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IntroductionTelomerase repairs the telomeric ends of chromosomes and is active in nearly all malignant cells. Hepatitis C virus (HCV) is known to be oncogenic and potential interactions with the telomerase system require further study. We determined the effects of HCV infection on human telomerase reverse transcriptase (TERT) expression and enzyme activity in primary human hepatocytes and continuous cell lines.ResultsPrimary human hepatocytes and Huh-7.5 hepatoma cells showed early de novo TERT protein expression 2–4 days after infection and these events coincided with increased TERT promoter activation, TERT mRNA, and telomerase activity. Immunoprecipitation studies demonstrated that NS3-4A protease-helicase, in contrast to core or NS5A, specifically bound to the C-terminal region of TERT through interactions between helicase domain 2 and protease sequences. Increased telomerase activity was noted when NS3-4A was transfected into cells, when added to reconstituted mixtures of TERT and telomerase RNA, and when incubated with high molecular weight telomerase ‘holoenzyme’ complexes. The NS3-4A catalytic effect on telomerase was inhibited with primuline or danoprevir, agents that are known to inhibit NS3 helicase and protease activities respectively. In HCV infected cells, NS3-4A could be specifically recovered with telomerase holoenzyme complexes in contrast to NS5A or core protein. HCV infection also activated the effector caspase 7 which is known to target TERT. Activation coincided with the appearance of lower molecular weight carboxy-terminal fragment(s) of TERT, chiefly sized at 45 kD, which could be inhibited with pancaspase or caspase 7 inhibitors.ConclusionsHCV infection induces TERT expression and stimulates telomerase activity in addition to triggering Caspase activity that leads to increased TERT degradation. These activities suggest multiple points whereby the virus can influence neoplasia. The NS3-4A protease-helicase can directly bind to TERT, increase telomerase activity, and thus potentially influence telomere repair and host cell neoplastic behavior.

Hepatitis C virus (HCV) infection is a common problem in patients treated with maintenance hemodialysis (HD) and is associated with an increased morbidity and mortality and lower quality of life. The major causes of HCV-associated mortality are liver and cardiovascular-related death. HCV-infected HD patients have a higher prevalence of inflammation-related metabolic and vascular diseases, leading to high rates of cardiovascular mortality in patients with end-stage renal disease. In the current era of highly effective direct-acting antiviral regimens, HCV treatment may also confer hepatic, cardiovascular and other morbidity and mortality benefits even to dialysis-dependent patients who do not qualify for kidney transplantation. Currently, the most accepted regimens in this patient population include elbasvir/grazoprevir and glecaprevir/pibrentasvir.

Norovirus (NoV) is recognized as one of the most common causative agent of diarrheal disease in young children worldwide. The current study was undertaken to determine the distribution of NoV genotypes in Japan. A total of 3,864 fecal specimens from children with acute gastroenteritis in five regions (Tokyo, Maizuru, Saga, Sapporo, and Osaka) of Japan from July 1995 to June 2001 were collected and then tested for the presence of NoV by RT-PCR. Three hundred sixty four were found to be positive for NoV, accounting for 11%. The highest prevalence of NoV infection was in November, December, and January as the early winter months in Japan. NoV was subjected to be further characterized to sequencing analysis. All NoVs belonged to two different genogroups I and II and these represented 3% and 97%, respectively. This finding indicated that NoV genogroup II was the dominant group causing acute gastroenteritis in Japan. Interestingly, NoV strains were classified into 16 distinct genotypes including genogroup II genotype 9 that was firstly identified in Japan. Of these, NoV genogroup II genotypes 3 and 4 dominated over other genotypes and became the leading strains in Japanese pediatric population. In conclusion, diarrhea due to NoV infection is still a health burden in Japan. This report also stresses the great genetic diversity as well as the importance of NoV causing the diarrhea in Japan.

The hepatitis B virus (HBV) genomes in Taiwanese aborigines, whose ancestors have lived in Taiwan for over 10,000 years, have not been characterized. In order to characterize of HBV in this special population, serum samples were obtained from serologically HBsAg-positive 27 Taiwanese aborigines. The pre-S1/S2 region and the full-length 3.2 kb of the HBV genome were amplified by PCR. Obtained amplicons were sequenced and confirmed the HBV genotypes by phylogenetic analysis. By phylogenetic analysis of the sequence of pre-S1/pre-S2 region, HBV/B2 (21/27: 78 %) was the most prevalent followed by genotype D (6/27: 22 %). Two strains of HBV/B2, each having 3,215 bp genomes, had recombination with genotype C in the pre-C/C gene which is characteristic of subgenotype B2 circulating in Southeast Asia. Interestingly, six strains of genotype D formed a distinct cluster between subgenotypes D1 and D2 suggesting a novel group of HBV. A similar finding could also be confirmed based on the entire 3,182 bp genome from four strains of HBV/D. This new cluster was supported by a branch with 99 % bootstrap value and 3.4-5.8 % nucleotide divergence over the entire genome from other known subgenotypes D1 to D9. Four strains of the new D subgenotype showed serotype ayw2, but had unique amino acid sequences consisting of N115 in the preS/S gene; P41 in the X gene; S239, K/E295, V567, and P708 in the P gene, respectively. From the above results, we provisionally proposed to designate it as novel quasi-subgenotype D2 identified in Taiwanese aborigines.

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Background: Antiviral actions of tetrapyrroles have been described in a number of systems. Our goal was to evaluate antagonism of the HCV NS3-4A protease by a variety of common porphyrins and characterize structure-activity relationships that may be useful for future drug design of HCV and related Flaviviruses. Methods: Using fluorometric assays, common metalloprotoporphyrins (MPP) all inhibited NS3-4A protease with IC 50 values in low micromolar ranges [CoPP (1.4 µM) < ZnPP = MnPP = SnPP < CuPP < FePP (6.5 µM) = protoporphyrin]. Results: Lineweaver-Burk plots confirmed that MPP: NS3 inhibition was basically competitive. All tested MPPs inhibited HCV genotype 1A, 1B, 2A and 3A recombinant proteases with the same fidelity suggesting wide antagonistic capabilities. However, when the MPPs were tested in cellular incubations with HCV replicons only Zn, Fe and free-base protoporphyrin showed comparable EC 50 and IC 50 values suggesting that there may be critical differences in MPP uptake and intracellular availability. Meso, deutero, and isohematoporphyrin derivatives, with or without metal substitution, all showed less anti-protease and antiviral activities as compared to protoporphyrins, suggesting that the planar, vinyl side chains are important for protease active site binding. MPPs were also active against three common protease mutants (T54A, A156T, and V36M) with equivalent or better IC 50 values as compared to wild type enzyme. Conclusion: These findings document the versatility of MPPs as antiviral agents with an expanded sensitivity for HCV genotypes and resistance to some common viral mutations. The results also suggest that further study of MPP structure and function will be useful for the development of new antiviral agents.

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