Shogo Ohkoshi

Hypervariable regions in the putative glycoprotein of hepatitis C virus

A comparison of the sequences of the putative glycoprotein region in three independent cDNA clones of hepatitis C virus and of sequences of four other clones revealed extensive genetic variation clustered and interspersed with highly conserved amino acid sequences. We obtained evidence for two hypervariable regions in the putative envelope glycoprotein, one region was assumed to be a potential antigenic site, as deduced from the hydrophilicity and analyses of secondary structures. These observations suggest the existence of a large pool of antigenic variants of hepatitis C virus, in Japan.

Comprehensive Analysis of the Effects of Ordinary Nutrients on Hepatitis C Virus RNA Replication in Cell Culture

ABSTRACT To date, only a limited number of studies have reported finding an influence of ordinary nutrients on hepatitis C virus (HCV) RNA replication. However, the effects of other nutrients on HCV RNA replication remain largely unknown. We recently developed a reporter assay system for genome-length HCV RNA replication in hepatoma-derived HuH-7 cells (OR6). Here, using this OR6 assay system, we comprehensively examined 46 nutrients from four nutrient groups: vitamins, amino acids, fatty acids, and salts. We found that three nutrients—β-carotene, vitamin D2, and linoleic acid—inhibited HCV RNA replication and that their combination caused additive and/or synergistic effects on HCV RNA replication. In addition, combined treatment with each of the three nutrients and interferon alpha or beta or fluvastatin inhibited HCV RNA replication in an additive manner, while combined treatment with cyclosporine synergistically inhibited HCV RNA replication. In contrast, we found that vitamin E enhanced HCV RNA replication and negated the effects of the three anti-HCV nutrients and cyclosporine but not those of interferon or fluvastatin. These results will provide useful information for the treatment of chronic hepatitis C patients who also take anti-HCV nutrients as an adjunctive therapy in combination with interferon. In conclusion, among the ordinary nutrients tested, β-carotene, vitamin D2, and linoleic acid possessed anti-HCV activity in a cell culture system, and these nutrients are therefore considered to be potential candidates for enhancing the effects of interferon therapy.

Reduced NKG2D ligand expression in hepatocellular carcinoma correlates with early recurrence

BACKGROUND & AIMSnThe activating receptor natural killer group 2, member D (NKG2D) and its ligands play a crucial role in immune response to tumors. NKG2D ligand expression in tumors has been shown to be associated with tumor eradication and superior patient survival, but the involvement of NKG2D ligands in the immune response against hepatocellular carcinoma (HCC) still remains to be elucidated.nnnMETHODSnWe investigated the expression of NKG2D ligands in HCC tissues collected from 54 patients and HCC cell lines. We also examined the proteasome expression and the effect of inhibition of proteasome activity on NKG2D ligand expression in HCC tissues and cell lines.nnnRESULTSnIn dysplastic nodules (DN), well-differentiated (well-HCC), and moderately-differentiated HCCs (mod-HCC), UL16-binding protein (ULBP) 1 was expressed predominantly in tumor cells, but not in poorly-differentiated HCCs (poor-HCC). Remarkably, recurrence-free survival of patients with ULBP1-negative HCC was significantly shorter than that of patients with ULBP1-positive HCC (p=0.006). Cox regression analysis revealed that loss of ULBP1 expression was an independent predictor of early recurrence (p=0.008). We confirmed that ULBP1 was expressed in the well- and mod-HCC cell lines, but not in the poor-HCC cell line KYN-2. However, inhibition of proteasome activity resulted in significant up-regulation of ULBP1 expression in KYN-2. Moreover, we found that 20S proteasome expression was more abundant in KYN-2 than that in the well- and mod-HCC cell lines.nnnCONCLUSIONSnULBP1 is prevalently expressed in DN to mod-HCC, but loss of its expression correlates with tumor progression and early recurrence.

Serum alpha-fetoprotein levels during and after interferon therapy and the development of hepatocellular carcinoma in patients with chronic hepatitis C.

The association between serum alpha-fetoprotein (AFP) levels during and after interferon (IFN) therapy and the development of hepatocellular carcinoma (HCC) was evaluated in patients with chronic hepatitis C (CHC). A total of 263 patients treated by IFN with or without ribavirin were enrolled in the study. Serum AFP levels during and after IFN therapy were investigated retrospectively, and statistical analysis was performed to identify the factors associated with HCC development. During IFN therapy, serum AFP levels significantly decreased, regardless of virologic response to treatment. Increased serum AFP levels (≥10xa0ng/ml) at the end of IFN therapy (EOT) was a close-to-significant variable affecting the development of HCC (Pxa0=xa00.057), and a significantly higher cumulative incidence of HCC was seen in patients with increased serum AFP levels at EOT (Pxa0=xa00.021). Serum AFP level at EOT is a possible predictor of HCC in CHC patients after IFN therapy.

Interferon Inhibits Progression of Liver Fibrosis and Reduces the Risk of Hepatocarcinogenesis in Patients with Chronic Hepatitis C: A Retrospective Multicenter Analysis of 652 Patients

A retrospective multicenter analysis of 652 patients with chronic hepatitis C who have been treated with interferon (IFN) was performed to assess the effects of IFN on the clinical course and development of HCC. During a mean follow-up of 54.8 months, hepatocellular carcinoma (HCC) developed in 7.0% of the patients. The rate was significantly higher in the patients who did not respond to IFN treatment than in those with sustained virological response and those who obtained a normalization of alanine aminotransferase levels despite the presence of HCV RNA (incomplete response) (P < 0.01). Using multivariate Coxs proportional hazard model, alcohol abuse (P < 0.05) and a higher level of fibrosis (P < 0.05) before treatment were the significant background factors associated with HCC development in the patients who did not respond to IFN. Interestingly, a significant increase in the rate of HCC development occurred in patients who had a histological finding of progressive fibrosis (F3). In addition, patients with low histological staging scores were likely to have an incomplete response, even if a sustained virological response was not obtained. IFN produced an improvement in histological activity and fibrosis stage in the second biopsy specimens irrespective of the clinical outcome when compared against untreated subjects.

Inhibition of hepatitis C virus infection and expression in vitro and in vivo by recombinant adenovirus expressing short hairpin RNA

Background and Aim:u2002 We have reported previously that synthetic small interfering RNA (siRNA) and DNA‐based siRNA expression vectors efficiently and specifically suppress hepatitis C virus (HCV) replication in vitro. In this study, we investigated the effects of the siRNA targeting HCV‐RNA in vivo.

Sustained biochemical remission after interferon treatment may closely be related to the end of treatment biochemical response and associated with a lower incidence of hepatocarcinogenesis

Abstract: Clinical background and incidence of hepatocellular carcinoma (HCC) of patients with chronic hepatitis C who obtained biochemical remission without eradication of virus (biochemical response) after interferon (IFN) treatment was retrospectively analyzed for 755 patients. Annual incidence of HCC was significantly lower in the patients with biochemical response and sustained response than that of the patients that did not show these responses. Logistic regression analysis showed that only the normalization of alanine aminotransferase (ALT) value at the end of IFN treatment was a significant factor for biochemical response. Annual incidence of HCC was significantly lower in the patients who obtained normalization of ALT values at the end of treatment than those who did not. Patients who were younger, who had a lower level of activity and fibrosis indices in histology, higher platelet count, and who were given more higher total dose of IFN were more likely to attain normalization of ALT levels at the end of treatment, and this was related to biochemical response. Low incidence of HCC in patients who obtained normalization of ALT values at the end of treatment was likely because they were in the earlier stage of chronic hepatitis. Active treatment of chronic hepatitis C with interferon in the early phase of the disease may bring about a biochemical response in some patients, even if sustained virological response is not obtained.

Blockade of ataxia telangiectasia mutated sensitizes hepatoma cell lines to sorafenib by interfering with Akt signaling

Sorafenib is a multi-kinase inhibitor applicable to hepatocellular carcinoma (HCC), but its limited therapeutic effects are a major problem to be solved. Here, we show that blockade of ataxia telangiectasia mutated (ATM) improves the antitumor effects of sorafenib. When hepatoma cell lines HepG2 and PLC/PRF/5 were treated with sorafenib plus ATM small inhibitory RNAs, ATM inhibitor KU55933 or caffeine, Akt signaling was suppressed and the cytotoxic effects were significantly potentiated. Moreover, ATM inhibition effectively suppressed the sorafenib-induced cell migration. Taken together, manipulation of ATM activity might be a useful strategy for improving sorafenib treatment of HCC.

Multicentric occurrence of hepatocellular carcinoma with nonalcoholic steatohepatitis

AIMnTo reveal the manner of hepatocellular carcinoma (HCC) development in patients with nonalcoholic steatohepatitis (NASH) focusing on multicentric occurrence (MO) of HCC.nnnMETHODSnWe compared clinicopathological characteristics between patients with and without MO of HCC arising from NASH background. The clinical features were implicated with reference to the literature available.nnnRESULTSnMO of HCC was identified with histological proof in 4 out of 12 patients with NASH-related HCC (2 males and 2 females). One patient had synchronous MO; an advanced HCC, two well-differentiated HCCs and a dysplastic nodule, followed by the development of metachronous MO of HCC. The other three patients had multiple advanced HCCs accompanied by a well-differentiated HCC or a dysplastic nodule. Of these three patients, one had synchronous MO, one had metachronous MO and the other had both synchronous and metachronous MO. There were no obvious differences between the patients with or without MO in terms of liver function tests, tumor markers and anatomical extent of HCC. On the other hand, all four patients with MO of HCC were older than 70 years old and had the comorbidities of obesity, type 2 diabetes mellitus (T2DM), hypertension and cirrhosis. Although these conditions were not limited to MO of HCC, all the conditions were met in only one of eight patients without MO of HCC. Thus, concurrence of these conditions may be a predisposing situation to synchronous MO of HCC. In particular, old age, T2DM and cirrhosis were suggested to be prerequisite for MO because these factors were depicted in common among two other cases with MO of HCC under NASH in the literature.nnnCONCLUSIONnThe putative predisposing factors and necessary preconditions for synchronous MO of HCC in NASH were suggested in this study. Further investigations are required to clarify the accurate prevalence and predictors of MO to establish better strategies for treatment and prevention leading to the prognostic improvement in NASH.

Characterization of elevated alanine aminotransferase levels during pegylated-interferon α-2b plus ribavirin treatment for chronic hepatitis C.

Aim:u2002 Elevation of alanine aminotransferase (ALT) levels during pegylated‐interferon (peg‐IFN) plus ribavirin therapy in patients with chronic hepatitis C [CHC] is a problem that cannot be disregarded. The aim of this study is to assess the frequency and to characterize clinical parameters of this phenomenon.