title: Fc-galactosylation modulates antibody-dependent cellular cytotoxicity of therapeutic antibodies.
abstract: The therapeutic activity of monoclonal antibodies can involve immune cell mediated effector functions including antibody-dependent cellular cytotoxicity (ADCC), an activity that is modulated by the structure of Fc-glycans, and in particular the lack of core fucose. The heterogeneity of these glycostructures and the inherent variability of traditional PBMC-based in vitro ADCC assays, have made it challenging to quantitatively assess the impact of other glycostructures on ADCC activity. We applied a quantitative NK cell based assay to generate a database consisting of Fc-glycostructure and ADCC data from 54 manufacturing batches of a CHO-derived monoclonal antibody. Explorative analysis of the data indicated that, apart from afucosylation, galactosylation levels could influence ADCC activity. We confirmed this hypothesis by demonstrating enhanced ADCC upon enzymatic hypergalactosylation of four different monoclonal antibodies derived using standard CHO manufacturing processes. Furthermore we quantitatively compare the effects of galactosylation and afucosylation in the context of glycan heterogeneity and demonstrate that while galactose can influence ADCC activity, afucosylation remains the primary driver of this activity.
authors.affiliation: Pharma Technical Development Analytics Biologics, Roche Diagnostics GmbH, Penzberg, Germany. Electronic address: marco.thomann@roche.com.
authors.forename: Marco
authors.initials: M
authors.lastname: Thomann
authors.affiliation: Pharma Biotech Manufacturing Penzberg, Roche Diagnostics GmbH, Penzberg, Germany.
authors.forename: Katharina
authors.initials: K
authors.lastname: Reckermann
authors.affiliation: Pharma Technical Development Analytics Biologics, Roche Diagnostics GmbH, Penzberg, Germany.
authors.forename: Dietmar
authors.initials: D
authors.lastname: Reusch
authors.affiliation: Pharma Biotech Manufacturing Penzberg, Roche Diagnostics GmbH, Penzberg, Germany.
authors.forename: Jessica
authors.initials: J
authors.lastname: Prasser
authors.affiliation: Analytical Development & Quality Control, Genentech Inc., South San Francisco, CA, USA.
authors.forename: Max L
authors.initials: ML
authors.lastname: Tejada
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000378:metabolism*; D000920:Antibody-Dependent Cell Cytotoxicity / Q000276:immunology*; D005643:Fucose / Q000378:metabolism; D005690:Galactose / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D007074:Immunoglobulin G / Q000737:chemistry / Q000378:metabolism*; D007694:Killer Cells, Natural / Q000276:immunology; D017452:Receptors, IgG / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D017452:Receptors, IgG; D005643:Fucose; D005690:Galactose
keywords: ADCC; Afucosylation; Antibody; CDC; Effector function; Fc-glycan; Fucosylation; Galactosylation; IgG1; Natural killer cell