Host sphingolipid biosynthesis is a promising therapeutic target for the inhibition of hepatitis B virus replication.

Serine palmitoyltransferase (SPT) catalyzes the first step in the sphingolipid biosynthetic pathway. Myriocin inhibits SPT and was shown to suppress the replication of hepatitis C virus (HCV) in vitro and in vivo. However, its effect on hepatitis B virus (HBV) replication is unknown. In this study, the HBV DNA levels in HuH7 cell culture supernatants were lowered successfully by using myriocin and it was found that the 50% inhibitory concentration of myriocin is approximately 5 µM. Myriocin and/or pegylated interferon (PEG-IFN) were also administered to chimeric mice for 2 weeks and the effects of these compounds on HBV DNA levels were determined. Myriocin alone did not reduce effectively the HBV DNA levels, whereas PEG-IFN alone reduced the DNA levels to 1/10th of the control levels. The combination of myriocin with PEG-IFN reduced the HBV levels to about 1/1,000 th of the control levels and induced a 1.0 log reduction in the levels of the HBV surface antigen and core protein. This latter effect was not observed in the other treatment groups. In conclusion, the combination of myriocin with PEG-IFN represses synergistically HBV replication in vivo without inducing hepatotoxicity.

A population-based cohort study for the risk factors of HCC among hepatitis B virus mono-infected subjects in Japan.

BACKGROUND: There have only been a few prospective studies investigating risk factors associated with the development of hepatocellular carcinoma (HCC) among chronic hepatitis B patients all over the world, and no study has been conducted in Japanese population. METHODS: A population-based cohort consisting of 19393 subjects (middle aged or older) with over 13 years' follow-up was investigated in Japan. RESULTS: Of 19393 subjects, 479 had hepatitis B virus (HBV) mono-infection (2.5%). During the 245923 person-years' follow-up (average follow-up period 12.7 years), 13 cases of newly diagnosed HCC were documented in the HBV mono-infected group. Several factors at baseline (male, smoking, alanine aminotransferase, the positivity of HBe antigen and HB core-related antigen, the proportion of HBV DNA ≥ 5 log copies/mL, T1753V mutation, and A1762T/G1764A double mutation) were significantly associated with HCC among HBV mono-infected subjects. Multivariate-adjusted Cox hazard model showed that A1762T/G1764A (hazard ratio 7.05 [95% confidence interval (CI) 1.03-48.12, P = 0.046]) was the only independent risk factor for the development of HCC. Kaplan-Meier method also showed that the probability of HCC occurrence-free was significantly lower in HBV mono-infected subjects with A1762T/G1764A double mutation than those without these mutations. CONCLUSION: HBV mono-infected subjects with A1762T/G1764A double mutation could be at high risk of HCC development during the natural course of HBV infection.

[Evaluation of high-sensitivity HBsAg quantitative assay for HBV genotype].

The clinical implication of the hepatitis B surface antigen (HBsAg) concentrations has been reported in HBV-infected patients during anti-viral treatment. HBV genotypes A and D are ubiquitous and scattered worldwide, especially northern America as well as Europe, whereas genotypes B and C are common in Asia. The aim of this study was to evaluate a new version of the Sysmex HBsAg quantitative kit based on Chemiluminescence Enzyme Immunoassay. Sera collected from 172 patients infected with any of the four major genotypes A to D (HBV/A, n = 18; B, n = 25; C, n = 84; D, n = 45), including the genotype D cases with weak reaction in the previous version of the kit. The new version of the kit having additional monoclonal antibody, showed improved sensitivity compared to the previous version as well as robust correlation with another quantitative HBsAg assay: the Abbot Architect. Observed during lamivudine therapy, increase in HBsAg and HBV DNA concentrations preceded the aminotransferase (ALT) elevation associated with drug-resistant HBV variant emergence (breakthrough hepatitis). In conclusion, reliability of the Sysmex HBsAg quantitative assay was confirmed for the four HBV genotypes common worldwide. Monitoring of serum HBsAg concentrations in addition to HBV DNA quantification, is helpful in evaluation of the response or resistance to anti-viral therapy.

A genetic variant of hepatitis B virus divergent from known human and ape genotypes isolated from a Japanese patient and provisionally assigned to new genotype J.

Hepatitis B virus (HBV) of a novel genotype (J) was recovered from an 88-year-old Japanese patient with hepatocellular carcinoma who had a history of residing in Borneo during the World War II. It was divergent from eight human (A to H) and four ape (chimpanzee, gorilla, gibbon, and orangutan) HBV genotypes, as well as from a recently proposed ninth human genotype I, by 9.9 to 16.5% of the entire genomic sequence and did not have evidence of recombination with any of the nine human genotypes and four nonhuman genotypes. Based on a comparison of the entire nucleotide sequence against 1,440 HBV isolates reported, HBV/J was nearest to the gibbon and orangutan genotypes (mean divergences of 10.9 and 10.7%, respectively). Based on a comparison of four open reading frames, HBV/J was closer to gibbon/orangutan genotypes than to human genotypes in the P and large S genes and closest to Australian aboriginal strains (HBV/C4) and orangutan-derived strains in the S gene, whereas it was closer to human than ape genotypes in the C gene. HBV/J shared a deletion of 33 nucleotides at the start of preS1 region with C4 and gibbon genotypes, had an S-gene sequence similar to that of C4, and expressed the ayw subtype. Efficient infection, replication, and antigen expression by HBV/J were experimentally established in two chimeric mice with the liver repopulated for human hepatocytes. The HBV DNA sequence recovered from infected mice was identical to that in the inoculum. Since HBV/J is positioned phylogenetically in between human and ape genotypes, it may help to trace the origin of HBV and merits further epidemiological surveys.

[Evaluation of hepatitis B virus genotyping EIA kit].

Clinical significance of Hepatitis B virus(HBV) genotyping is increasingly recognized. The aim of this study was to evaluate reproducibility, accuracy, and sensitivity of an enzyme immunoassay (EIA) based HBV genotyping kit, which designed to discriminate between genotypes to A, B, C, or D by detecting genotype-specific epitopes in PreS2 region. Using the four genotypes panels, the EIA demonstrated complete inter and intra-assay genotyping reproducibility. Serum specimens had stable results after 8 days at 4 degrees C, or 10 cycles of freezing-thawing. In 91 samples that have been genotyped by DNA sequencing, 87(95.6%) were in complete accordance with EIA genotyping. Of examined 344 HBsAg-positive serum specimens, genotypes A, B, C and D were determined in 26 (7.6%), 62 (18.0%), 228 (66.3%), and 9 (2.6%) cases, respectively. Of 19 (5.5%) specimens unclassified by the EIA, 13 were found to have low titer of HBsAg concentration (< 3 IU/ml), and the other 5 had amino acid mutations or deletions within targeted PreS2 epitopes. The EIA allowed genotyping even in HBV DNA negative samples (96.2%). In conclusion, HBV genotype EIA is reliable, sensitive and easy assay for HBV genotyping. The assay would be useful for clinical use.

Characteristics of hepatitis B virus genotype G coinfected with genotype H in chimeric mice carrying human hepatocytes.

Accumulated evidence indicated that hepatitis B virus genotype G (HBV/G) is present exclusively in coinfection with other HBV genotypes. In Mexico, HBV/G from 6 men who had sex with men were coinfected with HBV/H. Phylogenetically complete genomes of the 6 Mexican HBV/G strains were closely related to previous ones from the US/Europe. Using uPA/SCID mice with human hepatocytes, monoinfection with HBV/G did not result in detectable HBV DNA in serum, whereas superinfection with HBV/G at week 10 inoculated HBV/H when HBV/H DNA was elevated to >10(7) copies/mL has enhanced the replication of HBV/G. The HBV/G was enhanced in another 3 inoculated with a serum passage containing HBV/G with a trace of HBV/H. Coinfection of mice with HBV/G and H induced fibrosis in the liver. In conclusion, the replication of HBV/G can be enhanced remarkably when it is coinfected with HBV/H. Coinfection with HBV/G may be directly cytopathic in immunosuppressive conditions.