title: A novel antibody engineering strategy for making monovalent bispecific heterodimeric IgG antibodies by electrostatic steering mechanism.
abstract: Producing pure and well behaved bispecific antibodies (bsAbs) on a large scale for preclinical and clinical testing is a challenging task. Here, we describe a new strategy for making monovalent bispecific heterodimeric IgG antibodies in mammalian cells. We applied an electrostatic steering mechanism to engineer antibody light chain-heavy chain (LC-HC) interface residues in such a way that each LC strongly favors its cognate HC when two different HCs and two different LCs are co-expressed in the same cell to assemble a functional bispecific antibody. We produced heterodimeric IgGs from transiently and stably transfected mammalian cells. The engineered heterodimeric IgG molecules maintain the overall IgG structure with correct LC-HC pairings, bind to two different antigens with comparable affinity when compared with their parental antibodies, and retain the functionality of parental antibodies in biological assays. In addition, the bispecific heterodimeric IgG derived from anti-HER2 and anti-EGF receptor (EGFR) antibody was shown to induce a higher level of receptor internalization than the combination of two parental antibodies. Mouse xenograft BxPC-3, Panc-1, and Calu-3 human tumor models showed that the heterodimeric IgGs strongly inhibited tumor growth. The described approach can be used to generate tools from two pre-existent antibodies and explore the potential of bispecific antibodies. The asymmetrically engineered Fc variants for antibody-dependent cellular cytotoxicity enhancement could be embedded in monovalent bispecific heterodimeric IgG to make best-in-class therapeutic antibodies.
authors.affiliation: From the Departments of Therapeutic Discovery and Amgen Inc., Seattle, Washington 98119, phillipzhiliu@hotmail.com.
authors.forename: Zhi
authors.initials: Z
authors.lastname: Liu
authors.affiliation: From the Departments of Therapeutic Discovery and Amgen Inc., Burnaby, British Columbia V5A 1V7, Canada.
authors.forename: Esther C
authors.initials: EC
authors.lastname: Leng
authors.affiliation: From the Departments of Therapeutic Discovery and Amgen Inc., Thousand Oaks, California 91320, and.
authors.forename: Kannan
authors.initials: K
authors.lastname: Gunasekaran
authors.affiliation: From the Departments of Therapeutic Discovery and Amgen Inc., Seattle, Washington 98119.
authors.forename: Martin
authors.initials: M
authors.lastname: Pentony
authors.affiliation: From the Departments of Therapeutic Discovery and Amgen Inc., Seattle, Washington 98119.
authors.forename: Min
authors.initials: M
authors.lastname: Shen
authors.affiliation: From the Departments of Therapeutic Discovery and Amgen Inc., Seattle, Washington 98119.
authors.forename: Monique
authors.initials: M
authors.lastname: Howard
authors.affiliation: From the Departments of Therapeutic Discovery and Amgen Inc., Seattle, Washington 98119.
authors.forename: Janelle
authors.initials: J
authors.lastname: Stoops
authors.affiliation: From the Departments of Therapeutic Discovery and Amgen Inc., Burnaby, British Columbia V5A 1V7, Canada.
authors.forename: Kathy
authors.initials: K
authors.lastname: Manchulenko
authors.affiliation: Amgen Inc., Seattle, Washington 98119, Process and Product Development.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Razinkov
authors.affiliation: Amgen Inc., Seattle, Washington 98119, Therapeutic Innovation Unit.
authors.forename: Hua
authors.initials: H
authors.lastname: Liu
authors.affiliation: Amgen Inc., Seattle, Washington 98119, Therapeutic Innovation Unit.
authors.forename: William
authors.initials: W
authors.lastname: Fanslow
authors.affiliation: From the Departments of Therapeutic Discovery and Amgen Inc., Seattle, Washington 98119.
authors.forename: Zhonghua
authors.initials: Z
authors.lastname: Hu
authors.affiliation: From the Departments of Therapeutic Discovery and Amgen Inc., Seattle, Washington 98119.
authors.forename: Nancy
authors.initials: N
authors.lastname: Sun
authors.affiliation: From the Departments of Therapeutic Discovery and Amgen Inc., Seattle, Washington 98119.
authors.forename: Haruki
authors.initials: H
authors.lastname: Hasegawa
authors.affiliation: From the Departments of Therapeutic Discovery and Amgen Inc., Seattle, Washington 98119.
authors.forename: Rutilio
authors.initials: R
authors.lastname: Clark
authors.affiliation: From the Departments of Therapeutic Discovery and Amgen Inc., Burnaby, British Columbia V5A 1V7, Canada.
authors.forename: Ian N
authors.initials: IN
authors.lastname: Foltz
authors.affiliation: From the Departments of Therapeutic Discovery and Amgen Inc., Seattle, Washington 98119, fondofgo@yahoo.com.
authors.forename: Wei
authors.initials: W
authors.lastname: Yan
mesh_terms: D000596:Amino Acids / Q000737:chemistry; D000818:Animals; D000920:Antibody-Dependent Cell Cytotoxicity; D016466:CHO Cells; D002460:Cell Line; D003412:Cricetulus; D019281:Dimerization; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry* / Q000276:immunology; D051379:Mice; D015202:Protein Engineering; D055672:Static Electricity; D020349:Surface Plasmon Resonance
chemical_list: D000596:Amino Acids; D007074:Immunoglobulin G
keywords: Antibody; Antibody Engineering; Cancer Therapy; Fc-γ Receptor; Pancreatic Cancer