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Introduction: Atezolizumab and bevacizumab (AB) therapy was the first-line treatment for unresectable hepatocellular carcinoma (u-HCC). However, the predictive marker of therapeutic response that can be easily used in clinical practice is still unknown. We prospectively investigated the utility of time-intensity curve (TIC) analysis using contrast-enhanced ultrasound (CEUS) as a predictive indicator of therapeutic response after the start of AB therapy. Methods: Thirty-five patients who received AB therapy for u-HCC were included in this study. TIC analysis was performed in 28 patients who were able to undergo CEUS before and 3–7 days after administration. We analyzed prognostic factors related to the initial therapeutic response and long progression-free survival (PFS). Results: The initial therapeutic response using dynamic computed tomography or Gd-EOB magnetic resonance imaging at 8–12 weeks after administration was partial response/stable disease/progressive disease (PD) in 14/12/9 cases (40/34/26%). Cases with PD (n = 9) had more cases without decreased blood flow in TIC analysis compared with cases with non-PD (100 vs. 18%, p = 0.001). Cases without decreased blood flow in TIC analysis (n = 10) had more cases with PD compared with cases with decreased blood flow (60 vs. 0%, p = 0.001). PFS in patients without decreased blood flow early after the administration was shorter than that in those with decreased blood flow (9.1 vs. 28 weeks, p = 0.0051). Conclusion: Early evaluation by TIC analysis using CEUS may be useful in predicting the therapeutic response in patients treated with AB therapy for u-HCC.

Recently, with the advent of sofosbuvir/velpatasvir therapy, sustained virological response (SVR) can now be achieved even in patients with decompensated cirrhosis (dLC). However, the prognosis after SVR does not always improve in dLC, and appropriate indicators enabling prediction of prognosis is desired.Serum IP-10/CXCL10 levels were measured in 47 patients (15 chronic hepatitis [CH], 17 compensated cirrhosis [cLC], and 15 dLC) receiving direct acting antiviral (DAA) therapy, and their changes during the therapy were examined.All the patients achieved SVR. In patients with CH, the average IP-10 level was 367, 102, and 68 pg/ml respectively at baseline, at the end of therapy and at 12 weeks after SVR (SVR12), and was decreased upon DAA therapy (P 0.001). In patients with cLC, IP-10 was respectively 215, 91, and 77 pg/ml, and was decreased upon DAA therapy (P 0.001) while it was 283, 131, and 182 pg/ml in patients with dLC and there was no evident decrease (P = 0.55). When patients with dLC were further classified depending on the difference in Child-Pugh (CP) score improvement at SVR12, a significant decrease in IP-10 was observed after treatment in those with improvement (P = 0.023) while a significant increase was observed in those without improvement (P = 0.016).While serum IP-10 level was decreased in patients with CH/cLC and dLC with post-SVR-CP improvement following SVR, it was increased in patients with dLC without post-SVR CP improvement. The result indicates that IP-10 dynamics may be useful for predicting liver function after DAA therapy.

Objective Owing to advances in direct-acting antiviral (DAA) therapy, a considerable number of patients with hepatitis C virus (HCV)-positive hepatocellular carcinoma (HCC) are now able to achieve a sustained viral response (SVR) after curative treatment of HCC. However, the beneficial effect of a DAA-SVR on the survival remains unclear. Methods A total of 205 patients with HCC who were HCV-positive with Child-Pugh A at the onset from 2008 to 2018 were categorized into 2 groups: 140 patients untreated for HCV throughout the entire course after HCC development (untreated group) and 65 patients treated for HCV with DAAs following HCC treatment who achieved an SVR (SVR group). After propensity score matching, 63 patients from each group were selected. Using these patients, the survival and maintenance of Child-Pugh A after HCC treatment were compared between the untreated group and SVR group. Results There was a significant difference in the overall survival (p0.001) and the rate of maintaining Child-Pugh A (p0.001) between the groups. The 5-year survival rates were 96% (SVR group) and 60% (untreated group), and the proportions of patients with Child-Pugh A at 5 years after HCC treatment were 96% (SVR group) and 38% (untreated group). Conclusion In patients with HCV-positive HCC, achieving a DAA-SVR after HCC treatment significantly improved the overall survival rate compared with HCV-untreated patients. The contribution of DAA-SVR during the course of HCC treatment to a longer survival is mainly due to the prevention of the progression of Child-Pugh A to B/C. Further research is needed to determine whether aggressive antiviral therapy is also effective for HCC patients with Child-Pugh B/C.

The damping ratio (DR) and the loss modulus (G″) obtained by 3D MR elastography complex modulus analysis has been reported recently to reflect early intrahepatic inflammation, and is expected to be a noninvasive biomarker of inflammation in nonalcoholic fatty liver disease (NAFLD). However, the role of the DR and the G″ in Japanese NAFLD patients remains unclear.We enrolled 39 Japanese patients with NAFLD who underwent liver biopsy and 3D MR elastography within 1 month and analyzed the association between DR, G″, and histological activity.Regarding DR, no evident correlation was observed between the DR and histological activity (p = 0.14) when patients with all fibrosis stages were included. However, when patients were restricted up to stage F2 fibrosis, the association of the DR and inflammation became significant, the DR increasing with the degree of activity (p = 0.02). Among the constituents of fibrosis activity, ballooning correlated with the DR (p 0.01) while lobular inflammation did not. Regarding G″, it was correlated with histological activity (p 0.01), ballooning (p 0.01), and lobular inflammation (p 0.01) in patients with all fibrosis stages and in patients up to F2 fibrosis (p = 0.03 for activity and p = 0.04 for ballooning). The best cutoff value of DR for hepatitis activity in patients within the F2 stage was 0.094 (area under the receiver operating characteristic curve 0.775, 95% CI: 0.529-1.000) and G″ was 0.402 (area under the receiver operating characteristic curve 0.825, 95% CI: 0.628-1.000).The DR and G″ reflected the histological activity in Japanese patients with NAFLD during the early stage, indicating these values for noninvasive diagnosis of inflammation in Japanese patients with NAFLD.

AIM Recently, serum hepatitis B virus (HBV)-RNA has been reported to be detectable even when HBV particle production is inhibited by nucleot(s)ide analogues (NAs). However, the dynamics of the HBV-RNA sequence compared with those of HBV-DNA during the emergence of antiviral resistance are yet to be elucidated. METHODS First, we quantified serum HBV-RNA in 181 infected patients, and its relationships with clinical characteristics as well as HBV markers were investigated. Next, we undertook simultaneous deep sequencing of HBV-RNA/HBV-DNA and their dynamics among four patients receiving NA therapy who were experiencing viral breakthrough. RESULTS Serum HBV-RNA was detected in 25% (31/123) of cases among patients with HBV without NAs, and the detection rate was significantly high in hepatitis B e antigen-positive cases with high viral activity. In patients with chronic hepatitis, hepatitis B core-related antigen was significantly correlated with serum HBV-RNA irrespective of NA use. In the analysis of the four patients experiencing viral breakthrough, no NA resistance mutation was detected in the serum HBV-RNA immediately before the breakthrough. However, NA-resistant sequences appeared at the rates of 0%, 3%, 14%, and 100%, and the NA-resistant HBV-RNA sequence rate was correlated with the peak HBV-DNA titer multiplied by the HBV-DNA detection duration during the breakthrough (R2 = 0.978) observed before redisappearance of HBV-DNA following the addition of new NA. CONCLUSION Serum HBV-RNA could reflect the transcriptional activity of covalently closed circular DNA and hepatitis B core-related antigen. The dynamics of HBV-RNA could help understanding of the turnover process of HBV covalently closed circular DNA in the liver.

Liver damage presented as alanine aminotransferase (ALT) elevation and high ALT-caused treatment discontinuation occurs with high frequency in Japanese patients receiving daclatasvir plus asunaprevir (DCV/ASV) therapy for hepatitis C virus (HCV) infection, and its mechanism is unknown. A total of 247 Japanese patients consisting of two independent cohorts with genotype-1b HCV infection receiving DCV/ASV therapy were included. The association of ALT levels during therapy and single nucleotide polymorphisms (SNP) of five drug-metabolizing enzyme loci selected for their possible influence on NS3/4A and NS5A inhibitors was investigated. Among five SNPs, we found a significant correlation between the presence of the UGT1A1 rs4148323 A allele and ALT elevation (Grade 3 elevation in AA 57%, AG 18%, and GG 4%, P = 8.4E − 06) and drug discontinuation (AA 22%, AG 11%, and GG 2.5%, P = 8.7E − 04), while no association was observed with ALT values at baseline (Grade 3 elevation AA 0%, AG 4%, and GG 2%, P = 0.5). In contrast, patients with risk A allele for drug-induced ALT elevation had a tendency to respond more favorably to treatment (AA 100%, AG 93%, and GG 90%, P = 0.29). Through the analysis we suggest that the A allele in UGT1A1 rs4148323 (UGT1A1*6), which is highly prevalent in the Japanese population, should be considered a risk for the development of DCV/ASV therapy-induced ALT elevation. Pretreatment SNP testing of UGT1A1*6 might be beneficial for the prediction of liver damage induced by DCV/ASV or even by DCV/ASV plus beclabuvir.

AIM The landscape of cancer-related genetic aberrations in hepatocellular carcinoma (HCC) has gradually become clear through recent next-generation sequencing studies. However, it remains unclear how genetic aberrations correlate with imaging and histological findings. METHODS Using 117 formalin-fixed paraffin-embedded specimens of primary liver tumors, we undertook targeted next-generation sequencing of 50 cancer-related genes and digital polymerase chain reaction of hTERT. After classifying tumors into several imaging groups by hierarchal clustering with the information from gadoxetic acid enhanced magnetic resonance imaging, contrast-enhanced computed tomography, contrast-enhanced ultrasound, and diffusion-weighted imaging magnetic resonance imaging, the correlation between genetic aberrations and imaging and histology were investigated. RESULTS Most frequent mutations were hTERT (61.5%), followed by TP53 (42.7%), RB1 (24.8%), and CTNNB1 (18.8%). Liver tumors were classified into six imaging groups/grades, and the prevalence of hTERT mutations tended to increase with the advancement of imaging/histological grades (P = 0.026 and 0.13, respectively), whereas no such tendency was evident for TP53 mutation (P = 0.78 and 1.00, respectively). Focusing on the mutations in each tumor, although the variant frequency (VF) of hTERT did not change (P = 0.36 and 0.14, respectively) in association with imaging/histological grades, TP53 VF increased significantly (P = 0.004 and

AimDeletions are observed frequently in the preS1/S2 region of hepatitis B virus (HBV) genome, in association with liver disease advancement. However, the most significant preS1/S2 region and its influences on viral markers are unclear.MethodsThe preS1/S2 HBV regions of 90 patients without antiviral therapy were subjected to deep sequencing and deleted regions influencing viral markers were investigated.ResultsFrom the deletion frequency analysis in each patient, deletions were observed most frequently in the preS2 codon 132-141 region. When the patients were divided into three groups (0-0.1%: n = 27, 0.1%-10%: n = 34, 10-100%: n = 29), based on the deletion frequency, FIB-4 (p < 0.01), HBV DNA (p < 0.01), HBcrAg (p < 0.01) and preS1/S2 start codon mutations (p < 0.01, both) were significantly associated with the deletion. When clinical and viral markers were investigated by multivariate analysis for their association with the deletion, FIB-4 (p < 0.05), HBcrAg (p < 0.05), and preS1 start codon mutation (p < 0.01) were extracted as independent variables. When the influence of the preS codon 132-141deletions on HBsAg and HBcrAg, relative to HBV DNA, was investigated, the HBsAg/HBV DNA ratio was lower (0-10% vs. 10%-100%, pConclusionThe preS codon.132-141 deletions have a significant influence on the clinical characteristics and viral markers, even when present as a minor population. Importantly, the preS codon 132-141 deletions have a clear influence on the viral life cycle and pathogenesis.

AIM Although the viral markers hepatitis B surface antigen (HBsAg) and hepatitis B core-related antigen (HbcrAg) could reflect intrahepatic hepatitis B virus (HBV) replication activity and constitute important biomarkers for hepatocellular carcinoma (HCC), the value of using these two markers in combination for assessing HCC risk has not been clarified in detail. METHODS Four hundred and forty-nine consecutive patients with chronic HBV infection were included in the study and the association of HBsAg and HBcrAg with HCC risk was investigated cross-sectionally, as well as longitudinally. RESULTS When the high value cut-offs of HBsAg and HBcrAg were defined as 3.0 log IU/mL and 3.0 log U/mL, respectively, patients with a history of HCC were found frequently in the low HBsAg group (P = 0.002) and high HBcrAg group (P

Because of recent advances in deep sequencing technology, detailed analysis of hepatitis C virus (HCV) quasispecies and their dynamic changes in response to direct antiviral agents (DAAs) became possible, although the role of quasispecies is not fully understood. In this study, to clarify the evolution of viral quasispecies and the origin of drug-resistant mutations induced by interferon (IFN)-based protease inhibitor therapy, the nonstructural-3 (NS3) region of genotype 1b HCV in 34 chronic hepatitis patients treated with telaprevir (TVR)/pegylated interferon (PEG-IFN)/ribavirin (RBV) was subjected to a deep sequencing study coupled with phylogenetic analysis. Twenty-six patients (76.5%) achieved a sustained viral response (SVR), while 8 patients did not (non-SVR; 23.5%). When the complexity of the quasispecies was expressed as the mutation frequency or Shannon entropy value, a significant decrease in the IFNL3 (rs8099917) TT group and a marginal decrease in the SVR group were found soon (12 h) after the introduction of treatment, whereas there was no decrease in the non-SVR group and no significant decrease in mutation frequency in the IFNL3 TG/GG group. In the analysis of viral quasispecies composition in non-SVR patients, major populations greatly changed, accompanied by the appearance of resistance, and the compositions were unlikely to return to the pretreatment composition even after the end of therapy. Clinically TVR-resistant variants were observed in 5 non-SVR patients (5/8, 62.5%), all of which were suspected to have acquired resistance by mutations through phylogenetic analysis. In conclusion, results of the study have important implications for treatment response and outcome in interferon-based protease inhibitor therapy. IMPORTANCE In the host, hepatitis C virus (HCV) consists of a variety of populations (quasispecies), and it is supposed that dynamic changes in quasispecies are closely related to pathogenesis, although this is poorly understood. In this study, recently developed deep sequencing technology was introduced, and changes in quasispecies associated with telaprevir (TVR)/pegylated interferon (PEG-IFN)/ribavirin (RBV) triple therapy and their clinical significance were investigated extensively by phylogenetic tree analysis. Through this study, the associations among treatment response, changes in viral quasispecies complexity in the early stage of treatment, changes in the quasispecies composition, and origin of TVR-resistant variant HCV were elucidated.

Aim Liver fibrosis is a risk factor for hepatocellular carcinoma (HCC), but at what fibrotic stage the risk for HCC is increased has been poorly investigated quantitatively. This study aimed to determine the appropriate cut-off value of liver stiffness for HCC concurrence by FibroScan, and its clinical significance in hepatitis B virus (HBV), hepatitis C virus (HCV) and non-B, non-C (NBNC) liver disease. Methods Subjects comprised 1002 cases (246 with HCC and 756 without HCC) with chronic liver disease (HBV, 104; HCV, 722; and NBNC, 176). Results Liver stiffness was significantly greater in all groups with HCC, and the determined cut-off value for HCC concurrence was more than 12.0 kPa in those with HCV, more than 8.5 kPa in those with HBV and more than 12.0 kPa in those with NBNC. Liver stiffness of more than 12.0 kPa was an independent risk factor for new HCC development in HCV. For HCV, risk factors for HCC concurrence were old age, male sex, low albumin, low platelets and liver stiffness, while for HBV they were old age, low platelets and liver stiffness, and for NBNC they were old age, elevated α-fetoprotein and liver stiffness. Conclusion Liver stiffness cut-off values and their association with HCC concurrence were different depending on the etiology. In HCV, liver stiffness of more than 12.0 kPa was an independent risk factor for new HCC development. Collectively, determining the fibrotic cut-off values for HCC concurrence would be important in evaluating HCC risks.

Aim To investigate whether the patients with hypovascular liver nodules determined on the arterial phase and hypointensity on the hepatocyte phase gadoxetic acid-enhanced magnetic resonance imaging (hypovascular hypointense nodules) are at increased risk of hepatocarcinogenesis, we assessed subsequent typical hepatocellular carcinoma (HCC) development at any sites of the liver with and without such nodules. Methods One hundred and twenty-seven patients with chronic hepatitis B or C and without a history of HCC, including 68 with liver cirrhosis, were divided into those with (non-clean liver group, n = 18) and without (clean liver group, n = 109) hypovascular hypointense nodules. All the patients were followed up for 3 years, and HCC development rates and risk factors were analyzed with the Kaplan–Meier method and the Cox proportional hazard model, respectively. Results A total of 17 patients (10 in the non-clean liver group and seven in the clean liver group) developed typical HCC. Cumulative 3-year rates of HCC development were 55.5% in the non-clean liver group and 6.4% in the clean liver group (P

Aim Daclatasvir, a non-structural (NS)5A replication complex inhibitor, is a potent and promising direct antiviral agent (DAA) for hepatitis C virus (HCV), being most effective in genotype 1b infection. Although it is known that genotype 1b viruses with Y93H and/or L31M/V/F mutations have strong resistance to daclatasvir, it is not known whether there are some clinical background conditions that favor the occurrence of HCV carrying those NS5A mutations. Methods In this study, we carried out deep sequencing analysis of stored sera to determine the presence and significance of daclatasvir-resistant mutants in 110 genotype 1b HCV-infected patients with no previous daclatasvir treatment. Results Deep sequencing analysis revealed that the NS5A L31M/V/F and Y93H mutations were present in 13 (11.8%) and 34 (30.9%) of the 110 patients, respectively, and significantly more frequently than in the control plasmid. Simultaneous L31M/V/F and Y93H mutations were detected in four of the 110 patients (3.6%). When the clinical relevance of NS5A resistance was investigated, Y93H was significantly correlated with the IL28B major (TT) genotype of the host (P = 0.042). Conclusion Y93H was detected frequently by deep sequencing in daclatasvir treatment-naive patients. Importantly, it seems that the IL28B status of the patients may influence the presence of Y93H mutations, resulting in different treatment responses to daclatasvir.

Due to the recent development of various novel small molecule compounds called direct-acting antiviral (DAA) agents and their dramatic antiviral potency against hepatitis C virus (HCV), more than 90 % of patients can now eradicate HCV with DAA therapy, a drastic improvement over the days of interferon therapy both in terms of efficacy and tolerability. On the other hand, the emergence of resistance-associated variants (RAVs) during DAA-based therapy has become a major problem. Namely, due to their high potency and specificity, there is a risk that patients treated with DAAs might develop resistance to one or more classes of DAA. Because of high cross-resistance among the small number of approved DAAs and the limited number of targets, it is also possible for HCV strains resistant to all current DAAs to emerge. Although it is possible to some extent to predict the emergence of RAVs through screening for the presence of RAVs prior to therapy using direct sequencing or other methods, precise and accurate prediction is still not possible because the presence of RAVs at baseline does not guarantee treatment failure nor does the lack of preexisting RAVs preclude the emergence of resistance through de novo mutation during treatment. In order to determine whether the emergence of RAVs is predictable based on the composition of HCV quasispecies prior to treatment, in this study deep sequencing was used to correlate the presence of RAVs at baseline with the emergence of RAVs during treatment in patients treated with interferon-based DAA triple therapy versus interferon-free DAA therapy.

Aim: In the treatment of chronic hepatitis C, pegylated interferon (PEG-IFN) and ribavirin combination therapy must be continued for an adequate duration to improve the rate of sustained virological response. We attempted to predict the time point at which serum hepatitis C virus (HCV) RNA are undetectable during combination therapy. Methods: Patients with HCV genotype 1b were enrolled in a model preparation (n = 35) and a validation group (n = 70). All patients received PEG-IFN-α-2b/ribavirin combination therapy for at least 48 weeks, and serological samples were screened a minimum of 17 times during the therapy. Serum HCV RNA were measured by the Abbott RealTime HCV assay. Using the HCV dynamics model described by Neumann et al., we used multiple linear regression analysis to select factors that affected the undetectable time point. Results: Difference in viral load between weeks 1 and 2 was the only predictive factor for the undetectable time point of serum HCV RNA (r2 = 0.67, P

Aim: Hepatic steatosis is linked to development of hepatocellular carcinoma (HCC) in non-viral liver disease such as non-alcoholic steatohepatitis. The present study aimed to assess whether hepatic steatosis is associated with the development of HCC in chronic hepatitis C. Methods: We studied a retrospective cohort of 1279 patients with chronic hepatitis C who received interferon (IFN) therapy between 1994 and 2005 at a single regional hospital in Japan. Of these patients, 393 had a sustained virological response (SVR) and 886 had non-SVR to IFN therapy. After IFN therapy, these patients were screened for development of HCC every 6 months. The average period of observation was 4.5 years. Results: HCC developed in 68 patients. The annual incidence of HCC was 2.73% for patients with a steatosis grade of 10% or greater and 0.69% for patients with a steatosis grade of 0–9%. On multivariate analysis, higher grade of steatosis was a significant risk factor for HCC independent of older age, male sex, higher body mass index (BMI), advanced fibrosis stage and non-SVR to IFN therapy. The adjusted risk ratio of hepatic steatosis was 3.04 (confidence interval 1.82–5.06, P

An increase in the aging population is an impending problem. A large cohort study was carried out to determine the influence of aging and other factors on hepatocarcinogenesis in patients treated with interferon. Biopsy-proven 2547 chronic hepatitis C patients registered at our referral center since 1992 were included. Of these, 2166 were treated with interferon-based therapy. Incidences of hepatocellular carcinoma (HCC) associated with interferon were analyzed by Kaplan-Meier and person-years methods for an average follow-up of 7.5 years. Factors associated with HCC risk were determined by Cox proportional hazard analysis. HCC developed in 177 interferon-treated patients. The risk for HCC depended on age at primary biopsy and increased more than 15-fold after 65 years of age. Even when stratified by stage of fibrosis, the cumulative and annual incidences of HCC were significantly higher in older patients than in younger patients (P < 0.001) at the same stage of fibrosis, except for cirrhosis. Progression of fibrosis over time was significantly accelerated in older patients. The impact of viral eradication on HCC prevention was less significant in older patients than in younger patients. Multivariate analysis confirmed that age, gender, liver fibrosis, liver steatosis, total cholesterol level, fasting blood sugar level, baseline and postinterferon alpha-fetoprotein level, and virological response to interferon were independent risk factors associated with HCC. Aging was the strongest risk factor for a nonvirological response to interferon-based antiviral therapy. Conclusion: Elderly patients are at a higher risk for HCC. Hepatitis C viral eradication had a smaller effect on hepatocarcinogenesis in older patients. Patients should therefore be identified at an earlier age and treatment should be initiated. HEPATOLOGY 2010

症例は53歳の女性．2006年頃から腹部膨満感を自覚した．2008年5月，近医での腹部超音波にて巨大な腹腔内嚢胞性病変を指摘され当科を受診した．腫瘍マーカーは基準値内であり，画像検査では肝左葉由来の単房性嚢胞性病変を認め，多数の壁在結節を有していた．胆管嚢胞腺癌の術前診断で，当院外科にて肝左葉切除術を施行した．病理組織学的検査にて乳頭状隆起は粘液円柱上皮で構成され，分裂像，核異型を認め，胆管嚢胞腺癌と診断した．卵巣様間質や胆管との交通は明らかでなかった．免疫染色ではMUC5ACがびまん性に陽性，MUC1は一部陽性で，MUC2とMUC6は陰性であり，胃腺窩上皮細胞の粘液形質を有していた．過去の報告例の検討でも同様に胃腺窩上皮細胞の粘液形質を示す傾向を認めた．

症例1は44歳女性．人間ドックにて肝腫瘍を指摘され，精査目的に入院．HBV, HCVなどウイルスマーカー陰性で肝機能異常は認めず，各種画像診断から肝血管筋脂肪腫（AML）を考え，経皮的腫瘍生検にて診断確定した．経過観察中にて腫瘍径の増大は認めていない．症例2は40歳女性．健診にて肝腫瘍を指摘され，各種画像診断でも肝細胞癌が否定できず，短期間に腫瘍径が増大したため左葉外側切除術を施行した．病理組織より肝血管筋脂肪腫と診断された．症例1, 2は最終診断では肝血管筋脂肪腫であったが，症例1では脂肪成分に富み典型的なAML所見であったのに対し，症例2は脂肪成分に乏しく診断困難であった．両者ともに肝静脈が流出血管として描出され，造影超音波検査が有用であった．

Liver fibrosis is a risk factor for hepatocellular carcinoma (HCC), but at what fibrotic stage the risk for HCC is increased has been poorly investigated quantitatively. This study aimed to determine the appropriate cut-off value of liver stiffness for HCC concurrence by FibroScan, and its clinical significance in hepatitis B virus (HBV), hepatitis C virus (HCV) and non-B, non-C (NBNC) liver disease.Subjects comprised 1002 cases (246 with HCC and 756 without HCC) with chronic liver disease (HBV, 104; HCV, 722; and NBNC, 176).Liver stiffness was significantly greater in all groups with HCC, and the determined cut-off value for HCC concurrence was more than 12.0 kPa in those with HCV, more than 8.5 kPa in those with HBV and more than 12.0 kPa in those with NBNC. Liver stiffness of more than 12.0 kPa was an independent risk factor for new HCC development in HCV. For HCV, risk factors for HCC concurrence were old age, male sex, low albumin, low platelets and liver stiffness, while for HBV they were old age, low platelets and liver stiffness, and for NBNC they were old age, elevated α-fetoprotein and liver stiffness.Liver stiffness cut-off values and their association with HCC concurrence were different depending on the etiology. In HCV, liver stiffness of more than 12.0 kPa was an independent risk factor for new HCC development. Collectively, determining the fibrotic cut-off values for HCC concurrence would be important in evaluating HCC risks.

Daclatasvir, a non-structural (NS)5A replication complex inhibitor, is a potent and promising direct antiviral agent (DAA) for hepatitis C virus (HCV), being most effective in genotype 1b infection. Although it is known that genotype 1b viruses with Y93H and/or L31M/V/F mutations have strong resistance to daclatasvir, it is not known whether there are some clinical background conditions that favor the occurrence of HCV carrying those NS5A mutations.In this study, we carried out deep sequencing analysis of stored sera to determine the presence and significance of daclatasvir-resistant mutants in 110 genotype 1b HCV-infected patients with no previous daclatasvir treatment.Deep sequencing analysis revealed that the NS5A L31M/V/F and Y93H mutations were present in 13 (11.8%) and 34 (30.9%) of the 110 patients, respectively, and significantly more frequently than in the control plasmid. Simultaneous L31M/V/F and Y93H mutations were detected in four of the 110 patients (3.6%). When the clinical relevance of NS5A resistance was investigated, Y93H was significantly correlated with the IL28B major (TT) genotype of the host (P = 0.042).Y93H was detected frequently by deep sequencing in daclatasvir treatment-naïve patients. Importantly, it seems that the IL28B status of the patients may influence the presence of Y93H mutations, resulting in different treatment responses to daclatasvir.

To investigate whether the patients with hypovascular liver nodules determined on the arterial phase and hypointensity on the hepatocyte phase gadoxetic acid-enhanced magnetic resonance imaging (hypovascular hypointense nodules) are at increased risk of hepatocarcinogenesis, we assessed subsequent typical hepatocellular carcinoma (HCC) development at any sites of the liver with and without such nodules.One hundred and twenty-seven patients with chronic hepatitis B or C and without a history of HCC, including 68 with liver cirrhosis, were divided into those with (non-clean liver group, n = 18) and without (clean liver group, n = 109) hypovascular hypointense nodules. All the patients were followed up for 3 years, and HCC development rates and risk factors were analyzed with the Kaplan-Meier method and the Cox proportional hazard model, respectively.A total of 17 patients (10 in the non-clean liver group and seven in the clean liver group) developed typical HCC. Cumulative 3-year rates of HCC development were 55.5% in the non-clean liver group and 6.4% in the clean liver group (P 0.001), and those at the different sites from the initial nodules was also higher in the non-clean liver group (22.2%) than the clean liver group (6.4%) (P = 0.003). Multivariate analysis identified older age (P = 0.024), low platelet counts (P = 0.017) and a non-clean liver (P 0.001) as independent risk factors for subsequent HCC development.Patients with hypovascular hypointense liver nodules are at a higher risk for HCC development at any sites of the liver than those without such nodules.

The three syntypes of Herops munda De Vis (1884), the holotype of Dules marginatus boninensis Fowler (1907), and several Ryukyu specimens of Kuhlia boninensis were compared morphologically, and it was concluded that K. boninensis is a junior synonym of K. munda. Kuhlia munda is distinguished from congeners by the following combination of characteristics: number of lateral line scales 47–51, preorbital serrae 12–23, caudal fin with base and all margins broadly black, and the chevron-shaped central part pale.

In the treatment of chronic hepatitis C, pegylated interferon (PEG-IFN) and ribavirin combination therapy must be continued for an adequate duration to improve the rate of sustained virological response. We attempted to predict the time point at which serum hepatitis C virus (HCV) RNA are undetectable during combination therapy. Patients with HCV genotype 1b were enrolled in a model preparation (n = 35) and a validation group (n = 70). All patients received PEG-IFN-α-2b/ribavirin combination therapy for at least 48 weeks, and serological samples were screened a minimum of 17 times during the therapy. Serum HCV RNA were measured by the Abbott RealTime HCV assay. Using the HCV dynamics model described by Neumann et al., we used multiple linear regression analysis to select factors that affected the undetectable time point. Difference in viral load between weeks 1 and 2 was the only predictive factor for the undetectable time point of serum HCV RNA (r(2) = 0.67, P 0.0005), and we derived the following prediction equation: undetectable time point (week) = 13.495 × (viral load at day 14 [log IU/mL] - viral load at day 7 [log IU/mL]) + 25.456. The equation was applicable to the validation group. We created a formula for predicting the undetectable time point from viral load measurements early in PEG-IFN-α-2b/ribavirin combination therapy. An early response reflects sensitivity to therapy, and the estimation of an undetectable time point would be useful for determining the optimal duration of treatment for chronic hepatitis C patients.

Early disappearance of serum hepatitis C virus (HCV) RNA is the prerequisite for achieving sustained virological response (SVR) in peg-interferon (PEG-IFN) plus ribavirin (RBV) therapy for chronic hepatitis C. This study aimed to develop a decision tree model for the pre-treatment prediction of response.Genotype 1b chronic hepatitis C treated with PEG-IFN alpha-2b and RBV were studied. Predictive factors of rapid or complete early virological response (RVR/cEVR) were explored in 400 consecutive patients using a recursive partitioning analysis, referred to as classification and regression tree (CART) and validated.CART analysis identified hepatic steatosis (30%) as the first predictor of response followed by low-density-lipoprotein cholesterol (LDL-C) (/=100 mg/dL), age (50 and60 years), blood sugar (120 mg/dL), and gamma-glutamyltransferase (GGT) (40 IU/L) and built decision tree model. The model consisted of seven groups with variable response rates from low (15%) to high (77%). The reproducibility of the model was confirmed by the independent validation group (r(2) = 0.987). When reconstructed into three groups, the rate of RVR/cEVR was 16% for low probability group, 46% for intermediate probability group and 75% for high probability group.A decision tree model that includes hepatic steatosis, LDL-C, age, blood sugar, and GGT may be useful for the prediction of response before PEG-IFN plus RBV therapy, and has the potential to support clinical decisions in selecting patients for therapy and may provide a rationale for treating metabolic factors to improve the efficacy of antiviral therapy.

A 35-year-old woman was admitted to our hospital for right upper quadrant pain and multiple liver tumor were detected by diagnostic imaging. Tumors located near the surface of the liver which accompanied capsular retraction. Diagnostic laparoscopy showed multiple white tone tumors with retraction of the adjacent liver capsule. Tumor targeted biopsy was performed. The pathologic diagnosis of epithelioid hemangioendothelioma (EHE) was made by the positive staining of factor VIII-related antigen. EHE tend to locate in peripheral and extend to the liver capsule. Therefore, we face difficulties in getting biopsy sample safely. Here we report a useful case of laparoscopic examination and biopsy in the diagnosis of EHE.

Background & Aims: Clinical significance of molecules involving innate immunity in treatment response remains unclear. The aim is to elucidate the mechanisms underlying resistance to antiviral therapy and predictive usefulness of gene quantification in chronic hepatitis C (CH-C). Methods: We conducted a human study in 74 CH-C patients treated with pegylated interferon α-2b and ribavirin and 5 nonviral control patients. Expression of viral sensors, adaptor molecule, related ubiquitin E3-ligase, and modulators were quantified. Results: Hepatic RIG-I, MDA5, LGP2, ISG15, and USP18 in CH-C patients were up-regulated at 2- to 8-fold compared with nonhepatitis C virus patients with a relatively constitutive Cardif. Hepatic RIG-I, MDA5, and LGP2 were significantly up-regulated in nonvirologic responders (NVR) compared with transient (TR) or sustained virologic responders (SVR). Cardif and RNF125 were negatively correlated with RIG-I and significantly suppressed in NVR. Differences among clinical responses in RIG-I/Cardif and RIG-I/RNF125 ratios were conspicuous (NVR/TR/SVR = 1.3:0.6:0.4 and 2.3:1.3:0.8, respectively). Like viral sensors, ISG15 and USP18 were significantly up-regulated in NVR (4-fold and 2.3-fold, respectively). Multivariate and receiver operator characteristic analyses revealed higher RIG-I/Cardif ratio, ISG15, and USP18 predicted NVR. Lower Cardif in NVR was confirmed by its protein level in Western blot. Also, transcriptional responses in peripheral blood mononuclear cells to the therapy were rapid and strong except for Cardif in not only a positive (RIG-I, ISG15, and USP18) but also in a negative regulatory manner (RNF125). Conclusions: NVR may have adopted a different equilibrium in their innate immune response. High RIG-I/Cardif and RIG-I/RNF125 ratios and ISG15 and USP18 are useful in identifying NVR.

Background/Aims Interferon (IFN) therapy leads to regression of hepatic fibrosis in chronic hepatitis C patients who achieve a sustained virologic response (SVR), while the beneficial effect is limited in those who fail to do so. The aim of the present study was to define factors associated with progression of fibrosis in patients who do not achieve a SVR. Methods Fibrosis staging scores were compared between paired liver biopsies before and after IFN in 97 chronic hepatitis C patients who failed therapy. The mean interval between biopsies was 5.9 years. Factors associated with progression of fibrosis were analyzed. Results Fibrosis progressed in 23%, remained unchanged in 47% and regressed in 29%. Steatosis and a high average alanine aminotransferase (ALT) between biopsies were independent factors for progression of fibrosis with risk ratios of 5.53 and 4.48, respectively. Incidence and yearly rate of progression of fibrosis was 64% and 0.22±0.29 fibrosis units per year in those with both risk factors compared to 8% and −0.04±0.17 fibrosis units per year in those negative for both factors. Conclusions Hepatic steatosis and elevated ALT levels are risk factors for progression of fibrosis in chronic hepatitis C patients who fail to achieve a SVR to IFN therapy and therefore may be therapeutic targets to halt the potentially progressive disease.

Crystal structure prediction is one of the useful theoretical tools for designing and synthesizing new materials in pharmaceutical therapeutics and industrial electronics. Furthermore, the prediction can provide immense valuable scientific knowledge on a crystal growth, polymorphism and many properties of organic molecular crystals. Therefore, we have started the development of high-speed and high-accurate prediction method for organic molecular crystal structures [1,2]. In this work, we demonstrate the theoretical predictions of crystal structures of fourteen target molecules that were used in the international competitions known as CSP blind tests hosted by CCDC [3]. All strategies required for crystal lattice construction expanded to a given effective crystal radius, crystal energy calculation with the reliable molecular force field (MMFF94s) and exhaustive geometry search included a variety of crystal polymorphism are implemented into CONFLEX program [1]. As the results of the applications, we confirmed in all cases of target molecules that, at least, one calculated crystal structure in agreement with the corresponding observed ones can be found. Essential ability required for the prediction method to survive the CSP competitions is that the experimental crystal structure can computationally reproduce within top 3 of most stable structures in crystal energy evaluation. In these tests, only three applications to the target I (Orth. polymorph), II and VIII can successfully satisfy the demand. Details will be discussed in this conference.