title: Single-domain antibody-based and linker-free bispecific antibodies targeting FcγRIII induce potent antitumor activity without recruiting regulatory T cells.
abstract: Antibody-dependent cell-mediated cytotoxicity, one of the most prominent modes of action of antitumor antibodies, suffers from important limitations due to the need for optimal interactions with Fcγ receptors. In this work, we report the design of a new bispecific antibody format, compact and linker-free, based on the use of llama single-domain antibodies that are capable of circumventing most of these limitations. This bispecific antibody format was created by fusing single-domain antibodies directed against the carcinoembryonic antigen and the activating FcγRIIIa receptor to human Cκ and CH1 immunoglobulin G1 domains, acting as a natural dimerization motif. In vitro and in vivo characterization of these Fab-like bispecific molecules revealed favorable features for further development as a therapeutic molecule. They are easy to produce in Escherichia coli, very stable, and elicit potent lysis of tumor cells by human natural killer cells at picomolar concentrations. Unlike conventional antibodies, they do not engage inhibitory FcγRIIb receptor, do not compete with serum immunoglobulins G for receptor binding, and their cytotoxic activity is independent of Fc glycosylation and FcγRIIIa polymorphism. As opposed to anti-CD3 bispecific antitumor antibodies, they do not engage regulatory T cells as these latter cells do not express FcγRIII. Studies in nonobese diabetic/severe combined immunodeficient gamma mice xenografted with carcinoembryonic antigen-positive tumor cells showed that Fab-like bispecific molecules in the presence of human peripheral blood mononuclear cells significantly slow down tumor growth. This new compact, linker-free bispecific antibody format offers a promising approach for optimizing antibody-based therapies.
authors.affiliation: INSERM U1068, CRCM, France.
authors.forename: Caroline
authors.initials: C
authors.lastname: Rozan
authors.forename: Amélie
authors.initials: A
authors.lastname: Cornillon
authors.forename: Corinne
authors.initials: C
authors.lastname: Pétiard
authors.forename: Martine
authors.initials: M
authors.lastname: Chartier
authors.forename: Ghislaine
authors.initials: G
authors.lastname: Behar
authors.forename: Charlotte
authors.initials: C
authors.lastname: Boix
authors.forename: Brigitte
authors.initials: B
authors.lastname: Kerfelec
authors.forename: Bruno
authors.initials: B
authors.lastname: Robert
authors.forename: André
authors.initials: A
authors.lastname: Pèlegrin
authors.forename: Patrick
authors.initials: P
authors.lastname: Chames
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Teillaud
authors.forename: Daniel
authors.initials: D
authors.lastname: Baty
mesh_terms: D000818:Animals; D018033:Antibodies, Bispecific / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology*; D000920:Antibody-Dependent Cell Cytotoxicity / Q000276:immunology; D000970:Antineoplastic Agents / Q000737:chemistry / Q000494:pharmacology*; D002272:Carcinoembryonic Antigen / Q000276:immunology; D045744:Cell Line, Tumor; D016207:Cytokines / Q000096:biosynthesis; D004355:Drug Stability; D005260:Female; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology / Q000378:metabolism; D007694:Killer Cells, Natural / Q000276:immunology; D051379:Mice; D009369:Neoplasms / Q000188:drug therapy / Q000276:immunology; D011110:Polymorphism, Genetic; D011485:Protein Binding / Q000276:immunology; D055550:Protein Stability; D017452:Receptors, IgG / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D061905:Single-Domain Antibodies / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology*; D050378:T-Lymphocytes, Regulatory / Q000276:immunology* / Q000378:metabolism*; D014018:Tissue Distribution; D023041:Xenograft Model Antitumor Assays
chemical_list: D018033:Antibodies, Bispecific; D000970:Antineoplastic Agents; D002272:Carcinoembryonic Antigen; D016207:Cytokines; D007074:Immunoglobulin G; D017452:Receptors, IgG; D061905:Single-Domain Antibodies