title: Immunoglobulin domain crossover as a generic approach for the production of bispecific IgG antibodies.
abstract: We describe a generic approach to assemble correctly two heavy and two light chains, derived from two existing antibodies, to form human bivalent bispecific IgG antibodies without use of artificial linkers. Based on the knobs-into-holes technology that enables heterodimerization of the heavy chains, correct association of the light chains and their cognate heavy chains is achieved by exchange of heavy-chain and light-chain domains within the antigen binding fragment (Fab) of one half of the bispecific antibody. This "crossover" retains the antigen-binding affinity but makes the two arms so different that light-chain mispairing can no longer occur. Applying the three possible "CrossMab" formats, we generated bispecific antibodies against angiopoietin-2 (Ang-2) and vascular endothelial growth factor A (VEGF-A) and show that they can be produced by standard techniques, exhibit stabilities comparable to natural antibodies, and bind both targets simultaneously with unaltered affinity. Because of its superior side-product profile, the CrossMab(CH1-CL) was selected for in vivo profiling and showed potent antiangiogenic and antitumoral activity.
authors.affiliation: Biologics Research, Pharma Research and Early Development, Roche Diagnostics GmbH, D-82372 Penzberg, Germany. wolfgang.schaefer@roche.com
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Schaefer
authors.forename: Jörg T
authors.initials: JT
authors.lastname: Regula
authors.forename: Monika
authors.initials: M
authors.lastname: Bähner
authors.forename: Jürgen
authors.initials: J
authors.lastname: Schanzer
authors.forename: Rebecca
authors.initials: R
authors.lastname: Croasdale
authors.forename: Harald
authors.initials: H
authors.lastname: Dürr
authors.forename: Christian
authors.initials: C
authors.lastname: Gassner
authors.forename: Guy
authors.initials: G
authors.lastname: Georges
authors.forename: Hubert
authors.initials: H
authors.lastname: Kettenberger
authors.forename: Sabine
authors.initials: S
authors.lastname: Imhof-Jung
authors.forename: Manfred
authors.initials: M
authors.lastname: Schwaiger
authors.forename: Kay G
authors.initials: KG
authors.lastname: Stubenrauch
authors.forename: Claudio
authors.initials: C
authors.lastname: Sustmann
authors.forename: Markus
authors.initials: M
authors.lastname: Thomas
authors.forename: Werner
authors.initials: W
authors.lastname: Scheuer
authors.forename: Christian
authors.initials: C
authors.lastname: Klein
mesh_terms: D042702:Angiopoietin-2 / Q000276:immunology; D000818:Animals; D018033:Antibodies, Bispecific / Q000096:biosynthesis* / Q000737:chemistry* / Q000378:metabolism; D000915:Antibody Affinity; D000918:Antibody Specificity; D002460:Cell Line; D045744:Cell Line, Tumor; D005260:Female; D006801:Humans; D007074:Immunoglobulin G / Q000096:biosynthesis* / Q000737:chemistry* / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D016513:Mice, SCID; D008958:Models, Molecular; D018919:Neovascularization, Physiologic; D015202:Protein Engineering; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000276:immunology; D042461:Vascular Endothelial Growth Factor A / Q000276:immunology
chemical_list: D042702:Angiopoietin-2; D018033:Antibodies, Bispecific; D007074:Immunoglobulin G; D011994:Recombinant Proteins; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A