title: Monoclonal Antibodies against Epidermal Growth Factor Receptor Acquire an Ability To Kill Tumor Cells through Complement Activation by Mutations That Selectively Facilitate the Hexamerization of IgG on Opsonized Cells.
abstract: Triggering of the complement cascade induces tumor cell lysis via complement-dependent cytotoxicity (CDC) and attracts and activates cytotoxic cells. It therefore represents an attractive mechanism for mAb in cancer immunotherapy development. The classical complement pathway is initiated by IgG molecules that have assembled into ordered hexamers after binding their Ag on the tumor cell surface. The requirements for CDC are further impacted by factors such as Ab epitope, valency, and affinity. Thus, mAb against well-validated solid tumor targets, such as the epidermal growth factor receptor (EGFR) that effectively induces complement activation and CDC, are highly sought after. The potency of complement activation by IgG Abs can be increased via several strategies. We identified single-point mutations in the Fc domain (e.g., E345K or E430G) enhancing Fc:Fc interactions, hexamer formation, and CDC after Ab binds cell-surface Ag. We show that EGFR Abs directed against clinically relevant epitopes can be converted into mAb with unprecedented CDC activity. Alternative strategies rely on increasing the affinity of monomeric IgG for C1q by introduction of a quadruple mutation at the C1q binding site or via generation of an IgG1/IgG3 chimera. In this study we show that selective enhancement of C1q binding via avidity modulation is superior to the unattended increase in C1q binding via affinity approaches, particularly for target cells with reduced EGFR expression levels. Improving Fc:Fc interactions of Ag-bound IgG therefore represents a highly promising and novel approach for potentiating the anti-tumor activity of therapeutic mAb against EGFR and potentially other tumor targets.
authors.affiliation: Division of Stem Cell Transplantation and Immunotherapy, Department of Internal Medicine II, University Hospital Schleswig-Holstein, Christian Albrechts University, 24105 Kiel, Germany.
authors.forename: Annalina
authors.initials: A
authors.lastname: Tammen
authors.affiliation: Division of Stem Cell Transplantation and Immunotherapy, Department of Internal Medicine II, University Hospital Schleswig-Holstein, Christian Albrechts University, 24105 Kiel, Germany.|Medical Department 1, University Hospital Schleswig-Holstein, 23538 Lübeck, Germany.
authors.forename: Stefanie
authors.initials: S
authors.lastname: Derer
authors.affiliation: Division of Stem Cell Transplantation and Immunotherapy, Department of Internal Medicine II, University Hospital Schleswig-Holstein, Christian Albrechts University, 24105 Kiel, Germany.
authors.forename: Ralf
authors.identifier: 0000-0003-0925-929X
authors.initials: R
authors.lastname: Schwanbeck
authors.affiliation: Division of Stem Cell Transplantation and Immunotherapy, Department of Internal Medicine II, University Hospital Schleswig-Holstein, Christian Albrechts University, 24105 Kiel, Germany.
authors.forename: Thies
authors.initials: T
authors.lastname: Rösner
authors.affiliation: Division of Stem Cell Transplantation and Immunotherapy, Department of Internal Medicine II, University Hospital Schleswig-Holstein, Christian Albrechts University, 24105 Kiel, Germany.
authors.forename: Anna
authors.initials: A
authors.lastname: Kretschmer
authors.affiliation: Genmab, 3584 Utrecht, the Netherlands; and.
authors.forename: Frank J
authors.identifier: 0000-0002-7165-5771
authors.initials: FJ
authors.lastname: Beurskens
authors.affiliation: Genmab, 3584 Utrecht, the Netherlands; and.
authors.forename: Janine
authors.identifier: 0000-0002-9738-9926
authors.initials: J
authors.lastname: Schuurman
authors.affiliation: Genmab, 3584 Utrecht, the Netherlands; and.|Department of Immunohematology and Blood Transfusion, Leiden University Medical Center, 2333 Leiden, the Netherlands.
authors.forename: Paul W H I
authors.identifier: 0000-0002-4365-3859
authors.initials: PW
authors.lastname: Parren
authors.affiliation: Division of Stem Cell Transplantation and Immunotherapy, Department of Internal Medicine II, University Hospital Schleswig-Holstein, Christian Albrechts University, 24105 Kiel, Germany; t.valerius@med2.uni-kiel.de.
authors.forename: Thomas
authors.identifier: 0000-0001-9181-8067
authors.initials: T
authors.lastname: Valerius
mesh_terms: D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology* / Q000627:therapeutic use*; D000920:Antibody-Dependent Cell Cytotoxicity; D001665:Binding Sites; D045744:Cell Line, Tumor; D003167:Complement Activation; D015922:Complement C1q / Q000276:immunology / Q000378:metabolism; D066246:ErbB Receptors / Q000235:genetics / Q000276:immunology*; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism; D007167:Immunotherapy / Q000379:methods; D009154:Mutation; D017354:Point Mutation
chemical_list: D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D015922:Complement C1q; C512478:EGFR protein, human; D066246:ErbB Receptors