Inhibition of the binding of HCV serum particles to human hepatocytes by E1E2-specific D32.10 monoclonal antibody

J Med Virol. 2009 Oct;81(10):1726-33. doi: 10.1002/jmv.21562.

Abstract

The aim of this study was to determine the inhibition of binding activity of the monoclonal antibody (mAb) D32.10 which recognizes a highly conserved discontinuous antigenic determinant (E1:297-306, E2:480-494, and E2:613-621) expressed on the surface of serum-derived HCV particles (HCVsp) of genotypes 1a, 1b, 2a, and 3a. To this end, an in vitro direct cell-binding assay based on the attachment of radiolabeled HCVsp was developed, and Scatchard plots were used to analyze ligand-receptor binding data. HCV adsorption was also assessed by quantitating cell-associated viral RNA by a real-time RT-PCR method. Saturable concentration-dependent specific binding of HCVsp to Huh-7 or HepaRG cells was demonstrated. The Scatchard transformed data showed two-site interaction for Huh-7 and proliferative HepaRG cells: the high-affinity binding sites (K(d1) = 0.1-0.5 microg/ml) and the low-affinity binding sites (K(d1) = 5-10 microg/ml), and one-site high-affinity binding model between E1E2/D32.10-positive HCVsp and hepatocyte-like differentiated HepaRG cells. The E1E2-specific mAb D32.10 inhibited efficiently (>60%) and selectively the binding with an IC(50) <or=0.5 microg/ml in all the experimental approaches using serum HCV of genotype either 3 or 1b. This supports the involvement of the E1E2/D32.10 discontinuous antigenic determinant in the interactions between human hepatocytes and HCVsp, and suggests that D32.10-like antibodies present in sera from patients infected with HCV could play a protective role.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antibodies, Monoclonal / immunology*
  • Cell Line
  • Hepacivirus / immunology*
  • Hepacivirus / physiology*
  • Hepatitis C Antibodies / immunology*
  • Hepatocytes / virology*
  • Humans
  • Inhibitory Concentration 50
  • Viral Envelope Proteins / immunology
  • Virus Attachment*

Substances

  • Antibodies, Monoclonal
  • E1 protein, Hepatitis C virus
  • Hepatitis C Antibodies
  • Viral Envelope Proteins
  • glycoprotein E2, Hepatitis C virus