title: Microheterogeneity of therapeutic monoclonal antibodies is governed by changes in the surface charge of the protein.
abstract: It has previously been shown for individual antibodies, that the microheterogenity pattern can have a significant impact on various key characteristics of the product. The aim of this study to get a more generalized understanding of the importance of microheterogeneity. For that purpose, the charge variant pattern of various different commercially available therapeutic mAb products was compared using Cation-Exchange Chromatography with linear pH gradient antigen affinity, Fc-receptor affinity, antibody dependent cellular cytotoxicity (ADCC) and conformational stability. For three of the investigated antibodies, the basic charge variants showed a stronger binding affinity towards FcγRIIIa as well as an increased ADCC response. Differences in the conformational stability of antibody charge variants and the corresponding reference samples could not be detected by differential scanning calorimetry. The different biological properties of the mAb variants are therefore governed by changes in the surface charge of the protein and not by an altered structure. This can help to identify aspects of microheterogeneity that are critical for product quality and can lead to further improvements in the development and production of therapeutic antibody products.
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna, Austria.
authors.forename: Beate
authors.initials: B
authors.lastname: Hintersteiner
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna, Austria.
authors.forename: Nico
authors.initials: N
authors.lastname: Lingg
authors.affiliation: Apeiron Biologics AG, Campus-Vienna-Biocenter 5, Vienna, Austria.
authors.forename: Evelyne
authors.initials: E
authors.lastname: Janzek
authors.affiliation: Apeiron Biologics AG, Campus-Vienna-Biocenter 5, Vienna, Austria.
authors.forename: Oliver
authors.initials: O
authors.lastname: Mutschlechner
authors.affiliation: Apeiron Biologics AG, Campus-Vienna-Biocenter 5, Vienna, Austria.
authors.forename: Hans
authors.initials: H
authors.lastname: Loibner
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna, Austria.
authors.forename: Alois
authors.initials: A
authors.lastname: Jungbauer
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000378:metabolism* / Q000494:pharmacology; D061067:Antibodies, Monoclonal, Humanized; D000920:Antibody-Dependent Cell Cytotoxicity; D000068258:Bevacizumab / Q000737:chemistry; D002152:Calorimetry, Differential Scanning; D045744:Cell Line, Tumor; D000068818:Cetuximab / Q000737:chemistry; D002852:Chromatography, Ion Exchange / Q000379:methods; D004355:Drug Stability; D015395:Histocompatibility Antigens Class I / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007074:Immunoglobulin G / Q000737:chemistry / Q000378:metabolism; D007525:Isoelectric Focusing; D011961:Receptors, Fc / Q000737:chemistry / Q000378:metabolism; D017452:Receptors, IgG / Q000737:chemistry / Q000378:metabolism; D020349:Surface Plasmon Resonance
chemical_list: D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; C496124:FCGR3A protein, human; D015395:Histocompatibility Antigens Class I; D007074:Immunoglobulin G; D011961:Receptors, Fc; D017452:Receptors, IgG; D000068258:Bevacizumab; C527517:ofatumumab; D000068818:Cetuximab; C100688:Fc receptor, neonatal
keywords: ADCC; Bio-better; Cation exchange chromatography; Effector functions; FcγRIIIa; Therapeutic mAbs