Title: Will There Be an HCV Meeting in 2020? Summary of the 17th International Meeting on Hepatitis C Virus and Related Viruses
Abstract: Hepatitis C virus (HCV), which was discovered in 1989, is a major etiologic agent in human liver disease. Approximately 130 million people, or 2% of the population, worldwide are infected. The 17th International Meeting on Hepatitis C Virus and Related Viruses was held September 10–14, 2010, in Yokohama, Japan. The meeting was attended by almost 700 scientists from all over the world who are interested in the fundamental aspects of the molecular virology, immunology, pathogenesis, prevention, and treatment of HCV infection. Two special opening lectures given by Masaaki Komatsu and Takashi Gojobori focused attention on the related research fields of autophagy and genome biology, respectively. In the subsequent sessions, the latest research, original studies, and controversies were presented in 9 keynote lectures, 82 oral presentations, and 329 poster presentations. The opening scientific session of this meeting focused on the viral host cell entry processes. Thomas Baumert presented the keynote lecture, which included an overview of the HCV cell entry process and recent advances at his laboratory. These included the finding that HCV variants that reinfect the liver after transplantation demonstrate more efficient cell entry and are less susceptible to neutralization by host antibodies. He also described the isolation of monoclonal antibodies against claudin-1 that do not inhibit either extracellular or direct cell-to-cell HCV transfer. Alexander Ploss described the establishment of a mouse model for studying HCV cell entry. They utilized an HCV cell culture virus (HCVcc) expressing recombinase and transgenic mice bearing a recombinase-activatable fluorescent protein. Bioluminescent imaging indicated that only mice transduced with CD81 and occludin supported HCVcc entry. The presence of an intact immune system in these animals makes it particularly important for the testing of HCV vaccine candidates. Danyelle N. Martin described a role for transferrin receptor 1 (TfR1) in mediating HCV cell entry. The inhibition of HCV entry with TfR1 antibodies and silencing, suggest this factor should be added to the growing list of cellular proteins required for HCV cell entry. Joachim Lupberger presented results from a study showing an essential role for the epidermal growth factor receptor (EGFR) in HCV cell entry. He found that EGFR is required for both mediating the interactions between two other entry factors, CD81 and CLDN1, and catalyzing the fusion activity of viral glycoproteins. Volker Lohmann began the session by describing what is known of the functions of viral nonstructural proteins and their associated host cellular factors in viral translation and replication. He included an overview of viral isolates and model systems currently used, and presented data addressing the mechanisms for efficient replication of the JFH-1 isolate. Several reports have focused on the molecular basis of the architecture and composition of membrane-associated sites for HCV replication, which often induce membrane alterations, such as the so-called membranous web. Brenno Wolk demonstrated that NS4B is sufficient to direct all nonstructural proteins into the viral replication complex compartment, and that intragenotype-specific interactions are required for NS4B-dependent recruitment of NS5A. Ines Romero-Brey showed that the membranous web predominantly contains double-membrane vesicles with various diameters. These vesicle structures were connected to the endoplasmic reticulum (ER) through funnel-like structures. Several DDX DEAD-box RNA helicases were identified as host factors associated with HCV replication. Yasuo Ariumi presented the cross-talk of HCV with DDX proteins and the role of distinct DDX proteins in viral replication. Tetsuro Shimakami and Selena M. Sagan reported the importance of miR-122 to not only enhance IRES-mediated translation, but stabilize positive-strand HCV RNA by binding to its 5′ extremity. Enzymatic activity of host phosphatidyl-inositol-4 kinase III alpha was shown to be critically involved in HCV replication and the activity is regulated by HCV NS5A (Simon Reiss). Nam-Joon Cho reconstituted a functionally active full-length HCV polymerase on a biomimetic membrane platform. Deborah Harrus found that guanosine triphosphate specifically stimulates the initial step of de novo initiation by stimulating transition of newly formed linker primer. In the keynote lecture, Guangxiang G. Luo presented an overview of particle assembly and release, and the impact of apolipoprotein (Apo) E in the entry and assembly of HCV. He demonstrated the inhibition of HCVcc entry by treatment with anti-ApoE antibody and the direct interaction of ApoE with NS5A. Ann L. Wozniak showed an important role for p7 in the production of infectious particles. Their data suggest that p7 stimulates virus production through the alkalization of intracellular vesicles. Ophelia Granio showed that both p7 and NS2 are required for the recruitment of core from lipid droplets (LDs) to ER. Costin-Ioan I. Popescu showed that NS2 accumulated in dotted structures in the ER in juxtaposition with Core and LDs. They concluded that cross-talk among Core, E1, E2, p7, and NS2 was essential for virion assembly. Vlastimil Jirasko demonstrated point mutations in the transmembrane regions of NS2 impaired the particle production and suggested that NS2 serves as a platform of viral and cellular proteins that coordinates HCV assembly. Qisheng Li identified the proviral function of IKKα by genome wide siRNA screening. IKKα regulates lipid metabolism and biogenesis of LDs and may enhance production of virus particles. The very low-density lipoproteins are secreted via a Golgi-dependent pathway. Bryan R. Bishe demonstrated the important role of phosphatidylinositol-4-phosphate and its interacting protein GOLPH3 in HCV secretion in the trans-Golgi network. Roland Remenyi showed 3-dimensional visualization of the HCV life cycle in cultured cells by electron tomography. They detected virus-like particles at various cytoplasmic locations. Viral particles in the proximity of LDs and within sponge-like inclusion were observed. These results provide ultrastructural visualization of putative assembly sites close to LDs. In the invited lecture, Sara Cherry presented an overview of high-throughput screening toward the identification of host factors required for viral infection. The contribution of autophagy to the HCV life cycle was also presented in this section, most notably, host factors linked with lipids. Tsubasa Munakata showed that the fatty acid synthase is required for efficient HCV replication. They also suggested the importance of palmitate for HCV replication. Samantha L. Blackham presented both the thioredoxin-interacting protein and the peroxisome proliferator activated receptor-α have significant effects on HCV replication. The host factors functioning on infectious HCV particle production were also reported. Takayuki Hishiki demonstrated the isoform dependent binding affinities of ApoE for low-density lipoprotein receptors and they affect infectivity of HCV. Laurent Chatel-Chaix found that Y-box binding protein interacted with HCV NS3 protein and viral RNA and was relocalized from nucleocytoplasmics site to the core-containing surface of LDs. Mohsan Saeed reported that the ER-associated degradation pathway was activated by HCV infection in a viral envelope protein-dependent manner. Po-Yuan Ke showed that HCV infection induces the unfolded protein response and activates the autophagic pathway. They proposed that autophagy contributes to the suppression of HCV in an autolysosome formation-dependent manner. Hiroto Kambara did not find any effects on HCV replication by inhibition of autophagosome formation in replicon cells. They proposed a role for autophagy induced by HCV infection to avoid the generation of vacuolation harmful to cell survival. Qisheng Li reported the network map of cellular pathways and machineries that are associated with HCV life cycle. Very low-density lipoprotein is now considered to be one of a component of HCV particles. LDs are composed of fatty acid, triglyceride, and cholesterol, surrounded by several types of lipoproteins. In addition, Daniel J. Felmlee reported that chylomicron-associated viruses may be generated by virion association while in the vascular compartment. Francois Jean showed that the serine protease inhibitor protein Spn4A was modified to be directed to Site-1 protease specifically and was introduced into adenovirus vector to inhibit cholesterol and fatty acid syntheses for down-regulation of HCV propagation. The modified serpin could suppress Site-1 protease activity, reduce the LD, and block HCVcc infection. Nicolas Menzel tried to identify novel cellular factors involved in HCV assembly and release and found ERK inhibitor and cytosolic phospholipase A2 (cPLA2) inhibitor reduce viral production. cPLA2 inhibitor also reduced the amount of LD-associated core and supernatant ApoB/E. cPLA2 may be crucial for assembly of infectious HCV particles, possibly through participating in the formation of lipoproteins. Kohji Moriishi reported that the proteasome activator PA28γ participates in HCV propagation. PA28γ may participate in the propagation of HCV by regulating the degradation of Core in both ubiquitin-dependent and -independent manners. NS5A is regulated by phosphorylation of several host protein kinases. Takahiro Masaki identified 79 serine threonine protein kinases that were tightly associated with NS5A. Two of these may regulate the production of viral particles and/or viral replication. The early phase of host defense against viral infection has largely been delineated based on recent advances in innate immunity. In the invited lecture, Manoj N. Krishnan introduced his comprehensive study on the Toll-like receptor 3–TRIF (TICAM-1) pathway. Using RNAi and polyI:C, he screened the genes specifically up-regulated via the TRIF (TICAM-1) pathway. He expected that some viral infections are selectively blocked by the IPS-1 pathway, while others are blocked by the TRIF pathway. Michael Gale, Jr., identified IFITM1 inhibits HCV infection. IFITM1 assembles with CD81 and tranlocates to the tight junction. This translocation of CD81 hampers the receptor function of CD81. They also discovered a novel pathway for ISGF3 activation. A non-receptor type tyrosine kinase-1 triggers activation of ISGF3 independent from the classical IFNAR pathway. IP-10 is a chemokine and is a negative predictor for pegylated interferon (IFN)/ribavirin therapy. Matthew L. Albert indicated that there is a 2-amino-acid–deleted form of IP-10 that serves as an antagonist for intact IP-10, and this form abrogates an early virologic response. As this IP-10 truncation is mediated by dipeptidylpeptidase IV, they believed that dipeptidylpeptidase IV is a novel therapeutic target for HCV patients during IFN therapy. Joo Chun Yoon suggested that activation of natural killer cells is inhibited by HCV-infected hepatocytes. They claimed that the early phases of HCV infection may be established through the failure of virus-inducible natural killer cell activation. Shin-ichiro Nakagawa reported that polyI:C induces both type I IFN and IFN-λ in human hepatocytes. The antiviral effect appears to parallel the induction of IFN-λ. This, together with the report by Emmanuel Thomas, suggests that the IFN-λ system is activated in HCV infected hepatocytes. In a keynote lecture, Robert Thimme summarized the mechanisms of HCV-induced T-cell dysfunction. Multifaceted factors contribute to the hyporesponsiveness of T cells, including viral mutations, primary T-cell failure, lack of support from dendritic cells, expression of inhibitory molecules on T cells, and abundance of regulatory T cells (Tregs). Whether the ability of HCV-specific CTLs is comparable with that of CTLs having other specificities remains controversial. Bianca Seigel showed that HCV-specific CTLs are functionally impaired when compared with other CTLs, irrespective of their expression of inhibitory receptors or differentiation stages. CD161 is a C-type lectin that is expressed in HCV-specific CD8+ T cells with tissue homing phenotype. Vicki M. Fleming found that CD4+CD161+ T cells produce large amounts of inflammatory cytokines and accumulate in the liver, where they are thought to exert pro-inflammatory roles. Naruyasu Kakita reported that certain adaptive Tregs, known as interleukin (IL)-10–producing type 1 Tregs, are increased in HCV-positive hepatocellular carcinoma patients, and their significance in hepatocellular carcinoma was greater than that of natural Tregs. Even in patients who have attained a sustained virologic response, trace amounts of HCV RNA are sporadically detectable in plasma. Barbara Rehermann reported the inoculation studies of such plasma. Residual HCV RNA in patients was able to infect chimpanzees and induced broad, HCV-specific T-cell responses. HCV RNA levels continued to be high when T-cell responses declined, suggesting that such HCV remains transmissible as hepatotropic pathogens. In the invited lecture, Michael Diamond presented new mechanisms for West Nile virus immune evasion via 2'O methylation of viral RNA to subvert host innate immunity. Genome-wide analysis of quantitative data (transcriptomics, proteomics, and metabolomics) facilitates systems biology analysis of HCV infection. Deborah L. Diamond analyzed the pathways involved in the progression of chronic hepatitis, namely, fibrosis and carcinogenesis, and found that molecules relating to cell metabolism including fatty acid oxidation enzymes and antioxidant systems may be master regulators of liver disease progression in HCV infection. HCV core protein has been shown to play a key role in the development of steatosis in HCV infected liver, especially in patients with genotype 3a HCV infection. Sophie Clement-Leboube showed that PTEN expression was down-regulated in the HCV infected liver. Analysis of lipo-viral-particle from hepatitis C patients by Olivier Diaz revealed that empty lipo-viral-particle lacking HCV RNA outnumbers those with RNA. The presence of virus-modified lipoproteins in HCV-infected patients may play a role in the pathogenesis of hepatitis C. Massimiliano Pagani used serum miRNA signatures to monitor liver disease in HCV infection and found miRNome candidates that are specific for HCV disease progression. Shuhei Taguwa showed that Con1 replicon induces incomplete autophagy through the dysfunction of autolysosomal acidification, which results in the secretion of immature cathepsin B in cells. Because the secretion of the protein is enhanced in many types of tumors, this observation may be associated with the pathogenesis of liver tumorigenesis in HCV infection. The existence of extrahepatic manifestations is another issue of interest. Essential mixed cryoglobulinemia, membranoproliferative glomerulonephritis, and Sjögren syndrome are conditions that have been shown to correlate with HCV infection. Nicola A. Fletcher reported that brain microvascular endothelial cells express all the recognized entry factors for HCV, and brain microvascular endothelial cells actually support infection by HCVpp and HCVcc. This suggests potential disorders of the central nervous system in HCV infection. In the keynote lecture, Masashi Mizokami presented "Genome-wide association study and its application for HCV treatment." He emphasized that the functional relevance of IL-28B single nucleotide polymorphisms should be elucidated to further advance the progress of research on the mechanisms of chronic HCV infection and treatment. Yasuhiro Asahina presented that genetic variation in IL-28B is associated with gene expression involving innate immunity. Minor alleles of IL-28B, as well as a higher RIG-I/IPS-1 ratio are associated with null viral response. Martin Laggins correlated IL-28B genetic variation with pretreatment levels of IP-10 and HCV RNA throughout therapy. The favorable genetic variation of IL-28B single nucleotide polymorphisms (major allele) was significantly associated with lower baseline IP-10. Masao Honda revealed that hepatic IFN-stimulated genes (ISGs) are associated with genetic variation in IL-28B and the outcome of IFN therapy for chronic hepatitis C using microarray gene expression profiling of the biopsied liver samples. Multivariate logistic regression analysis showed that ISGs, fibrosis stage, and ISDR mutations were strongly associated with viral response. Hepatic ISGs were associated with the IL-28B polymorphism and expression was significantly higher in patients with the minor genotype than in those with the major genotype. Takashi Motomura also analyzed ISG expression using liver transplantation samples. Expression of ISGs in recipients' liver carrying the minor allele of IL-28B was significantly up-regulated when compared with the major allele. Surprisingly, IFN sensitivity for recurrent hepatitis C after liver transplantation is influenced by IL-28B genetic variation not only in recipients, but also in donors. This session opened with a keynote lecture by Raffaele De Francesco describing the current state of drug development for patients with chronic hepatitis C. Because of the rapid development of NS3/4A, NS5A, and NS5B inhibitors, he finally presented the hopeful message "Will there be an HCV meeting in 2020?". Lotte Coelmont characterized an NS5A D320E variant showing low-level resistance to DEB025, a cyclophilin (Cyp)-binding molecule. This study suggests that DEB025 presents a high barrier to resistance, and that D320E confers low-level resistance to DEB025 by reducing the need for CypA-dependent isomerization of NS5A. Paul Targett-Adams reported that NS5A inhibitors stimulated redistribution of NS5A from the ER to ring-like structures in the cytoplasm, and disrupted colocalization with NS5B. This study suggests that NS5A inhibitors perturb formation of new replication complexes rather than acting on preformed complexes. Luis M. Schang developed a family of small synthetic rigid amphiphiles with large hydrophilic heads and small, planar and rigid hydrophobic tails, called RAFIs (rigid amphipathic fusion inhibitors), which inhibit the infectivity of enveloped virions including HCV. Emmanuel Thomas screened host genes involving the anti-HCV activity of ribavirin. Among 64 host genes, several candidate genes were identified as host factors involving ribavirin's anti-HCV activity. Interestingly, silencing of the ITPA gene increased the anti-HCV activity of ribavirin. Pablo Gastaminza identified a novel family of 1,2-diamines as an anti-HCV reagent from a chemical library. The analysis of ∼300 derivatives identified several compounds with enhanced potency and low cytotoxicity. HCV therapeutic vaccines are aimed to induce effective T-cell responses. Marianne Mikkelsen reported that vaccination of mice with recombinant adenovirus expressing HCV NS3 fused to the MHC class II chaperon protein invariant chain significantly enhanced NS3 specific CD8+ T-cell responses, and protected mice against NS3-expressing vaccinia virus challenge. This vaccination induced polyfunctional CD8+ memory T cells. Lars Frelin aimed to restore immunologic function through vaccination in a transgenic mouse model with impaired HCV-specific T-cell responses owing to a persistent presence of hepatic HCV NS3/4A antigens. They found that heterologous sequences improved activation and expansion of NS3/4A-specific T cells in a wild-type host, as well as in a tolerant NS3/4A-transgenic mouse model. The authors also suggested an important role for Tregs in the impaired HCV-specific T-cell responses. Livia M.G. Rossi examined antibody cross-immunoreactivity against different HVR1 variants to identify antigens with a possible application of HCV vaccine development. The authors identified a small set of HVR1 variants that cross-immunoreacted with a large number of HVR1 peptides, thus suggesting their potential use in the development of HCV vaccine candidates. HCV2010 in Yokohama was successful and contributed to the progress of research in the field. HCV infection remains one of the most serious worldwide health problems. The goals of this symposium were to increase the scientific understanding of this virus and gain insights applicable to future efforts to control its infection. From this point of view, we gained further fundamental understanding about HCV at the meeting. The discovery of IL-28B as a new host factor involved in HCV treatment and pathogenesis had a major impact on HCV research. New treatment advances have been made in recent years and will continue in the near future. We would like to conclude that this meeting was successful in providing opportunities for exchanging up-to-date information and international collaboration. The next meeting will take place in Seattle, Washington, from September 8–12, 2011 (http://www.hcv2011.org/).