Hepatitis C virus infection and related liver disease: the quest for the best animal model

Hepatitis C virus (HCV) is a major cause of cirrhosis and hepatocellular carcinoma (HCC) making the virus the most common cause of liver failure and transplantation. HCV is estimated to chronically affect 130 million individuals and to lead to more than 350,000 deaths per year worldwide. A vaccine is currently not available. The recently developed direct acting antivirals (DAAs) have markedly increased the efficacy of the standard of care but are not efficient enough to completely cure all chronically infected patients and their toxicity limits their use in patients with advanced liver disease, co-morbidity or transplant recipients. Because of the host restriction, which is limited to humans and non-human primates, in vivo study of HCV infection has been hampered since its discovery more than 20 years ago. The chimpanzee remains the most physiological model to study the innate and adaptive immune responses, but its use is ethically difficult and is now very restricted and regulated. The development of a small animal model that allows robust HCV infection has been achieved using chimeric liver immunodeficient mice, which are therefore not suitable for studying the adaptive immune responses. Nevertheless, these models allowed to go deeply in the comprehension of virus-host interactions and to assess different therapeutic approaches. The immunocompetent mouse models that were recently established by genetic humanization have shown an interesting improvement concerning the study of the immune responses but are still limited by the absence of the complete robust life cycle of the virus. In this review, we will focus on the relevant available animal models of HCV infection and their usefulness for deciphering the HCV life cycle and virus-induced liver disease, as well as for the development and evaluation of new therapeutics. We will also discuss the perspectives on future immunocompetent mouse models and the hurdles to their development

Animal models to study hepatitis C virus infection

With more than 71 million chronically infected people, the hepatitis C virus (HCV) is a major global health concern. Although new direct acting antivirals have significantly improved the rate of HCV cure, high therapy cost, potential emergence of drug-resistant viral variants, and unavailability of a protective vaccine represent challenges for complete HCV eradication. Relevant animal models are required, and additional development remains necessary, to effectively study HCV biology, virus-host interactions and for the evaluation of new antiviral approaches and prophylactic vaccines. The chimpanzee, the only non-human primate susceptible to experimental HCV infection, has been used extensively to study HCV infection, particularly to analyze the innate and adaptive immune response upon infection. However, financial, practical, and especially ethical constraints have urged the exploration of alternative small animal models. These include different types of transgenic mice, immunodeficient mice of which the liver is engrafted with human hepatocytes (humanized mice) and, more recently, immunocompetent rodents that are susceptible to infection with viruses that are closely related to HCV. In this review, we provide an overview of the currently available animal models that have proven valuable for the study of HCV, and discuss their main benefits and weaknesses

Animal models for hepatitis E virus

Hepatitis E virus (HEV) is an underdiagnosed pathogen with approximately 20 million infections each year and currently the most common cause of acute viral hepatitis. HEV was long considered to be confined to developing countries but there is increasing evidence that it is also a medical problem in the Western world. HEV that infects humans belongs to the Orthohepevirus A species of the Hepeviridae family. Novel HEV-like viruses have been observed in a variety of animals and some have been shown to be able to cross the species barrier, causing infection in humans. Several cell culture models for HEV have been established in the past years, but their efficiency is usually relatively low. With the circulation of this virus and related viruses in a variety of species, several different animal models have been developed. In this review, we give an overview of these animal models, indicate their main characteristics, and highlight how they may contribute to our understanding of the basic aspects of the viral life cycle and cross-species infection, the study of pathogenesis, and the evaluation of novel preventative and therapeutic strategies

Impact of lipids and lipoproteins on hepatitis C virus infection and virus neutralization

Hepatitis C virus (HCV) infections represent a major global health problem. End-stage liver disease caused by chronic HCV infection is a major indication for liver transplantation. However, after transplantation the engrafted liver inevitably becomes infected by the circulating virus. Direct acting antivirals are not yet approved for use in liver transplant patients, and limited efficacy and severe side effects hamper the use of pegylated interferon combined with ribavirin in a post-transplant setting. Therefore, alternative therapeutic options need to be explored. Viral entry represents an attractive target for such therapeutic intervention. Understanding the mechanisms of viral entry is essential to define the viral and cellular factors involved. The HCV life cycle is dependent of and associated with lipoprotein physiology and the presence of lipoproteins has been correlated with altered antiviral efficacy of entry inhibitors. In this review, we summarise the current knowledge on how lipoprotein physiology influences the HCV life cycle. We focus especially on the influence of lipoproteins on antibodies that target HCV envelope proteins or antibodies that target the cellular receptors of the virus. This information can be particularly relevant for the prevention of HCV re-infection after liver transplantation

Blocking entry of hepatitis B and D viruses to hepatocytes as a novel immunotherapy for treating chronic infections

Background. Chronic hepatitis B and D virus (HBV/HDV) infections can cause cancer. Current HBV therapy using nucleoside analogues (NAs) is life-long and reduces but does not eliminate the risk of cancer. A hallmark of chronic hepatitis B is a dysfunctional HBV-specific T-cell response. We therefore designed an immunotherapy driven by naive healthy T cells specific for the HDV antigen (HDAg) to bypass the need for HBV-specific T cells in order to prime PreS1-specific T cells and PreS1 antibodies blocking HBV entry. Methods. Ten combinations of PreS1 and/or HDAg sequences were evaluated for induction of PreS1 antibodies and HBV- and HDV-specific T cells in vitro and in vivo. Neutralization of HBV by PreS1-specific murine and rabbit antibodies was evaluated in cell culture, and rabbit anti-PreS1 were tested for neutralization of HBV in mice repopulated with human hepatocytes. Results. The best vaccine candidate induced T cells to PreS1 and HDAg, and PreS1 antibodies blocking HBV entry in vitro. Importantly, adoptive transfer of PreS1 antibodies prevented, or modulated, HBV infection after a subsequent challenge in humanized mice. Conclusions. We here describe a novel immunotherapy for chronic HBV/HDV that targets viral entry to complement NAs and coming therapies inhibiting viral maturation

Challenge pools of hepatitis C virus genotypes 1-6 prototype strains: replication fitness and pathogenicity in chimpanzees and human liver-chimeric mouse models

Chimpanzees represent the only animal model for studies of the natural history of hepatitis C virus (HCV). To generate virus stocks of important HCV variants, we infected chimpanzees with HCV strains of genotypes 1-6 and determined the infectivity titer of acute-phase plasma pools in additional animals. The courses of first- and second-passage infections were similar, with early appearance of viremia, HCV RNA titers of >10(4.7) IU/mL, and development of acute hepatitis; the chronicity rate was 56%. The challenge pools had titers of 10(3)-10(5) chimpanzee infectious doses/mL. Human liver-chimeric mice developed high-titer infections after inoculation with the challenge viruses of genotypes 1-6. Inoculation studies with different doses of the genotype 1b pool suggested that a relatively high virus dose is required to consistently infect chimeric mice. The challenge pools represent a unique resource for studies of HCV molecular virology and for studies of pathogenesis, protective immunity, and vaccine efficacy in vivo

Development and characterization of a human monoclonal antibody for prevention of HCV recurrence in liver transplant patients

More than 170 million people worldwide are chronically infected with hepatitis C virus (HCV) and are at risk of developing liver fibrosis, cirrhosis and hepatocellular carcinoma. Liver transplantation is the only option for patients with HCV-induced end-stage liver diseases. Nevertheless, infection of the newly grafted liver occurs immediately and universally after transplantation. Despite the recent progress in HCV therapy, a prophylactic vaccine is still not available. The role of neutralizing monoclonal antibodies (mAbs) in protection from different viral infections including HCV, HIV and Ebola has been reported. In the last few years, several mAbs with neutralizing activity have been described but only few mAbs have been evaluated in vivo. In the present study, we describe the development of a mAb, designated 2A5, isolated from HCV genotype 1b chronic patient. ELISA results indicated high affinity of mAb 2A5 towards HCV envelope glycoprotein (E1E2). The binding activity was completely lost against denatured E1E2 protein indicating that it targets a conformational epitope within the envelope region. Epitope mapping using alanine mutants of E1E2 proteins defined critical binding residues within the regions 419-447 and 612-617. Results of pseudoparticles (HCVpp) and cell culture produced virus (HCVcc) neutralization showed broad neutralizing activity of mAb 2A5 against all HCV genotypes. The efficacy study of mAb 2A5 in immune-deficient mice of which the liver is repopulated with human hepatocytes (humanized mice) showed complete protection from HCV challenge for genotypes 1a and 4a, while partial protection was achieved for genotypes 1b and 6a. Sequence analysis of E1E2 protein from non-protected mice did not revealed resistance mutations at interaction residues of mAb 2A5. In conclusion, mAb 2A5 shows potent anti-HCV neutralizing activity both in vitro and in vivo and could hence provide an effective strategy to prevent HCV recurrence in chronically infected HCV liver transplant patients. In addition, the broad neutralizing activity of this mAb presents a valuable epitope for the design of HCV vaccine with cross-protection activity

Artemisinin analogues as potent inhibitors of in vitro hepatitis C virus replication

We reported previously that Artemisinin (ART), a widely used anti-malarial drug, is an inhibitor of in vitro HCV subgenomic replicon replication. We here demonstrate that ART exerts its antiviral activity also in hepatoma cells infected with full length infectious HCV JFH-1. We identified a number of ART analogues that are up to 10-fold more potent and selective as in vitro inhibitors of HCV replication than ART. The iron donor Hemin only marginally potentiates the anti-HCV activity of ART in HCV-infected cultures. Carbon-centered radicals have been shown to be critical for the anti-malarial activity of ART. We demonstrate that carbon-centered radicals-trapping (the so-called TEMPO) compounds only marginally affect the anti-HCV activity of ART. This provides evidence that carbon-centered radicals are not the main effectors of the anti-HCV activity of the Artemisinin. ART and analogues may possibly exert their anti-HCV activity by the induction of reactive oxygen species (ROS). The combined anti-HCV activity of ART or its analogues with L-N-Acetylcysteine (L-NAC) [a molecule that inhibits ROS generation] was studied. L-NAC significantly reduced the in vitro anti-HCV activity of ART and derivatives. Taken together, the in vitro anti-HCV activity of ART and analogues can, at least in part, be explained by the induction of ROS; carbon-centered radicals may not be important in the anti-HCV effect of these molecules