title: Tumour antigen targeted monoclonal antibodies incorporating a novel multimerisation domain significantly enhance antibody dependent cellular cytotoxicity against colon cancer.
abstract: Tumour antigen targeted antibodies (mAbs) can induce natural killer (NK) cells to kill tumours through antibody dependent cellular cytotoxicity (ADCC) upon engagement of NK cell expressed FcγRIIIa. FcγRIIIa polymorphisms partially dictate the potency of the ADCC response. The high affinity FcγRIIIa-158-valine (V) polymorphism is associated with more potent ADCC response than the low affinity FcγRIIIa-158-phenylalanine (F) polymorphism. Because approximately 45% of patients are homozygous for the FcγRIIIa-158-F polymorphism (FF genotype), their ability to mount ADCC is impaired. We investigated whether a novel mAb capable of binding multiple antigen specific targets and engaging multiple low affinity FcγRIIIa receptors could further enhance ADCC against colon cancer in vitro. Specifically, we generated a novel anti-epidermal growth factor receptor (EGFR) antibody (termed a stradobody) consisting of an unmodified Fab sequence and two Immunoglobulin G, subclass 1 (IgG1) Fc domains separated by an isoleucine zipper domain and the 12 amino-acid IgG2 hinge. The stradobody framework induced multimerisation and was associated with increased binding to the EGFR and FcγRIIIa. From a functional perspective, when compared to an unmodified anti-EGFR mAb with a sequence identical to cetuximab (a commercially available anti-EGFR mAb), stradobodies significantly enhanced ADCC. These effects were observed using both KRAS wild type HT29 and KRAS mutant SW480 colon cancer cells as targets, and by NK cells obtained from healthy donors and a cohort of patients with colon cancer. These data suggest that high avidity cross-linking of multiple tumour surface antigens and multiple NK cell associated FcγRIIIa molecules can enhance ADCC and partially overcome impaired ADCC by FF genotype individuals in vitro.
authors.affiliation: Baltimore Veterans Administration Medical Center, Section of Surgical Oncology, and Research and Development Service, 10 N. Greene Street, 5C Surgical Services Area, Baltimore, MD 21201, USA; Division of General and Oncologic Surgery, University of Maryland School of Medicine, 22 S. Greene Street, Room S4B12, Baltimore, MD 21201, USA; Department of Otorhinolaryngology-Head and Neck Surgery, University of Maryland School of Medicine, 16 S. Eutaw Street, Baltimore, MD 21201, USA. Electronic address: ajain@smail.umaryland.edu.
authors.forename: Ajay
authors.initials: A
authors.lastname: Jain
authors.forename: Bhawna
authors.initials: B
authors.lastname: Poonia
authors.forename: Edward C
authors.initials: EC
authors.lastname: So
authors.forename: Ravi
authors.initials: R
authors.lastname: Vyzasatya
authors.forename: Erin E
authors.initials: EE
authors.lastname: Burch
authors.forename: Henrik S
authors.initials: HS
authors.lastname: Olsen
authors.forename: Emmanuel Y
authors.initials: EY
authors.lastname: Mérigeon
authors.forename: David S
authors.initials: DS
authors.lastname: Block
authors.forename: Xiaoyu
authors.initials: X
authors.lastname: Zhang
authors.forename: Dan H
authors.initials: DH
authors.lastname: Schulze
authors.forename: Nader N
authors.initials: NN
authors.lastname: Hanna
authors.forename: William S
authors.initials: WS
authors.lastname: Twadell
authors.forename: Harris G
authors.initials: HG
authors.lastname: Yfantis
authors.forename: Siaw L
authors.initials: SL
authors.lastname: Chan
authors.forename: Ling
authors.initials: L
authors.lastname: Cai
authors.forename: Scott E
authors.initials: SE
authors.lastname: Strome
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000494:pharmacology*; D000920:Antibody-Dependent Cell Cytotoxicity; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000235:genetics / Q000276:immunology* / Q000628:therapy*; D066246:ErbB Receptors / Q000235:genetics / Q000276:immunology*; D005838:Genotype; D019073:HT29 Cells; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000276:immunology; D007694:Killer Cells, Natural / Q000276:immunology*; D020641:Polymorphism, Single Nucleotide; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary
chemical_list: D000911:Antibodies, Monoclonal; D007141:Immunoglobulin Fc Fragments; D066246:ErbB Receptors
keywords: ADCC; Colon cancer; Fc; Fc modification; FcγRIIIa; Immunoglobulin; NK cell; Polyvalent