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. 2007 Aug;81(15):8063-71.
doi: 10.1128/JVI.00193-07. Epub 2007 May 16.

High-avidity monoclonal antibodies against the human scavenger class B type I receptor efficiently block hepatitis C virus infection in the presence of high-density lipoprotein

Maria Teresa Catanese  1 , Rita Graziani, Thomas von Hahn, Martine Moreau, Thierry Huby, Giacomo Paonessa, Claudia Santini, Alessandra Luzzago, Charles M Rice, Riccardo Cortese, Alessandra Vitelli, Alfredo Nicosia
Affiliations

High-avidity monoclonal antibodies against the human scavenger class B type I receptor efficiently block hepatitis C virus infection in the presence of high-density lipoprotein

Maria Teresa Catanese et al. J Virol. 2007 Aug.

Abstract

The human scavenger class B type 1 receptor (SR-B1/Cla1) was identified as a putative receptor for hepatitis C virus (HCV) because it binds to soluble recombinant HCV envelope glycoprotein E2 (sE2). High-density lipoprotein (HDL), a natural SR-B1 ligand, was shown to increase the in vitro infectivity of retroviral pseudoparticles bearing HCV envelope glycoproteins and of cell culture-derived HCV (HCVcc), suggesting that SR-B1 promotes viral entry in an HDL-dependent manner. To determine whether SR-B1 participates directly in HCV infection or facilitates HCV entry through lipoprotein uptake, we generated a panel of monoclonal antibodies (MAbs) against native human SR-B1. Two of them, 3D5 and C167, bound to conformation-dependent SR-B1 determinants and inhibited the interaction of sE2 with SR-B1. These antibodies efficiently blocked HCVcc infection of Huh-7.5 hepatoma cells in a dose-dependent manner. To examine the role of HDL in SR-B1-mediated HCVcc infection, we set up conditions for HCVcc production and infection in serum-free medium. HCVcc efficiently infected Huh-7.5 cells in the absence of serum lipoproteins, and addition of HDL led to a twofold increase in infectivity. However, the HDL-induced enhancement of infection had no impact on the neutralization potency of MAb C167, despite its ability to inhibit both HDL binding to cells and SR-B1-mediated lipid transfer. Of note, MAb C167 also potently blocked Huh-7.5 infection by an HCV strain recovered from HCVcc-infected chimpanzees. These results demonstrate that SR-B1 is essential for infection with HCV produced in vitro and in vivo and suggest the possible use of anti-SR-B1 antibodies as therapeutic agents.

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Figures

FIG. 1.
FIG. 1.
Binding properties of anti-SR-B1 MAbs and neutralization of sE2 binding to stable cell line CHO/SR-B1. (A) Binding of anti-SR-B1 MAbs to Huh-7.5 cells as measured by flow cytometry. Values on the vertical axis are MFIs. Values on the horizontal axis are concentrations (ng/ml) of anti-SR-B1 MAbs on a logarithmic scale. Saturation curves for 3D5 (closed circles), C167 (open circles), 6B8 (closed triangles), and C11 (open triangles) were fitted to experimental data with the Sigma Plot program. MFIs obtained with IgG isotypic controls were subtracted. Apparent Kds are shown in the inset. (B) Inhibition of binding of sE2 to stable cell line CHO/SR-B1 by anti-SR-B1 MAbs as measured by flow cytometry. Concentrations (ng/ml) of 3D5, C167, and C11 are indicated on the horizontal axis on a logarithmic scale. On the vertical axis are reported the MFIs of sE2 binding to the CHO/SR-B1 cell line. Neutralization-of-binding curves for MAbs 3D5 (closed circles), C167 (open circles), and C11 (closed triangles) were fitted to experimental data with the Sigma Plot program, and IC50s of MAbs are shown in the inset.
FIG. 2.
FIG. 2.
Cross-competition between anti-SR-B1 antibodies. 3D5 (A) and C11 (B) binding to Huh-7.5 cells was measured by flow cytometry in the absence of competing antibodies (black bars) or in the presence of either isotypic control IgG (Ctrl IgG; gray bars) or an anti-SR-B1 antibody (MAb C167; open bars) as a competitor. Values are percentages of MAb binding, relative to the binding of the MAb in the absence of a competing MAb.
FIG. 3.
FIG. 3.
Influence of anti-SR-B1 MAbs on HDL binding to SR-B1 and on SR-B1-mediated cholesterol transfer. (A) CHO or CHO/hSR-B1 cells were preincubated in PBS-BSA (no IgG; open bars) or in the presence of control IgG (Ctrl IgG; gray bars) or anti-SR-B1 MAbs (black bars, preincubation with 6B8; hatched bars, preincubation with C11; dotted bars, preincubation with C167; checked bars, preincubation with 3D5) at 5 μg/ml prior to incubation with DiI-labeled HDL (50 μg protein/ml) for 1 h at 4°C. Data are the percentages of DiI-HDL-positive cells. Values are the means ± standard deviations of three independent experiments. (B) CHO and CHO/hSR-B1 cells were labeled with [3H]cholesterol, washed, and incubated for 30 min with SFM containing increasing amounts (open bars, 0.3 μg/ml; gray bars, 1 μg/ml; black bars, 3 μg/ml; hatched bars, 9 μg/ml) of control IgG (Ctrl IgG) or anti-SR-B1 MAbs (6B8, C11, C167, and 3D5). After an additional 2 h of incubation in the same medium in the presence HDL (20 μg protein/ml), the amount of [3H]cholesterol present in the medium or remaining in the cells was determined. Data are the percentages of SR-B1-mediated cellular cholesterol efflux and were calculated as the differences between the efflux values determined with CHO/hSR-B1 and those determined with parental CHO cells. Values are the means ± standard deviations of three independent experiments.
FIG. 4.
FIG. 4.
Effects of human serum and HDL on HCVcc infection. (A) J6/JFH HCVcc produced in SFM was used to infect Huh-7.5 cells in SFM (gray bar) or in SFM complemented with 10% FBS (black bar) or human serum (HS) from three different healthy donors (open bar, HS I; light gray bar, HS II; hatched bar, HS III;). Total cellular RNA was analyzed by quantitative RT-PCR for HCV content at day 3 postinfection. On the vertical axis is shown the percentage of HCV copies (% HCV infection) relative to the number of HCV copies measured in cells infected in the presence of 10% FBS. Data are averages and standard deviations of triplicate wells. (B) J6/JFH HCVcc produced in SFM was used to infect Huh-7.5 cells in SFM alone or in the presence of increasing concentrations of HDL. Total cellular RNA was analyzed by quantitative RT-PCR for HCV content at day 3 postinfection. On the vertical axis is shown the percentage of HCV copies (% HCV infection) relative to the number of HCV copies measured in cells infected in SFM. Data are averages and standard deviations of triplicate wells.
FIG. 5.
FIG. 5.
Neutralization of J6/JFH HCVcc infection by anti-SR-B1 MAb in SFM and in the presence of HDL. Huh-7.5 cells were infected with HCVcc produced in SFM upon preincubation with increasing concentrations (pg/ml) of anti-SR-B1 MAb C167 (indicated on the horizontal axis on a logarithmic scale) in SFM or in SFM plus 1.6 μg/ml HDL. Total cellular RNA was analyzed by quantitative RT-PCR for HCV content at day 3 postinfection. On the vertical axis is shown the percentage of HCV copies (% HCV infection) measured in cells infected in the presence of C167, relative to the number of HCV copies measured in control antibody-treated cells either in SFM (open circles) or in the presence of 1.6 μg/ml HDL (closed circles). Neutralization curves were extrapolated by fitting experimental data with the Sigma Plot program, and the IC50s of the MAbs are shown in the inset.
FIG. 6.
FIG. 6.
Anti-SR-B1 MAbs inhibit infection of Huh-7.5 with chimpanzee-derived HCV. (A) Infection of Huh-7.5 cells with chimpanzee-derived HCV (ex vivo HCVcc) was performed in the presence of anti-SR-B1 antibody (MAb C167; gray bars), anti-human-CD81 (MAb 1.3.3.22; open bars), or control isotypic IgG (Ctrl IgG; black bar) at the concentrations (μg/ml) indicated on the horizontal axis. Total cellular RNA was analyzed for HCV content by quantitative RT-PCR at 3 days postinfection. On the vertical axis is shown the percentage of HCV copies (% HCV infection) relative to the number of HCV copies measured in control antibody-treated cells. Data are averages and standard deviations of triplicate wells. (B) Infection of Huh-7.5 cells in the presence of increasing concentrations (pg/ml) of anti-SR-B1 MAb C167 (indicated on the horizontal axis on a logarithmic scale) was performed either with chimpanzee-derived HCV (ex vivo HCVcc, open circles) or with cell culture-derived HCV (HCVcc, closed circles). RNA was analyzed for HCV content by quantitative RT-PCR at day 3 postinfection. On the vertical axis is shown the percentage of HCV copies (% HCV infection) relative to the number of HCV copies measured in control antibody-treated cells. Neutralization curves were extrapolated by fitting experimental data with the Sigma Plot program, and the IC50s of the MAbs are shown in the inset.
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References

    1. Acton, S., A. Rigotti, K. T. Landschulz, S. Xu, H. H. Hobbs, and M. Krieger. 1996. Identification of scavenger receptor SR-BI as a high density lipoprotein receptor. Science 271:518-520. - PubMed
    1. Agnello, V., G. Abel, M. Elfahal, G. B. Knight, and Q. X. Zhang. 1999. Hepatitis C virus and other Flaviviridae viruses enter cells via low density lipoprotein receptor. Proc. Natl. Acad. Sci. USA 96:12766-12771. - PMC - PubMed
    1. André, P., F. Komurian-Pradel, S. Deforges, M. Perret, J. L. Berland, M. Sodoyer, S. Pol, C. Brechot, G. Paranhos-Baccala, and V. Lotteau. 2002. Characterization of low- and very-low-density hepatitis C virus RNA-containing particles. J. Virol. 76:6919-6928. - PMC - PubMed
    1. Barth, H., C. Schafer, M. I. Adah, F. Zhang, R. J. Linhardt, H. Toyoda, A. Kinoshita-Toyoda, T. Toida, T. H. Van Kuppevelt, E. Depla, F. Von Weizsacker, H. E. Blum, and T. F. Baumert. 2003. Cellular binding of hepatitis C virus envelope glycoprotein E2 requires cell surface heparan sulfate. J. Biol. Chem. 278:41003-41012. - PubMed
    1. Bartosch, B., A. Vitelli, C. Granire, C. Goujon, J. Dubuisson, S. Pascale, E. Scarselli, R., Cortese, A. Nicosia, and F. L. Cosset. 2003. Cell entry of hepatitis C virus requires a set of coreceptors that include the CD81 tetraspanin and the SR-B1 scavenger receptor. J. Biol. Chem. 278:41624-41630. - PubMed

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