBV-HCC, plasma cells (hazard ratio [HR] = 1.05; 95% CI 1.00-1.10; p = 0.034) and activated dendritic cells (HR = 1.08; 95% CI 1.01-1.17; p = 0.03) were significantly associated with OS, whereas in HCV-HCC, monocytes (HR = 1.21) were significantly associated with OS. Furthermore, for RFS, CD8+ T cells (HR = 0.98) and M0 macrophages (HR = 1.02) were potential biomarkers in HBV-HCC, whereas neutrophils (HR = 1.01) were an independent predictor in HCV-HCC. Lastly, in both HBV-HCC and HCV-HCC, CD8+ T cells (HR = 0.97) and activated dendritic cells (HR = 1.09) had a significant association with OS, while γ delta T cells (HR = 1.04), monocytes (HR = 1.05), M0 macrophages (HR = 1.04), M1 macrophages (HR = 1.02), and activated dendritic cells (HR = 1.15) were highly associated with RFS. Conclusions: These findings demonstrated that TILs are potential survival predictors in HCC and different kinds of TILs are observed according to the virus type. Therefore, further investigations are warranted to elucidate the role of TILs in HCC, which may improve immunotherapy outcomes., Competing Interests: The authors declare no conflict of interest.

Maspin suppresses tumor progression by promoting cell adhesion and apoptosis and by inhibiting cell motility. However, its role in tumorigenesis of hepatocellular carcinoma (HCC) remains unclear. The gene regulation of maspin and its relationship with HCC patient prognosis were investigated in this study. Maspin expression was specifically reduced in HBV-associated patients and correlated with their poor prognosis. Maspin downregulation in HCC cells was induced by HBx to promote their motility and resistance to anoikis and chemotherapy. HBx-dependent induction of microRNA-7, -107, and -21 was further demonstrated to directly target maspin mRNA, leading to its protein downregulation. Higher expressions of these microRNAs also correlated with maspin downregulation in HBV-associated patients, and were associated with their poor overall survival. These data not only provided new insights into the molecular mechanisms of maspin deficiency by HBx, but also indicated that downregulation of maspin by microRNAs confers HBx-mediated aggressiveness and chemoresistance in HCC.

Our previous studies demonstrated that autocrine motility factor/phosphoglucose isomerase (AMF/PGI) possesses tumorigenic activities through the modulation of intracellular signaling. We then investigated the effects of ursolic acid (UA), oleanolic acid (OA), tangeretin, and nobiletin against AMF/PGI-mediated oncogenesis in cultured stable Huh7 and Hep3B cells expressing wild-type or mutated AMF/PGI and in a mouse model in this study. The working concentrations of the tested compounds were lower than their IC10 , which was determined by Brdu incorporation and colony formation assay. Only UA efficiently suppressed the AMF/PGI-induced Huh7 cell migration and MMP-3 secretion. Additionally, UA inhibited the AMF/PGI-mediated protection against TGF-β-induced apoptosis in Hep3B cells, whereas OA, tangeretin, and nobiletin had no effect. In Huh7 cells and tumor tissues, UA disrupted the Src/RhoA/PI 3-kinase signaling and complex formation induced by AMF/PGI. In the Hep3B system, UA dramatically suppressed AMF/PGI-induced anti-apoptotic signaling transmission, including Akt, p85, Bad, and Stat3 phosphorylation. AMF/PGI enhances tumor growth, angiogenesis, and pulmonary metastasis in mice, which is correlated with its enzymatic activity, and critically, UA intraperitoneal injection reduces the tumorigenesis in vivo, enhances apoptosis in tumor tissues and also prolongs mouse survival. Combination of sub-optimal dose of UA and cisplatin, a synergistic tumor cell-killing effects was found. Thus, UA modulates intracellular signaling and might serve as a functional natural compound for preventing or alleviating hepatocellular carcinoma., (© 2012 Wiley Periodicals, Inc.)

Prognosis of hepatocellular carcinoma (HCC) is generally poor. The role of modifiable lifestyle factors on HCC survival has been less studied. To examine whether prediagnosis smoking and alcohol affected HCC survival stratified by viral etiology, we conducted a prospective cohort study of 2,273 (1990 with viral hepatitis and 283 without) incident HCC cases aged 20-75 years who were enrolled between 1997 and 2004 from a Taiwanese multicenter study, and followed up through 2007. Information on habitual smoking and alcohol consumption was obtained at baseline through personal interview. After follow-up to a maximum of 10 years, 1,757 participants died and 1,488 (84.7%) were attributed to HCC. Prediagnosis smoking and alcohol worsened prognosis independent of each other and clinical predictors. The effects of both risky behaviors were limited to viral hepatitis-related HCC and more profound among those with early-stage HCC. Risk for HCC-specific mortality increased with increasing pack-years smoked and ethanol intake (all p < 0.001 for trend), with an additive effect shown for the two habits [hazard ratio (HR) for alcohol ≥ 46.2 g/day and ≥ 10 pack-years = 1.72, 95% confidence interval (CI) = 1.45-2.05]. For either habit, cessation reduced HCC-specific mortality, but a significant mortality benefit occurred 10 years after abstinence (quitting smoking ≥ 10 years vs. continuing smokers: HR = 0.77, 95% CI = 0.61-0.97; quitting drinking ≥ 10 years vs. continuing drinkers: HR = 0.74, 95% CI = 0.56-0.98). In conclusion, among patients with viral hepatitis-related HCC, prediagnosis smoking and alcohol have a deleterious effect on HCC survival. Quitting smoking or drinking alcohol could reduce the excess risk, but only after a long interval of cessation., (Copyright © 2012 UICC.)

This study investigated the potential effects of natural products ursolic acid (UA) and oleanolic acid (OA) against HBx-mediated tumorigenic activities in vitro and in vivo. HBx transactivated Sp-1 and Smad 3/4 in Huh7 and FL83B hepatocytes and induced cell migration of Huh7 and HepG2. HBx also induced MMP-3 secretion in Huh7 and acted against TGF-β-induced apoptosis in Hep3B. UA almost completely blocked the HBx-mediated effects, while OA had a partial inhibitive effect. Utilization of specific MAPK inhibitors and immunoblotting demonstrated that UA selectively activated MAPK signaling in certain tested cells. Preintraperitoneal injection of UA fully prevented the tumor growth of HBV-containing 2.2.15 cells, while OA-treated mice had smaller tumors than the control group. Our results suggested that UA possesses a hepatoprotective ability and illustrated the evident effects against HBx-mediated tumorigenic activities without toxicity in a mouse model.

A region on chromosome 4q25 has recently been highlighted as linked to hepatocellular carcinoma (HCC). In this study, we performed a family-based association analysis with 67 single-nucleotide polymorphisms (SNPs) to map this linkage region in 240 families with HCC, 212 (88.3%) of which were ascertained through hepatitis B virus surface antigen (HBsAg)-positive index cases. Individual SNP analysis with correction for multiple testing identified 10 SNPs in two correlated haplotype blocks, located in or around the 3'-phosphoadenosine 5'-phosphosulfate synthetase-1 (PAPSS1) gene (all P-values: <0.0075). Our linkage data and GIST (Genotype identity-by-descent sharing test) indicate that 6 of these 10 SNPs contributed to the linkage signal. The haplotype block of the strongest association with HCC extended from the intron 5 to the 3'-flanking region of PAPSS1; multiple consecutive three-SNP haplotypes in this region were significant. The most significant haplotype showed odd ratios of 3.41 (95% confidence interval (CI)=1.36-8.53) for homozygous individuals in a case-unaffected sibling analysis. This haplotype also revealed an association with elevated serum alpha-fetoprotein and with poor survival in familial cases and an independent series of HBsAg-positive cases with small tumor present at the time of hospital admission. These results implicate PAPSS1 as a candidate HCC-susceptibility gene in hepatitis B carriers.

Purpose: To determine prospectively whether body-mass index (BMI) is associated with liver-related morbidity and mortality among male hepatitis B virus (HBV) carriers., Patients and Methods: We performed a prospective study of 2,903 male HBV surface antigen-positive government employees who were free of cancer at enrollment between 1989 and 1992. Main outcome measures included ultrasonography, biochemical tests, incident hepatocellular carcinoma (HCC), and liver-related death., Results: During mean follow-up of 14.7 years, 134 developed HCC and 92 died as a result of liver-related causes. In Cox proportional hazards models adjusting for age, number of visits, diabetes, and use of alcohol and tobacco, the hazard ratios for incident HCC were 1.48 (95% CI, 1.04 to 2.12) in overweight men (BMI between 25.0 and 29.9 kg/m(2)) and 1.96 (95% CI, 0.72 to 5.38) in obese men (BMI >or= 30.0 kg/m(2)), compared with normal-weight men (BMI between 18.5 and 24.9 kg/m(2)). Liver-related mortality had adjusted hazard ratios of 1.74 (95% CI, 1.15 to 2.65) in overweight men and 1.50 (95% CI, 0.36 to 6.19) in obese men. Excess BMI was also associated with the occurrence of fatty liver and cirrhosis detected by ultrasonography, as well as elevated ALT and gamma-glutamyltransferase (GGT) activity during follow-up. The association of BMI with GGT was stronger than with ALT, and elevated GGT activity and cirrhosis were the strongest predictors for incident HCC and liver-related death., Conclusion: This longitudinal cohort study indicates that excess body weight is involved in the transition from healthy HBV carrier state to HCC and liver-related death among men.

We have previously shown that AMF/PGI induces hepatoma cell migration through the induction of MMP-3. This work investigates how AMF/PGI activates the MMP-3 gene. We demonstrated that AMF/PGI transactivates the MMP-3 gene promoter through AP-1. The transactivation and induction of cell migration effect of AMF/PGI directly correlates with its enzymatic activity. Various analyses showed that AMF/PGI stimulated the Src-RhoA-PI3-kinase signaling pathway, and these three signaling molecules could form a complex. Our results demonstrate a new mechanism of AMF/PGI-induced cell migration and a link between Src-RhoA-PI3-kinase, AP-1, MMP-3 and hepatoma cell migration.

Background and Aims: To investigate the temporal relationship between sequence variation in the enhancer II (EnhII), basal core promoter (BCP), and precore regions of hepatitis B virus (HBV) and the risk of hepatocellular carcinoma (HCC), we conducted a nested case-control study within a cohort of 4841 male HBV carriers who were recruited during the period 1988-1992., Methods: The HBV DNA sequence was determined in baseline blood samples taken from 132 incident cases and 204 controls. Base exchanges during follow-up in 71 cases were compared with 81 controls with samples taken during a similar length of follow-up., Results: Nine single nucleotide polymorphisms in the EnhII/BCP regions (six of which were genotype C HBV related) were associated with subsequent risk of HCC. The strength of these associations decreased as the lag time between baseline measurement and diagnosis increased over 3 years. However, an increased disease risk in subjects with BCP double variants (mostly T1762/A1764) or genotype C HBV-related variants was evident 9 years or more before diagnosis. The BCP double variants (odds ratio, 1.92 (95% confidence interval, 1.14 to 3.25)) were statistically significantly associated with HCC risk even after adjusting for alanine aminotransferase levels, antibodies against HBV e antigen, HBV genotype, HBV viral load, and other sequence variants. Longitudinal analysis indicated that the increased HCC risks for at-risk sequence variants were attributable to the persistence of these variants., Conclusions: HCC risk is associated with sequence variation in the EnhII/BCP regions of HBV, and persistence of at-risk sequence variants is critical for HCC development.

Little is known about the longitudinal course of hepatitis B virus (HBV) load and its relationship with the development of hepatocellular carcinoma (HCC). We conducted a case-cohort study nested within a cohort of 2874 HBV surface antigen-positive male Taiwanese government employees aged 30 years or older. HBV genotype and DNA levels (i.e. viral load) were tested using polymerase chain reaction-based assays on plasma samples from 112 cases and 1031 non-cases. Prediagnostic plasma levels of HBV DNA were measured in multiple samples collected from each man (total 7706 samples), taken over periods of up to 16 years before diagnosis. Baseline viral load influenced HBV genotype-specific HCC risks and predicted the persistence of high viral load (>/=4.39 log copies/ml) that can cause HCC. Moderate to high tracking of viral load was observed within 9 years. Hepatitis B e antigen (P < 0.0001), genotype C HBV infection (P = 0.0369) and longitudinal alanine aminotransferase (ALT) elevation (defined as ALT abnormality in >/=50% of the visits) (P = 0.0005) were positively related to longer duration of persistence for high viral load. After multivariate adjustment, HBV genotype C [odds ratio (OR) = 5.97, 95% confidence interval (CI) = 3.44-10.34], high viral load detected at >/=50% of the visits (compared with sustained low viral load: OR = 5.04, 95% CI = 2.31-11.00) and longitudinal ALT elevation (compared with sustained normal ALT levels: OR = 2.84, 95% CI = 1.46-5.51) accounted for 43.5, 57.2 and 24.9% of HCCs, respectively. The results suggest that maintenance of viral load <4.39 log copies/ml was associated with sustained normalization of ALT levels and decreased risk of HCC.

Chromosome 4q is one of the most common regions with a high frequency of allelic loss in hepatocellular carcinoma (HCC). To identify the HCC-susceptibility locus on chromosome 4q, we have performed linkage and family-based association analyses on Chinese families with HCC from Taiwan, where hepatitis B is hyperendemic. Using 77 microsatellite markers spanning chromosome 4q on 52 multiplex families, we found suggestive evidence of linkage to 4q22.3-28.1 with a maximum two-point heterogeneity LOD (HLOD) score of 2.55 at marker D4S3240 on chromosome 4q25. Multipoint analyses with microsatellite markers in the region 4q22.3-28.1 resulted in a maximum HLOD score of 3.12 and a maximum nonparametric linkage (NPL) Z score of 1.98 (pointwise P=0.0080; region-wide empirical P=0.021) for D4S3240. The evidence for linkage to D4S3240 was seen mostly in a subset of 28 families lacking affected parents, which showed multipoint HLOD and NPL scores of 3.25 and 2.79 (pointwise P=0.0028; region-wide empirical P=0.008), respectively. Family-based association analyses of the 77 microsatellite markers in 191 families (53 multiplex plus 138 singleton families) using the pedigree disequilibrium test provide further support for observed linkage. Additional genotyping in the 52 multiplex families informative for linkage analyses was performed for 29 single-nucleotide polymorphisms around D4S3240. A common haplotype (at markers rs7442180 and rs221330) positioned approximately 873 kb away from D4S3240 was associated with HCC, with P=0.0074.

Background/aims: Hepatitis B virus (HBV), a major causative agent of hepatocellular carcinoma (HCC), encodes an oncogenic X protein (HBx) that transcriptionally activates multiple viral and cellular promoters. How this regulation influences HCC is unclear., Methods: Global gene expression in HBx-expressing and non-expressing hepatoma cells was analyzed using cDNA microarrays., Results: Genes that were markedly up- or down-regulated in the presence of HBx included those involved in signal transduction, metabolism, apoptosis, cytokine production, cell cycle, cell adhesion and oncogenesis. Other genes of ill-defined function were also affected. Trans-activation proficient HBx up-regulated the transcription, translation and secretion of matrix metalloproteinase-3 (MMP-3), manifest as a cell migratory phenotype. This HBx effect was abrogated in the presence of a MMP-3 specific peptide inhibitor., Conclusions: HBx exerts selective transcriptional control in hepatoma cells and induces cellular migration through the activation of MMP-3.

Background: Although chronic infection with hepatitis B virus (HBV) has been established as a cause of hepatocellular carcinoma (HCC), the roles of viral load and HBV genotype remain unclear., Methods: From 1988 through 1992, baseline blood samples were collected from 4841 Taiwanese men who were HBV carriers but had not been diagnosed with HCC. We used real-time polymerase chain reaction assays of plasma DNA samples to quantify HBV DNA levels (a measure of viral load) and determine HBV genotypes for 154 case patients who were diagnosed with HCC during 14 years of follow-up and 316 control subjects. Unconditional logistic regression was used to assess odds ratios (ORs) of HCC for HBV-related factors. All statistical tests were two-sided., Results: The risk of HCC increased with increasing HBV viral load (adjusted OR for the highest versus the lowest quintile of HBV DNA copies/mL = 7.26, 95% confidence interval [CI] = 3.54 to 14.89; P(trend)<.001). Genotype C HBV was associated with an increased risk of HCC compared with other HBV genotypes (adjusted OR = 5.11, 95% CI = 3.20 to 8.18). Both viral load and genotype were positively associated with HCC within 10-year age categories among subjects aged 30 years old to older than 60 years. Genotype C HBV was associated with increased viral load, and associations of HBV genotype and viral load with HCC risk were additive. The adjusted OR of HCC for those carrying genotype C HBV and with viral load in the highest quintile was 26.49 (95% CI = 10.41 to 67.42) compared with HBV carriers with other HBV genotypes and viral load in the lowest two quintiles., Conclusions: Measurements of HBV viral load and genotype may help to define which male HBV carriers aged 30 years or older are at high risk for HCC.

Phosphoglucose isomerase/autocrine motility factor (PGI/AMF) catalyzes the isomerization between glucose-6-phosphate and fructose-6-phosphate, and is involved in cytokine activity, mitogenesis, differentiation, oncogenesis, and tumor metastasis. Presently, we demonstrate that exogenous PGI/AMF stimulates the migration of Huh7 and HepG2 hepatoma cells, but not Hep3B cells. Inhibition of PGI/AMF by PGI/AMF specific inhibitor 5-phospho-D-arabinonate markedly repressed the cellular migration. RT-PCR was used to examine the expression profile of matrix metalloproteinases (MMPs). MMP-3 transcripts, protein level, and secreted form were significantly upregulated in PGI/AMF-treated Huh7 and HepG2 cells, but not in Hep3B cells. MMP-3 inhibition abolished the PGI/AMF-induced cell motility. The observations are consistent with a downstream mediation role of MMP-3 in PGI/AMF-stimulated tumor cell metastasis.