Variations in the viral NS5B region in Japanese patients with chronic hepatitis C virus genotype 1b infection. No specific amino acid substitution was identified as determinants of treatment response to interferon/ribavirin combination therapy.

OBJECTIVE: A recent study suggested that the substitution of amino acid 415 of HCV NS5B from phenylalanine to tyrosine in patients with HCV genotype 1a infection is induced by ribavirin and responsible for resistance to ribavirin therapy. The aim of this study was to evaluate whether specific variations in the HCV NS5B sequence in Japanese patients with HCV genotype 1b (HCV/1b) infection are associated with treatment response or selected by treatment with interferon-alpha/ribavirin combination therapy. METHODS: Eighteen Japanese patients with HCV/1b infection receiving interferon-alpha/ribavirin combination therapy for 24 weeks were studied. Five patients treated with interferon-alpha monotherapy for 24 weeks were also studied as controls. The entire HCV NS5B sequence before and after therapy was determined. RESULTS: All HCV isolates had tyrosine at position 415 of NS5B before and after therapy. Further analysis showed that no specific amino acid substitutions were identified to associate with clinical response and no specific amino acid substitutions were induced/selected by the clinical treatment. CONCLUSION: No specific HCV NS5B nucleotide/amino acid sequence variations, including amino acid 415 of NS5B, were identified as being associated with clinical treatment response or selected by the combination therapy in Japanese patients with HCV/1b infection.

Detection of anti-hepatitis C virus effects of interferon and ribavirin by a sensitive replicon system.

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 microg/ml, respectively. In clinical concentrations, replications were reduced to 0.09% and 53.74% by interferon (100 IU/ml) and ribavirin (3 microg/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.

Distribution of HBV genotypes among HBV carriers in Benin:phylogenetic analysis and virological characteristics of HBV genotype E.

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.

HBV/HCV Infection, Alcohol, Tobacco and Genetic Polymorphisms for Hepatocellular Carcinoma in Nagoya, Japan.

A population-based case-control study was carried out to investigate risk factors for hepatocellular carcinoma (HCC) in Nagoya, Japan, including hepatitis virus infections, drinking and smoking habits and genetic polymorphisms in aldehyde dehydrogenase2 (ALDH2) and cytochrome P4502E1 (CYP2E1). A total of 84 patients with HCC and 84 sex, age and residence pair-matched controls were recruited for this study. By univariate analysis, hepatitis B virus (HBV) (OR=5.14; 95%CI=2.29-11.6) and hepatitis C virus (HCV)(OR=32.00; 95%CI=7.83-130.7) infections, having a history of blood transfusion (OR=5.25; 95%CI=1.80-15.29), and habitual smoking (OR=2.36; 95%CI=1.17-4.78) were significantly linked to cases; by multivariate analysis, HCV infection (OR=23.5; 95%CI=5.07-108.9) and habitual smoking (OR=5.41; 95%CI=1.10-26.70) were still associated with a significantly increased risk. The c1/c1 genotype of CYP2E1 (odds ratio [OR]= 0.45; 95% confidence interval [CI]=0.21-0.99), detected by Pstl and Rsal digestion was significantly more prevalent in the control group, while 1-1 genotype of ALDH2 (OR=1.24; 95%CI=0.70-2.20) did not demonstrate variation. There were no statistically significant interactions between habitual smoking/drinking and genetic polymorphisms of ALDH2/P4502E1 with reference to HCC development. These findings suggest that viruses, especially HCV infection, and habitual smoking are major independent risk factors, while genetic polymorphisms of ALDH2 and CYP2E1 have only limited contribution to the risk of HCC in Nagoya, Japan.