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Kazuki Komase,
Shinya Maekawa,
Mika Miura,
Ryota Sueki,
Makoto Kadokura,
Hiroko Shindo,
Kuniaki Shindo,
Fumitake Amemiya,
Yasuhiro Nakayama, Taisuke Inoue,
Minoru Sakamoto,
Atsuya Yamashita,
Kohji Moriishi,
Nobuyuki Enomoto
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ABSTRACT: AIM: Prediction of treatment responses to pegylated interferon (PEG IFN) plus ribavirin (RBV) therapy is uncertain for genotype 1b chronic hepatitis C. METHODS: In this study, 96 patients were investigated for the correlation between 36 pretreatment serum chemokine/cytokine levels and PEG IFN/RBV treatment efficacy by a sandwich enzyme-linked immunoassay (ELISA) and a bead array. RESULTS: First, chemokines/cytokines were measured semiquantitatively by sandwich ELISA in 31 randomly-selected patients and the serum regulated on activation normal T-cell expressed and secreted (RANTES) level was found to be significantly higher in the sustained virological response (SVR) group than the non-SVR group (P = 0.048). Precise RANTES measurement in all 96 patients using a bead array confirmed this correlation (P = 0.002). However, the genetic RANTES haplotype was not significantly related to the serum level. The serum RANTES level was extracted by multivariate analysis (odds ratio = 4.09, 95% confidence interval = 1.02-16.5, P = 0.048) as an independent variable contributing to SVR. CONCLUSION: The serum RANTES level is an important determinant influencing the virological response to PEG IFN/RBV therapy in chronic hepatitis C.
Hepatology Research 11/2012; · 2.20 Impact Factor
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Ryota Sueki,
Shinya Maekawa,
Mika Miura,
Makoto Kadokura,
Kazuki Komase,
Hiroko Shindo,
Asuka Kanayama,
Takako Ohmori,
Kuniaki Shindo,
Fumitake Amemiya,
Yasuhiro Nakayama,
Tomoyoshi Uetake, Taisuke Inoue,
Minoru Sakamoto,
Nobuyuki Enomoto
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ABSTRACT: The emergence of amino acid or nucleotide substitutions leads to lamivudine resistance in hepatitis B virus (HBV) infected patients. The aim of this study was to investigate whether viral sequences help predict the emergence of lamivudine resistance. The study subjects comprised 59 consecutive patients infected with HBV treated with daily therapy of 100 mg lamivudine. Among those, 32 patients with adequate pretreatment serum preservation were investigated for the correlation between viral amino acid substitutions and the appearance of lamivudine resistance with consideration of clinical background by determining dominant HBV full open reading frames. Viral resistance to lamivudine emerged in 28 of 59 patients (47%) in a median period of 2.45 years. Sequence comparisons of HBV genomes between patients who later developed lamivudine resistance and patients who did not revealed the existence of significant differences between the two groups in the pre-S1 84 (P = 0.042), pre-S2 1 (P = 0.017) and 22 (P = 0.015), and polymerase tp 95 (P = 0.046), judged by a log-rank test. Viral sequence analyses revealed the presence of amino acid substitutions in HBV pre-S1 and pre-S2 that may be associated with the emergence of lamivudine resistance during chronic HBV infection.
Journal of Medical Virology 09/2012; 84(9):1360-8. · 2.82 Impact Factor
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Hiroko Shindo,
Shinya Maekawa,
Kazuki Komase,
Ryota Sueki,
Mika Miura,
Makoto Kadokura,
Kuniaki Shindo,
Fumitake Amemiya,
Takatoshi Kitamura,
Yasuhiro Nakayama, Taisuke Inoue,
Minoru Sakamoto,
Shun-Ichi Okada,
Yasuhiro Asahina,
Namiki Izumi,
Masao Honda,
Shuichi Kaneko,
Nobuyuki Enomoto
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ABSTRACT: BACKGROUND AND AIMS: Protease inhibitor (PI)-resistant hepatitis C virus (HCV) variants may be present in substantial numbers in PI-untreated patients according to recent reports. However, influence of these viruses in the clinical course of chronic hepatitis C has not been well characterized. METHODS: The dominant HCV nonstructural 3 (NS3) amino acid sequences were determined in 261 HCV genotype 1b-infected Japanese patients before pegylated interferon plus ribavirin (PEG-IFN/RBV) therapy, and investigated the patients' clinical characteristics as well as treatment responses including sustained virological response (SVR) rate. HCV-NS3 sequences were also determined in 39 non-SVR patients after completion of the therapy. RESULTS: Four single mutations (T54S, Q80K, I153V, and D168E) known to confer PI resistance were found in 35 of 261 patients (13.4%), and double mutations (I153V plus T54S/D168E) were found in 6 patients (2.3%). Responses to PEG-IFN/RBV therapy did not differ between patients with and without PI-resistance mutations (mutation group, SVR 48%; wild-type group, SVR 40%; P = 0.38). On the other hand, two mutations appeared in two non-SVR patients after PEG-IFN/RBV therapy (I153V and E168D, 5.1%). CONCLUSIONS: PI-resistance-associated NS3 mutations exist in a substantial proportion of untreated HCV-1b-infected patients. The impact of these mutations in the treatment of PIs is unclear, but clinicians should pay attention to avoid further development of PI resistance.
Hepatology International 08/2011; · 2.64 Impact Factor
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Mika Miura,
Shinya Maekawa,
Makoto Kadokura,
Ryota Sueki,
Kazuki Komase,
Hiroko Shindo,
Takako Ohmori,
Asuka Kanayama,
Kuniaki Shindo,
Fumitake Amemiya,
Yasuhiro Nakayama,
Takatoshi Kitamura,
Tomoyoshi Uetake, Taisuke Inoue,
Minoru Sakamoto,
Shunichi Okada,
Nobuyuki Enomoto
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ABSTRACT: BACKGROUND AND AIMS: The association between hepatitis C virus (HCV) sequences with interleukin 28B (IL28B) single-nucleotide polymorphism (SNP) in the development of hepatocellular carcinoma (HCC) has not been well clarified. METHODS: Complete HCV open-reading frame sequences were determined in 20 patients developing HCC and 23 non-HCC patients with HCV-1b infection in two distant time points. An additional 230 patients were studied cross-sectionally for core and NS5A sequences with HCC development. Among them, 98 patients with available samples were investigated for changes in viral core sequences over time. Finally, IL28B SNPs and HCC development were investigated in 228 patients. RESULTS: During observation period (HCC for 10.8 years, and non-HCC for 11.1 years), changes in core a.a. 70 and three amino acid positions in NS5A were characteristics of the patients developing HCC. In 230 patients, Q (glutamine) or H (histidine) to R (arginine) ratio at core a.a. 70 was significantly higher in the HCC group (HCC group 43:22 vs. non-HCC group 66:99, p = 0.001). A change in core R70Q was observed over time in 11 patients associated with a decrease in platelets (p = 0.005) and albumin (p = 0.005), while a Q70R change was observed in 4 patients without associated changes in platelets (nonsignificant) and albumin (nonsignificant). IL28B SNP showed significant correlation with the core a.a. 70 residue. There was no evident link between IL28B SNPs and the occurrence of HCC. CONCLUSIONS: Hepatitis C virus core a.a. 70 residue is associated with liver disease progression and is independent factor for HCC development in genotype-1b infection. IL28B SNPs are related to core a.a. 70 residue, but not to HCC. The functional relevance of core a.a. 70 residue in hepatitis C pathogenesis should be further investigated.
Hepatology International 08/2011; · 2.64 Impact Factor
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Makoto Kadokura,
Shinya Maekawa,
Ryota Sueki,
Mika Miura,
Kazuki Komase,
Hiroko Shindo,
Fumitake Amemiya,
Tomoyoshi Uetake, Taisuke Inoue,
Minoru Sakamoto,
Mina Nakagawa,
Naoya Sakamoto,
Mamoru Watanabe,
Nobuyuki Enomoto
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ABSTRACT: A proportion of patients infected with genotype 2a hepatitis C virus (HCV) cannot achieve a sustained virological response (SVR) to pegylated-interferon plus ribavirin therapy (PEG-IFN/RBV) but the reason remains unclear. The present study aimed to clarify the possible correlation between viral sequence variations and final outcome.
The pretreatment complete open reading frame (ORF) sequences of genotype 2a HCV were determined by direct sequencing for two independent groups of patients (43 patients as test; group 1 and 35 as validation; group 2), and the correlation with the final outcome was explored.
Patients with SVR (n = 58) and with non-SVR (n = 20) differed significantly in pretreatment HCV RNA level (p = 0.002), fibrosis score (p = 0.047), and cumulative RBV dosage (p = 0.003). By comparison of all amino acid positions in the complete HCV ORFs, threonine at amino acid (aa) 110 in the core region was remarkably frequent in SVR (p = 0.01 for group 1, p = 0.004 for group 2, and p = 5E-05 for combined). A sliding window analysis revealed that the total number of amino acid variations within the NS5A aa 2258-2306 region were significantly high in SVR compared to non-SVR patients (p = 0.01 for group 1, p = 0.006 for group 2, and p = 0.0006 for combined). Multivariate analyses revealed that core aa 110 (p = 0.02), NS5A aa 2258-2306 (p = 0.03), and cumulative RBV dosage (p = 0.02) were identified as independent variables associated with the final outcome.
The outcome of PEG-IFN/RBV therapy is significantly influenced by variation in the core and NS5A regions in genotype 2a HCV infection.
Hepatology International 03/2011; 5(3):789-99. · 2.64 Impact Factor
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Makoto Kadokura,
Shinya Maekawa,
Ryota Sueki,
Mika Miura,
Kazuki Komase,
Hiroko Shindo,
Fumitake Amemiya,
Tomoyoshi Uetake, Taisuke Inoue,
Minoru Sakamoto,
Mina Nakagawa,
Naoya Sakamoto,
Mamoru Watanabe,
Nobuyuki Enomoto
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ABSTRACT: Patients infected with genotype 2b hepatitis C virus (HCV) generally can achieve favorable responses to pegylated-interferon plus ribavirin therapy (PEG-IFN/RBV). However, a proportion of patients show poorer responses and the correlation between viral sequence variation and treatment outcome remains unclear.
The pretreatment complete open reading frame (ORF) sequences of genotype 2b HCV determined by direct sequencing were investigated for correlation with the final outcome in a total of 60 patients.
In this study group, 87.5% (14/16) of non-sustained virological response (non-SVR) patients (n = 16) were relapsers. Compared to sustained virological response (SVR) patients (n = 44), non-SVR patients were older and could not achieve prompt viral clearance after the therapy induction. Comparing each viral protein between the two groups, viral sequences were more diverse in SVR patients and that diversity was found primarily in the E1, p7, and NS5A proteins. In searching for specific viral regions associated with the final outcome, several regions in E2, p7, NS2, NS5A, and NS5B were extracted. Among these regions, part of the interferon sensitivity determining region (ISDR) was included. In these regions, amino acid substitutions were associated with the final outcome in an incremental manner, depending upon the number of substitutions.
Viral sequences are more diverse in SVR patients than non-SVR patients receiving PEG-IFN/RBV therapy for genotype-2b HCV infection. Through systematic comparison of viral sequences, several specific regions, including part of the ISDR, were extracted as having significant correlation with the final outcome.
PLoS ONE 01/2011; 6(9):e24514. · 4.09 Impact Factor
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Akira Matsui,
Tatsuya Yamaguchi,
Shinya Maekawa,
Chikako Miyazaki,
Shinichi Takano,
Tomoyoshi Uetake, Taisuke Inoue,
Masahiko Otaka,
Hiroyuki Otsuka,
Tadashi Sato,
Atsuya Yamashita,
Yuka Takahashi,
Nobuyuki Enomoto
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ABSTRACT: To comprehensively screen for genetic events underlying colorectal cancer, we performed suppression subtraction hybridization analysis on an advanced colon cancer. Because Dickkopf-4, a member of the Dickkopf family acting as a Wnt-signaling modulator, was identified as one of the upregulated genes in this specimen, we investigated expression profiles of all the Dickkopf family members in 55 colorectal tumors (21 cancers and 34 adenomas). We also investigated mechanisms regulating the expression of Dickkopf-4 in these cancers in vitro and in vivo. Compared with normal adjacent mucosae, Dickkopf-4 (median 27.4, P < 0.01) and -2 (median 51.4, P < 0.01) were strongly expressed in colorectal cancers. The level of Dickkopf-4 was positively correlated with fibroblast growth factor-20 (rs = 0.61, P = 0.00017), a representative β-catenin transcriptional target gene, and with the degree of nuclear accumulation of β-catenin in colorectal tumors. Dickkopf-4 was induced by activated β-catenin in vitro. Reciprocally, recombinant Dickkopf-4 significantly inhibited T-cell factor/lymphocyte enhancer factor reporter activity stimulated by recombinant Wnt3a in human embryonic kidney 293 cells. We conclude that Dickkopf-4 and -2 are significantly upregulated in most colorectal tumors, and that Dickkopf-4 upregulation reflects activation of the Wnt/canonical pathway. (Cancer Sci 2009; 100: 1923–1930)
Cancer Science 07/2009; 100(10):1923 - 1930. · 3.33 Impact Factor
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Akira Matsui,
Tatsuya Yamaguchi,
Shinya Maekawa,
Chikako Miyazaki,
Shinichi Takano,
Tomoyoshi Uetake, Taisuke Inoue,
Masahiko Otaka,
Hiroyuki Otsuka,
Tadashi Sato,
Atsuya Yamashita,
Yuka Takahashi,
Nobuyuki Enomoto
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[hide abstract]
ABSTRACT: To comprehensively screen for genetic events underlying colorectal cancer, we performed suppression subtraction hybridization analysis on an advanced colon cancer. Because Dickkopf-4, a member of the Dickkopf family acting as a Wnt-signaling modulator, was identified as one of the upregulated genes in this specimen, we investigated expression profiles of all the Dickkopf family members in 55 colorectal tumors (21 cancers and 34 adenomas). We also investigated mechanisms regulating the expression of Dickkopf-4 in these cancers in vitro and in vivo. Compared with normal adjacent mucosae, Dickkopf-4 (median 27.4, P < 0.01) and -2 (median 51.4, P < 0.01) were strongly expressed in colorectal cancers. The level of Dickkopf-4 was positively correlated with fibroblast growth factor-20 (r(s) = 0.61, P = 0.00017), a representative beta-catenin transcriptional target gene, and with the degree of nuclear accumulation of beta-catenin in colorectal tumors. Dickkopf-4 was induced by activated beta-catenin in vitro. Reciprocally, recombinant Dickkopf-4 significantly inhibited T-cell factor/lymphocyte enhancer factor reporter activity stimulated by recombinant Wnt3a in human embryonic kidney 293 cells. We conclude that Dickkopf-4 and -2 are significantly upregulated in most colorectal tumors, and that Dickkopf-4 upregulation reflects activation of the Wnt/canonical pathway.
Cancer Science 07/2009; 100(10):1923-30. · 3.33 Impact Factor
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Fumitake Amemiya,
Shinya Maekawa,
Yoshie Itakura,
Asuka Kanayama,
Akira Matsui,
Shinichi Takano,
Tatsuya Yamaguchi,
Jun Itakura,
Takatoshi Kitamura, Taisuke Inoue,
Minoru Sakamoto,
Kozue Yamauchi,
Shunichi Okada,
Atsuya Yamashita,
Naoya Sakamoto,
Masahiko Itoh,
Nobuyuki Enomoto
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ABSTRACT: Recently, microdomains of organelle membranes rich in sphingomyelin and cholesterol (called "lipid rafts") have been considered to act as a scaffold for the hepatitis C virus (HCV) replication complex. Using the HCV cell culture system, we investigated the effect of myriocin, a sphingomyelin synthesis inhibitor, on HCV replication. We also investigated the combined effect of myriocin with interferon (IFN) and myriocin with simvastatin. Myriocin suppressed replication of both a genotype 1b subgenomic HCV replicon (Huh7/Rep-Feo) and genotype 2a infectious HCV (JFH-1 HCV) in a dose-dependent manner (for subgenomic HCV-1b, maximum of 79% at 1000 nmol/L; for genomic HCV-2a, maximum of 40% at 1000 nmol/L). Combination treatment with myriocin and IFN or myriocin and simvastatin attenuated HCV RNA replication synergistically in Huh7/Rep-Feo cells. Our data demonstrate that the sphingomyelin synthesis inhibitor strongly suppresses replication of both the subgenomic HCV-1b replicon and the JFH-1 strain of genotype 2a infectious HCV, indicating that lipid metabolism could be a novel target for HCV therapy.
The Journal of Infectious Diseases 03/2008; 197(3):361-70. · 6.41 Impact Factor
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ABSTRACT: An unenveloped DNA virus named TT virus (TTV) has been reported in association with acute and chronic hepatitis of unknown etiology. The effect of interferon on TTV was evaluated in the patients with chronic hepatitis C who were coinfected with TTV. TTV DNA was determined by a polymerase chain reaction with heminested primers in the 96 patients with chronic hepatitis C who received interferon-α (516 million units in 26 weeks) and followed for 24 months thereafter. TTV DNA was detected in 31 (32%) patients before therapy. TTV DNA became undetectable during interferon therapy and remained absent in 14 (45% of the 31 patients) through 24 months thereafter. The four patients with pretreatment TTV DNA titer ≥103/ml did not respond. These results indicate that TTV is sensitive to interferon, and the response would be inversely correlated with pretreatment viral titers. J. Med. Virol. 58:196–200, 1999. © 1999 Wiley-Liss, Inc.
Journal of Medical Virology 05/1999; 58(3):196 - 200. · 2.82 Impact Factor
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ABSTRACT: A second-generation method of genotyping hepatitis C virus (HCV) was developed by the polymerase chain reaction (PCR) with sense as well as antisense primers deduced from the core gene. HCV RNA specimens extracted from sera were reverse-transcribed and amplified with universal primers in the first round of PCR to obtain fragments of 433 base pairs representing nucleotides 319–751. In the second round of PCR, portions of PCR products were amplified separately with sense and antisense primers specific for each of the five common genotypes prevailing across the world i.e., I/1a, II/1b, III/2a, IV/2b and V/3a. The specificity of the method was verified by a panel of 177 HCV isolates of various genotypes in the genetic groups 1–9. It allowed clear differentiation of genotype I/1a from Il/1b which was not always accomplished by the previous method. When 501 sera from blood donors and hepatitis patients with HCV viremia from various countries were genotyped by the second-generation method, 478 (95.4%) were classified into the five genotypes. HCV RNA samples from 23 (4.6%) sera were not classifiable into any of the five common genotypes and, by sequence analysis, 22 were found to be of four genotypes in group 4 and one of genotype 1c in Simmond's classification.
Journal of Virological Methods.
