All‐trans retinoic acid for treatment of chronic hepatitis C

Background/Aims: In vitro studies in the subgenomic hepatitis C virus (HCV) replicon system have identified all‐trans retinoic acid (ATRA) as a potential therapeutic against hepatitis C. Thus, the antiviral potential of this drug should be assessed in vivo. Methods: Twenty highly treatment experienced serotype 1 patients with non‐response to conventional or pegylated interferon‐α (Peg‐/IFN‐α) and ribavirin were randomly assigned to 12 weeks of monotherapy with ATRA (group A) or a combination of ATRA and PegIFN‐α2a (group B). HCV RNA was assessed by bDNA assay and if negative by highly sensitive polymerase chain reaction. Results: During treatment, five of 10 patients in group A had a drop of viraemia >1log, while in group B after 8 weeks five of 10 dropped >2log, and three of 10 cleared HCV RNA from serum. Viraemia relapsed after treatment cessation. ATRA was rather well tolerated, with transient headache, dry skin and mucosa representing the most common side effects. Conclusions: The viral load reduction under ATRA monotherapy, although limited and transient, supports the antiviral activity of ATRA. However, the rapid loss of HCV RNA in three of 10 previous non‐responders under ATRA and PegIFN‐α2a treatment demonstrates a strong additive or synergistic ATRA effect and calls for a controlled trial to assess the therapeutic potential of this drug.     

Mutagenic effect of ribavirin on hepatitis C nonstructural 5B quasispecies in vitro and during antiviral therapy

BACKGROUND AND AIMS: Addition of ribavirin to interferon alfa treatment has substantially increased sustained virologic response rates in patients with chronic hepatitis C (CHC). Ribavirin acts as an RNA virus mutagen in vitro, thereby leading to error catastrophe. However, data in CHC are controversial. METHODS: The nonstructural (NS) 5B quasi-species heterogeneity was analyzed in Huh7 cells harboring a subgenomic hepatitis C virus (HCV) replicon system treated with ribavirin or levovirin. Accordingly, NS5B quasi-species were studied in 14 patients with CHC who received ribavirin alone or combined with pegylated interferon alfa both at baseline and during the first weeks of therapy. Analysis of NS3 quasi-species served as control. RESULTS: Cultivation of HCV replicon cells with ribavirin led to higher NS5B mutational frequencies compared with levovirin-treated or untreated cells (P < .05). Patients receiving ribavirin monotherapy showed higher overall mutational frequencies within NS3 and NS5B during therapy as compared with baseline (P < .01). Proportions of ribavirin-specific G-to-A and C-to-T transitions increased (P < .01). Paired analysis confirmed significant mean increases of mutational frequencies of approximately 5%. Ribavirin serum concentrations were positively correlated with mutational frequency changes (P < .05). In patients receiving combination therapy, a decrease of NS5B mutational frequencies ( approximately 10%) and lower proportions of G-to-A and T-to-C mutations (P < .01) were detectable. CONCLUSIONS: Ribavirin, but not its L-enantiomer levovirin, is a mutagen in HCV replicon cells. In patients with CHC, ribavirin monotherapy exhibits a moderate mutagenic effect early during therapy that is not detectable in combination with pegylated interferon alfa.     

IL‐28B (IFN‐λ3) and IFN‐α synergistically inhibit HCV replication

Genetic variation in the IL‐28B (interleukin‐28B; interferon lambda 3) region has been associated with sustained virological response (SVR) rates in patients with chronic hepatitis C treated with peginterferon‐α and ribavirin. However, the mechanisms by which polymorphisms in the IL‐28B gene region affect host antiviral responses are not well understood. Using the HCV 1b and 2a replicon system, we compared the effects of IFN‐λs and IFN‐α on HCV RNA replication. The anti‐HCV effect of IFN‐λ3 and IFN‐α in combination was also assessed. Changes in gene expression induced by IFN‐λ3 and IFN‐α were compared using cDNA microarray analysis. IFN‐λs at concentrations of 1 ng/mL or more exhibited concentration‐ and time‐dependent HCV inhibition. In combination, IFN‐λ3 and IFN‐α had a synergistic anti‐HCV effect; however , no synergistic enhancement was observed for interferon‐stimulated response element (ISRE) activity or upregulation of interferon ‐ stimulated genes (ISGs). With respect to the time course of ISG upregulation, the peak of IFN‐λ3‐induced gene expression occurred later and lasted longer than that induced by IFN‐α. In addition, although the genes upregulated by IFN‐α and IFN‐λ3 were similar to microarray analysis, interferon‐stimulated gene expression appeared early and was prolonged by combined administration of these two IFNs. In conclusion, IFN‐α and IFN‐λ3 in combination showed synergistic anti‐HCV activity in vitro. Differences in time‐dependent upregulation of these genes might contribute to the synergistic antiviral activity.     

Ribavirin mode of action in chronic hepatitis C: from clinical use back to molecular mechanisms

Ribavirin is an old broad‐spectrum antiviral that is highly effective when used in combination with interferon‐α and also as part of triple therapies containing new inhibitors of the hepatitis C virus (HCV) non‐structural (NS)3/4 protease or HCV NS5B polymerase for the treatment of patients with chronic hepatitis C. However, the molecular mechanisms by which ribavirin enhances early and sustained virological response rates during interferon‐based antiviral HCV therapy are still unknown. Several mechanisms including (i) immunomodulatory properties, (ii) inhibition of the inosine monophosphate dehydrogenase, (iii) direct inhibition of the HCV‐encoded NS5B RNA polymerase, (iv) induction of lethal mutagenesis and (v) modulation of interferon‐stimulated gene expression are currently proposed. Here, we discuss recent advances from in vitro data and their importance for the situation in patients with chronic hepatitis C. Furthermore, theoretical aspects from mathematical modelling of ribavirin action in chronic hepatitis C are reviewed.     

Amino acid substitutions of hepatitis C virus core protein are not associated with intracellular antiviral response to interferon‐α in vitro

Background: Studies on patients with hepatitis C virus (HCV) of genotype 1b have suggested that amino acids (aa) 70 and/or 91 of the HCV core protein affect the outcome of interferon (IFN)‐α and ribavirin (RBV) therapy, although there are no clear supporting data in vitro. Aims: This study was designed to determine the differences among the antiviral activities of HCV core proteins with various substitutions at aa70 and/or aa91. Methods: The retroviral vectors expressing the HCV core proteins with substitutions of arginine/leucine, arginine/methionine, glutamine/leucine or glutamine/methionine at aa70/aa91 were transiently transfected or stably transducted into an immortalized hepatocyte line (PH5CH8), hepatoma cell lines and an HCV‐RNA replicating cell line (sOR) to evaluate antiviral responses to IFN‐α or IFN‐α/RBV. Sequence analysis was performed using genome‐length HCV‐RNA replicating cells (OR6 and AH1) to evaluate HCV core mutations during IFN‐α treatment. Results: The promoter activity levels of IFN‐stimulated genes in the transiently transfected cells or the mRNA levels of 2′‐5′‐oligoadenylate synthetase in the stably transducted PH5CH8 cells were not associated with the HCV core aa70 and/or aa91 substitutions during IFN‐α treatment. Antiviral responses to IFN‐α or IFN‐α/RBV treatment were enhanced in sOR cells stably transducted with the HCV core, although there were no differences in antiviral responses among the cells expressing different core types. Sequence analysis showed no aa mutations after IFN‐α treatment. Conclusions: Antiviral activities were enhanced by HCV core transduction, but they were not associated with the HCV core aa70 and/or aa91 substitutions by in vitro analysis.     

Effect of ribavirin and amantadine on early hepatitis C virus RNA rebound and clearance in serum during daily high-dose interferon

The early rebound in serum HCV RNA during HCV dynamic studies with high-dose interferon may be due to de novo infection with interferon escape quasispecies. We simultaneously measured serum alanine aminotransferase (ALT) and HCV RNA at rapid intervals in chronic HCV liver disease patients during interferon therapy alone or in combination with ribavirin and amantadine. HCV RNA declined rapidly between 0 and 48 hr in all patients (phase 1). Ribavirin and amantadine significantly increased this phase 1 decline. In all four monotherapy patients with viral rebound, the increasing levels of HCV RNA were associated with a parallel increase in serum ALT, consistent with a hepatitis flare or de novo infection. By contrast, in the four monotherapy patients without viral rebound, and all eight patients receiving combination therapy, the slow progressive phase two decay was associated with declining serum ALT levels. Ribavirin or ribavirin and amantadine significantly and incrementally increased the phase two HCV RNA clearance. Dynamic sequencing in the HVR1 region in one rebound patient confirmed the potential for rapid evolutionary changes during interferon therapy. These preliminary data suggest that early viral rebound might be associated with de novo infection with interferon escape HCV variants, which in turn are attenuated by ribavirin and amantadine.     

Firstline treatment for hepatitis C: combination interferon/ribavirin versus interferon monotherapy

In the initial treatment of chronic hepatitis C, interferon‐alfa (IFN‐α) monotherapy for 24–48 weeks induces sustained response rates of only 10–20%. Combination therapy with IFN‐α plus ribavirin induces a sustained response in 40–50% of patients, and can be now recommended as the firstline therapy for chronic hepatitis C. Stopping therapy at week 12 because of persistent viraemia is unnecessary with the combination therapy because later clearance of HCV RNA can still occur with a sustained response. Patients with HCV genotype 1 should receive 48 weeks of combination therapy, in contrast to 24 weeks for patients with genotypes 2 or 3. For patients who cannot tolerate the side effects of ribavirin, such as anaemia, IFN‐α at 3 MU three times weekly for 48 weeks is preferred as the initial therapy. The long‐acting pegylated IFN can be expected to enhance the efficacy of combination therapy in the treatment of chronic hepatitis C and appears to be much more potent as monotherapy. Further studies are needed to improve the current ‘half‐full’ status of chronic hepatitis C treatment.     

Modulation of replication efficacy of the hepatitis C virus replicon Con1 by site‐directed mutagenesis of an NS4B aminoterminal basic leucine zipper

Summary. The hepatitis C virus (HCV) nonstructural protein 4B (NS4B) is assumed to function as a membrane anchor and protein hub for the viral replication complex. The aim of the current work was to modulate HCV replication efficacy in the subgenomic Con1 replicon by mutations of specific sites within the aminoterminal‐located basic leucine zipper (bZIP), a candidate motif for protein–protein interactions involving NS4B. Mutational sites and amino acid substitutes were determined by in‐silico sequence analyses of the NS4B‐bZIP motif in 357 isolates of HCV genotype 1b from the euHCVdB and LosAlamos database and consecutive analysis of conserved physico‐chemical properties at bZIP specific positions. Mutants with predicted minor, medium or major reduction of replication efficacy were tested in the pFKI389neo/NS3‐3′/ET plasmid replicon model. Four sites (L25, T29, V39 and W43) of crucial importance for bZIP‐mediated protein interaction with predicted apolarity of respective amino acid positions were selected for mutational studies. Substitutes with physico‐chemical properties matching the predicted requirements either well (T29A), moderately (L25W, V39W), or insufficiently (T29E, W43E) were associated with slightly improved, moderate and marked decreased replication efficacy, respectively. Spontaneous (T29G) and adaptive (A28G, E40G) mutations occurred in the T29E mutation isolate only and were associated with marked reduction of replication efficacy. The bZIP motif region of NS4B is crucial for RNA replication in the subgenomic Con1 replicon system. RNA replication efficacy can be modulated by site‐directed mutagenesis at specific bZIP functional sites. New adaptive amino acid mutations were identified within the HCV NS4B protein.     

Evolution of hepatitis C virus genome in chronically infected patients receiving ribavirin monotherapy

Recent results of clinical trials suggest that combination of interferon and ribavirin exhibits an enhanced antiviral effect in the treatment of chronic hepatitis C. To investigate the effect of ribavirin on hepatitis C virus (HCV) infection, we analysed the evolution of the genetic heterogeneity of HCV in relation to the anti-HCV humoral response in patients treated by ribavirin alone. The study population included 35 patients with liver biopsy proven chronic hepatitis C infected with HCV genotype 1. Among them, 26 were treated with ribavirin for at least 12 months and nine untreated patients served as a control group. Serum samples were analysed before and at 6 and 12 months of therapy. Three regions of the HCV genome, i.e. HVR1, a domain of NS5A including part of the interferon sensitivity determining region (ISDR), and a segment of NS5B, were amplified by RT-PCR using specific primers. The PCR products were then studied using single-strand conformation polymorphism (SSCP) analysis followed by either direct sequencing, or cloning and sequencing. In parallel, the humoral anti-E1 response was studied using an ELISA (Innotest HCV E1Ab, Innogenetics). The results of HCV genome analysis showed no significant effect on the amino acid sequence evolution of the HVR1, NS5A and NS5B regions of HCV. Analysis of a phylogenetic tree from the major quasispecies variants showed the absence of correlation with ribavirin response, and the absence of selection of viral strains during ribavirin treatment. A trend towards a decrease in the anti-E1 Ab response was also observed. Altogether these results suggest that ribavirin may not exhibit a direct antiviral effect, but may trigger a favourable response to interferon by modulating the immune response against HCV.     

Antiviral therapy: inhibition of Hepatitis C Virus expression by RNA interference directed against the NS5B region of the viral genome

Background: Hepatitis C virus (HCV) is a major public health problem with 170 million chronically infected people throughout the world. Currently, the only treatment available consists of a combination of pegylated interferon (INF-a) and ribavirin, but only half of the patients treated show a sufficient antiviral response. Thus there is a great need for the development of new treatments for HCV infections. RNA interference (RNAi) represents a new promising approach to develop effective antiviral drugs and has been extremely effective against HCV gene expression in short-term cell culture. Our aim was to determine the effect of RNAi directed against the NS5B-HCV region on HCV expression in a human hepatoma cell line that expresses HCV-subgenomic replicon (Huh7 HCV replicon cells). Methods: We transfected Huh7 HCV replicon cells with different concentrations of RNAi (100-200 nM) targeting the NS5B region of the viral genome. 2-6 days post-transfection HCV-RNA was quantified by semiquantitative and real-time RT-PCR, and HCV NS5B protein levels were assayed by western blot. Cell viability was also quantified by MTT assay. Results: Our results indicate that the NS5B-siRNAs used in this study can specifically inhibit HCV-RNA replication and protein expression (more than 90%) compared to control cells. Conclusions: Synthetic siRNA against NS5B-HCV inhibited HCV replication and viral proteins levels and thereby becomes a powerful strategy to combat hepatitis C virus.     

Interferon-α suppressed granulocyte colony stimulating factor production is reversed by CL097, a TLR7/8 agonist

Background and Aim: Neutropenia, a major side‐effect of interferon‐α (IFN‐α) therapy can be effectively treated by the recombinant form of granulocyte colony stimulating factor (G‐CSF), an important growth factor for neutrophils. We hypothesized that IFN‐α might suppress G‐CSF production by peripheral blood mononuclear cells (PBMCs), contributing to the development of neutropenia, and that a toll‐like receptor (TLR) agonist might overcome this suppression. Methods: Fifty‐five patients who were receiving IFN‐α/ribavirin combination therapy for chronic hepatitis C virus (HCV) infection were recruited. Absolute neutrophil counts (ANC), monocyte counts and treatment outcome data were recorded. G‐CSF levels in the supernatants of PBMCs isolated from the patients and healthy controls were assessed by enzyme‐linked immunosorbent assay following 18 h of culture in the absence or presence of IFN‐ α or the TLR7/8 agonist, CL097. Results: Therapeutic IFN‐α caused a significant reduction in neutrophil counts in all patients, with 15 patients requiring therapeutic G‐CSF. The reduction in ANC over the course of IFN‐α treatment was paralleled by a decrease in the ability of PBMCs to produce G‐CSF. In vitro G‐CSF production by PBMCs was suppressed in the presence of IFN‐α; however, co‐incubation with a TLR7/8 agonist significantly enhanced G‐CSF secretion by cells obtained both from HCV patients and healthy controls. Conclusions: Suppressed G‐CSF production in the presence of IFN‐α may contribute to IFN‐α‐induced neutropenia. However, a TLR7/8 agonist elicits G‐CSF secretion even in the presence of IFN‐α, suggesting a possible therapeutic role for TLR agonists in treatment of IFN‐α‐induced neutropenia.     

Hepatitis C virus NS3/4A quasispecies diversity in acute hepatitis C infection in HIV‐1 co‐infected patients

The growing number of cases of acute hepatitis C (AHC) infections among human immunodeficiency virus type 1 (HIV‐1)‐positive men who have sex with men (MSM) in the last 10 years has promoted the search for predictors of AHC clearance as well as for epidemiological networks of viral transmission. We characterized the diversity and catalytic efficiency of HCV NS3/4A protease quasispecies in AHC patients coinfected with HIV‐1. Plasma samples obtained at HCV diagnosis from 18 MSM HIV‐coinfected patients with AHC were studied. Five HCV monoinfected patient samples with AHC were also investigated. An average of 39 clones from each sample was analysed. The catalytic efficiency of the dominant quasispecies (i.e. the most abundant) from each quasispecies was also assayed for mitochondrial antiviral signalling protein (MAVS) cleavage. Phylogenetic analysis identified two clusters of patients with highly related viruses, suggesting a common source of HCV infection. None of the 18 MSM HIV‐coinfected patients spontaneously cleared HCV, although 78% of the treated patients achieved a sustained virological response after early treatment with pegylated interferon (pegIFN) plus ribavirin (RBV). The synonymous‐nonsynonymous (ds/dn) mutation ratio, a marker of selective pressure, was higher in AHC compared to 26 HIV‐1‐infected men with genotype 1a chronic hepatitis C (CHC) (P < 0.0001). NS3/4A proteases from AHC patients also exhibited higher catalytic efficiency compared to CHC patients (P < 0.0001). No differences were found when ds/dn mutation ratios and NS3/4A protease catalytic efficiencies from AHC HIV‐coinfected patients were compared with AHC monoinfected patients. The presence of epidemiological networks of HCV transmission was confirmed among HIV‐1‐positive MSM. In addition, substantial genetic diversity was demonstrated in AHC. NS3/4A protease efficiency cleaving MAVS may be associated with virus transmission and response to pegIFN/RBV treatment.     

High incidence of type 1 diabetes mellitus during or shortly after treatment with pegylated interferon α for chronic hepatitis C virus infection

Background: Development of diabetes mellitus (DM) during or shortly after treatment with interferon α (IFN‐α) in patients with chronic hepatitis C virus (HCV) infection has been reported sporadically. We prospectively screened for DM during and after IFN‐α therapy for chronic HCV infection. Methods: Blood glucose levels of patients with chronic HCV infection were routinely assessed at all outpatient visits during and after treatment with pegylated‐IFN‐α (Peg‐IFN‐α) and ribavirin (Riba). Results: Between December 2002 and October 2005, 189 non‐diabetic patients were treated with Peg‐IFN‐α/Riba, of whom five developed type 1 DM (2.6%), three type 2 DM (1.6%) and one an indeterminate type of DM. Classical symptoms of DM were present in three patients who developed DM shortly after cessation of Peg‐IFN‐α/Riba. In the other patients, symptoms of DM were either indistinguishable from side effects caused by Peg‐IFN‐α/Riba or absent. Conclusion: Our study showed a high incidence of type 1 DM during Peg‐IFN‐α/Riba therapy for chronic HCV infection. Symptoms of DM may be absent or mistaken for Peg‐IFN‐α/Riba‐associated side effects. To diagnose DM without delay, we propose routine assessment of blood glucose at all outpatient visits during and after Peg‐IFN‐α/Riba treatment in chronic HCV patients.     

Current issues in the management of paediatric viral hepatitis

Viral hepatitis poses important problems for children. In preschoolers, hepatitis A virus (HAV) infection frequently causes acute liver failure. Vaccinating toddlers against HAV in countries with high endemicity is expected to decrease mortality. HAV vaccine demonstrates efficacy (comparable to immunoglobulin) as post‐exposure prophylaxis. A recently developed vaccine against hepatitis E virus (HEV) may benefit fetal health, because pregnant women are most prone to acute liver failure as a result of HEV. Hepatitis B vaccine continues to demonstrate value and versatility for preventing serious liver disease. With chronic infection, undetectable levels of serum HBV DNA complement e‐seroconversion as the preferred outcome measure; suppressed viral load correlates with long‐term complications better than HBeAg status. Among Taiwanese children, low pretreatment HBV DNA (<2 × 10⁸ copies/ml) strongly predicted response to interferon‐α. Future paediatric studies must incorporate HBV DNA levels. The rationale for routine treatment of immunotolerant hepatitis B during childhood remains uncertain. Any treatment of chronic hepatitis B in childhood requires consideration of the risks and benefits. Childhood hepatitis C virus (HCV) infection results mainly from mother‐to‐infant transmission. Babies of HCV‐infected women should be tested for serum HCV RNA at 1 month of age. If negative, confirmatory anti‐HCV antibody testing may be performed between 12 and 15 months of age. Children with chronic hepatitis C may develop progressive fibrosis/cirrhosis, particularly in the setting of obesity and insulin resistance. Treatment of children chronically infected with genotype 2 or 3 is highly successful: combination therapy of pegylated interferon‐α and ribavirin is well tolerated and superior to pegylated interferon‐α alone.     

Identification of a ribavirin-resistant NS5B mutation of hepatitis C virus during ribavirin monotherapy

Ribavirin (RBV), a guanosine analogue, has been suggested to exert an antiviral action against hepatitis C virus (HCV) by causing lethal mutations and suppressing RNA polymerase in vitro, but the mechanism of its clinical therapeutic effects is currently unknown. To test the hypothesis that RBV could act both as an RNA mutagen and inhibit viral RNA synthesis in vivo, we studied the evolution of the nucleotide sequences of HCV RNA at the nonstructural (NS) 5B region in patients receiving RBV, placebo, or interferon alfa (IFN-alpha) monotherapy. The RBV group showed a slightly more accelerated evolution rate of HCV RNA quasispecies than either the IFN-alpha or placebo group. RBV caused preferentially A-to-G and U-to-A mutations. Interestingly, an NS5B amino acid 415 Phe-to-Tyr (F415Y) mutation emerged in all (5 of 5) patients infected with HCV genotype 1a during the RBV treatment. Subsequently, the parental 415F strain reemerged in some patients after the treatment was discontinued. The effect of the amino acid substitution at NS5B415 on HCV RNA replication was then investigated using an HCV subgenomic replicon in Huh7 cells. We showed that treatment of replicon cells with RBV reduced the HCV RNA level of NS5B415F replicon, but not NS5B415Y, in a dose-dependent manner. Thus, NS5B F415Y mutation represents an RBV-resistant variant. The 3-dimensional modeling and structure analysis of NS5B protein revealed that the 415th amino acid is located at the P helix region of the thumb subdomain, which may interact with the minor groove of the template-primer duplex in the putative RNA-binding cleft. In conclusion, RBV could work as a weak mutagen for HCV RNA in HCV-infected patients. Furthermore, the selection of an RBV-resistant variant with a single amino acid substitution in NS5B suggested that RBV may directly interact with HCV RNA polymerase, thus interfering with its enzymatic activity.     

Impact of high‐dose interferon induction and ribavirin therapy in patients with chronic hepatitis C relapsing after or not responding to interferon monotherapy

Background/aims: Initial high‐dose interferon‐α induction therapy in combination with ribavirin improves sustained response rates in treatment‐naïve patients. This prospective, randomized, controlled study tested whether non‐responders or relapsers to interferon monotherapy also benefit from induction therapy. Methods: Patients with chronic hepatitis C who had not responded to (n=75) or relapsed (n=80) after previous interferon therapy were randomized to receive three different interferon doses during the first 14 weeks of therapy (A: 10 MU IntronA®/day for 2 weeks, followed by 10 MU/2 days for 12 weeks; B:5 MU/d for 14 weeks; C: 5 MU/2 days for 14 weeks) followed in all by 5 MU/2 days for 24 weeks. All patients received 1–1.2 g ribavirin/day throughout the whole study. Results: The rates of viral clearance at any time on treatment were similar in all groups. Sustained response rates were also not different among the groups in interferon nonresponders (A 32%, B 29%, C 31%) and relapsers (A 64%, B 68%, C 71%), respectively, as well as in patients with different genotypes. As expected, sustained response rates were higher in patients with genotype non‐1 than in those with genotype 1. Conclusion: High‐dose induction therapy does not improve the outcome of interferon/ribavirin therapy in interferon nonresponders or relapsers.     

Effect of combination therapy with ribavirin and high-dose interferon-α2b for 24 weeks in chronic hepatitis C

Background and Aim: The aim of the present study was to determine whether a 24‐week course of combination therapy with ribavirin and high‐dose interferon‐α2b (IFN‐α2b) could provide an acceptable treatment efficacy in chronic hepatitis C (CHC). Methods: Seventy‐six patients with CHC whose serum hepatitis C virus (HCV) RNA levels were more than 100 kIU/mL on quantitative polymerase chain reaction (PCR) assay were included. The patients were assigned to two different dose groups of IFN‐α2b: group A (n = 39) received 6 MU and group B (n = 37) received 10 MU. Each group received the dose daily for 14 days then three times per week for a total of 24 weeks. In addition, HCV genotype 1b patients in group A and group B were classified into group C (n = 20) and D (n = 29), respectively. All patients received 600 or 800 mg ribavirin per day. Results: Sustained response rates in group A were significantly higher than those in group B (66.7%vs 35.1%, intent‐to‐treat, P = 0.0060). However, sustained response rates in group C were not different from those in group D (45.0%vs 20.7%, intent‐to‐treat, P = 0.0696). The proportion of patients who discontinued the treatment or reduced drug dosage because of adverse events was significantly higher in group B than in group A (27.0%vs 7.69%, P = 0.0224). Conclusion: A 24‐week course of combination therapy with ribavirin and 6 MU IFN‐α2b had an acceptable efficacy with fewer adverse events than that with ribavirin and 10 MU IFN‐α2b in CHC.     

Peginterferon alpha-2a (40 KD) plus ribavirin for the treatment of chronic hepatitis C in Japanese patients

Background: The efficacy and safety of peginterferon alpha‐2a (40 KD) (peg‐IFNα‐2a) plus ribavirin has not been reported for Japanese patients with chronic hepatitis C. The aim of this study was to evaluate this combination in treatment‐naïve patients and in non‐responders or relapsers to interferon monotherapy. Methods: Overall, 201 treatment‐naïve patients with hepatitis C virus (HCV) genotype‐1b were randomly assigned to 180 µg peg‐IFNα‐2a once‐weekly plus ribavirin 600–1000 mg/day or peg‐IFNα‐2a plus placebo for 48 weeks. Additionally, peg‐IFNα‐2a plus ribavirin was administered for 48 weeks to 100 non‐responders or relapsers (85% genotype‐1) to previous interferon monotherapy. Results: A sustained virological response (SVR) was attained among significantly more treatment‐naïve patients receiving combination therapy than monotherapy (61% vs 26%, P < 0.001). For patients with high baseline HCV RNA, the SVR rate was 59% with peg‐IFNα‐2a plus ribavirin versus 24% with peg‐IFNα‐2a monotherapy. Among non‐responders or relapsers to previous interferon monotherapy, 54% attained an SVR. Adverse events were generally mild, and discontinuations rates due to adverse events or laboratory abnormalities were low. Conclusion: In Japanese patients, peg‐IFNα‐2a plus ribavirin provided significant improvement in SVR rates compared with peg‐IFNα‐2a alone in treatment‐naïve patients, and was effective as re‐treatment for non‐responders or relapsers to previous treatment with interferon monotherapy.     

Mitochondrial electron transport inhibition in full genomic hepatitis C virus replicon cells is restored by reducing viral replication

Background/Aim: Hepatitis C virus (HCV) core protein has been shown to inhibit mitochondrial electron transport and to increase reactive oxygen species (ROS) in vitro and in vivo. The aim of this study was to investigate whether inhibiting HCV replication could restore the mitochondrial redox state and electron transport activity. Methods: We measured ROS, mitochondrial reduced glutathione content, and mitochondrial complex I, II, III and IV activities and protein expression in full genomic HCV replicon cells and cured cells that had been prepared by eliminating HCV RNA from replicon cells by interferon (IFN)‐α treatment. Results: Cured cells had significantly lower ROS production and greater mitochondrial glutathione content than replicon cells. Complete inhibition of HCV replication by IFN‐α restored complex I and IV activities by 20–30% (P<0.01) and complex I expression (P<0.05). Treatment with fluvastatin, one of the 3‐hydroxy‐3‐methylglutaryl co‐enzyme A reductase inhibitors, which is known to have anti‐HCV activity, partially inhibited core protein expression and restored complex I activity in full genomic HCV replicon cells to a lesser degree (P<0.05). Conclusions: Our results show that the mitochondrial redox state and electron transport activity can be restored by reducing HCV replication.     

Impact of the peginterferon‐α 2a and ribavirin plasma levels on viral kinetics and sustained virological response in genotype 1 HCV/HIV‐co‐infected patients with the unfavourable non‐CC IL28B genotypes

Studies on the association between the peginterferon‐α and ribavirin levels and sustained virological response (SVR) have shown yielded conflicting results, but most of them were performed before the influence of IL28B polymorphisms was known. Our aim was to assess the effects of peginterferon‐α 2a and ribavirin plasma levels on viral kinetics and SVR in hepatitis C virus genotype 1 HCV‐1/HIV‐co‐infected patients according to IL28B genotype. This was a cohort study of HCV‐1/HIV‐co‐infected patients who were HCV‐treatment naïve and for whom the efficacy of peginterferon‐α 2a plus ribavirin was evaluated by per‐protocol analysis. The peginterferon‐α 2a and ribavirin levels were measured by ELISA and HPLC‐UV, respectively. The relationships among host and viral factors, the trough drugs levels and virological responses were analysed by multivariate regression analyses. A total of 131 Caucasian patients were included (cirrhosis:38.9%). Overall, SVR rate was 39.6%. In patients with CC IL28B genotype, SVR was related neither to peginterferon‐α 2a nor to ribavirin plasma levels, while higher levels of both drugs were the only variables independently associated with SVR in individuals with CT/TT IL28B genotypes (OR, 5.02; CI₉₅, 1.45–17.1; P = 0.001 and 4.0; CI₉₅, 1.08–14.7; P = 0.038, respectively). Moreover, faster viral declines were observed in CT/TT patients when pegIFN‐α 2a and ribavirin plasma levels were greater than 3400 pg/mL and 1.6 μg/mL, respectively. In contrast to the results for CC patients, the results in patients carrying the unfavourable CT/TT IL28B genotypes showed that plasma levels of both drugs have significant effects on viral kinetics and SVR.