title: Efficient generation of stable bispecific IgG1 by controlled Fab-arm exchange.
abstract: The promise of bispecific antibodies (bsAbs) to yield more effective therapeutics is well recognized; however, the generation of bsAbs in a practical and cost-effective manner has been a formidable challenge. Here we present a technology for the efficient generation of bsAbs with normal IgG structures that is amenable to both antibody drug discovery and development. The process involves separate expression of two parental antibodies, each containing single matched point mutations in the CH3 domains. The parental antibodies are mixed and subjected to controlled reducing conditions in vitro that separate the antibodies into HL half-molecules and allow reassembly and reoxidation to form highly pure bsAbs. The technology is compatible with standard large-scale antibody manufacturing and ensures bsAbs with Fc-mediated effector functions and in vivo stability typical of IgG1 antibodies. Proof-of-concept studies with HER2×CD3 (T-cell recruitment) and HER2×HER2 (dual epitope targeting) bsAbs demonstrate superior in vivo activity compared with parental antibody pairs.
authors.affiliation: Genmab, 3584 CM, Utrecht, The Netherlands.
authors.forename: Aran F
authors.initials: AF
authors.lastname: Labrijn
authors.forename: Joyce I
authors.initials: JI
authors.lastname: Meesters
authors.forename: Bart E C G
authors.initials: BE
authors.lastname: de Goeij
authors.forename: Ewald T J
authors.initials: ET
authors.lastname: van den Bremer
authors.forename: Joost
authors.initials: J
authors.lastname: Neijssen
authors.forename: Muriel D
authors.initials: MD
authors.lastname: van Kampen
authors.forename: Kristin
authors.initials: K
authors.lastname: Strumane
authors.forename: Sandra
authors.initials: S
authors.lastname: Verploegen
authors.forename: Amitava
authors.initials: A
authors.lastname: Kundu
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Gramer
authors.forename: Patrick H C
authors.initials: PH
authors.lastname: van Berkel
authors.forename: Jan G J
authors.initials: JG
authors.lastname: van de Winkel
authors.forename: Janine
authors.initials: J
authors.lastname: Schuurman
authors.forename: Paul W H I
authors.initials: PW
authors.lastname: Parren
mesh_terms: D000818:Animals; D018033:Antibodies, Bispecific / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D057809:HEK293 Cells; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D007141:Immunoglobulin Fc Fragments / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D007074:Immunoglobulin G / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D019169:Jurkat Cells; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics
chemical_list: D018033:Antibodies, Bispecific; D007140:Immunoglobulin Fab Fragments; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D011994:Recombinant Proteins