Human and Host Species Transferrin Receptor 1 Use by North American Arenaviruses

ABSTRACT At least five New World (NW) arenaviruses cause hemorrhagic fevers in South America. These pathogenic clade B viruses, as well as nonpathogenic arenaviruses of the same clade, use transferrin receptor 1 (TfR1) of their host species to enter cells. Pathogenic viruses are distinguished from closely related nonpathogenic ones by their additional ability to utilize human TfR1 (hTfR1). Here, we investigate the receptor usage of North American arenaviruses, whose entry proteins share greatest similarity with those of the clade B viruses. We show that all six North American arenaviruses investigated utilize host species TfR1 orthologs and present evidence consistent with arenavirus-mediated selection pressure on the TfR1 of the North American arenavirus host species. Notably, one of these viruses, AV96010151, closely related to the prototype Whitewater Arroyo virus (WWAV), entered cells using hTfR1, consistent with a role for a WWAV-like virus in three fatal human infections whose causative agent has not been identified. In addition, modest changes were sufficient to convert hTfR1 into a functional receptor for most of these viruses, suggesting that a minor alteration in virus entry protein may allow these viruses to use hTfR1. Our data establish TfR1 as a cellular receptor for North American arenaviruses, highlight an "arms race" between these viruses and their host species, support the association of North American arenavirus with fatal human infections, and suggest that these viruses have a higher potential to emerge and cause human diseases than has previously been appreciated. IMPORTANCE hTfR1 use is a key determinant for a NW arenavirus to cause hemorrhagic fevers in humans. All known pathogenic NW arenaviruses are transmitted in South America by their host rodents. North American arenaviruses are generally considered nonpathogenic, but some of these viruses have been tentatively implicated in human fatalities. We show that these North American arenaviruses use the TfR1 orthologs of their rodent host species and identify TfR1 polymorphisms suggesting an ongoing "arms race" between these viruses and their hosts. We also show that a close relative of a North American arenavirus suggested to have caused human fatalities, the Whitewater Arroyo species complex virus AV96010151, uses human TfR1. Moreover, we present data that imply that modest changes in other North American arenaviruses might allow these viruses to infect humans. Collectively, our data suggest that North American arenaviruses have a higher potential to cause human disease than previously assumed

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

Plasmodium falciparum Plasmepsin 2 Duplications, West Africa

Dihydroartemisinin/piperaquine (DHA/PPQ) is increasingly deployed as antimalaria drug in Africa. We report the detection in Mali of Plasmodium falciparum infections carrying plasmepsin 2 duplications (associated with piperaquine resistance) in 7/65 recurrent infections within 2 months after DHA/PPQ treatment. These findings raise concerns about the long-term efficacy of DHA/PPQ treatment in Africa.This work was supported by a Swedish Research Council Grant (no. VR-2014-3134). The WANECAM study is funded by the European and Developing Countries Clinical Trial Partnership and by the Medicines for Malaria Venture (Geneva, Switzerland) and is co-funded by the United Kingdom Medical Research Councils, the Swedish International Development Cooperation Agency, the German Ministry for Education and Research, the University Claude Bernard (Lyon, France), the University of Science, Techniques, and Technologies of Bamako (Bamako, Mali), the Centre National de Recherche et de Formation sur le Paludisme (Burkina Faso), the Institut de Recherche en Sciences de la Sante (Bobo-Dioulasso, Burkina Faso), and the Centre National de Formation et de Recherche en Sante Rurale (Guinea).J.I. was supported by EuroInkaNet/Erasmus Mundus Program. Fundacao para a Ciencia e Tecnologia supports M.S. (grant no. SFRH/BD/129769/2017), M.I.V. (grant no. SFRH/BPD/76614/2011), and P.E.F. (grant no. IF/00143/2015)

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

Stimulatory MAIT cell antigens reach the circulation and are efficiently metabolised and presented by human liver cells.

OBJECTIVE Mucosal-associated invariant T (MAIT) cells are the most abundant T cells in human liver. They respond to bacterial metabolites presented by major histocompatibility complex-like molecule MR1. MAIT cells exert regulatory and antimicrobial functions and are implicated in liver fibrogenesis. It is not well understood which liver cells function as antigen (Ag)-presenting cells for MAIT cells, and under which conditions stimulatory Ags reach the circulation. DESIGN We used different types of primary human liver cells in Ag-presentation assays to blood-derived and liver-derived MAIT cells. We assessed MAIT cell stimulatory potential of serum from healthy subjects and patients with portal hypertension undergoing transjugular intrahepatic portosystemic shunt stent, and patients with inflammatory bowel disease (IBD). RESULTS MAIT cells were dispersed throughout healthy human liver and all tested liver cell types stimulated MAIT cells, hepatocytes being most efficient. MAIT cell activation by liver cells occurred in response to bacterial lysate and pure Ag, and was prevented by non-activating MR1 ligands. Serum derived from peripheral and portal blood, and from patients with IBD stimulated MAIT cells in MR1-dependent manner. CONCLUSION Our findings reveal previously unrecognised roles of liver cells in Ag metabolism and activation of MAIT cells, repression of which creates an opportunity to design antifibrotic therapies. The presence of MAIT cell stimulatory Ags in serum rationalises the observed activated MAIT cell phenotype in liver. Increased serum levels of gut-derived MAIT cell stimulatory ligands in patients with impaired intestinal barrier function indicate that intrahepatic Ag-presentation may represent an important step in the development of liver disease

Synergy of entry inhibitors with direct-acting antivirals uncovers novel combinations for prevention and treatment of hepatitis C

Objective: Although direct-acting antiviral agents (DAAs) have markedly improved the outcome of treatment in chronic HCV infection, there continues to be an unmet medical need for improved therapies in difficult-to-treat patients as well as liver graft infection. Viral entry is a promising target for antiviral therapy. Design: Aiming to explore the role of entry inhibitors for future clinical development, we investigated the antiviral efficacy and toxicity of entry inhibitors in combination with DAAs or other host-targeting agents (HTAs). Screening a large series of combinations of entry inhibitors with DAAs or other HTAs, we uncovered novel combinations of antivirals for prevention and treatment of HCV infection. Results: Combinations of DAAs or HTAs and entry inhibitors including CD81-, scavenger receptor class B type I (SR-BI)- or claudin-1 (CLDN1)-specific antibodies or small-molecule inhibitors erlotinib and dasatinib were characterised by a marked and synergistic inhibition of HCV infection over a broad range of concentrations with undetectable toxicity in experimental designs for prevention and treatment both in cell culture models and in human liver-chimeric uPA/SCID mice. Conclusions: Our results provide a rationale for the development of antiviral strategies combining entry inhibitors with DAAs or HTAs by taking advantage of synergy. The uncovered combinations provide perspectives for efficient strategies to prevent liver graft infection and novel interferon-free regimens

Organoid Models of Human Liver Cancers Derived from Tumor Needle Biopsies

Hepatocellular carcinoma (HCC) is the most common primary liver cancer and the second most frequent cause of cancer-related mortality worldwide. The multikinase inhibitor sorafenib is the only treatment option for advanced HCC. Due to tumor heterogeneity, its efficacy greatly varies between patients and is limited due to adverse effects and drug resistance. Current in vitro models fail to recapitulate key features of HCCs. We report the generation of long-term organoid cultures from tumor needle biopsies of HCC patients with various etiologies and tumor stages. HCC organoids retain the morphology as well as the expression pattern of HCC tumor markers and preserve the genetic heterogeneity of the originating tumors. In a proof-of-principle study, we show that liver cancer organoids can be used to test sensitivity to sorafenib. In conclusion, organoid models can be derived from needle biopsies of liver cancers and provide a tool for developing tailored therapies

Prevention of hepatitis C virus reinfection during liver transplantation using monoclonal anti-receptor antibodies

Le virus de l'hépatite C (VHC) représente un problème de santé publique majeur. Environ 170 millions de personnes sont infectées dans le monde, et plus de 70% développeront une infection chronique. Le traitement actuel qui associe l'interféron- pégylé etHepatitis C virus (HCV) is a major problem of public health. Approximatively 170 million individuals are infected worldwide, and more than 70% of infected individuals will develop chronic hepatitis. The current treatment based on pegylated interferon- an

Prévention de la réinfection par le virus de l'hépatite C lors de la transplantation hépatique par l'utilisation d'anticorps monoclonaux anti-récepteur

Le virus de l hépatite C (VHC) représente un problème de santé publique majeur. Environ 170 millions de personnes sont infectées dans le monde, et plus de 70% développeront une infection chronique. Le traitement actuel qui associe l interféron- pégylé et la ribavirine ne permet de guérir que 50% des patients tous génotypes confondus. Les complications de l hépatite C chronique, la cirrhose et le carcinome hépatocellulaire, représentent des indications majeures de transplantation hépatique (TH). Malheureusement la récurrence de l infection par le VHC sur le greffon est systématique et évolue vers une cirrhose en moins de 5 ans chez une proportion importante de patients. A l heure actuelle, il n y a pas de moyen de prévention de la réinfection du greffon par le VHC. Une meilleure connaissance des phases précoces de la réinfection permettrait le développement de stratégies préventives de l infection par le VHC. Durant ma thèse nous nous sommes intéressés aux mécanismes de la réinfection par le VHC au cours de la TH en nous focalisant sur l étape de l entrée virale dans l hépatocyte. Nous avons montré pour la première fois qu au cours de la TH, et contrairement aux variants non sélectionnés après la greffe, les variants du VHC sélectionnés après la greffe (qui réinfectent le greffon) entrent plus efficacement dans la cellule hôte et échappent à la neutralisation induite par les anticorps des sérums autologues collectés avant la greffe. Nous avons, par la suite, identifié les mutations impliquées dans l entrée et l échappement viral aux anticorps neutralisants. Ces mutations favorisent l entrée virale en modulant la dépendance du VHC à un des facteurs d entrée virale, le CD81. La protéine de jonction Claudine 1 (CLDN1) est un autre facteur d entrée essentiel à l infection par le VHC. Nous avons produit des anticorps monoclonaux anti-CLDN1 et montré que ces anticorps inhibent de manière spécifique tous les génotypes du VHC ainsi que les variants viraux résistants à la neutralisation par le sérum autologue lors de la TH. L ensemble de ces résultats suggère que l étape d entrée du virus pourrait constituer une cible prometteuse pour le développement de stratégies préventives de la réinfection du greffon par le VHC.Hepatitis C virus (HCV) is a major problem of public health. Approximatively 170 million individuals are infected worldwide, and more than 70% of infected individuals will develop chronic hepatitis. The current treatment based on pegylated interferon- and ribavirin can only cure 50 % of patients. HCV-related cirrhosis and hepatocellular carcinoma are a leading indication for liver transplantation (LT). The major limitation of LT is the universal re-infection of the liver graft. To date, there is no prophylactic strategy to prevent re-infection. A better understanding of the early phases of re-infection would allow developing of preventive strategies. During my PhD thesis, we focussed on the viral entry step to study the molecular mechanisms of re-infection of the liver graft. For the first time, we have demonstrated that during LT, selected variants re-infecting the liver graft show enhanced entry and escape from antibody-mediated neutralization. We identified two adaptive mutations in envelope glycoprotein E2 mediating enhanced entry and evasion of an highly infectious escape variant. These mutations markedly modulated CD81 receptor dependency resulting in enhanced viral entry. Among the host entry factors involved in viral entry, claudin 1 (CLDN1) is essential for HCV infection. We have produced monoclonal anti-CLDN1 antibodies and have shown that these antibodies specifically inhibit HCV entry from all major genotypes, as well as cell entry of the highly infectious escape variants of HCV that were resistant to autologous neutralizing antibodies. Taken together, these results suggest that targeting HCV entry into hepatocytes with specific monoclonal antibodies, as anti-CLDN1 antibodies, may constitute a novel antiviral approach to prevent primary HCV infection, such as after LT.STRASBOURG-Sc. et Techniques (674822102) / SudocSudocFranceF