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藤原 圭フジワラ ケイ

所属部署医学研究科消化器・代謝内科学分野
職名講師
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Last Updated :2019/05/24

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  • Characterization of hepatitis B virus with complex structural variations., Fujiwara K, Matsuura K, Matsunami K, Iio E, Nojiri S, BMC microbiology, 18, (1) ,   2018年12月, 査読有り
  • TLL1 variant associated with development of hepatocellular carcinoma after eradication of hepatitis C virus by interferon-free therapy., Iio E, Matsuura K, Shimada N, Atsukawa M, Itokawa N, Abe H, Kato K, Takaguchi K, Senoh T, Eguchi Y, Nomura H, Yoshizawa K, Kang JH, Matsui T, Hirashima N, Kusakabe A, Miyaki T, Fujiwara K, Matsunami K, Tsutsumi S, Iwakiri K, Tanaka Y, Journal of gastroenterology,   2018年10月, 査読有り
  • Evaluation of antiviral effects of novel NS5A inhibitors in hepatitis C virus cell culture system, with full-genome infectious clones., Murayama A, Fujiwara K, Yamada N, Shiina M, Masaki T, Muramatsu M, Wakita T, Kato T, Antiviral research,   2018年08月, 査読有り
  • Clinical Evaluation of Hepatocarcinogenesis and Outcome Using a Novel Glycobiomarker Wisteria floribunda Agglutinin-Positive Mac-2 Binding Protein (WFA+-M2BP) in Chronic Hepatitis C with Advanced Fibrosis., Inoue T, Tsuzuki Y, Iio E, Shinkai N, Matsunami K, Fujiwara K, Matsuura K, Nojiri S, Tanaka Y, Japanese journal of infectious diseases, 71, (3) 177 - 183,   2018年05月, 査読有り
  • Gut Dysbiosis Associated With Hepatitis C Virus Infection., Inoue T, Nakayama J, Moriya K, Kawaratani H, Momoda R, Ito K, Iio E, Nojiri S, Fujiwara K, Yoneda M, Yoshiji H, Tanaka Y, Clinical infectious diseases : an official publication of the Infectious Diseases Society of America,   2018年05月, 査読有り
  • Changes in serum lipid profiles caused by three regimens of interferon-free direct-acting antivirals for patients infected with hepatitis C virus., Inoue T, Goto T, Iio E, Matsunami K, Fujiwara K, Shinkai N, Matsuura K, Matsui T, Nojiri S, Tanaka Y, Hepatology research : the official journal of the Japan Society of Hepatology, 48, (3) E203 - E212,   2018年02月, 査読有り
  • Novel non-canonical genetic rearrangements termed “complex structural variations” in HBV genome, Kei Fujiwara, Kayoko Matsunami, Etsuko Iio, Shunsuke Nojiri, Takashi Joh, Virus Research, 238, 84 - 93,   2017年06月15日, © 2017 Elsevier B.V. Backgrounds and aims: Chronic hepatitis B virus (HBV) infection is an important worldwide public health issue. Further knowledge on the characteristics of HBV will facilitate its eradication. Genome structural variations (SVs) are defined by its canonical form such as duplication, deletion, and insertion. However, recent studies have reported complex SVs that cannot be explained by those canonical SVs. A HBV strain (UK2) with an unusual genome structure rearrangement that was completely different from known mutations or rearrangements was previously reported. Thus, this study was conducted to confirm the rearrangement in UK2 as a novel complex SV, and to find additional HBV strains with complex SVs. Further, the contribution of complex SVs in hepadnavirus variability was investigated. Methods The genome rearrangement pattern in UK2 was analyzed. Further, a search of online databases retrieved additional HBV strains which were candidates to harbor complex SVs. The architecture of each rearrangement in the candidate strains was analyzed by bioinformatical tools. In addition, alignment of woolly monkey hepatitis virus (WMHV) and HBV from human and non-human primates was performed to investigate the contribution of complex SVs to variability of hepadnavirus. Results The rearrangement in UK2 was confirmed as a complex SV. An additional 15 HBV strains were retrieved from databases, and confirmed as harboring complex SVs. Complex combinations of deletion, insertion, and duplication characterized the novel rearrangements. The complex SVs in six strains (37.5%) were composed of deletion, insertion, and duplication. The complex SVs in another six strains (37.5%) consisted of deletion and insertion, followed by insertions and duplication in three strains (18.8%), and deletion and duplication in one strain (6.3%). In addition, unique preS1 promoter insertions, which contained the hepatocyte nuclear factor 1 binding site, were observed in seven (43.8%) of 16 strains. Further, analysis of the genetic sequences of WMHV and HBV from human and non-human primates showed that complex combinations of deletions and insertions accounted for their genetic differences. Conclusions Non-canonical genetic rearrangements termed complex SVs were observed in HBV. Further, complex SVs accounted for the genetic differences of WMHV and HBV from human and non-human primates.
  • Clinical evaluation of sofosbuvir/ledipasvir in patients with chronic hepatitis C genotype 1 with and without prior daclatasvir/asunaprevir therapy, Etsuko Iio, Noritomo Shimada, Koichi Takaguchi, Tomonori Senoh, Yuichiro Eguchi, Masanori Atsukawa, Akihito Tsubota, Hiroshi Abe, Keizo Kato, Keizo Kato, Atsunori Kusakabe, Tomokatsu Miyaki, Kentaro Matsuura, Kayoko Matsunami, Noboru Shinkai, Kei Fujiwara, Shunsuke Nojiri, Yasuhito Tanaka, Hepatology Research,   2017年01月01日, © 2017 The Japan Society of Hepatology. Aim: This study explored treatment outcomes of sofosbuvir (SOF)/ledipasvir (LDV) therapy for chronic hepatitis C patients with and without prior daclatasvir (DCV)/asunaprevir (ASV) therapy. Methods: Overall, 530 Japanese patients who were infected with hepatitis C virus genotype 1 received SOF/LDV therapy for 12 weeks, and resistance-associated variants (RAVs) in the hepatitis C virus non-structural protein (NS)5A and NS5B regions were assessed at baseline and virological relapse by direct sequencing. Results: Sustained virological response (SVR) rates did not significantly differ between patients with and without NS5A Y93H/N (94.2% [113/120] vs. 97.7% [345/353] ), but the SVR rate was significantly lower in patients with prior DCV/ASV therapy compared to those without (69.2% [18/26] vs. 98.4% [496/504] , P < 0.001). Among 26 patients with prior DCV/ASV therapy, the prevalence of NS5A multi-RAVs (≥2) was similar between responders and non-responders (61% [11/18] vs. 75% [5/8] ), but all patients without RAVs achieved SVR. Multivariate analysis showed that prior DCV/ASV therapy and history of hepatocellular carcinoma were independently associated with treatment failure (odds ratio, 37.55; 95% confidence interval, 10.78-130.76; P < 0.001 for prior DCV/ASV therapy; odds ratio, 4.42; 95% confidence interval, 1.09-18.04; P=0.03 for the history of HCC). All SOF/LDV failure patients (n=8) with prior DCV/ASV treatment had two or more factors of cirrhosis, IL28B unfavorable genotype, and baseline NS5A multi-RAVs. The multiple NS5A RAVs had increased but NS5B substitutions, C316N/A207T/A218S or L159F, had not changed at the time of relapse. Conclusions: Prior DCV/ASV therapy is associated with failure of SOF/LDV therapy due to multiple RAVs.
  • Efficacy of daclatasvir/asunaprevir according to resistance-associated variants in chronic hepatitis C with genotype 1, Etsuko Iio, Noritomo Shimada, Hiroshi Abe, Masanori Atsukawa, Kai Yoshizawa, Koichi Takaguchi, Yuichiro Eguchi, Hideyuki Nomura, Tomoyuki Kuramitsu, Jong Hon Kang, Takeshi Matsui, Noboru Hirashima, Akihito Tsubota, Atsunori Kusakabe, Izumi Hasegawa, Tomokatsu Miyaki, Noboru Shinkai, Kei Fujiwara, Shunsuke Nojiri, Yasuhito Tanaka, Journal of Gastroenterology, 52, (1) ,   2017年01月01日, © 2016, Japanese Society of Gastroenterology. Background: The present study explored the treatment outcome of daclatasvir (DCV) and asunaprevir (ASV) therapy combining oral direct-acting antiviral agents (DAAs) for chronic hepatitis C (HCV) including liver cirrhosis according to resistance-associated variants (RAVs) in NS3/NS5A region. Methods: Overall, 641 patients enrolled in Japan with HCV-1b received DCV and ASV for 24 weeks. Baseline drug-resistant mutations L31F/I/M/V, Q54H, P58S, A92K, and Y93H in the HCV NS5A region and V36A, T54A/S, Q80K/L/R, R155K/T/Q, A156S/V/T, and D168A/E/H/T/V in the HCV NS3/4A region were assessed by direct sequencing. Results: Overall, 86.9 % (543/625) of patients had SVR12, which was significantly higher in NS5A 93Y (wild) (88.3 %) compared with NS5A 93H at baseline (48.0 %), indicating the SVR12 rate was significantly lower in patients with 93H mutations. Additionally, 66.7 % (18/27) of patients with prior triple therapy including simeprevir (SMV) failure had virological failure. The virological failure rate of DCV/ASV therapy after SMV failure was significantly higher in those with preexisting NS3/4A 168 substitutions compared with without substitutions at baseline [84.2 % (16/19) vs. 28.6 % (2/7), p = 0.014] . The number of patients with multiple RAVs or deletions in NS5A increased from 0 to 85 % in failed patients. Alanine aminotransferase elevation was a frequent adverse event causing discontinuation of DCV/ASV therapy, although 87.5 % (14/16) patients achieved SVR12, subsequently. Conclusions: History of SMV therapy and pre-existing NS5A Y93H were associated with virological failure of DCV/ASV therapy, resulting in the emergence of multiple RAVs. Patients with RAVs at baseline should be assessed to optimize future DAA therapies.
  • Effects of branched-chain amino acid supplementation after radiofrequency ablation for hepatocellular carcinoma: A randomized trial, Shunsuke Nojiri, Kei Fujiwara, Noboru Shinkai, Etsuko Iio, Takashi Joh, Nutrition, 33, 20 - 27,   2017年01月01日, © 2016 Elsevier Inc. Objective Maintenance of liver function is important for better outcomes after radiofrequency ablation (RFA) for hepatocellular carcinoma (HCC). The aim of this study was to examine the effects of oral branched-chain amino acid (BCAA) supplementation on liver function, intrahepatic recurrence rate, and incidence of complications after RFA for HCC. Methods Patients with cirrhosis who underwent RFA were enrolled between August 2009 and April 2012, randomized to oral supplementation with Aminoleban EN (BCAA group) or diet alone (control group), and followed to determine changes in serum parameters and health status. Patients in the BCAA group were instructed to ingest a packet of Aminoleban EN twice daily. Levels of physical and mental stress were assessed using the Short Form-8 health survey. Oral BCAA and dietary interventions were initiated 2 wk before local therapy, and contrast-enhanced computed tomography was performed every 3 mo to assess recurrence. Results We evaluated 25 patients in the BCAA group and 26 in the control group. The median follow-up period was 3.9 y (736–1818 d). There were no significant differences between the two groups in basal characteristics. Complications were less frequent in the BCAA group (P = 0.03). Event-free survival was significantly higher in the BCAA group, whereas the intrahepatic recurrence rate was significantly lower (P = 0.04 and 0.036, respectively). A significant improvement in the Short Form-8 mental component score was observed in the BCAA group only (P  <  0.01). Conclusion Aminoleban EN may be beneficial for cirrhotic patients after RFA to relieve mental stress and reduce the risks for intrahepatic recurrence and complications.
  • A novel glycobiomarker, Wisteria floribunda agglutinin macrophage colony-stimulating factor receptor, for predicting carcinogenesis of liver cirrhosis, Etsuko Iio, Makoto Ocho, Makoto Ocho, Akira Togayachi, Akira Togayachi, Masanori Nojima, Atsushi Kuno, Atsushi Kuno, Yuzuru Ikehara, Yuzuru Ikehara, Izumi Hasegawa, Hiroshi Yatsuhashi, Hiroshi Yatsuhashi, Kazumi Yamasaki, Kazumi Yamasaki, Noritomo Shimada, Tatsuya Ide, Noboru Shinkai, Shunske Nojiri, Kei Fujiwara, Takashi Joh, Masashi Mizokami, Masashi Mizokami, Hisashi Narimatsu, Hisashi Narimatsu, Yasuhito Tanaka, Yasuhito Tanaka, International Journal of Cancer, 138, (6) 1462 - 1471,   2016年01月, © 2015 UICC. Recently, we identified a novel liver fibrosis glycobiomarker, Wisteria floribunda agglutinin (WFA)-reactive colony stimulating factor 1 receptor (WFA + -CSF1R), using a glycoproteomics-based strategy. The aim of this study was to assess the value of measuring WFA + -CSF1R levels for the prognosis of carcinogenesis and outcome in liver cirrhosis (LC) patients with hepatitis C virus (HCV). WFA + -CSF1R and Total-CSF1R levels were measured in serum samples from 214 consecutive HCV-infected patients to evaluate their impact on carcinogenesis and the survival of LC patients. Serum WFA + -CSF1R levels were significantly higher in LC patients than chronic hepatitis (CH) patients (p < 0.001). The AUC of WFA + -CSF1R for predicting overall survival, calculated by time-dependent ROC analysis, was 0.691 and the HR (per 1-SD increase) was 1.80 (95% CI, 1.23-2.62, p < 0.001). Furthermore, the survival rate of LC patients with high WFA + -CSF1R levels (≥310 ng/ml) was significantly worse than those with lower levels (p < 0.01). The AUC of WFA + /total-CSF1R percentage (WFA + -CSF1R%) for predicting the cumulative carcinogenesis rate was 0.760, with an HR of 1.66 (95% CI 1.26-2.20, p < 0.001). In fact, the carcinogenesis rate was significantly higher in LC patients with a high WFA + -CSF1R% (≥ 35%, p = 0.006). Assessing serum levels of WFA + -CSF1R has diagnostic value for predicting carcinogenesis and the survival of LC patients. What's new? To find better biomarkers of liver fibrosis the authors developed a novel assay to detect serum levels of Wisteria floribunda agglutinin (WFA)-reactive colony-stimulating factor 1 receptor (WFA + -CSF1R). CSF1R is a membrane glycoprotein associated with cell proliferation and found elevated in a variety of cancers. They report that in hepatitis C virus-infected patients who developed liver cirrhosis, the survival rate of patients with high WFA + -CSF1R levels was significantly worse. Similarly, the carcinogenesis rate was higher in patients with high percentage of WFA + -CSF1R, suggesting that this glycobiomarker may have clinical applications beyond the mere evaluation of liver fibrosis.
  • Influence of genes suppressing interferon effects in peripheral blood mononuclear cells during triple antiviral therapy for chronic hepatitis C, Sayuki Iijima, Kentaro Matsuura, Kentaro Matsuura, Kentaro Matsuura, Tsunamasa Watanabe, Koji Onomoto, Takashi Fujita, Kyoko Ito, Etsuko Iio, Etsuko Iio, Tomokatsu Miyaki, Kei Fujiwara, Noboru Shinkai, Atsunori Kusakabe, Mio Endo, Shunsuke Nojiri, Takashi Joh, Yasuhito Tanaka, PLoS ONE, 10, (2) ,   2015年02月23日, © 2015 Iijima et al. The levels of expression of interferon-stimulated genes (ISGs) in liver are associated with response to treatment with pegylated interferon (PEG-IFN) plus ribavirin (RBV). However, associations between the responses of ISGs to IFN-based therapy and treatment efficacy or interleukin-28B (IL28B) genotype have not yet been determined. Therefore, we investigated the early responses of ISGs and interferon-lambdas (IFN-λs) in peripheral blood mononuclear cells (PBMCs) during PEG-IFN/RBV plus NS3/4 protease inhibitor (PI) therapy. We prospectively enrolled 50 chronic hepatitis C patients with HCV genotype 1, and collected PBMCs at baseline, 8 and 24 h after the initial administration of PEG-IFN/RBV/PI. Levels of mRNAs for selected ISGs and IFN-λs were evaluated by real-time PCR. All 31 patients with a favorable IL28B genotype and 13 of 19 with an unfavorable genotype achieved sustained virological responses (SVR). Levels of mRNA for A20, SOCS1 , and SOCS3, known to suppress antiviral activity by interfering with the IFN signaling pathway, as well as IRF1 were significantly higher at 8 h in patients with an unfavorable IL28B genotype than in those with a favorable one (P = 0.007, 0.026, 0.0004, 0.0006, respectively), especially in the non-SVR group. Particularly, the fold-change of IRF1 at 8 h relative to baseline was significantly higher in non-SVR than in SVR cases with an unfavorable IL28B genotype (P = 0.035). In conclusion, levels of several mRNAs of genes suppressing antiviral activity in PBMCs during PEG-IFN/RBV/PI differed according to IL28B genotypes, paralleling treatment efficacy.
  • Evaluation of hepatocellular carcinoma development in patients with chronic hepatitis C by EOB-MRI, Shunsuke Nojiri, Kei Fujiwara, Noboru Shinkai, Mio Endo, Takashi Joh, World Journal of Hepatology, 6, (12) 930 - 938,   2014年01月01日, © 2014 Baishideng Publishing Group Inc. Aim: To evaluate the efficacy of ethoxibenzyl-magnetic resonance imaging (EOB-MRI) as a predictor of hepatocellular carcinoma (HCC) development. Methods: Between August 2008 and 2009, we studied 142 hepatitis C virus-infected patients (male 70, female 72), excluding those with HCC or a past history, who underwent EOB-MRI in our hospital. The EOB-MRI index [liver-intervertebral disc ratio (LI)] was calculated as: (post-liver intensity/post-intervertebral disc intensity)/(pre-liver intensity/pre-intervertebral disc intensity). Results: The median follow-up period was 3.1 years and the patients were observed until the end of the study period (31 December, 2012). In the follow-up period, HCC occurred in 21 patients. The cumulative occurrence rates were 2.1%, 9.1%, and 14.1% at 1, 2, and 3 years, respectively. Using the optimal cut-off value of LI 1.46, on univariate analysis, age, aspartate amino transferase (AST), α-fetoprotein (AFP) ≥ 10, albumin, total cholesterol, prothrombin time, platelets, and LI < 1.46 were identified as independent factors, but on multivariate analysis, LI < 1.46: risk ratio 6.05 (1.34-27.3, P = 0.019) and AFP ≥ 10: risk ratio 3.1 (1.03-9.35, P = 0.045) were identified as independent risk factors. LI and Fib-4 index have higher area under the receiver operating characteristic curves than other representative fibrosis evaluation methods, such as Forn's index and AST-to-platelet ratio index. Conclusion: LI is associated with the risk of HCC occurrence in hepatitis C patients. LI may be a substitute for liver biopsy when evaluating this risk and its combined use with Fib-4 is a better predictive method of HCC progression.
  • Serum interferon-gamma-inducible protein-10concentrations and IL28B genotype associated with responses to pegylated interferon plus ribavirin with and without telaprevir for chronic hepatitis C, Kentaro Matsuura, Tsunamasa Watanabe, Sayuki Iijima, Shuko Murakami, Kei Fujiwara, Etsuro Orito, Etsuko Iio, Mio Endo, Atsunori Kusakabe, Noboru Shinkai, Tomokatsu Miyaki, Shunsuke Nojiri, Takashi Joh, Yasuhito Tanaka, Hepatology Research, 44, (12) 1208 - 1216,   2014年01月, © 2013 The Japan Society of Hepatology. Aim: Several studies have shown that high pretreatment concentrations of serum interferon-γ-inducible protein-10 (IP-10) are correlated with non-response to pegylated interferon (PEG-IFN) plus ribavirin (RBV) for chronic hepatitis C (CHC). However, there are few reports on their effect on the Asian population. Methods: We enrolled 104 Japanese genotype 1 CHC individuals treated with PEG-IFN/RBV and 45 with PEG-IFN/RBV/telaprevir, and evaluated the impact of pretreatment serum IP-10 concentrations on their virological responses. Results: The pretreatment serum IP-10 concentrations were not correlated with IL28B genotype. The receiver-operator curve analysis determined the cut-off value of IP-10 for predicting a sustained virological response (SVR) as 300 pg/mL. In multivariate analysis, the IL28B favorable genotype and IP-10 concentration of less than 300 pg/mL were independent factors for predicting SVR. In a subgroup of patients with the IL28B favorable genotype, the SVR rate was higher in the patients with IP-10 of less than 300 than in those with 300 pg/mL or more, whereas no patient with the IL28B unfavorable genotype and IP-10 of 300 pg/mL or more achieved SVR. Among the patients treated with PEG-IFN/RBV/telaprevir, low pretreatment concentrations of serum IP-10 were associated with a very rapid virological response, defined as undetectable HCV RNA at week 2 after the start of therapy. Conclusion: Pretreatment serum IP-10 concentrations are associated with treatment efficacy in PEG-IFN/RBV and with early viral kinetics of hepatitis C virus in PEG-IFN/RBV/telaprevir therapy.
  • A case of a HBV carrier with HDV superinfection treated by PEG-IFN, Atsunori Kusakabe, Yasuhito Tanaka, Etsuko Iio, Shuko Murakami, Kentaro Matsuura, Noboru Shinkai, Tomokatsu Miyaki, Kei Fujiwara, Shunsuke Nojiri, Etsuro Orito, Takashi Joh, Acta Hepatologica Japonica, 55, (11) 653 - 660,   2014年01月01日, © 2014 The Japan Society of Hepatology. A 26-year-old Mongolian woman was admitted to our hospital because of liver dysfunction As she has been a HBV carrier, acute exacerbation of chronic hepatitis B seemed to be the cause of liver dysfunction at first. However, the loads of serum HBV-DNA on admission were low (2.8 log copy/ml). As she was pregnant, she was observed without treatment. Liver function once improved without treatment, but it became worse again after delivery. At this time, it was suspected HDV superinfection could affect the liver dysfunction. For HDV-RNA was positive in stored sera by RT-PCR, she was diagnosed as HBV and HDV superinfection and started treatment with Peg-IFN. Although HDV infection is rare in Japan, in case of acute exacerbation of a HBV carrier with low serum HBV-DNA level, HBV and HDV superinfection should be considered.
  • ITPA genetic variants influence efficacy of PEG-IFN/RBV therapy in older patients infected with HCV genotype 1 and favourable IL28B type, K. Matsuura, Y. Tanaka, T. Watanabe, K. Fujiwara, E. Orito, M. Kurosaki, N. Izumi, N. Sakamoto, N. Sakamoto, N. Enomoto, H. Yatsuhashi, A. Kusakabe, N. Shinkai, S. Nojiri, T. Joh, M. Mizokami, Journal of Viral Hepatitis, 21, (7) 466 - 474,   2014年01月, Summary Inosine triphosphatase (ITPA) genetic variants are strongly associated with ribavirin (RBV)-induced anaemia during pegylated interferon (PEG-IFN) plus RBV therapy. However, the treatment efficacy of ITPA genetic variants has not been fully explored. We enrolled 309 individuals infected with hepatitis C virus genotype 1, who were treated with PEG-IFN plus RBV for 48 weeks. The ITPA SNP: rs1127354 and IL28B SNP: rs8099917 were genotyped. We examined the risk factors for severe anaemia up to week 12 after the start of treatment and treatment efficacy. The incidence of severe anaemia, ≥3 g/dL reduction or < 10 g/dL of haemoglobin (Hb) up to week 12, was more frequent in patients with CC at rs1127354 [65% (145/224), 33% (73/224)] than in those with CA/AA [25% (21/85), 6% (8/85)] (P < 0.0001). ITPA genotype, pretreatment Hb level and age were independent predictive factors for severe anaemia: Hb < 10 g/dL. In IL28B favourable type, the sustained virologic response rate was higher in ≥60-year-old patients with CA/AA than in those with CC [71% (22/31) vs 40% (26/65), P = 0.005], although there was no significant difference in treatment efficacy according to ITPA genetic variants in the < 60-year-old patients. The proportion of patients administered ≥80% of the dosage of RBV was significantly higher in the patients with CA/AA than in those with CC (P = 0.025), resulting in a lower relapse rate. In conclusion, ITPA genetic variants were associated with severe RBV-induced anaemia and could influence the efficacy of PEG-IFN plus RBV treatment among elderly patients with IL28B favourable type. © 2013 John Wiley & Sons Ltd.
  • Investigation of residual hepatitis C virus in presumed recovered subjects, Kei Fujiwara, Robert D. Allison, Robert D. Allison, Richard Y. Wang, Patricia Bare, Patricia Bare, Kentaro Matsuura, Cathy Schechterly, Krishna Murthy, Francesco M. Marincola, Harvey J. Alter, Hepatology, 57, (2) 483 - 491,   2013年02月, Recent studies have found hepatitis C virus (HCV) RNA in peripheral blood mononuclear cells (PBMCs) of the majority of presumed recovered subjects. We investigated this unexpected finding using samples from patients whose HCV RNA and anti-HCV status had been serially confirmed. HCV RNA was detected in PBMCs from 66 of 67 chronic HCV carriers. Subpopulation analysis revealed that the viral load (log copies/10 6 cells) in B cells (4.14 ± 0.71) was higher than in total PBMCs (3.62 ± 0.71; P < 0.05), T cells (1.67 ± 0.88; P < 0.05), and non-B/T cells (2.48 ± 1.15; P < 0.05). HCV negative-strand RNA was not detected in PBMCs from any of 25 chronically infected patients. No residual viral RNA was detected in total PBMCs or plasma of 59 presumed recovered subjects (11 spontaneous and 48 treatment induced) using nested real-time polymerase chain reaction with a detection limit of 2 copies/μg RNA (from ∼1 × 10 6 cells). PBMCs from 2 healthy HCV-negative blood donors became HCV RNA positive, with B-cell predominance, when mixed in vitro with HCV RNA-positive plasma, thus passively mimicking cells from chronic HCV carriers. No residual HCV was detected in liver or other tissues from 2 spontaneously recovered chimpanzees. Conclusion: (1) HCV RNA was detected in PBMCs of most chronic HCV carriers and was predominant in the B-cell subpopulation; (2) HCV detected in PBMCs was in a nonreplicative form; (3) HCV passively adsorbed to PBMCs of healthy controls in vitro, becoming indistinguishable from PBMCs of chronic HCV carriers; and (4) residual HCV was not detected in plasma or PBMCs of any spontaneous or treatment-recovered subjects or in chimpanzee liver, suggesting that the classic pattern of recovery from HCV infection is generally equivalent to viral eradication. © 2012 American Association for the Study of Liver Diseases.
  • Noninvasive evaluation of hepatic fibrosis in hepatitis C virus-infected patients using ethoxybenzyl-magnetic resonance imaging, Shunsuke Nojiri, Atsunori Kusakabe, Kei Fujiwara, Noboru Shinkai, Kentaro Matsuura, Etsuko Iio, Tomokatsu Miyaki, Takashi Joh, Journal of Gastroenterology and Hepatology (Australia), 28, (6) 1032 - 1039,   2013年01月01日, Background and Aims: Liver biopsy is the gold standard test to determine the grade of fibrosis, but there are associated problems. Because gadolinium-ethoxybenzyl-diethylenetriamine pentaacetic acid is secreted partially in hepatocytes and bile, it is possible that ethoxybenzyl-magnetic resonance imaging (EOB-MRI) correlates with liver function and liver fibrosis. The aim of this study was to compare the fibrosis seen in liver biopsy samples to the signal intensity of the hepatobiliary phase measured on EOB-MRI in hepatitis C virus (HCV)-infected patients. Methods: Two hundred twenty-four (estimation 149, validation 75) HCV-infected patients with histologically proven liver tissue who underwent EOB-MRI were studied. Overall, fibrosis staging was 15/24/19/46/45 for F0/F1/F2/F3/F4, respectively. A 1.5-Tesla magnetic resonance system was used, and the regions of interest of the liver were measured. Four methods were used: (i) relative enhancement: (post-enhanced signal intensity [SI] - pre-enhanced intensity)/pre-enhanced intensity; (ii) liver-to-intervertebral disk ratio (LI): post-enhanced (liver SI/interdisc SI)/pre-enhanced (liver SI/inter disc SI); (iii) liver-to-muscle ratio: post-enhanced (liver SI/muscle SI)/pre-enhanced (liver SI/muscle SI); and (iv) liver-to-spleen ratio: post-enhanced (liver SI/spleen SI)/pre-enhanced (liver SI/spleen SI). Results: To discriminate F0-1 versusF2-4 or F0-2 versusF3-4 or F0-3 versusF4, LI at 25min (LI25) had the highest area under receiver operating characteristic (0.88, 0.87, and 0.87, respectively) in these four methods and also in the validation set. Conclusion: LI at 25min seems to be a useful method to determine the staging of fibrosis as a non-invasive method in HCV-infected hepatitis or cirrhosis patients. © 2013 Journal of Gastroenterology and Hepatology Foundation and Wiley Publishing Asia Pty Ltd.
  • Clinical factors related to long-term administration of sorafenib in patients with hepatocellular carcinoma, Shunsuke Nojiri, Atsunori Kusakabe, Kei Fujiwara, Noboru Shinkai, Kentaro Matsuura, Etsuko Iio, Tomokatsu Miyaki, Tomoyuki Nomura, Satoshi Sobue, Hitoshi Sano, Izumi Hasegawa, Tomoyoshi Ohno, Yoshitsugu Takahashi, Etsuro Orito, Takashi Joh, Cancer Management and Research, 4, (1) 423 - 429,   2012年12月17日, Background: Sorafenib has been approved in the indication of unresectable hepatocellular carcinoma, but there are many cases in which administration of the drug is discontinued due to severe side effects. In this study, we compared the characteristics of patients who continued and discontinued sorafenib. Methods: Ninety-six patients (75 men and 21 women) were initiated on sorafenib from July 2009 through September 2011. The patient characteristics of interest included gender, age, etiology, Child-Pugh classification, treatment history and frequency, and levels of α-fetoprotein, des-gamma-carboxy prothrombin, aspartate amino acid transferase, and alanine aminotransferase. Duration of administration of sorafenib and reasons for its discontinuation were compared. Results: Median overall survival was 11.8 months. Discontinuation of sorafenib within 90 days was identified as an independent prognostic factor for overall survival on multivariate analysis (P < 0.0001). Transarterial chemoembolization performed six times or more (P = 0.013) was also identified as an independent factor contributing to discontinuation of sorafenib within 90 days in multivariate analysis. Patients who received sorafenib for ≥90 days had significantly longer overall survival than those who discontinued it (P < 0.0001). Conclusion: Prolonged treatment with sorafenib is an important factor in achieving extended overall survival. We recommend starting sorafenib before latent liver damage has occurred as a result of too many transarterial chemoembolization procedures. © 2012 Nojiri et al, publisher and licensee Dove Medical Press Ltd.
  • Quantitation of HBsAg predicts response to entecavir therapy in HBV genotype C patients, Etsuro Orito, Kei Fujiwara, Hiroshi Kanie, Tesshin Ban, Tomonori Yamada, Katsumi Hayashi, World Journal of Gastroenterology, 18, (39) 5570 - 5575,   2012年12月01日, AIM: To analysis the factors that predict the response to entecavir therapy in chronic hepatitis patients with hepatitis B virus (HBV) genotype C. METHODS: Fifty patients [hepatitis B e antigen (HBeAg)- negative:HBeAg-positive = 26:24] with HBV genotype C, who received naïve entecavir therapy for > 2 years, were analyzed. Patients who showed HBV DNA levels ≥ 3.0 log viral copies/mL after 2 years of entecavir therapy were designated as slow-responders, while those that showed < 3.0 log copies/mL were termed rapidresponders. Quantitative hepatitis B surface antigen (HBsAg) levels (qHBsAg) were determined by the Architect HBsAg QT immunoassay. Hepatitis B core-related antigen was detected by enzyme immunoassay. Pre-C and Core promoter mutations were determined using by polymerase chain reaction (PCR). Drug-resistance mutations were detected by the PCR-Invader method. RESULTS: At year 2, HBV DNA levels in all patients in the HBeAg-negative group were < 3.0 log copies/mL. In contrast, in the HBeAg-positive group, 41.7% were slow-responders, while 58.3% were rapid-responders. No entecavir-resistant mutants were detected in the slow-responders. When the pretreatment factors were compared between the slow- and rapid-responders; the median qHBsAg in the slow-responders was 4.57 log IU/mL, compared with 3.63 log IU/mL in the rapidresponders (P < 0.01). When the pretreatment factors predictive of HBV DNA-negative status at year 2 in all 50 patients were analyzed, HBeAg-negative status, low HBV DNA levels, and low qHBsAg levels were significant (P < 0.01). Multivariate analysis revealed that the low qHBsAg level was the most significant predictive factor (P = 0.03). CONCLUSION: Quantitation of HBsAg could be a useful indicator to predict response to entecavir therapy. © 2012 Baishideng. All rights reserved.
  • Combination of hepatitis B viral antigens and DNA for prediction of relapse after discontinuation of nucleos(t)ide analogs in patients with chronic hepatitis B, Akihiro Matsumoto, Eiji Tanaka, Yoshiyuki Suzuki, Mariko Kobayashi, Yasuhito Tanaka, Noboru Shinkai, Shuhei Hige, Hiroshi Yatsuhashi, Shinya Nagaoka, Kazuaki Chayama, Masataka Tsuge, Osamu Yokosuka, Fumio Imazeki, Shuhei Nishiguchi, Masaki Saito, Kei Fujiwara, Nobuyuki Torii, Naoki Hiramatsu, Yoshiyasu Karino, Hiromitsu Kumada, Hepatology Research, 42, (2) 139 - 149,   2012年02月01日, Aim: The factors associated with hepatitis recurrence after discontinuation of nucleos(t)ide analogs (NAs) in patients with chronic hepatitis B were analyzed to predict the risk of relapse more accurately. Methods: A total of 126 patients who discontinued NA therapy were recruited retrospectively. The clinical conditions of a successful discontinuation were set as alanine aminotransferase (ALT) below 30IU/L and serum hepatitis B virus (HBV) DNA below 4.0 log copies/mL. Result s: Relapse of hepatitis B were judged to occur when maximal serum ALT became higher than 79IU/L or when maximal serum HBV DNA surpassed 5.7 log copies/mL following NA discontinuation since these values corresponded with mean values of ALT (30IU/L) and HBV DNA (4.0 log copies/mL), respectively. At least 90% of patients with either detectable hepatitis B e antigen or serum HBV DNA higher than 3.0 log copies/mL at the time of NA discontinuation relapsed within one year. In the remaining patients, higher levels of both hepatitis B surface and core-related antigens at the time of discontinuation, as well as a shorter course of NA treatment, were significantly associated with relapse by multivariate analysis. Conclusions: It appears that negative results for hepatitis B e antigen and serum HBV DNA lower than 3.0 log copies/mL are essential for successful NA discontinuation, which may be attained by a longer treatment period. Levels of hepatitis B surface and core-related antigens are also significant factors independently associated with relapse of hepatitis. © 2011 The Japan Society of Hepatology.
  • Efficacy and safety of S-1 plus docetaxel combination therapy for patients with advanced gastric cancer, Hirotaka Nishiwaki, Tomonori Yamada, Keiichirou Tachi, Hiroyasu Iwasaki, Yasuki Hori, Shuuichirou Umemura, Yoshihiro Yamakawa, Takuya Kurimoto, Tesshin Ban, Kei Fujiwara, Katsumi Hayashi, Akira Nemoto, Junko Shiroko, Etsurou Orito, Japanese Journal of Cancer and Chemotherapy, 36, 1663 - 1666,   2009年01月01日, Recently, the significant efficacy of S-1 monotherapy or S-1 plus CDDP combination therapy has been reported. Docetaxel also has been reported to have favorable efficacy in gastric cancer. In addition, docetaxel can be administered in outpatient clinics. We investigated the efficacy and safety of S-1 plus docetaxel combination therapy for 35 naive patients with advanced gastric cancer. Docetaxel was administered at a dose of 40 mg/m 2 on day 1, and oral S-1 was administered at the full dose of 80 mg/m 2 twice daily for two weeks followed by one week rest. MST was 300 days, and the response rate was 42.9%. Although leucopenia was observed in 31%, all patients were able to be continue this therapy. In conclusion, we considered that this S-1 plus docetaxel combination therapy was effective and safe in advanced gastric cancer, and convenient for outpatients.
  • Comparison of complete sequences of hepatitis B virus genotype C between inactive carriers and hepatocellular carcinoma patients before and after seroconversion, Kiyoaki Ito, Yasuhito Tanaka, Michio Kato, Kei Fujiwara, Kei Fujiwara, Fuminaka Sugauchi, Tomoyuki Sakamoto, Tomoyuki Sakamoto, Noboru Shinkai, Noboru Shinkai, Etsuro Orito, Masashi Mizokami, Journal of Gastroenterology, 42, (10) 837 - 844,   2007年10月, Background: Most patients who acquire chronic hepatitis B virus (HBV) infection by perinatal transmission become inactive carriers (IC) after hepatitis B e (HBe) antigen seroconversion, whereas some patients have persistent abnormal serum transaminase levels and develop hepatocellular carcinoma (HCC) in the anti-HBe-positive phase. The aim of this study was to investigate the HCC-related mutations of HBV. Methods: Complete sequences of HBV were examined among eight IC and eight HCC patients infected with HBV genotype C before and after seroconversion. Results: The frequency of the T1653 mutation tended to be higher among HCC patients after seroconversion (16.7% vs. 62.5%; P = 0.086). The prevalence of a basal core promoter double mutation (T1762/A1764) was high among both IC and HCC patients after seroconversion (83.3% vs. 87.5%; P = 0.825). Among the HCC patients, a pre-S deletion mutant was detected in 62.5% patients before seroconversion, and in 37.5% patients after seroconversion. The core deletion mutant was also detected in 50% of HCC patients only before seroconversion. Deletion mutants of the pre-S or core region before seroconversion were significantly associated with HCC patients (0% vs. 62.5%; P = 0.007, 0% vs. 50%; P = 0.021, respectively). Conclusions: Our data showed a significant association of pre-S and core deletion mutants before seroconversion with HCC development. The T1653 mutation after seroconversion was frequently found in HCC patients infected with HBV genotype C. These results suggest that mutations may be predictive factor for development of HCC. © Springer-Verlag Tokyo 2007.
  • Impaired cytotoxic T lymphocyte inductivity by dendritic cells derived from patients with hepatitis C virus-positive hepatocellular carcinoma, Tomoyoshi Ohno, Noboru Hirashima, Etsuro Orito, Izumi Hasegawa, Kei Fujiwara, Kiyoaki Itoh, Atsushi Ozasa, Noboru Shinkai, Yasuhito Tanaka, Takanobu Kato, Ryuzo Ueda, Kenji Sakakibara, Hepatology Research, 37, (4) 276 - 285,   2007年04月01日, Aim: Peptide-based therapeutic vaccines are being developed. The aim of this study was to determine the feasibility of immunotherapy to hepatitis C virus (HCV)-positive hepatocellular carcinoma (HCC) by assessing the inductivity of peptide-specific cytotoxic T lymphocyte (CTL) by dendritic cells. Methods: The inductivity of CTL was characterized in six patients with HCV-positive HCC, and compared to seven healthy volunteers and six patients with chronic HCV hepatitis (control). Results: Peptide-specific CTL was comparably induced in controls, but not induced in any patients with HCC. To characterize this, the cytokine profile and the expression of surface molecules interacting between dendritic and T cells were evaluated. Among the cytokines, production of interferon (IFN)-γ was found to be impaired and closely related to the results of CTL assays, while the expression of surface molecules showed no significant changes. Conclusions: In HCV-positive HCC patients, CTL inductivity by dendritic cells is impaired. This may be related to the impaired production of IFN-γ. © 2007 The Japan Society of Hepatology.
  • Molecular epidemiology of hepatitis B virus in the United Republic of Tanzania, Izumi Hasegawa, Yasuhito Tanaka, Fuat Kurbanov, Namiko Yoshihara, Ahmed El-Gohary, Eligius Lyamuya, Mecky Matee, Pius Magessa, Kei Fujiwara, Kei Fujiwara, Atsushi Ozasa, Atsushi Ozasa, Fuminaka Sugauchi, Etsuro Orito, Ryuzo Ueda, Masashi Mizokami, Masashi Mizokami, Journal of Medical Virology, 78, (8) 1035 - 1042,   2006年08月, In the United Republic of Tanzania, 457 voluntary blood donors were enrolled in hepatitis B virus (HBV) serological screening; 4.8% (22/457) carried HBsAg, 13.6% (3/22) of whom were HBeAg-positive. The mean age among HBeAg-negative carriers was 31 years, HBV DNA was detectable in 81.8% (18/22), the mean level was 3.67 (±1.77) log copies/ml. Genotype A was determined in 90.9% (20/22) and 18/20 were classified into subgenotype Aa (Asia/Africa), The basal core promoter, precore and partial core nucleotide sequences were analyzed in the 18 strains; T1809/T1812 ("Kozak" sequence) and A/T1888 (encapsidation signal) variants were identified in 100% and 78%, respectively. The complete genome sequencing for one of the Tanzanian strains revealed no recombination. In conclusion, HBV seroprevalence is high among general population in Tanzania, and the HBV/Aa-infection is predominant. The indicated tendency to early HBeAg seroconversion and declining of the viral load should be confirmed further in case-control studies. © 2006 Wiley-Liss, Inc.
  • A case-control study of response to lamivudine therapy for 2 years in Japanese and Chinese patients chronically infected with hepatitis B virus of genotypes Bj, Ba and C, Etsuro Orito, Kei Fujiwara, Yasuhito Tanaka, Man Fung Yuen, Ching Lung Lai, Takanobu Kato, Fuminaka Sugauchi, Atsunori Kusakabe, Michio Sata, Takeshi Okanoue, Hirofumi Niitsuma, Hiroshi Sakugawa, Izumi Hasegawa, Masashi Mizokami, Hepatology Research, 35, (2) 127 - 134,   2006年06月01日, Background/aims: In eastern Asian countries, hepatitis B virus (HBV) genotype Ba (HBV/Ba), HBV/Bj and HBV/C are prevalent. The aim was to investigate the response or resistance to lamivudine therapy among patients with different HBV genotypes. Methods: Of 67 Japanese and Chinese patients with chronic hepatitis B, 18 patients with HBV/Bj, 15 with HBV/Ba and 34 with HBV/C were selected for a case-control study matched according to gender and age. All the patients were treated with lamivudine for 2 years and evaluated the response or emergence of the YMDD mutation at year 2 during the treatment. HBV genotypes were detected by the restriction fragment length polymorphism. The YMDD mutation was detected by the direct sequencing after amplification by PCR. Results: At year 2 during therapy, 44.8% of the patients showed normalization of ALT and undetectable HBV DNA (favorable response), 35.8% developed the YMDD mutation. There was no significant difference of response to the therapy among the three genotype groups. The emergence of the YMDD mutation was associated with HBV/C. By the multiple logistic regression analysis, however, the significant factor of a favorable response was a higher pretreatment ALT level and negative HBeAg status and the significant factor of the emergence of the YMDD mutation was HBV/C. Conclusions: Higher pretreatment ALT level, HBeAg status or HBV genotype may affect the response or resistance to lamivudine therapy. © 2006 Elsevier Ireland Ltd. All rights reserved.
  • T1653 mutation in the box α increases the risk of hepatocellular carcinoma in patients with chronic hepatitis B virus genotype C Infection, Kiyoaki Ito, Yasuhito Tanaka, Etsuro Orito, Masaya Sugiyama, Kei Fujiwara, Fominaka Sugauchi, Takanobu Kato, Hajime Tokita, Namiki Izumi, Michio Kato, Man Fung Yuen, Ching Lung Lai, Robert G. Gish, Ryuzo Ueda, Masashi Mizokemi, Masashi Mizokemi, Clinical Infectious Diseases, 42, (1) 1 - 7,   2006年01月, Background. Most patients with chronic hepatitis B virus infection become carriers of inactive virus after hepatitis B e antigen seroconversion; however, a subgroup of patients have persistent abnormal transaminase levels and develop hepatocellular carcinoma after seroconversion. Methods. In an age-matched case-control study, 40 carriers of inactive virus (mean age ± standard deviation [SD], 50.9 ± 11.1 years), 40 parents with chronic hepatitis (mean age ± SD, 50.2 ± 8.9 years), and 40 patients with hepatocellular carcinoma (mean age ± SD, 50.7 ± 9.4 years) who were infected with hepatitis B virus genotype C and had test results positive for antibody to hepatitis B e antigen were analyzed. Results. The prevalence of T1653 in the box α was significantly higher among patients with hepatocellular carcinoma than among carriers|of inactive virus who did not have hepatocellular carcinoma (70% vs. 25%; P < .0001) or chronic hepatitis (70% vs. 35%; P = .003). Mutations in the basic core promoter region (T1762/A1764) were frequently found in all groups, regardless of clinical status (in 77.5% of carriers of inactive virus, 77.5% of patients with chronic hepatitis, and 90% of patients with hepatocellular carcinoma). In the multivariate analysis, the presence of T1653, an alanine aminotransferase level of ≥ 37 U/L, and a platelet count of < 18 × 10 4 platelets/ mm 3 were independent predictive values for hepatocellular carcinoma (odds ratio [95% confidence interval], 5.05 [1.56-16.35] , 12.56 [3.05-51.77], and 11.5 [3.47-38.21] , respectively). High α-fetoprotein level was the only independent predictive value for T1653 in patients with hepatocellular carcinoma (odds ratio, 12.67; 95% confidence interval, 1.19-134.17]). Among patients with test results positive for antibody to hepatitis B e antigen who had hepatocellular carcinoma and were infected with different genotypes of hepatitis B virus, the prevalence of T1653 was 40%, 15%, 25%, 25%, 67%, and 23% in patients infected with hepatitis B virus genotypes Aa, Ae, Ba, Bj, C, and D, respectively (P < .05 for genotype C vs. genotypes Ae, Ba, Bj, or D). Conclusions. Our data indicate that the addition of T1653 mutation in the box a to the basic core promoter mutation increases the risk of hepatocellular carcinoma in patients with hepatitis B virus genotype C. © 2005 by the Infectious Diseases Society of America. All rights reserved.
  • Distribution of HBV genotypes among HBV carriers in Benin: Phyologenetic analysis and virological characteristics of HBV genotype E, Kei Fujiwara, Yasuhito Tanaka, Etsuro Orito, Tomoyoshi Ohno, Takanobu Kato, Kanji Sugihara, Izumi Hasegawa, Mayumi Sakurai, Kiyoaki Ito, Atsushi Ozasa, Yuko Sakamoto, Isao Arita, Ahmed El-Gohary, Agossou Benoit, Sophie I. Ogoundele-Akplogan, Namiko Yoshihara, Ryuzo Ueda, Masashi Mizokami, World Journal of Gastroenterology, 11, 6410 - 6415,   2005年11月07日, Aim: To determine the distribution of Hepatitis B virus (HBV) genotypes in Benin, and to clarify the virological characteristics of the dominant genotype. Methods: Among 500 blood donors in Benin, 21 HBsAg-positive donors were enrolled in the study. HBV genotypes were determined by enzyme immunoassay and restriction fragment length polymorphism. Complete genome sequences were determined by PCR and direct sequencing. Results: HBV genotype E (HBV/E) was detected in 20/21 (95.2%), and HBV/A in 1/21 (4.8%). From the age-specific prevalence of HBeAg to anti-HBe seroconversion (SC) in 19 HBV/E subjects, SC was estimated to occur frequently in late teens in HBV/E. The comparison of four complete HBV/E genomes from HBeAg-positive subjects in this study and five HBV/E sequences recruited from the database revealed that HBV/E was distributed throughout West Africa with very low genetic diversity (nucleotide homology 96.7-99.2%). Based on the sequences in the basic core promoter (BCP) to precore region of the nine HBV/E isolates compared to those of the other genotypes, a nucleotide substitution in the BCP, G1757A, was observed in HBV/E. Conclusion: HBV/E is predominant in the Republic of Benin, and SC is estimated to occur in late teens in HBV/E. The specific nucleotide substitution G1757A in BCP, which might influence the virological characteristics, is observed in HBV/E. © 2005 The WJG Press and Elsevier Inc. All rights reserved.
  • Novel type of hepatitis B virus mutation: Replacement mutation involving a hepatocyte nuclear factor 1 binding site tandem repeat in chronic hepatitis B virus genotype E, Kei Fujiwara, Yasuhito Tanaka, Emma Paulon, Etsuro Orito, Masaya Sugiyama, Kiyoaki Ito, Ryuzo Ueda, Masashi Mizokami, Masashi Mizokami, Nikolai V. Naoumov, Journal of Virology, 79, (22) 14404 - 14410,   2005年11月, The genetic diversity of hepatitis B virus (HBV) strains has evolved through mutations such as point mutations, deletions or insertions, and recombination. We identified and characte rized a novel type of mutation which is a complex of external insertion, deletion, and internal duplication in sequences from one of six patients with chronic hepatitis B virus genotype E (HBV/E). We provisionally named this mutation a "replacement mutation"; the core promoter upstream regulatory sequence/basic core promoter was replaced with a part of the S1 promoter covering the hepatocyte nuclear factor 1 (HNF1) binding site, followed by a tandem repeat of the HNF1 site. A longitudinal analysis of the HBV population over 6 years showed the clonal change from wild-type HBV/E to replacement-mutant type, resulting in a lower hepatitis B (HB) e antigen titer, a high HBV DNA level in serum, and progression of liver fibrosis. In an in vitro study using a replication model, the replacement-mutant HBV showed higher replication levels than the wild-type HBV/E replicon, probably mediated by altered transcription factor binding. Additionally, this HNF1 site replacement mutation was associated with excessive HB nucleocapsid protein expression in hepatocytes, in both in vivo and in vitro studies. This novel mutation may be specific to HBV genotype E, and its prevalence requires further investigation. Copyright © 2005, American Society for Microbiology. All Rights Reserved.
  • Detection of anti-hepatitis C virus effects of interferon and ribavirin by a sensitive replicon system, Takanobu Kato, Tomoko Date, Michiko Miyamoto, Masaya Sugiyama, Yasuhito Tanaka, Etsuro Orito, Tomoyoshi Ohno, Kanji Sugihara, Izumi Hasegawa, Kei Fujiwara, Kiyoaki Ito, Atsushi Ozasa, Masashi Mizokami, Takaji Wakita, Takaji Wakita, Journal of Clinical Microbiology, 43, (11) 5679 - 5684,   2005年11月, Although combination therapy with interferon and ribavirin has improved the treatment for chronic hepatitis C virus (HCV) infection, the detailed anti-HCV effect of ribavirin in clinical concentrations remains uncertain. To detect the anti-HCV effect of ribavirin in lower concentrations, a sensitive and accurate assay system was developed using the reporter replicon system with an HCV genotype 2a subgenomic replicon (clone JFH-1) that exhibits robust replication in various cell lines. This reporter replicon was generated by introducing the luciferase reporter gene (instead of the neomycin resistance gene) into the subgenomic JFH-1 replicon. To assess the replication of this reporter replicon, luciferase activity was measured serially up to day 3 after transient transfection of Huh7 cells. The luciferase activity increased exponentially over the time course of the experiment. After adjustment for transfection efficiency and transfected cell viability, the impacts of interferon and ribavirin were determined. The administration of interferon and ribavirin resulted in dose-dependent suppression of replicon RNA replications. The 50% inhibitory concentration of interferon and ribavirin was 1.80 IU/ml and 3.70 μg/ml, respectively. In clinical concentrations, replications were reduced to 0.09% and 53.74% by interferon (100 IU/ml) and ribavirin (3 μg/ml), respectively. Combination use of ribavirin and interferon enhanced the anti-HCV effect of interferon by 1.46- to 1.62-fold. In conclusion, we developed an accurate and sensitive replicon system, and the antivirus effect of interferon and ribavirin was easily detected within their clinical concentrations by this replicon system. This system will provide a powerful tool for screening new antiviral compounds against HCV. Copyright © 2005, American Society for Microbiology. All Rights Reserved.
  • Classifying genotype F of hepatitis B virus into F1 and F2 subtypes, Hideaki Kato, Kei Fujiwara, Robert G. Gish, Hiroshi Sakugawa, Hiroshi Yoshizawa, Fuminaka Sugauchi, Etsuro Orito, Ryuzo Ueda, Yasuhito Tanaka, Takanobu Kato, Yuzo Miyakawa, Masashi Mizokami, World Journal of Gastroenterology, 11, 6295 - 6304,   2005年10月28日, Aim: To explore the propriety of providing hepatitis B Virus (HBV) genotypes F and H with two distinct genotypes. Methods: Eleven HBV isolates of genotype F (HBV/F) were recovered from pati ents living in San Francisco, Japan, Panama, and Venezuela, and their full-length sequences were determined. Phylogenetic analysis was carried out among them along with HBV isolates previously reported. Results: Seven of them clustered with reported HBV/F isolates in the phylogenetic tree constructed on the entire genomic sequence. The remaining four flocked on another branch along with three HBV isolates formerly reported as genotype H. These seven HBV isolates, including the four in this study and the three reported, had a sequence divergence of 7.3-9.5% from the other HBV/F isolates, and differed by > 13.7% from HBV isolates of the other six genotypes (A-E and G). Based on a marked genomic divergence, falling just short of > 8% separating the seven genotypes, these seven HBV/F isolates were classified into F2 subtype and the former seven into F1 subtype provisionally. In a pairwise comparison of the S-gene sequences among the 7 HBV/F2 isolates and against 47 HBV/F1 isolates as well as 136 representing the other six genotypes (A-E and G), two clusters separated byrdistinct genetic distances emerged. Conclusion: Based on these analyses, classifying HBV/F isolates into two subtypes (F1 and F2) would be more appropriate than providing them with two distinct genotypes (F and H). © 2005 The WJG Press and Elsevier Inc. All rights reserved.
  • A new subtype (subgenotype) Ac (A3) of hepatitis B virus and recombination between genotypes A and E in Cameroon, Fuat Kurbanov, Yasuhito Tanaka, Kei Fujiwara, Fuminaka Sugauchi, Dora Mbanya, Leopold Zekeng, Nicaise Ndembi, Charlotte Ngansop, Lazare Kaptue, Tomoyuki Miura, Eiji Ido, Masanori Hayami, Hiroshi Ichimura, Masashi Mizokami, Journal of General Virology, 86, (7) 2047 - 2056,   2005年07月01日, Blood samples (n=544) from two different populations (Pygmies and Bantus) in Cameroon, West Africa, were analysed. Serological tests indicated that the anti-hepatitis C virus (HCV) prevalence in Bantus (20.3 %) was higher than that in Pygmies (2.3 %, P < 0.0001), whereas the distribution of hepatitis B virus (HBV) serological markers was equally high in both populations: in total, 9.4, 17.3 and 86.8 % for HBsAg, anti-HBs and anti-HBc, respectively. HBV genotype A (HBV/A) and HBV/E were predominant (43.5 % each) in both populations, and HBV/D was found in a minority (13 %). The preS/S region was sequenced in nine cases (five HBV/A and four HBV/E) and the complete genome in six cases (four HBV/A and two HBV/E). Subsequent phylogenetic analysis revealed that the HBV/A strains were distinct from the subtypes (subgenotypes) described previously, Ae (A2) and Aa (A1), and in the preS/S region they clustered with previously reported sequences from Cameroon. Based on the nucleotide difference from Aa (A1) and Ae (A2), more than 4 % in the complete genome, the Cameroonian strains were suggested to represent a new subtype (subgenotype), designated HBV/Ac (A3). A high (3.9 %) nucleotide divergence in HBV/Ac (A3) strains suggested that the subtype (subgenotype) has a long natural history in the population of Cameroon. One of the HBV/Ac (A3) strains was found to be a recombinant with an HBV/E-specific sequence in the polymerase reverse transcriptase domain. Further cohort studies will be required to assess detailed epidemiological, virological and clinical characteristics of HBV/Ac (A3), as well as its recombinant form. © 2005 SGM.
  • New combination test for hepatitis C virus genotype and viral load determination using Amplicor GT HCV MONITOR test v2.0, Motokazu Mukaide, Yasuhito Tanaka, Hirokazu Kakuda, Kei Fujiwara, Fuat Kurbanov, Eturo Orito, Kentaro Yoshioka, Kiyotaka Fujise, Shoji Harada, Takazumi Kozaki, Kazuo Takemura, Kazumasa Hikiji, Masashi Mizokami, World Journal of Gastroenterology, 11, 469 - 475,   2005年01月28日, Aim: To develop a new sensitive and inexpensive hepatitis C virus (HCV) combination test (HCV Guideline test) that enables the determination of HCV genotypes 1, 2 and 3, and simultaneous determination of HCV viral load using commercial Amplicor GT HCV MONITOR test v2.0 (microwell version). Methods: The HCV Guideline test used the PCR product generated in commercial Amplicor GT HCV Monitor test v2.0 for viral load measurement using microwell plate version of Amplicor HCV Monitor and also captured on separate plates containing capture probes and competitive oligonucleotide probes specific for HCV genotypes 1, 2 and 3, The HCV genotype was subsequently determined using the biotin-labeled PCR product and five biotin-labeled HCV-specific probes. Results: The sensitivity of the HCV Guideline test was 0.5 KIU/mL. Specificity of the HCV Guideline test was confirmed by direct sequencing of HCV core region and molecular evolutionary analyses based on a panel of 31 samples. The comparison of the HCV Guideline test and an in-house HCV core genotyping assay using 252 samples from chronic hepatitis C patients indicated concordant results for 97.2% of samples (59.5% genotype 1, 33.7% genotype 2, 6.0% genotype 3, and 0.8% mixed genotypes). Similarly, the HCV Guideline test showed concordance with a serological test, and the serological test failed to assign any serotype in 12.7% of the samples, indicating a better sensitivity of the HCV Guideline test. Conclusion: Clinically, both viral load and genotypes (1, 2 and 3) have been found to be major predictors of antiviral therapy outcome regarding chronic hepatitis C based on guidelines and they are, in normal circumstances, performed as separate stand-alone assays. The HCV Guideline test is a useful method for screening large cohorts in a routine clinical setting for determining the treatment regimen and for predicting the outcome of antiviral therapy of chronic hepatitis C. © 2005 The WJG Press and Elsevier Inc. All rights reserved.
  • Genotype and phylogenetic characterization of hepatitis B virus among multi-ethnic cohort in Hawaii, Mayumi Sakurai, Fuminaka Sugauchi, Naoky Tsai, Seiji Suzuki, Izumi Hasegawa, Kei Fujiwara, Etsuro Orito, Ryuzo Ueda, Masashi Mizokami, World Journal of Gastroenterology, 10, 2218 - 2222,   2004年08月01日, Aim: Hepatitis B virus (HBV) genomes in carriers from Hawaii have not been evaluated previously. The aim of the present study was to evaluate the distribution of HBV genotypes and their clinical relevance in Hawaii. Methods: Genotyping of HBV among 61 multi-ethnic carriers in Hawaii was performed by genetic methods. Three complete genomes and 61 core promoter/precore regions of HBV were sequenced directly. Results: HBV genotype distribution among the 61 carriers was 23.0% for genotype A, 14.7% for genotype B and 62.3% for genotype C. Genotypes A, B and C were obtained from the carriers whose ethnicities were Filipino and Caucasian, Southeast Asian, and various Asian and Micronesian, respectively. All cases of genotype B were composed of recombinant strains with genotype C in the precore plus core region named genotype Ba. HBeAg was detected more frequently in genotype C than in genotype B (68.4% vs 33.3%, P < 0.05) and basal core promoter (BCP) mutation (T1762/A1764) was more frequently found in genotype C than in genotype B. Twelve of the 38 genotype C strains possessed C at nucleotide (nt) position 1858 (C-1858). However there was no significant difference in clinical characteristics between C-1858 and T-1858 variants. Based on complete genome sequences, phylogenetic analysis revealed one patient of Micronesian ethnicity as having C-1858 clustered with two isolates from Polynesia with T-1858. In addition, two strains from Asian ethnicities were clustered with known isolates in carriers from Southeast Asia. Conclusion: Genotypes A, B and C are predominant types among multi-ethnic HBV carriers in Hawaii, and distribution of HBV genotypes is dependent on the ethnic background of the carriers in Hawaii. Copyright © 2004 by The WJG Press.
  • Lack of association between occult hepatitis B virus DNA viral load and aminotransferase levels in patients with hepatitis C virus-related chronic liver disease, Kei Fujiwara, Yasuhito Tanaka, Etsuro Orito, Tomoyoshi Ohno, Takanobu Kato, Fuminaka Sugauchi, Seiji Suzuki, Yuko Hattori, Mayumi Sakurai, Izumi Hasegawa, Takashi Ozasa, Futoshi Kanie, Hideyuki Kano, Ryuzo Ueda, Masashi Mizokami, Masashi Mizokami, Journal of Gastroenterology and Hepatology (Australia), 19, (12) 1343 - 1347,   2004年01月, Background and Aim: Occult hepatitis B virus (HBV) infection in hepatitis C virus (HCV)-infected patients might enhance the severity of chronic liver disease (CLD). To elucidate the correlation between occult HBV infection and the clinical course of HCV-related CLD, we evaluated whether the fluctuation of occult HBV-DNA directly affects the serum alanine aminotransferase (ALT) level. Methods: Forty-one patients with HCV-related CLD who received regular outpatient treatment and 42 age-, sex-, and antibody to hepatitis B core antigen positivity-matched healthy volunteers were enrolled. Serum HBV-DNA was quantitatively detected using real-time detection polymerase chain reaction (RTD-PCR). Serial serum samples in three patients were measured for HBV-DNA, ALT and HCV core antigen. Results: Hepatitis B virus DNA was amplified in eight of the HCV-related CLD patients (19.5%), which was significantly higher than that of healthy volunteers (2.4%). No significant difference between the genotype 1 HCV-related CLD group and the genotype 2 group was found. Based on the analyses using serial serum samples, the elevation of HBV-DNA did not occur before the ALT flares, but occurred at the same time or after the ALT flares. Conclusions: The prevalence of occult HBV infection of HCV-related CLD is significantly higher than that of control. Occult HBV infection has no influence on ALT flares among patients with HCV-related CLD. © 2004 Blackwell Publishing Asia Pty Ltd.
  • High stability of enzyme immunoassay for hepatitis C virus core antigen - Evaluation before and after incubation at room temperature, Yasuhito Tanaka, Kazumi Takagi, Toshiya Fujihara, Katsuhiko Kitsugi, Kei Fujiwara, Kumiko Hiramatsu, Yoshinori Ito, Yoshimitsu Takasaka, Masaji Sakai, Masashi Mizokami, Masashi Mizokami, Hepatology Research, 26, (4) 261 - 267,   2003年08月, Hepatitis C virus (HCV) RNA is thought to be less stable than HCV core antigen (HCV-Ag), however there have been few studies on comparing the stability of HCV-Ag with that of HCV-RNA in vitro. The aim of this study is to evaluate serial levels of HCV-Ag and HCV-RNA in serum before and after incubation at 4 or 25 °C for 7 days to estimate an assay suitable for general laboratory use. In this study, we demonstrate that HCV-Ag levels are highly reproducible (coefficients of variation (CVs); 0.89-6.92%) and stable (84.8% of the initial level) with incubation of even 25 °C for 7 days, whereas HCV-RNA levels are much less reproducible (CVs; 9.13-29.66%) and decrease dramatically (15.1% of the initial level) after incubation, particularly at 25 °C. The measurement of the HCV-Ag level was found to be suitable for HCV quantification with serum samples stored either at 4 °C or under unknown conditions. Additionally, it successfully eliminated inhibitors such as heparin from plasma and could be applied to a variety of clinical specimens. Our data suggest the significance of measuring the HCV-Ag level during clinical management independently of the HCV-RNA level, particularly because of its high stability. © 2003 Elsevier Science B.V. All rights reserved.
  • Transforming growth factor-beta-1 genetic polymorphism in Japanese patients with chronic hepatitis C virus infection, Seiji Suzuki, Yasuhito Tanaka, Etsuro Orito, Fuminaka Sugauchi, Fuminaka Sugauchi, Izumi Hasegawa, Izumi Hasegawa, Mayumi Sakurai, Mayumi Sakurai, Kei Fujiwara, Kei Fujiwara, Tomoyoshi Ohno, Ryuzo Ueda, Masashi Mizokami, Masashi Mizokami, Journal of Gastroenterology and Hepatology (Australia), 18, (10) 1139 - 1143,   2003年01月, Background and Aim: Transforming growth factor beta-1 (TGF-β1) is one of the most dominant fibrogenic cytokines in hepatic fibrosis. The aim of the present study was to examine the effects of TGF-β1 polymorphisms in Japanese patients with chronic hepatitis C virus (HCV) infection and in healthy control subject s. Methods: The TGF-β1 genotypes at codon 10 and codon 25 were determined in 206 Japanese patients with chronic HCV infection and in 101 Japanese healthy control subjects. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was used for the detection of these polymorphisms. The degree of hepatic fibrosis was assessed by liver biopsy and graded according to the New Inuyama Classification for chronic hepatitis graded F0-4. Results: The authors found no significant differences in genotype distributions and allele frequency between the HCV patients and the healthy control subjects. The frequencies of the TT, TC, and CC genotypes of codon 10 were 24%, 42% and 35%, respectively, among the patients of the F0-2 group, and 31%, 40% and 29%, respectively, among those of the F3-4 group. No significant differences were shown between the TGF-β1 polymorphism at codon 10 and the stage of hepatic fibrosis. In contrast, no genetic alteration of codon 25 was found in healthy controls and patients with chronic HCV infection. Conclusion: These results suggest that there may not be a significant relationship between polymorphism at codon 10 and the development of progressive hepatic fibrosis in the Japanese population. © 2003 Blackwell Publishing Asia Pty Ltd.
  • A case report of malignant peritoneal mesothelioma diagnosed by electron microscopic ultrastructure, Kei Fujiwara, Hideaki Kato, Atsunori Kusakabe, Naka Nakamura, Hiroyuki Takai, Takashi Tohyama, Masamichi Yokoe, Akira Nemoto, Kanji Sugihara, Katsuo Hayashi, Masataka Ogino, Toyonori Tsuzuki, Japanese Journal of Gastroenterology, 99, 1114 - 1118,   2002年09月01日
  • A case report of ulcerative colitis accompanied by goblet cell carcinoid and adenocarcinoma, Hideaki Kato, Katsuo Hayashi, Kanji Sugihara, Akira Nemoto, Kei Fujiwara, Takashi Tohyama, Masamichi Yokoe, Hiroyuki Takai, Naka Nakamura, Atsunori Kusakabe, Masataka Ogino, Hiroshi Hasegawa, Toyonori Tsuzuki, Japanese Journal of Gastroenterology, 99, 500 - 504,   2002年06月01日
  • Effect of interferon therapy on Japanese chronic hepatitis C virus patients with anti-liver/kidney microsome autoantibody type 1, Yoshihiko Iijima, Takanobu Kato, Hiroshi Miyakawa, Masataka Ogino, Makoto Mizuno, Kanji Sugihara, Takahiro Ando, Kei Fujiwara, Etsuro Orito, Ryuzo Ueda, Masashi Mizokami, Masashi Mizokami, Journal of Gastroenterology and Hepatology (Australia), 16, (7) 782 - 788,   2001年01月01日, Aim: The aim of this study was to determine the prevalence of anti-liver/kidney microsome autoantibody type 1 (anti-LKM-1) among hepatitis C virus (HCV)-infected Japanese patients at various stages (chronic hepatitis, liver cirrhosis and hepatocellular carcinoma), and to assess the influence of anti-LKM-1 on interferon therapy. Methods: A total of 390 serum samples from 215 HCV-infected patients with chronic hepatitis (HCV-CH), 81 HCV-infected patients with liver cirrhosis (HCV-LC), and 94 HCV-HCC infected patients were subjected to examination. Ninety-one HBsAg-positive patients and 137 healthy subjects served as controls. Anti-liver/kidney microsome autoantibody type 1 was determined by using a newly developed ELISA using recombinant cytochrome P450 IID6 as the antigen. Resu lts: Anti-liver/kidney microsome autoantibody type 1 was detected in six of the 390 (1.5%) chronic HCV-infected patients (four were HCV-CH and two were HCV-LC); in contrast, it was not detected in control groups. Among the 110 HCV-CH patients treated with interferon (IFN), four were positive for anti-LKM-1. No change in anti-LKM-1 immunoreactivity from negative to positive during interferon therapy was observed. Moreover, no increase in the serum alanine aminotransferase level was observed in these four patients with anti-LKM-1. Conclusion: Our study indicates that: (i) anti-LKM-1 does not aggravate the liver disease associated with HCV infection; and (ii) no change in anti-LKM-1 immunoreactivity from negative to positive or no aggravations of liver dysfunction were observed among HCV-CH patients during the IFN therapy for Japanese patients with liver disease. © 2001 Blackwell Science Asia Pty Ltd.

受賞

  •   2013年10月, 日本肝臓学会, 冠Award、第6回CHUGAI Award

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