title: Increasing the affinity for tumor antigen enhances bispecific antibody cytotoxicity.
abstract: We tested the hypothesis that bispecific Abs (Bsab) with increased binding affinity for tumor Ags augment retargeted antitumor cytotoxicity. We report that an increase in the affinity of Bsab for the HER2/neu Ag correlates with an increase in the ability of the Bsab to promote retargeted cytotoxicity against HER2/neu-positive cell lines. A series of anti-HER2/neu extracellular domain-directed single-chain Fv fragments (scFv), ranging in affinity for HER2/neu from 10(-7) to 10(-11) M, were fused to the phage display-derived NM3E2 human scFV: NM3E2 associates with the extracellular domain of human FcgammaRIII (CD16). The resulting series of Bsab promoted cytotoxicity of SKOV3 human ovarian carcinoma cells overexpressing HER2/neu by human PBMC preparations containing CD16-positive NK cells. The affinity for HER2/neu clearly influenced the ability of the Bsab to promote cytotoxicity of (51)Cr-labeled SKOV3 cells. Lysis was 6.5% with an anti-HER2/neu K(D) = 1.7 x 10(-7) M, 14.5% with K(D) = 5.7 x 10(-9) M, and 21.3% with K(D) = 1.7 x 10(-10) M at 50:1 E:T ratios. These scFv-based Bsab did not cross-link receptors and induce leukocyte calcium mobilization in the absence of tumor cell engagement. Thus, these novel Bsab structures should not induce the dose-limiting cytokine release syndromes that have been observed in clinical trials with intact IgG BSAB: Additional manipulations in Bsab structure that improve selective tumor retention or facilitate the ability of Bsab to selectively cross-link tumor and effector cells at tumor sites should further improve the utility of this therapeutic strategy.
authors.affiliation: Department of Medical Oncology, Fox Chase Cancer Center, Philadelphia, PA 19111, USA.
authors.forename: A M
authors.initials: AM
authors.lastname: McCall
authors.forename: L
authors.initials: L
authors.lastname: Shahied
authors.forename: A R
authors.initials: AR
authors.lastname: Amoroso
authors.forename: E M
authors.initials: EM
authors.lastname: Horak
authors.forename: H H
authors.initials: HH
authors.lastname: Simmons
authors.forename: U
authors.initials: U
authors.lastname: Nielson
authors.forename: G P
authors.initials: GP
authors.lastname: Adams
authors.forename: R
authors.initials: R
authors.lastname: Schier
authors.forename: J D
authors.initials: JD
authors.lastname: Marks
authors.forename: L M
authors.initials: LM
authors.lastname: Weiner
mesh_terms: D000276:Adjuvants, Immunologic / Q000378:metabolism* / Q000633:toxicity*; D018033:Antibodies, Bispecific / Q000378:metabolism* / Q000633:toxicity*; D000918:Antibody Specificity; D000920:Antibody-Dependent Cell Cytotoxicity; D000951:Antigens, Neoplasm / Q000276:immunology* / Q000378:metabolism*; D001666:Binding Sites, Antibody; D020013:Calcium Signaling / Q000276:immunology; D003601:Cytotoxicity Tests, Immunologic; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000378:metabolism; D007424:Intracellular Fluid / Q000276:immunology / Q000378:metabolism; D007700:Kinetics; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000276:immunology / Q000378:metabolism; D017452:Receptors, IgG / Q000276:immunology / Q000378:metabolism; D014407:Tumor Cells, Cultured
chemical_list: D000276:Adjuvants, Immunologic; D018033:Antibodies, Bispecific; D000951:Antigens, Neoplasm; D007135:Immunoglobulin Variable Region; D018719:Erb-b2 Receptor Tyrosine Kinases; D017452:Receptors, IgG