Neutralizing Antibody-Resistant Hepatitis C Virus Cell-to-Cell Transmission

Abstract

Hepatitis C virus (HCV) can initiate infection by cell-free particle and cell-cell contact-dependent transmission. In this study we use a novel infectious coculture system to examine these alternative modes of infection. Cell-to-cell transmission is relatively resistant to anti-HCV glycoprotein monoclonal antibodies and polyclonal immunoglobulin isolated from infected individuals, providing an effective strategy for escaping host humoral immune responses. Chimeric viruses expressing the structural proteins representing the seven major HCV genotypes demonstrate neutralizing antibody-resistant cell-to-cell transmission. HCV entry is a multistep process involving numerous receptors. In this study we demonstrate that, in contrast to earlier reports, CD81 and the tight-junction components claudin-1 and occludin are all essential for both cell-free and cell-to-cell viral transmission. However, scavenger receptor BI (SR-BI) has a more prominent role in cell-to-cell transmission of the virus, with SR-BI-specific antibodies and small-molecule inhibitors showing preferential inhibition of this infection route. These observations highlight the importance of targeting host cell receptors, in particular SR-BI, to control viral infection and spread in the liver. Hepatitis C virus (HCV) establishes chronic infection in 3%of the world's population, resulting in a progressive liver disease that is one of the leading indications for liver transplantation. HCV has evolved several immune evasion strategies in order to persist within the infected host (15,20,40), including genetic escape from humoral immune responses (25,46). However, functional constraints may restrict antigenic change in some regions of the virally encoded E1E2 envelope glycoproteins, such as the CD81 receptor binding site (9,11,33). The observation that glycoprotein-specific antibodies from chronically infected subjects neutralize the infectivity of laboratory prototype HCV strains yet demonstrate a limited ability to control HCV replication in vivo (40) suggest that additional means of evading antibody responses may exist.How virus particles disseminate within an immune-competent host has been a relatively neglected area of study; however, it is becoming increasingly clear that viruses employ multiple strategies to infect new target cells. Diffusion through the pericellular environment or the vascular circulation introduces a rate-limiting step in virus entry and exposes particles to the humoral immune system. Consequently, a number of viruses have evolved direct cell-to-cell modes of transmission that maximize particle delivery, often in a neutralizing antibody (nAb)-resistant manner (reviewed in reference 30).We (44) and others (48) previously reported that HCV strain JFH-1 could be transmitted via cell-free and cell-to-cell routes in vitro. We extend these observations and show that disruption of HCV particle assembly or physical separation of target and producer cells ablates transmission, demonstrating that intact virions are transferred via cell-cell conta...

Apolipoprotein E Mediates Evasion From Hepatitis C Virus Neutralizing Antibodies

“…32 To investigate whether apoE association is relevant in cell-to-cell transmission, we used an anti-apoE antibody (1D7) that binds the low-density lipoprotein receptorÀbinding domain of apoE 16 and inhibits HCV infection ( Figure 7A). Jc1 demonstrated an approximately 3-fold higher capacity to spread by direct cell-to-cell transmission than VL:JFH1 in this assay ( Figure 6A).…”

Impact of Intra- and Interspecies Variation of Occludin on Its Function as Coreceptor for Authentic Hepatitis C Virus Particles

“…Whether the mechanisms of cell entry during cell-to-cell spread are distinct from those during cell entry by an extracellular virion is unclear. Most notably, there is conflicting data on whether cell-to-cell spread is CD81 independent (22,38,41) or not (7). OCLN knockdown has been reported to reduce cell-to-cell transmission by 2-to 3-fold in an assay in which spread, despite the presence of neutralizing antibodies in the supernatant, was defined as occurring cell to cell (7).…”

“…Most notably, there is conflicting data on whether cell-to-cell spread is CD81 independent (22,38,41) or not (7). OCLN knockdown has been reported to reduce cell-to-cell transmission by 2-to 3-fold in an assay in which spread, despite the presence of neutralizing antibodies in the supernatant, was defined as occurring cell to cell (7). We used an agarose overlay to prevent cell-free spread and a HCV-NS3/4A-sensitive fluorescent reporter to monitor spread of infection in living cells and found that the absence of OCLN in target cells abolished cell-to-cell spread, indicating that this route is strictly OCLN dependent.…”

Entry and replication of recombinant hepatitis C viruses in cell culture

“…Infected events in the second populations are considered as indicator of cell-to-cell transmission if the cell-free infectivity is blocked. This last condition can be 13 achieved by using saturating concentrations of neutralizing antibodies [171,186,187], particular virus mutants or target cells [101], or by culturing the cells under an agarose overlay that prevents Brownian motion of cell-free viruses [102,171]. The dual cell-based reporter system developed by Jones et al…”