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Background and Aims: Molecular processes driving immune-active chronic hepatitis B (CHB) with and without hepatitis B e antigen (HBeAg) remain incompletely understood. This study aimed to investigate expression profiles of serum and intrahepatic HBV markers and replicative activity of HBV in CHB patients with or without HBeAg., Methods: This study recruited 111 untreated immune-active CHB (60 HBeAg-positive and 51 HBeAg-negative) patients and quantified intrahepatic covalently closed circular DNA (cccDNA), pre-genomic RNA (pgRNA), total HBV DNA (tDNA), and replicative intermediates as well as serum HBV markers (HBV DNA, hepatitis B surface antigen, hepatitis B core-related antigen). Correlations between HBV markers and clinico-virological factors influencing expression levels of HBV markers were analysed., Results: Levels of all serum markers and intrahepatic cccDNA/tDNA as well as cccDNA transcriptional activity and virion productivity were significantly reduced in HBeAg-negative patients compared to those in HBeAg-positive patients. Additionally, correlations between intrahepatic cccDNA/pgRNA and serum markers were impaired in HBeAg-negative individuals. Aminotransferase levels were positively correlated with cccDNA transcriptional activity in HBeAg-positive patients, but not in HBeAg-negative patients. Notably, among HBeAg-positive patients, there was a progressive decline in pgRNA level, transcriptional activity, and serum HBV markers as liver fibrosis advanced, which was not observed in HBeAg-negative patients., Conclusions: HBeAg loss is correlated with diminished intrahepatic HBV reservoirs and cccDNA transcription, leading to decreased serum HBV marker levels. Circulating HBV markers are not reliable indicators of intrahepatic HBV replicative activity for HBeAg-negative patients. Our findings reveal distinct disease phenotypes between immune-active CHB with and without HBeAg, highlighting the need to establish optimal surrogate biomarkers that can accurately mirror intrahepatic viral activity to aid in decision-making for antiviral therapy for immune-active CHB., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background & Aims: Nucleos(t)ide analogue (NA) therapy in chronic hepatitis B (CHB) patients reduces liver-related mortality. However, long-term outcomes after pegylated interferon (PEG-IFN) therapy remain to be elucidated. Therefore, we aimed to investigate the long-term effectiveness and clinical outcomes of PEG-IFN therapy., Methods: A total of 190 patients treated with PEG-IFN for CHB or compensated cirrhosis were consecutively enrolled between 2005 and 2014, and 122 patients who completed the treatment were analysed. The initial response was assessed at 6 months post-treatment and defined as achieving both <2000 IU/mL HBV DNA and HBeAg loss or seroconversion in the HBeAg-positive group, and <2000 IU/mL HBV DNA in the HBeAg-negative group. The rates of HBsAg loss, disease progression to cirrhosis or HCC, and sustained off-therapy response, defined as not requiring further NAs because of low viremia and liver enzymes, were analysed., Results: The median follow-up period was 7.2 years. Forty-three (35.2%) patients achieved an initial response and 53 patients (43.4%) achieved a sustained response. Initial responders displayed higher rates of sustained response than noninitial responders (69.6% vs 32.5%, P < .001). A higher rate of HBsAg loss was observed in patients who achieved a sustained response than in non-sustained responders (16.2% vs 2.5%, P = .01). Disease progression to cirrhosis or HCC was observed in eight patients (6.6%) who were nonsustained responders., Conclusions: During long-term follow-up after PEG-IFN treatment, nearly half of patients achieved sustained response without the need of further NA and these patients displayed favourable outcomes, including HBsAg loss and no disease progression., (© 2020 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: The Gut and Obesity Asia (GO ASIA) workgroup was formed to study the relationships between obesity and gastrointestinal diseases in the Asia Pacific region., Aim: To study factors associated with nonalcoholic steatohepatitis (NASH) and advanced fibrosis, and medical treatment of biopsy-proven nonalcoholic fatty liver disease (NAFLD) patients., Methods: Retrospective study of biopsy-proven NAFLD patients from centres in the GO ASIA Workgroup. Independent factors associated with NASH and with advanced fibrosis on binary logistic regression analyses in a training cohort were used for the development of their corresponding risk score, which were validated in a validation cohort., Results: We included 1008 patients from nine centres across eight countries (NASH 62.9%, advanced fibrosis 17.2%). Independent predictors of NASH were body mass index ≥30 kg/m 2 , diabetes mellitus, dyslipidaemia, alanine aminotransferase ≥88 U/L and aspartate aminotransferase ≥38 U/L, constituting the Asia Pacific NASH risk score. A high score has a positive predictive value of 80%-83% for NASH. Independent predictors of advanced fibrosis were age ≥55 years, diabetes mellitus and platelet count <150 × 10 9 /L, constituting the Asia-Pacific NAFLD advanced fibrosis risk score. A low score has a negative predictive value of 95%-96% for advanced fibrosis. Only 1.7% of patients were referred for structured lifestyle program, 4.2% were on vitamin E, and 2.4% were on pioglitazone., Conclusions: More severe liver disease can be suspected or ruled out based on factors identified in this study. Utilisation of structured lifestyle program, vitamin E and pioglitazone was limited despite this being a cohort of biopsy-proven NAFLD patients with majority of patients having NASH., (© 2018 John Wiley & Sons Ltd.)

Background & Aims: Treatment approaches for hepatocellular carcinoma (HCC) vary across countries, but these differences and their potential impact on outcomes have not been comprehensively assessed. Data from the multinational GIDEON (Global Investigation of therapeutic DEcisions in HCC and Of its treatment with sorafeNib) registry evaluated differences in patient characteristics, practice patterns and outcomes in HCC across geographical regions in patients who received sorafenib., Methods: GIDEON is a non-randomised, observational registry study conducted in 39 countries across five global regions. HCC patients in whom a decision to treat with sorafenib was made in clinical practice and according to local practices were included., Results: 3202 patients were evaluable for safety analysis: Asia-Pacific (n = 928), Japan (n = 508), Europe (n = 1113), USA (n = 563) and Latin America (n = 90). Patients in Japan had earlier-stage disease at initial diagnosis compared with patients in other regions (Barcelona Clinic Liver Cancer stage A; 43.7% vs 9.1-24.3%). Use of locoregional therapies before sorafenib, including transarterial chemoembolisation, was more common in Japan (84.4%) and Asia-Pacific (67.2%) compared with the USA (49.4%) and Europe (43.5%). Treatment patterns with respect to sorafenib also differed, with a shorter duration of treatment reported in the USA and Asia-Pacific. Time from initial diagnosis to death was longer in Japan compared with other regions (median, 79.6 months vs 14.8-25.0 months)., Conclusions: Data from GIDEON highlight regional variations in the management of HCC and patient outcomes. Greater standardisation of management may help optimise outcomes for HCC patients., (© 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background & Aims: Sequential therapy posed a high risk of emergence of multidrug resistance and presented a management issue in chronic hepatitis B (CHB) treatment. We evaluated the antiviral efficacy and safety of entecavir (ETV) plus tenofovir (TDF) combination therapy in multidrug-resistant (MDR) CHB patients., Methods: In this prospective, multicentre study, MDR CHB patients, defined as measurable serum HBV DNA (≥60 IU/ml) while on any rescue treatment regimen for at least 24 weeks and the presence of documented prior genotypic resistance to both nucleoside analogue(s) and nucleotide analogue, were treated with ETV 1.0 mg and TDF 300 mg combination therapy for 48 weeks., Results: A total of 64 eligible patients who had previously failed to a median three lines of antiviral therapy (range, 2-6) were included. At baseline, median age was 47.0 years, 89.1% were HBeAg(+), and median HBV DNA was 4.24 (range, 2.11-6.73) log10 IU/ml. By week 4, 12, 24 and 48, 15/64 (23.4%), 36/64 (56.3%), 43/64 (67.2%) and 55/64 (85.9%) patients achieved a HBV DNA <60 IU/ml respectively. The mean reduction of HBV DNA from baseline to 4 and 48 weeks was 1.23 log10 IU/ml and 2.38 log10 IU/ml respectively. Although five patients experienced virological breakthrough, all were transient and no resistant mutation to TDF or novel mutation was detected in any patients., Conclusions: In difficult-to-treat MDR CHB patients with a high exposure to multiple antiviral drugs, ETV plus TDF combination therapy can provide a very high rate of viral suppression through 48 weeks of treatment., (© 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background & Aims: Here, we evaluated the safety and immunogenicity of hepatitis B virus (HBV) DNA vaccine, HB-110, in mice and Korean patients with chronic hepatitis B (CHB) undergoing adefovir dipivoxil (ADV) treatment., Methods: For animal study, mice (BALB/c or HBV transgenic) were immunized with mHB-110, and T-cell and antibody responses were evaluated. For clinical study, 27 patients randomly received either ADV alone or ADV in combination with HB-110. Liver function tests, serum HBV DNA levels and the presence of HBeAg/anti-HBe were analysed. T-cell responses were estimated by ELISPOT and FACS analysis., Results: mHB-110 induced higher T-cell and antibody responses than mHB-100 in mice. No adverse effects were observed by HB-110 cotreated with ADV. HBV-specific T-cell responses were induced in a portion of patients in medium to high dose of HB-110. Interestingly, HB-110 exhibited positive effects on ALT normalization and maintenance of HBeAg seroconversion. One patient, who received high dose of HB-110 exhibited HBeAg seroconversion during vaccination, which correlated with vaccine-induced T-cell responses without ALT elevation., Conclusions: HB-110 was safe and tolerable in CHB patients. In contrast to results in animal models, HB-110 in Korean patients exhibited weaker capability of inducing HBV-specific T-cell responses and HBeAg seroconversion than HB-100 in Caucasian patients. As Asian patients, who are generally infected via vertical transmission, appeared to have higher level of immune tolerance than Caucasian, novel approaches for breaking immune tolerance rather than enhancing immunogenicity may be more urgently demanded to develop effective therapeutic HBV DNA vaccines., (© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

To investigate the effect of chemical chaperones on the production and aggregation of flag-tagged cartilage oligomeric matrix protein-Angiopoietin1 (FCA1) in recombinant Chinese hamster ovary (CHO) cells, CHO cells were cultivated in serum-free media with various chemical chaperones, 1 mM 4-phenylbutyrate (4-PBA), 200 mM proline, 3% glycerol, 2% dimethyl sulfoxide (DMSO), and without chemical chaperone as control. The addition of chemical chaperones enhanced FCA1 production and specific FCA1 productivity, q(FCA)(1). For example, the q(FCA)(1) at 200 mM proline was fourfold higher than that at control. Unlike q(FCA)(1), the aggregation of FCA1 was strongly affected by which chemical chaperone was added. The addition of 2% DMSO and 200 mM proline significantly reduced the proportion of aggregates, but the addition of 1 mM 4-PBA and 3% glycerol was hardly effective. The proportions of aggregates were 29.5 and 55.6% at 2% DMSO and 200 mM proline, respectively, whereas it was 79.6% at control. The exact mechanism how chemical chaperones affected the aggregation of FCA1 was not investigated in this study, and therefore, more extensive works will be needed to clarify why different chemical chaperones behaved differently in reducing the aggregation of FCA1. Among chemical chaperones tested, DMSO was the most effective one in regard to enhancing the production and reducing the aggregation of FCA1 in CHO cells., (Copyright © 2011 American Institute of Chemical Engineers (AIChE).)

Background: Long-term results after downstaging hepatocellular carcinoma (HCC) prior to liver transplantation (LT) remain unknown., Aims: To investigate dropouts and post-transplant outcome among patients with downstaged HCC by transarterial chemo-lipiodolization (TACL)., Methods: Between 2000 and 2007, 386 patients with HCC initially exceeding Milan criteria underwent TACL for tumour downstaging and were consecutively enrolled., Results: Overall, 160 (41.5%) patients achieved successful downstaging of HCC to within Milan criteria. During the follow-up, 82 eventually dropped off the waiting list for LT, with estimated dropout rates at 1, 2 and 5 years of 46.7%, 70.2%, and 87.2%, respectively. The overall post-transplant survival rates at 1, 2 and 5 years were 89.2%, 70.3% and 54.6% and the corresponding rates for recurrence-free survival were 74.7%, 71.8% and 66.3% respectively. Multivariate analysis indentified alpha-fetoprotein (AFP) levels > or = 100 ng/mL at LT (P = 0.003), maximum tumour size > or = 7 cm (P = 0.002) and the lack of complete necrosis by TACL (P = 0.048) as independent predictors of HCC recurrence after LT. Patients with none of these risk factors had an excellent post-transplant outcome, with an 87.5% probability of recurrence-free survival up to 6 years., Conclusions: These long-term results may contribute to the database for optimizing management of LT candidates with downstaged HCC. Based on our data, patients with a maximum tumour size <7 cm who achieve complete necrosis together with AFP levels <100 ng/mL at LT may be the best candidates for LT following downstaging using TACL.

Background/aims: In patients with hepatocellular carcinoma (HCC) exceeding the Milan criteria, the recurrence rate after liver transplantation is over 50%. We investigated pretransplant factor(s) that could predict recurrence after living donor liver transplantation (LDLT) in patients with HCC exceeding the Milan criteria., Methods: Pre-operative imaging showed that, of the 111 HCC patients who underwent LDLT between June 1995 and January 2006, 37 exceeded the Milan criteria. Clinical factors before LDLT were evaluated., Results: The 1- and 3-year cumulative recurrence rates were 35 and 55% respectively. Pretransplant risk factors for HCC recurrence were large tumour size (>6 cm, P=0.001), tumour exposed to the liver surface (P=0.014) and progressive disease after pretransplant treatment (P=0.038). The 2-year HCC recurrence rates in patients with 0, 1, 2 and 3 factors were 0% (0/4), 9% (1/16), 80% (8/10) and 100% (7/7) respectively (P<0.001). The 2-year survival rate was significantly higher in patients with 0 or 1 factor than in patients with two or more factors (P=0.022)., Conclusions: In patients with HCC exceeding the Milan criteria, the three pretransplant factors that may be useful for identifying those with high HCC recurrence potential after LDLT are tumour size >6 cm, progressive disease after pretransplant treatment and tumour exposed to the liver surface.

Background: Although transarterial chemotherapy is used to retard tumour progression for hepatocellular carcinoma (HCC) patients awaiting orthotopic liver transplantation (OLT), information regarding the acceptable waiting time and appropriate patient selection for the therapy is lacking., Aim: To examine dropout times and determine the best candidates for pre-transplant transarterial therapy in a cohort study., Methods: In total, 180 consecutive HCC candidates receiving pre-transplant chemo-lipiodolization were included in the study., Results: Overall, 70 (38.9%) patients dropped off the waiting list during the median follow-up of 19 months. According to the Child-Pugh (C-P) classification, the estimated dropout rates at 1 and 2 years were 17.2% and 44.8% for the C-P A group and 33.4% and 81.3% for the C-P B/C group, respectively. C-P B/C patients experienced more frequent dropouts than C-P A patients (P < 0.001). Risk factor analysis identified C-P classification to be the strongest predictor of dropout (P < 0.001). On multivariate analysis, alpha-fetoprotein (AFP) >100 ng/mL, tumour size >3 cm and multiple nodules remained independently predictive of dropout for C-P A group (all P < 0.05). Candidates with none of these factors were found to be at the lowest risk of dropout, with only a 22.5% dropout rate up to 41 months., Conclusions: This study suggests that Child-Pugh A patients with one nodule <3 cm and AFP < 100 ng/mL may be the best candidates for pre-transplant chemo-lipiodolization, with the lowest dropout rate. However, comparative studies with other therapeutic options are needed to assess the definitive role of transarterial therapy in this setting.

Background/aim: Recent data have shown that the clinical outcome of hepatitis C virus (HCV) infection may be influenced by the host genetic factor. The aim of this study was to investigate whether particular human leukocytes antigen (HLA) molecules are associated with the susceptibility to HCV infection in the Korean population., Methods: One hundred and thirty-seven patients with chronic HCV infection and 206 normal individuals were examined for HLA class I and II molecules., Results: In class I antigens, the frequencies of HLA-A3 (relative risk (RR)=3.5, P<0.04), HLA-B35 (RR=2.0, P<0.03), and HLA-B46 (RR=2.5, P<0.02) significantly increased in chronic HCV carriers compared with the controls. The frequencies of DRB1*0803, DQB1*0601 and DQB1*0604 were significantly higher in chronic HCV carriers than in controls (RR=2.5, P<0.005; RR=1.8, P<0.05; RR=1.9, P<0.04, respectively). On the other hand, the frequencies of DRB1*0301, DQA1*0501 and DQB1*0201 were significantly lower in chronic HCV carriers than in normal controls (RR=0.2, P<0.03; RR=0.4, P<0.004; RR=0.5, P<0.02, respectively). The haplotype DRB1*0803-DQB1*0601 significantly increased (RR=2.5, P<0.02) while the DQA1*0501-DQB1*0201 significantly decreased (RR=0.2, P<0.03) in chronic HCV carriers compared with normal controls. In stratification analysis to investigate the interrelationships among the associated alleles, DRB1*0803 and DQB1*0601 were associated with HLA-B46, particularly in patients with chronic HCV carriers., Conclusions: These results suggest that particular HLA alleles may have an influence on chronic HCV infection as a host genetic factor in the Korean population.

Purpose: The aim of this prospective study was to evaluate the effectiveness of preconditioning molecular adsorbent recirculating system (MARS) treatment on patients with acute-on-chronic liver failure (AoCLF), who were awaiting living donor liver transplantation (LDLT)., Patients and Methods: Between January and December 2001, 10 consecutive AoCLF patients (with progressive hyperbilirubinemia (>20 mg/dl) and hepatic encephalopathy grade > or =2) were studied. MARS was used in eight of these patients who were evaluated for LDLT during 2001. Three AoCLF patients who received LDLT before clinical use of MARS were used as historical controls., Results: Because of a shortage of donors, only five out of 10 patients considered for LDLT could receive transplants. Three patients were treated with MARS for 8 h the day before receiving LDLT, and all three survived. The remaining two patients who received transplants, and who were not pretreated with MARS, died from sepsis and multi-organ failure within 2 weeks. Four of the patients who did not receive transplants because of donor shortage died despite 1 or 3 MARS treatments, however bilirubin levels and grade of encephalopathy were significantly reduced in these patients., Conclusions: Results of this small pilot study suggest that MARS, by reducing the severity of jaundice and encephalopathy, might be effective as a bridging option in AoCLF patients awaiting LDLT.

To understand the different responses of recombinant Chinese hamster ovary (rCHO) cells to low culture temperature regarding specific productivity (q), 12 parental clones and their corresponding amplified clones producing a humanized antibody were cultivated at 32 and 37 degrees C. The specific growth rate of all clones, including both parental and amplified clones, decreased by 30-63% at 32 degrees C, compared to rates at 37 degrees C. In contrast, their specific antibody productivity (qAb) was significantly enhanced at 32 degrees C. Furthermore, the degree of qAb enhancement at 32 degrees C varied a lot from 4- to 25-fold among the parental clones. At 32 degrees C, most of the amplified clones, regardless of methotrexate (MTX) levels, also showed enhanced qAb but to a lesser extent than their parental clones. However, clone 14 amplified at 0.32 microM MTX (clone 14-0.32) and clone 20 amplified at 1 microM MTX (clone 20-1.00), unlike their parental clones, did not show enhanced qAb at 32 degrees C. Thus, it was found that the enhancing effect of low culture temperature on q of rCHO cells depends on clones. Taken together, the results obtained here emphasize the importance of clonal selection for the successful application of low culture temperature to the enhanced foreign protein production in rCHO cells.

A single stressful culture condition induced by hypoosmotic stress (210 mOsm kg(-1)), low culture temperature (32 degrees C), or NaBu addition (1 mM) resulted in a 1.8- to 2.2-fold enhancement of specific erythropoietin (EPO) productivity (qEPO) of recombinant Chinese hamster ovary (rCHO) cells compared to normal culture condition (37 degrees C and 310 mOsm kg(-1)). Simultaneous application of these stressful conditions further enhanced qEPO up to approximately 5-fold. However, the quality of EPO was affected by stressful culture conditions. The proportion of acidic isoforms of EPO under a single stressful condition was 2.8-13.8% lower than that under normal culture condition. Simultaneous application of the stressful conditions further decreased the portion of acidic isoforms but not significantly. Despite 5-fold enhancement of q(EPO), the portion of acidic isoforms under the simultaneous application of stressful culture conditions was 12.9-21.6% lower than that under normal culture condition. Taken together, these results suggest the potential of simultaneous application of different stressful culture conditions to the production phase of two-stage culture, where cell growth and production phases are separated, for improved EPO production.

Lowering the culture temperature has been suggested as a useful tool for improving the production of recombinant proteins in Chinese hamster ovary (CHO) cells. In an effort to improve anti-4-1BB antibody production in recombinant CHO (rCHO) cells, rCHO cells producing anti-4-1BB antibody (LGA31-56) were cultivated at three different temperatures, 30, 33, and 37 degrees C. Lowering the culture temperature led to suppressed cell growth, cell cycle arrest in G(0)/G(1) phase, and improved cell viability for a longer period. However, antibody production and q(Ab) were not increased at low culture temperature. The maximum antibody concentration and q(Ab) at 37 degrees C were 110.6 +/- 2.6 microg mL(-)(1) and 0.43 +/- 0.03 microg (10(6) cells h)(-)(1), respectively, whereas those at 30 degrees C were 28.3 +/- 3.8 microg mL(-)(1) and 0.44 +/- 0.07 (10(6) cells h)(-)(1), respectively. Northern blot analysis revealed that lowering the culture temperature did not increase the transcription level of heavy and light chains. These results were quite in contrast with the improved production of erythropoietin, which is expressed in the same CHO host and driven by the same CMV promoters, by lowering the temperature. Taken together, the results obtained imply that the beneficial effect of low culture temperature on recombinant protein production in rCHO cells is cell-line-specific.