Apolipoprotein E mediates evasion from hepatitis C virus−neutralizing antibodies

Background & Aims Efforts to develop an effective vaccine against hepatitis C virus (HCV) have been hindered by the propensity of the virus to evade host immune responses. HCV particles in serum and in cell culture associate with lipoproteins, which contribute to viral entry. Lipoprotein association has also been proposed to mediate viral evasion of the humoral immune response, though the mechanisms are poorly defined. Methods We used small interfering RNAs to reduce levels of apolipoprotein E (apoE) in cell culture−derived HCV−producing Huh7.5-derived hepatoma cells and confirmed its depletion by immunoblot analyses of purified viral particles. Before infection of naïve hepatoma cells, we exposed cell culture−derived HCV strains of different genotypes, subtypes, and variants to serum and polyclonal and monoclonal antibodies isolated from patients with chronic HCV infection. We analyzed the interaction of apoE with viral envelope glycoprotein E2 and HCV virions by immunoprecipitation. Results Through loss-of-function studies on patient-derived HCV variants of several genotypes and subtypes, we found that the HCV particle apoE allows the virus to avoid neutralization by patient-derived antibodies. Functional studies with human monoclonal antiviral antibodies showed that conformational epitopes of envelope glycoprotein E2 domains B and C were exposed after depletion of apoE. The level and conformation of virion-associated apoE affected the ability of the virus to escape neutralization by antibodies. Conclusions In cell-infection studies, we found that HCV-associated apoE helps the virus avoid neutralization by antibodies against HCV isolated from chronically infected patients. This method of immune evasion poses a challenge for the development of HCV vaccines

The functional role of sodium taurocholate cotransporting polypeptide NTCP in the life cycle of hepatitis B, C and D viruses

Chronic hepatitis B, C and D virus (HBV, HCV and HDV) infections are a major cause of liver disease and cancer worldwide. Despite employing distinct replication strategies, the three viruses are exclusively hepatotropic, and therefore depend on hepatocyte-specific host factors. The sodium taurocholate co-transporting polypeptide (NTCP), a transmembrane protein highly expressed in human hepatocytes that mediates the transport of bile acids, plays a key role in HBV and HDV entry into hepatocytes. Recently, NTCP has been shown to modulate HCV infection of hepatocytes by regulating innate antiviral immune responses in the liver. Here, we review the current knowledge of the functional role and the molecular and cellular biology of NTCP in the life cycle of the three major hepatotropic viruses, highlight the impact of NTCP as an antiviral target and discuss future avenues of research

Neutralizing antibodies and pathogenesis of hepatitis C virus infection.

Hepatitis C virus (HCV) infection is a major cause of chronic liver disease worldwide. The interplay between the virus and host innate and adaptive immune responses determines the outcome of infection. There is increasing evidence that host neutralizing responses play a relevant role in the resulting pathogenesis. Furthermore, viral evasion from host neutralizing antibodies has been revealed to be an important contributor in leading both to viral persistence in acute liver graft infection following liver transplantation, and to chronic viral infection. The development of novel model systems to study HCV entry and neutralization has allowed a detailed understanding of the molecular mechanisms of virus-host interactions during antibody-mediated neutralization. The understanding of these mechanisms will ultimately contribute to the development of novel antiviral preventive strategies for liver graft infection and an urgently needed vaccine. This review summarizes recent concepts of the role of neutralizing antibodies in viral clearance and protection, and highlights consequences of viral escape from neutralizing antibodies in the pathogenesis of HCV infection

Hepatitis C virus cell-cell transmission and resistance to direct-acting antiviral agents

Hepatitis C virus (HCV) is transmitted between hepatocytes via classical cell entry but also uses direct cell-cell transfer to infect neighboring hepatocytes. Viral cell-cell transmission has been shown to play an important role in viral persistence allowing evasion from neutralizing antibodies. In contrast, the role of HCV cell-cell transmission for antiviral resistance is unknown. Aiming to address this question we investigated the phenotype of HCV strains exhibiting resistance to direct-acting antivirals (DAAs) in state-of-the-art model systems for cell-cell transmission and spread. Using HCV genotype 2 as a model virus, we show that cell-cell transmission is the main route of viral spread of DAA-resistant HCV. Cell-cell transmission of DAA-resistant viruses results in viral persistence and thus hampers viral eradication. We also show that blocking cell-cell transmission using host-targeting entry inhibitors (HTEIs) was highly effective in inhibiting viral dissemination of resistant genotype 2 viruses. Combining HTEIs with DAAs prevented antiviral resistance and led to rapid elimination of the virus in cell culture model. In conclusion, our work provides evidence that cell-cell transmission plays an important role in dissemination and maintenance of resistant variants in cell culture models. Blocking virus cell-cell transmission prevents emergence of drug resistance in persistent viral infection including resistance to HCV DAAs

In vivo combination of human anti-envelope glycoprotein E2 and -Claudin-1 monoclonal antibodies for prevention of hepatitis C virus infection

Despite the development of direct-acting antivirals (DAAs), hepatitis C virus (HCV) infection remains a major cause for liver disease and cancer worldwide. Entry inhibitors block virus host cell entry and, therefore, prevent establishment of chronic infection and liver disease. Due to their unique mechanism of action, entry inhibitors provide an attractive antiviral strategy in organ transplantation. In this study, we developed an innovative approach in preventing HCV infection using a synergistic combination of a broadly neutralizing human monoclonal antibody (HMAb) targeting the HCV E2 protein and a host-targeting anti-claudin 1 (CLDN1) humanized monoclonal antibody. An in vivo proof-of-concept study in human liver-chimeric FRG-NOD mice proved the efficacy of the combination therapy at preventing infection by an HCV genotype 1b infectious serum. While administration of individual antibodies at lower doses only showed a delay in HCV infection, the combination therapy was highly protective. Furthermore, the combination proved to be effective in preventing infection of primary human hepatocytes by neutralization-resistant HCV escape variants selected during liver transplantation, suggesting that a combination therapy is suited for the neutralization of difficult-to-treat variants. In conclusion, our findings suggest that the combination of two HMAbs targeting different steps of virus entry improves treatment efficacy while simultaneously reducing treatment duration and costs. Our approach not only provides a clinical perspective to employ HMAb combination therapies to prevent graft re-infection and its associated liver disease but may also help to alleviate the urgent demand for organ transplants by allowing the transplantation of organs from HCV-positive donors

Approches de génomique fonctionnelle pour la caractérisation de facteurs hépatiques impliqués dans l’infection par le virus de l’hépatite B

Chronic infection by hepatitis B virus (HBV) is a major public health issue and remains the principal cause of progressive chronic liver disease and hepatocellular carcinoma. While the genome and structure of the virus have been studied in great details, the lack of efficient cell culture models has impaired the understanding of molecular interactions between virus and its host cells. However, a better understanding of virus-host interactions is crucial for the development of new efficient antiviral strategy. Recent advances in the characterization of viral entry paved the way to the development of novel infection models available for innovative high-throughput functional genomic studies a unique opportunity to discover new therapeutic targets. The development of these models which were not available in the laboratory and their validation in virus-host interactions studies allowed me the investigation of poorly known viral mechanism as viral detection and escape of HBV from the innate immune response. These models were subsequently used in a functional high-throughput screen that identified CDKN2C as a cellular host factor involved in HBV viral replication. Thus, these works highlight new mechanisms of virus host interactions. They are the starting points of novel research programs contributing to the development of more physiological infection system and characterization of new therapeutic target to eradicate HBV.L’infection chronique par le virus de l’hépatite B (HBV) est un problème majeur de santé publique et est la principale cause de développement de maladies hépatiques chroniques progressives et de carcinome hépatocellulaire. Alors que le génome et la structure du virus ont été étudiés en détail, le manque de modèles d’étude in vitro efficaces a été un frein pour la découverte et la compréhension des interactions moléculaires entre le virus et son hôte. Or, une meilleure compréhension des interactions virus-hôtes est primordiale pour ledéveloppement de nouveaux antiviraux efficaces. Les récents progrès sur la compréhension de l’entrée virale ouvrent la voie au développement de nouveaux modèles d’infection adaptés aux études de génomiquesfonctionnelles à haut débit, opportunité unique pour découvrir de nouvelles cibles thérapeutiques à grande échelle.La mise au point de modèles d’infection non disponibles au laboratoire au début de ma thèse et leur validation dans des études sur les interactions virus-hôte, a permis l’étude d’un mécanisme viral encore mal compris qu’est la détection et l’échappement du HBV à la réponse immunitaire innée. Ces modèles ont par la suite permis demettre au point et de réaliser un criblage fonctionnel à haut débit qui a identifié CDKN2C comme facteur d’hôte impliqué la réplication virale du HBV. Ainsi, les travaux réalisés dans le cadre de cette thèse ont permis la mise en évidence et la compréhension de différents mécanismes d’interactions du virus avec la cellule hôte lors del’infection. Ils ouvrent la voie au développement de nouveaux systèmes d’infection plus physiologiques et la caractérisation de nouvelles cibles thérapeutiques permettant d’éradiquer le VHB

Approches de génomique fonctionnelle pour la caractérisation de facteurs hépatiques impliqués dans l’infection par le virus de l’hépatite B

Chronic infection by hepatitis B virus (HBV) is a major public health issue and remains the principal cause of progressive chronic liver disease and hepatocellular carcinoma. While the genome and structure of the virus have been studied in great details, the lack of efficient cell culture models has impaired the understanding of molecular interactions between virus and its host cells. However, a better understanding of virus-host interactions is crucial for the development of new efficient antiviral strategy. Recent advances in the characterization of viral entry paved the way to the development of novel infection models available for innovative high-throughput functional genomic studies a unique opportunity to discover new therapeutic targets. The development of these models which were not available in the laboratory and their validation in virus-host interactions studies allowed me the investigation of poorly known viral mechanism as viral detection and escape of HBV from the innate immune response. These models were subsequently used in a functional high-throughput screen that identified CDKN2C as a cellular host factor involved in HBV viral replication. Thus, these works highlight new mechanisms of virus host interactions. They are the starting points of novel research programs contributing to the development of more physiological infection system and characterization of new therapeutic target to eradicate HBV.L’infection chronique par le virus de l’hépatite B (HBV) est un problème majeur de santé publique et est la principale cause de développement de maladies hépatiques chroniques progressives et de carcinome hépatocellulaire. Alors que le génome et la structure du virus ont été étudiés en détail, le manque de modèles d’étude in vitro efficaces a été un frein pour la découverte et la compréhension des interactions moléculaires entre le virus et son hôte. Or, une meilleure compréhension des interactions virus-hôtes est primordiale pour ledéveloppement de nouveaux antiviraux efficaces. Les récents progrès sur la compréhension de l’entrée virale ouvrent la voie au développement de nouveaux modèles d’infection adaptés aux études de génomiquesfonctionnelles à haut débit, opportunité unique pour découvrir de nouvelles cibles thérapeutiques à grande échelle.La mise au point de modèles d’infection non disponibles au laboratoire au début de ma thèse et leur validation dans des études sur les interactions virus-hôte, a permis l’étude d’un mécanisme viral encore mal compris qu’est la détection et l’échappement du HBV à la réponse immunitaire innée. Ces modèles ont par la suite permis demettre au point et de réaliser un criblage fonctionnel à haut débit qui a identifié CDKN2C comme facteur d’hôte impliqué la réplication virale du HBV. Ainsi, les travaux réalisés dans le cadre de cette thèse ont permis la mise en évidence et la compréhension de différents mécanismes d’interactions du virus avec la cellule hôte lors del’infection. Ils ouvrent la voie au développement de nouveaux systèmes d’infection plus physiologiques et la caractérisation de nouvelles cibles thérapeutiques permettant d’éradiquer le VHB

Characterization of hepatic factors involved in hepatitis B virus infection using functional genomic approache

L’infection chronique par le virus de l’hépatite B (HBV) est un problème majeur de santé publique et est la principale cause de développement de maladies hépatiques chroniques progressives et de carcinome hépatocellulaire. Alors que le génome et la structure du virus ont été étudiés en détail, le manque de modèles d’étude in vitro efficaces a été un frein pour la découverte et la compréhension des interactions moléculaires entre le virus et son hôte. Or, une meilleure compréhension des interactions virus-hôtes est primordiale pour ledéveloppement de nouveaux antiviraux efficaces. Les récents progrès sur la compréhension de l’entrée virale ouvrent la voie au développement de nouveaux modèles d’infection adaptés aux études de génomiquesfonctionnelles à haut débit, opportunité unique pour découvrir de nouvelles cibles thérapeutiques à grande échelle.La mise au point de modèles d’infection non disponibles au laboratoire au début de ma thèse et leur validation dans des études sur les interactions virus-hôte, a permis l’étude d’un mécanisme viral encore mal compris qu’est la détection et l’échappement du HBV à la réponse immunitaire innée. Ces modèles ont par la suite permis demettre au point et de réaliser un criblage fonctionnel à haut débit qui a identifié CDKN2C comme facteur d’hôte impliqué la réplication virale du HBV. Ainsi, les travaux réalisés dans le cadre de cette thèse ont permis la mise en évidence et la compréhension de différents mécanismes d’interactions du virus avec la cellule hôte lors del’infection. Ils ouvrent la voie au développement de nouveaux systèmes d’infection plus physiologiques et la caractérisation de nouvelles cibles thérapeutiques permettant d’éradiquer le VHB.Chronic infection by hepatitis B virus (HBV) is a major public health issue and remains the principal cause of progressive chronic liver disease and hepatocellular carcinoma. While the genome and structure of the virus have been studied in great details, the lack of efficient cell culture models has impaired the understanding of molecular interactions between virus and its host cells. However, a better understanding of virus-host interactions is crucial for the development of new efficient antiviral strategy. Recent advances in the characterization of viral entry paved the way to the development of novel infection models available for innovative high-throughput functional genomic studies a unique opportunity to discover new therapeutic targets. The development of these models which were not available in the laboratory and their validation in virus-host interactions studies allowed me the investigation of poorly known viral mechanism as viral detection and escape of HBV from the innate immune response. These models were subsequently used in a functional high-throughput screen that identified CDKN2C as a cellular host factor involved in HBV viral replication. Thus, these works highlight new mechanisms of virus host interactions. They are the starting points of novel research programs contributing to the development of more physiological infection system and characterization of new therapeutic target to eradicate HBV