Generation and characterization of a novel tetravalent bispecific antibody that binds to hepatitis B virus surface antigens

• Published in: Molecular immunology Vol. 37; no. 18; pp. 1123 - 1130
• Main Authors: Park, Sung Sup, Ryu, Chun Jeih, Kang, Young Jun, Kashmiri, S.V.S., Hong, Hyo Jeong
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01.12.2000
• Subjects: Antibodies, Bispecific - genetics; Antibodies, Bispecific - immunology; Antibodies, Monoclonal - genetics; Antibodies, Monoclonal - immunology; Antibody Affinity; Antibody engineering; Bispecific antibody; Hepatitis B Antibodies - genetics; Hepatitis B Antibodies - immunology; hepatitis B surface antigen; Hepatitis B Surface Antigens - immunology; Hepatitis B virus; Immunoglobulin Fc Fragments - genetics; Immunoglobulin Fc Fragments - immunology; Immunoglobulin Variable Region - genetics; Immunoglobulin Variable Region - immunology; Models, Molecular; Precipitin Tests; S antigen; Single-chain antibody; Single-chain antibody; CHO, Chinese hamster ovary; HBV, hepatitis B virus; BSA, bovine serum albumin; mAb, monoclonal antibody; FBS, fetal bovine serum; PBS, phosphate buffered saline; V H, heavy-chain variable domain; Bispecific antibody; ELISA, enzyme-linked immunosorbent assay; PAGE, polyacrylamide gel electrophoresis; C γ1, gamma 1 heavy chain constant region; V L, light-chain variable domain; Ag, antigen; bp, base pair; PCR, polymerase chain reaction; Antibody engineering; Hepatitis B virus; ScFv, single-chain antibody fragment
• ISSN: 0161-5890; 1872-9142
• DOI: 10.1016/S0161-5890(01)00027-X

Hepatitis B virus (HBV) infection is a worldwide public health problem affecting about 350 million people. HBV envelope contains three surface antigens, called pre-S1, pre-S2 and S. For the prophylaxis of HBV infection, only an anti-S monoclonal antibody was tested for the protective efficacy against HBV infection, but it was shown to be incomplete. In addition, some immune escape mutants carrying mutations on the S antigen were reported. Therefore, a multivalent bispecific antibody rather than a single monoclonal antibody would be more beneficial for the prophylaxis of HBV infection. We have generated a novel tetravalent bispecific antibody with two binding sites for each of the S and pre-S2 antigens. Each of the antigen-binding sites was composed of a single-chain Fv (ScFv). The tetravalent antibody was generated by constructing a single gene encoding a single-chain protein. This protein consisted of an anti-S ScFv whose carboxyl end was tethered, through a 45 amino acid linker, to the amino terminus of anti-preS2 ScFv that in turn was joined to the hinge region of human γ1 constant region. The single-chain protein was expressed in Chinese hamster ovary cells and secreted in culture supernatant as a homodimeric molecule. The tetravalent bispecific antibody showed both anti-S and anti-pre-S2 binding activities. In addition, the binding affinity of the bispecific antiboy for HBV particles was greater than that of either parental antibody. The tetravalent bispecific antibody is a potentially useful reagent for the prevention and treatment of HBV infection.