Protein disulfide isomerase inhibition synergistically enhances the efficacy of sorafenib for hepatocellular carcinoma.

Sorafenib is the only approved targeted drug for hepatocellular carcinoma (HCC), but its effect on patients' survival gain is limited and varies over a wide range depending on pathogenetic conditions. Thus, enhancing the efficacy of sorafenib and finding a reliable predictive biomarker are crucial to achieve efficient control of HCCs. In this study, we utilized a systems approach by combining transcriptome analysis of the mRNA changes in HCC cell lines in response to sorafenib with network analysis to investigate the action and resistance mechanism of sorafenib. Gene list functional enrichment analysis and gene set enrichment analysis revealed that proteotoxic stress and apoptosis modules are activated in the presence of sorafenib. Further analysis of the endoplasmic reticulum stress network model, combined with in vitro experiments, showed that introducing an additional stress by treating the orally active protein disulfide isomerase (PDI) inhibitor (PACMA 31) can synergistically increase the efficacy of sorafenib in vitro and in vivo, which was confirmed using a mouse xenograft model. We also found that HCC patients with high PDI expression show resistance to sorafenib and poor clinical outcomes, compared to the low-PDI-expression group. CONCLUSION: These results suggest that PDI is a promising therapeutic target for enhancing the efficacy of sorafenib and can also be a biomarker for predicting sorafenib responsiveness. (Hepatology 2017;66:855-868).

Targeted Proteomics Predicts a Sustained Complete-Response after Transarterial Chemoembolization and Clinical Outcomes in Patients with Hepatocellular Carcinoma: A Prospective Cohort Study.

This study was aimed to identify blood-based biomarkers to predict a sustained complete response (CR) after transarterial chemoembolization (TACE) using targeted proteomics. Consecutive patients with HCC who had undergone TACE were prospectively enrolled (training (n = 100) and validation set (n = 80)). Serum samples were obtained before and 6 months after TACE. Treatment responses were evaluated using the modified Response Evaluation Criteria in Solid Tumors (mRECIST). In the training set, the MRM-MS assay identified five marker candidate proteins (LRG1, APCS, BCHE, C7, and FCN3). When this five-marker panel was combined with the best-performing clinical variables (tumor number, baseline PIVKA, and baseline AFP), the resulting ensemble model had the highest area under the receiver operating curve (AUROC) value in predicting a sustained CR after TACE in the training and validation sets (0.881 and 0.813, respectively). Furthermore, the ensemble model was an independent predictor of rapid progression (hazard ratio (HR), 2.889; 95% confidence interval (CI), 1.612-5.178; P value < 0.001) and overall an unfavorable survival rate (HR, 1.985; 95% CI, 1.024-3.848; P value = 0.042) in the entire population by multivariate analysis. Targeted proteomics-based ensemble model can predict clinical outcomes after TACE. Therefore, this model can aid in determining the best candidates for TACE and the need for adjuvant therapy.

Ultrasound-guided percutaneous portal transplantation of peripheral blood monocytes in patients with liver cirrhosis.

BACKGROUND/AIMS: Liver transplantation offers the only definite cure for cirrhosis but lacking donors is problem. Stem cell therapy is attractive in this setting. In this study, we aimed to explore the safety and efficacy of ultrasound-guided percutaneous portal transplantation of peripheral blood monocyte cell (PBMC) in cirrhotic patients. METHODS: A total of nine decompensated cirrhotic patients were randomized into three groups: group 1 (n = 3) was control group, group 2 (n = 3) received granulocyte-colony stimulating factor (G-CSF) mobilization for 3 days, and group 3 (n = 3) received G-CSF mobilized PBMCs by leukapheresis and PBMC transplantation through ultrasound-guided percutaneous portal vein puncture. Liver function and clinical features were evaluated. RESULTS: At baseline, the Child-Turcotte-Pugh and the model for end-stage liver disease scores were comparable in study groups. Compared with group 1, there was a tendency to improve liver function in group 3 at 6 months after treatment. Treatment was tolerable and no complications were encountered related to the G-CSF mobilization or percutaneous portal administration of PBMCs. Imaging studies showed patent portal veins at the end of the study period. CONCLUSIONS: Autologous PBMC transplantation through ultrasound-guided percutaneous portal vein puncture could be considered as a safe alternative treatment for decompensated cirrhotic patients.

Differential sensitivity of hepatocellular carcinoma cells to suppression of hepatocystin transcription under hypoxic conditions.

Mutations in the gene encoding hepatocystin/80 K-H (PRKCSH) cause autosomal dominant polycystic liver disease. Hepatocystin deficiency impairs glucosidase II activity, which is critical for processing and folding glycoproteins in the endoplasmic reticulum (ER). Hypoxia is known as a strong stimulus for generating survival signals in hepatocellular carcinoma (HCC) cells. However, hypoxia may induce cell apoptosis under conditions of severe ER stress. Thus, we hypothesized that suppression of hepatocystin transcription induces HCC cell death under hypoxic conditions due to excessive ER stress. A new human HCC cell line, SNU-3058, was established following primary culture of tumor cells harvested from a Korean patient with rapidly growing hypovascular HCC. In cell culture, human HCC cells (Huh-7, SNU-761, and SNU-3058) were treated with control siRNA or hepatocystin siRNA with or without doxorubicin under hypoxic conditions. Cell viability, ER stress, unfolded protein response (UPR), and apoptosis were assessed using the MTS assay, immunoblot assay, and RT-PCR. Suppression of hepatocystin transcription attenuated proliferation in Huh-7 and SNU-761 cells, while proliferation was amplified in SNU-3058 cells. Similar results were observed following treatment with doxorubicin. Hepatocystin siRNA transfection increased cell death in Huh-7 and decreased cell death in SNU-3058. In SNU-3058, hepatocystin siRNA amplified GRP78, known as a pro-survival and cyto-protective signal, and attenuated the pro-apoptotic signal CHOP. These findings suggest that suppression of hepatocystin transcription induce the UPR, which alleviates damage associated with ER stress in SNU-3058. UPR had a limited role in protecting SNU-761 cells, resulting in cell death through apoptosis. In addition, blocking of pro-survival UPR signal by bacitracin or GRP78 knockdown, attenuated hepatocystin siRNA-induced proliferation in SNU-3058 cells under hypoxia. In this study, we demonstrated that different sensitivities to hepatocystin siRNA among human HCC cell lines are dependent on appropriate UPRs to hypoxia-induced ER stress following hepatocystin siRNA transfection. Because UPR is the main evasive mechanism for apoptosis induced by suppression of hepatocystin, targeting hepatocystin via UPR suppression could be a strategy for treating HCC.

Percutaneous ethanol injection therapy is comparable to radiofrequency ablation in hepatocellular carcinoma smaller than 1.5 cm: A matched case-control comparative analysis.

Although percutaneous ethanol injection therapy (PEIT) is best indicated for patients with small hepatocellular carcinoma (HCC), the survival advantage of PEIT needs confirmation in real-world practice. This study was approved by the institutional review board, and the informed consent was waived. The study included 535 consecutive patients with newly diagnosed early stage (Barcelona Clinic Liver Cancer [BCLC] 0 or A) HCC who underwent initially radiofrequency ablation (RFA) (n = 288) or PEIT (n = 247) from January 2005 to December 2010. The primary outcome was overall survival (OS) and the secondary outcome was time to progression (TTP). The longest diameters of tumors of the groups differed significantly and larger for RFA group than PEIT group (P < 0.001; 1.94 ±â€Š0.65 cm vs 1.60 ±â€Š0.50 cm, respectively). The 5-year OS rates were 72.2% in the RFA group and 67.4% in the PEIT group (P = 0.608). Even after propensity score matching, OS rates between the 2 groups were similar (5-year OS: 72.8% with RFA [n = 175] and 68.0% with PEIT [n = 175]) (P = 0.709). Moreover, in patients with the longest diameter of tumors (≤1.5 cm), multivariate Cox regression analysis showed that the treatment modality was not a significant prognosticator for OS (hazard ratio [HR], 1.690; 95% confidence interval [CI], 0.828-3.449; P = 0.149) and time to progression (HR, 1.160; 95% CI, 0.773-1.740; P = 0.474). PEIT and RFA show equal effectiveness in treating HCCs <1.5 cm in terms of OS and time to progression.

Fucoidan-induced ID-1 suppression inhibits the in vitro and in vivo invasion of hepatocellular carcinoma cells.

Hepatocellular carcinoma (HCC) is a fast growing tumor associated with a high tendency for vascular invasion and distant metastasis. Recently, we reported that fucoidan displays inhibitory effect on proliferation and invasion of HCC cells. In this study, we investigated the anti-metastatic effect of fucoidan on HCC cells and the key signal that modulates metastasis. The anti-metastatic effect of fucoidan was evaluated in vitro using an invasion assay with human HCC cells (Huh-7, SNU-761, and SNU-3085) under both normoxic (20% O2 and 5% CO2, at 37°C) and hypoxic (1% O2, 5% CO2, and 94% N2, at 37°C) conditions. Complementary DNA (cDNA) microarray analysis was performed to find the molecule which is significantly suppressed by fucoidan. In vivo study using a distant metastasis model by injecting SNU-761 cells into spleen via portal vein was performed to confirm the inhibitory effect by small interfering RNA (siRNA) transfection. Immunoblot analyses were used to investigate the signaling pathway. Fucoidan significantly suppressed the invasion of human HCC cells (Huh-7, SNU-761, and SNU-3085). Using cDNA microarray analysis, we found the molecule, ID-1, which was significantly suppressed by fucoidan treatment. Downregulation of ID-1 by siRNA significantly decreased invasion of HCC cells, both in vitro and in vivo (both P<0.05) in a NDRG-1/CAP43-dependent manner. In immunoblot assay, downregulation of ID-1 by siRNA decreased the expressions of epithelial-mesenchymal transition markers including CK19, vimentin, MMP2, and fibronectin. Immunofluorescence study also revealed that actin rearrangement was inhibited when ID-1 was down-regulated in HCC cells. Interestingly, in SNU-761 cells, the ID-1 expressions under hypoxic conditions were lower as compared to those under normoxic conditions. Under hypoxic conditions, HIF-1α up-regulated NDRG-1/CAP43, while HIF-2α down-regulated ID-1, which might be a compensatory phenomenon against hypoxia-induced HCC invasion. In conclusion, NDRG-1/CAP43-dependent down-regulation of ID-1 suppressed HCC invasion both in vitro and in vivo, which was modulated by fucoidan treatment. Moreover, the compensatory down-regulation of ID-1 against hypoxia-induced HCC invasion was observed. ID-1 is a novel therapeutic target for the treatment of metastatic HCC.

Hypoxia Enhances Tumor-Stroma Crosstalk that Drives the Progression of Hepatocellular Carcinoma.

BACKGROUND: Crosstalk between tumor cells and their microenvironment plays a crucial role in the progression of hepatocellular carcinoma (HCC). Hypoxia, a common feature of advanced HCC, has been shown to modulate the evolution of the tumor microenvironment. In this study, we investigated the effect of hypoxia on tumor-stroma crosstalk in HCC. METHODS: Human HCC cell lines (Huh-BAT, SNU-475) were cocultured with an activated human hepatic stellate cell line (HSCs; LX-2) under either normoxic or hypoxic conditions. Cell growth was evaluated with the MTS assay. Apoptotic signaling cascades were assessed by immunoblot analysis. Expression of CD31 and phosphorylated (p-) Akt in HCC tissues was detected by immunohistochemistry. RESULTS: Coculturing HCC cells with HSCs under hypoxic conditions enhanced their proliferation, migration, and resistance to bile acid (BA)-induced apoptosis compared to coculturing under normoxic conditions. Under hypoxia, of various HSC-derived growth factors, PDGF-BB was the most up-regulated, leading to the activation of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway in HCC cells. Immunohistochemical study also revealed that p-Akt was highly expressed in hypoxic, hypovascular HCC as compared to hypervascular HCC. Neutralizing antisera to PDGF-BB or a PI3K inhibitor attenuated the proliferation of HCC cells cocultured with HSCs, and sensitized HCC cells to BA-induced apoptosis, especially under hypoxic conditions. CONCLUSIONS: In conclusion, hypoxic HSC-derived PDGF-BB stimulates the proliferation of HCC cells through activation of the PI3K/Akt pathway, while the inhibition of PDGF-BB or PI3K/Akt pathways enhances apoptotic cell death. Targeting tumor-stroma crosstalk might be a novel therapy in the management of human HCCs.

Body Fat Distribution and Risk of Incident and Regressed Nonalcoholic Fatty Liver Disease.

BACKGROUND & AIMS: Some studies have examined correlations between visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT) with nonalcoholic fatty liver disease (NAFLD) or between VAT and NAFLD. We investigated the longitudinal association between body fat distribution (VAT vs SAT) and incidence and regression of NAFLD, adjusting for risk factors, in a large population-based cohort. METHODS: We collected data from adults who underwent abdominal ultrasonography (to identify liver fat), abdominal fat computed tomography scan, and blood tests from March 2007 through December 2008. Each patient underwent an anthropometric assessment and completed a questionnaire about their medical history, physical activity, and diet. Our final analysis involved 2017 subjects from the initial cohort who participated in a voluntary follow-up health screen performed in 2011 and 2013. The median follow-up time was 4.43 years. RESULTS: We found 288 incident cases of NAFLD; 159 patients had NAFLD regression during the follow-up period. An increasing area of VAT was associated with higher incidence of NAFLD in the multivariable analysis (highest quintile vs lowest quintile of VAT hazard ratio [HR], 2.23; 95% confidence interval [CI], 1.28-3.89; P for trend = .002; HR, 1.36 [per 1 standard deviation]; 95% CI, 1.16-1.59). An increased area of SAT was significantly associated with regression of NAFLD (highest quintile vs lowest quintile of SAT HR, 2.30; 95% CI, 1.28-4.12; P for trend = .002; HR, 1.36 [per 1 standard deviation]; 95% CI, 1.08-1.72). CONCLUSIONS: In a large cohort study, larger areas of VAT were longitudinally associated with higher risk of incident NAFLD (during a period of approximately 4 years). In contrast, larger areas of SAT were longitudinally associated with regression of NAFLD. These data indicate that certain types of body fat are risk factors for NAFLD, whereas other types could reduce risk for NAFLD.

Fucoidan protects hepatocytes from apoptosis and inhibits invasion of hepatocellular carcinoma by up-regulating p42/44 MAPK-dependent NDRG-1/CAP43.

Fucoidan is a traditional Chinese medicine suggested to possess anti-tumor effects. In this study the anti-metastatic effects of fucoidan were investigated in vitro in human hepatocellular carcinoma (HCC) cells (Huh-7 and SNU-761) under normoxic and hypoxic conditions and in vivo using a distant liver metastasis model involving injection of MH134 cells into spleen via the portal vein. Its ability to protect hepatocytes against bile acid (BA)-induced apoptosis was investigated in primary hepatocytes. Fucoidan was found to suppress the invasion of HCC cells through up-regulation of p42/44 MAPK-dependent NDRG-1/CAP43 and partly, under normoxic conditions, through up-regulation of p42/44 MAPK-dependent VMP-1 expression. It also significantly decreased liver metastasis in vivo. As regards its hepatoprotective effect, fucoidan decreased BA-induced hepatocyte apoptosis as shown by the attenuation of caspase-8, and -7 cleavages and suppression of the mobilization of caspase-8 and Fas associated death domain (FADD) into the death-inducing signaling complex. In summary, fucoidan displays inhibitory effects on proliferation of HCC cells and protective effects on hepatocytes. The results suggest fucoidan is a potent suppressor of tumor invasion with hepatoprotective effects.

Visceral Obesity Predicts Significant Fibrosis in Patients With Nonalcoholic Fatty Liver Disease.

Nonalcoholic fatty liver disease (NAFLD) is associated with visceral obesity. However, the association between visceral adipose tissue (VAT) area and fibrosis in NAFLD patients has not been completely established. This study was aimed to determine the relationship between the computed tomography-measured VAT area and significant fibrosis in NAFLD patients. A total of 324 NAFLD patients and 132 controls were evaluated by liver biopsy. NAFLD was diagnosed based on histological examinations and alcohol consumption <20 g/day. The NAFLD patients showed a higher age and gender-adjusted VAT area than the control group (86.1 ± 2.3 vs 56.7 ± 3.7, P < 0.001). The VAT area increased across the control, NAFLD without significant fibrosis, and NAFLD with significant fibrosis groups (54.9 ± 3.5, 80.6 ± 2.4, and 123.4 ± 6.4, P < 0.001). This association persisted after adjusting for multiple confounders (P for trend = 0.028). A multivariate regression analysis demonstrated the VAT area was independently associated with NAFLD with significant fibrosis (F2-F4) (odds ratio [OR] 1.21 95% confidence interval [CI] 1.07-1.37 per 10 cm(2) increase of VAT area; OR 2.62 [per 1 - standard deviation (SD)] 95% CI 1.41-4.86). Moreover, a multivariate logistic regression analysis revealed the VAT area was independently associated with nonalcoholic steatohepatitis (NASH) in NAFLD (OR 1.17 95% CI 1.05-1.32 per 10 cm increase of VAT area; OR 2.21 [per 1 - SD] 95% CI 1.25-3.89). Increased VAT area is independently associated with NASH or significant fibrosis and VAT might be a central target for lifestyle modifications in NAFLD patients.

Hepatoprotective effect of vitamin C on lithocholic acid-induced cholestatic liver injury in Gulo(-/-) mice.

Prevention and restoration of hepatic fibrosis from chronic liver injury is essential for the treatment of patients with chronic liver diseases. Vitamin C is known to have hepatoprotective effects, but their underlying mechanisms are unclear, especially those associated with hepatic fibrosis. Here, we analyzed the impact of vitamin C on bile acid induced hepatocyte apoptosis in vitro and lithocholic acid (LCA)-induced liver injury in vitamin C-insufficient Gulo(-/-) mice, which cannot synthesize vitamin C similarly to humans. When Huh-BAT cells were treated with bile acid, apoptosis was induced by endoplasmic reticulum stress-related JNK activation but vitamin C attenuated bile acid-induced hepatocyte apoptosis in vitro. In our in vivo experiments, LCA feeding increased plasma marker of cholestasis and resulted in more extensive liver damage and hepatic fibrosis by more prominent apoptotic cell death and recruiting more intrahepatic inflammatory CD11b(+) cells in the liver of vitamin C-insufficient Gulo(-/-) mice compared to wild type mice which have minimal hepatic fibrosis. However, when vitamin C was supplemented to vitamin C-insufficient Gulo(-/-) mice, hepatic fibrosis was significantly attenuated in the liver of vitamin C-sufficient Gulo(-/-) mice like in wild type mice and this hepatoprotective effect of vitamin C was thought to be associated with both decreased hepatic apoptosis and necrosis. These results suggested that vitamin C had hepatoprotective effect against cholestatic liver injury.

Synergistic anti-tumor efficacy of doxorubicin and flavopiridol in an in vivo hepatocellular carcinoma model.

PURPOSE: A previous study showed that flavopiridol increased doxorubicin sensitivity in hypoxic hepatocellular carcinoma (HCC) cells by increasing apoptosis through suppressing hypoxia-inducible N-myc downstream-regulated gene-1 (NDRG1) expression. However, this has not been investigated in an in vivo HCC model. Therefore, we aimed to elucidate whether the combination of doxorubicin and flavopiridol has a synergistic anti-tumor effect in an in vivo HCC model. METHODS: An HCC mouse model was established by implanting C3H/He mouse with MH134 cells. Then, doxorubicin with or without flavopiridol was injected. The anti-tumor efficacy was assessed by evaluating tumor volumes, and the underlying mechanism was investigated by quantifying apoptotic cells, the Ki-67 proliferation index, and microvessel densities (MVDs). Immunohistochemistry of NDRG1 was performed to determine the underlying mechanism. RESULTS: Tumor growth was significantly suppressed in the doxorubicin + flavopiridol combination group compared to the other three groups. The percentage of apoptotic cells was significantly higher, and Ki-67-positive proliferating cells were significantly lower in the combination group compared to the other groups; however, MVDs were not significantly different across the groups. Increased apoptosis by flavopiridol occurred by suppressing hypoxia-inducible NDRG1 expression. CONCLUSIONS: These results show that a combination of doxorubicin and flavopiridol has a synergistic anti-tumor effect in an in vivo HCC model. This synergistic effect of combination therapy was attributed to increased apoptosis and decreased proliferation of tumor cells rather than decreased angiogenesis. These findings suggest that flavopiridol might be an effective adjuvant therapy to doxorubicin-resistant HCC cells by inducing apoptosis through suppression of NDRG1 expression.

Forns index predicts recurrence and death in patients with hepatitis B-related hepatocellular carcinoma after curative resection.

BACKGROUND & AIMS: Advanced liver fibrosis is associated with recurrence after curative resection of hepatocellular carcinoma (HCC). This study aimed to investigate whether noninvasive fibrosis indices could predict intrahepatic recurrence and death after curative resection of HCC. METHODS: Patients who underwent curative resection for hepatitis B virus (HBV)-related HCC between 2006 and 2010 at a single tertiary hospital were included. This study analysed the association of noninvasive fibrosis indices with recurrence and overall survival. RESULTS: A total of 303 patients were included. During a median follow-up period of 56.0 (interquartile range, 42.0-70.0) months, 151 (49.8%) patients experienced HCC recurrence and 54 (17.8%) died. Based on multivariate analysis, Forns index [hazard ratio (HR), 1.238; 95% confidence interval (CI), 1.097-1.398; P = 0.001] was independently associated with tumour recurrence after adjustment for anti-HBe positivity, histological cirrhosis, tumour size and number. Patients with Forns index <6.9 had a significantly longer recurrence-free survival rate than patients with Forns index ≥6.9 (P < 0.001 by log-rank test). Forns index (HR, 1.246; 95% CI, 1.034-1.501; P = 0.02) could also predict overall survival after adjustment for tumour size and number. Forns index detected cirrhosis with an AUROC of 0.700 (95% CI, 0.641-0.758). Aspartate aminotransferase-to-platelet ratio index, cirrhosis discriminant score, FIB-4 index, P2/MS and Lok index detected cirrhosis comparably to Forns index, but were not associated with tumour recurrence or death. CONCLUSIONS: Our data suggest that Forns index could be a useful predictor of recurrence and overall survival after curative resection of HBV-related HCC.

Tenofovir monotherapy versus tenofovir plus lamivudine or telbivudine combination therapy in treatment of lamivudine-resistant chronic hepatitis B.

Tenofovir disoproxil fumarate (TDF) monotherapy is a therapeutic option for chronic hepatitis B (CHB) patients infected with hepatitis B virus (HBV) variants resistant to lamivudine (LAM). We evaluated the antiviral efficacy and safety of TDF alone compared to those of TDF plus LAM or telbivudine (LdT) combination in patients harboring HBV variants with genotypic resistance to LAM. This multicenter retrospective study included consecutive patients who had LAM-resistant HBV variants and were treated with TDF alone (monotherapy group) or TDF combined with LAM or LdT (combination therapy group) for at least 6 months. Inverse probability of treatment weighting (IPTW) for the entire cohort was applied to control for treatment selection bias. Overall, 153 patients (33 in the monotherapy group and 120 in the combination therapy group) were analyzed. The overall probability of achieving complete virologic suppression at month 12 was 91.6%: 88.6% in the monotherapy group and 92.6% in the combination therapy group. Combination therapy was not superior to monotherapy in viral suppression before and after IPTW (P=0.562 and P=0.194, respectively). Hepatitis B e antigen (HBeAg) loss, biochemical response, and virologic breakthrough did not differ between treatment groups. The probabilities of complete virologic suppression were comparable between treatment groups in the subsets according to HBeAg status and HBV DNA levels at baseline. No patient experienced any significant renal dysfunction during the treatment period. In conclusion, TDF monotherapy has antiviral efficacy comparable to that of TDF plus LAM or LdT combination therapy, with a favorable safety profile in CHB patients with LAM-resistant HBV variants.

Comparison of the effects of transarterial chemoembolization for advanced hepatocellular carcinoma between patients with and without extrahepatic metastases.

BACKGROUND/AIMS: Sorafenib is a standard treatment for advanced hepatocellular carcinoma (HCC) (Barcelona Clinic Liver Cancer [BCLC] stage C). However, transarterial chemoembolization (TACE) has also been widely used as a treatment for patients with advanced HCC, even if they have extrahepatic metastases (EHM). The aim of this study was to determine the efficacy of TACE for advanced HCC patients with EHM upon initial diagnosis, as compared with those patients without EHM. METHODS: This cohort study involved consecutive patients who underwent TACE as an initial treatment for advanced HCC. One hundred seventy-seven patients with EHM (the EHM group) and 205 with portal vein invasion without EHM (the non-EHM group) were included. A survival analysis was performed to compare overall survival between the two groups. RESULTS: The mean age was 54.5±9.9 years, and median follow-up duration was 13.1 months (range, 0.5-111.0). Overall survival was significantly shorter in the EHM group than the non-EHM group (median, 8.3 vs. 19.1 months; P<0.001). A multivariate analysis showed that the presence of EHM was an independent poor prognostic factor for shorter overall survival (adjusted hazard ratio, 1.74; 95% confidence interval, 1.39-2.17; P<0.001) after adjustment for Child-Pugh classification, intrahepatic tumor T classification, tumor response to TACE, and serum alpha-fetoprotein level. Patients administered TACE and systemic therapy demonstrated a better survival rate than those administered TACE alone in both the EHM (median, 13.5 vs. 7.2 months) and non-EHM groups (median, 27.9 vs. 18.2 months) (both, P<0.05). CONCLUSIONS: The prognosis of advanced HCC patients with EHM is significantly worse than those without EHM administered repeated TACE treatments, even if their tumor stage was similar to BCLC stage C. These results suggest that EHM presence means aggressive tumor biology and that BCLC stage C might be subclassified according to EHM presence.

A novel prognostic factor for hepatocellular carcinoma: protein disulfide isomerase.

BACKGROUND/AIMS: Protein disulfide isomerase (PDI) has been implicated in the survival and progression of some cancer cells, by compensating for endoplasmic reticulum stress by upregulating the protein-folding capacity. However, its prognostic role in patients with hepatocellular carcinoma (HCC) has not been investigated. METHODS: We collected HCC tissues from 83 HCC patients who underwent surgical resection for an immunohistochemical study of PDI. Overall survival (OS) was measured from the date of surgical resection until the date of death from any cause. Radiological progression was evaluated using the modified Response Evaluation Criteria in Solid Tumors in an independent radiological assessment. RESULTS: PDI expression was found to be increased in human HCC compared to adjacent nontumor tissues. Increased immunopositivity for PDI was associated with a high Edmondson-Steiner grade (p = 0.028). Univariate analysis of patients who had undergone surgical resection for HCC showed that tumor PDI upregulation is a significant risk factor for poor OS (p = 0.016; hazard ratio [HR], 1.980) and time to progression (TTP; p = 0.007; HR, 1.971). Multivariate analyses revealed that high PDI expression was an independent predictor of a shorter TTP (p = 0.015; HR, 1.865) and poor OS (p = 0.012; HR, 2.069). CONCLUSIONS: Upregulated PDI expression is associated with aggressive clinicopathological features of HCC; thus, PDI might serve as an independent prognostic factor and a potential therapeutic target for HCC patients.

Efficacy of entecavir-tenofovir combination therapy for chronic hepatitis B patients with multidrug-resistant strains.

The emergence of multidrug-resistant (MDR) strains of hepatitis B virus (HBV) is a major concern. This study aimed to investigate the efficacy and safety of combination therapy with entecavir (ETV) plus tenofovir disoproxil fumarate (TDF) against MDR HBV. To adjust for differences in baseline characteristics, inverse probability weighting (IPW) using propensity scores for the entire cohort and weighted Cox proportional hazards models were applied. Ninety-three consecutive patients who were treated with ETV-TDF combination therapy for >6 months were included; at baseline, 45 were infected with HBV strains with genotypic resistance to lamivudine (LAM) and ETV (the LAM/ETV-R group), 28 with strains resistant to LAM and adefovir (ADV) (the LAM/ADV-R group), and 20 with strains resistant to LAM, ETV, and ADV (the LAM/ETV/ADV-R group). The median duration of rescue therapy was 13.0 (range, 6.7 to 31.7) months. Seventy-four of 93 patients (79.6%) achieved complete virologic suppression, after a median of 4.5 (95% confidence interval, 3.0 to 6.0) months. The cumulative probability of complete virologic suppression at month 6 was 63.6% (55.7%, 75.0%, and 65.0% in the LAM/ETV-R, LAM/ADV-R, and LAM/ETV/ADV-R groups, respectively). During the treatment period, these probabilities were not significantly different across the resistance profiles before and after IPW (P = 0.072 and P = 0.510, respectively). In multivariate analysis, a lower baseline HBV DNA level, but not resistance profiles, was an independent predictor of complete virologic suppression. Renal dysfunction was not observed during the treatment period. In conclusion, rescue therapy with ETV-TDF combination is efficient and safe in patients infected with MDR HBV strains regardless of the antiviral drug resistance profiles.

Tauroursodeoxycholic acid attenuates progression of steatohepatitis in mice fed a methionine-choline-deficient diet.

BACKGROUND AND AIM: Endoplasmic reticulum (ER) stress has been implicated in the development of nonalcoholic steatohepatitis. A methionine-choline-deficient (MCD) diet induces robust ER stress response and steatohepatitis, but the effects of ER stress modulation on the course of steatohepatitis remain uncertain. The present study evaluated whether reducing ER stress using the chemical chaperone tauroursodeoxycholic acid (TUDCA) could limit hepatocyte lipoapoptosis and progression of MCD diet-induced steatohepatitis. METHODS: HuH7 cells stably transfected with sodium taurocholate cotransporting polypeptide (HuH-Ntcp cells) and palmitate (PA) were used. Experimental steatohepatitis was induced in male C57BL/6 mice using an MCD diet, and three different doses of TUDCA (500, or 1,000 mg/kg, once daily; or 500 mg/kg twice daily) were administered by gavage from the start of the MCD diet regimen or after 4 weeks. RESULTS: TUDCA reduced PA-induced ER stress as manifested by decreased eIF2α phosphorylation, XBP1 splicing and expression of BiP, ATF4, and CHOP in HuH-Ntcp cells. TUDCA also decreased PA-induced JNK phosphorylation, Puma up-regulation and Bax activation, which in turn suppressed caspase-dependent hepatocyte lipoapoptosis. Mice given TUDCA did not show a significant decrease in the intrahepatic triglyceride contents and steatosis. However, TUDCA treatment significantly reduced hepatic damage compared to controls for both early and late treatment groups. TUDCA treatment reduced the expression of ER stress markers and pro-apoptotic proteins, leading to decreased apoptosis and oxidative stress. Finally, TUDCA reduced histological fibrosis along with the down-regulation of pro-fibrotic gene expression in both early and late treatment groups. CONCLUSIONS: These results show that TUDCA attenuates the progression of MCD diet-induced steatohepatitis by reducing ER stress.

Serum insulin-like growth factor-1 predicts disease progression and survival in patients with hepatocellular carcinoma who undergo transarterial chemoembolization.

Insulin like-growth factor-1 (IGF-1) reflects hepatic synthetic function and plays a major role in the development and progression of various cancers. In the present study, we investigated whether baseline serum IGF-1 levels predict time-to-progression (TTP) and overall survival (OS) in hepatocellular carcinoma (HCC) patients treated with transarterial chemoembolization (TACE). A total of 155 consecutive treatment-naive patients with HCC who had undergone TACE as initial treatment were included from a prospective cohort. Baseline serum IGF-1 levels were analyzed with regard to their associations with disease progression and survival. During a median follow-up period of 41.8 months, patients with low IGF-1 levels showed significantly shorter TTP (median, 6.0 months; 95% confidence interval [CI], 4.5-7.6) than patients with high IGF-1 levels (median, 16.5 months; 95% CI, 4.9-28.1; p = 0.003). In the multivariate analysis, BCLC stage, serum vascular endothelial growth factorlevels, and IGF-1 levels were independent risk factors for disease progression. The hazard ratio (HR) of progression for each 10 ng/mL decrease in IGF-1 level was 1.072 (95% CI, 1.029-1.117; p = 0.001). Furthermore, together with tumor size, stage, and treatment response, IGF-1 levels were an independent predictor of poorer survival (for each 10 ng/mL decrease in IGF-1 level; HR, 1.057; 95% CI, 1.001-1.115; p = 0.045). In conclusion, low baseline IGF-1 levels independently correlated with shorter TTP and poorer OS in patients with HCC who underwent TACE.

Nonalcoholic fatty liver disease is a negative risk factor for prostate cancer recurrence.

Nonalcoholic fatty liver disease (NAFLD) is closely related to the metabolic syndrome, which is associated with an increased risk of various malignancies. In this study, we investigated the association between NAFLD and prostate cancer biochemical recurrence (BCR) after radical prostatectomy. Consecutive prostate cancer patients who underwent radical prostatectomy were enrolled from two hospitals in Korea and randomly assigned to the training (n=147) or validation set (n=146). The presence of NAFLD, BMI, preoperative prostate-specific antigen, and histological findings including Gleason score (GSc) were analyzed in regard to their association with BCR. NAFLD was diagnosed based on ultrasonography or unenhanced computed tomography images. BCR-free survival rates were calculated using the Kaplan-Meier method. In the training set, 32 (21.8%) patients developed BCR during a median follow-up period of 51 (inter-quartile range, 35-65) months. In the multivariate analysis, the presence of NAFLD (hazard ratio (HR), 0.36; 95% CI, 0.14-0.97; P=0.04) was an independent negative predictive factor of BCR after adjustment for pathological GSc. Applied to the validation set, the presence of NAFLD maintained its prognostic value for longer time-to-BCR (HR, 0.17; 95% CI, 0.06-0.49; P=0.001). In the subgroup analysis of patients with NAFLD, NAFLD fibrosis score was a single independent negative predictor for BCR (HR, 0.54; 95% CI, 0.30-0.98; P=0.04). Our study demonstrated that NAFLD may play a protective role against BCR after radical prostatectomy for prostate cancer. Further study is warranted to elucidate the mechanism of protective effect in patients with NAFLD.