Production, characterization and pharmacokinetic properties of antibodies with N-linked Mannose-5 glycans
| Autor: | Chae Janeka Reed, Jeff Lutman, Darren Brown, Robert Bayer, Shan Chung, Anne Wong, Eric Stefanich, Marcella Yu, Jean-Philippe Stephan |
|---|---|
| Rok vydání: | 2012 |
| Předmět: |
Mannosidase
Spectrometry Mass Electrospray Ionization Glycan Glycosylation Cell Survival Metabolic Clearance Rate Immunology Mice Nude Mannose Enzyme-Linked Immunosorbent Assay CHO Cells Binding Competitive Mice chemistry.chemical_compound Alkaloids Cricetulus Polysaccharides Cell Line Tumor Cricetinae Report Mannosidases Animals Humans Immunology and Allergy Cells Cultured Antibody-dependent cell-mediated cytotoxicity biology Receptors IgG Antibody-Dependent Cell Cytotoxicity Antibodies Monoclonal Molecular biology In vitro Immunoglobulin Fc Fragments Biochemistry chemistry Kifunensine Area Under Curve biology.protein Female Antibody |
| Zdroj: | mAbs. 4:475-487 |
| ISSN: | 1942-0870 1942-0862 |
| Popis: | The effector functions of therapeutic antibodies are strongly affected by the specific glycans added to the Fc domain during post-translational processing. Antibodies bearing high levels of N-linked mannose-5 glycan (Man5) have been reported to exhibit enhanced antibody-dependent cell-mediated cytotoxicity (ADCC) compared with antibodies with fucosylated complex or hybrid glycans. To better understand the relationship between antibodies with high levels of Man5 and their biological activity in vivo, we developed an approach to generate substantially homogeneous antibodies bearing the Man5 glycoform. A mannosidase inhibitor, kifunensine, was first incorporated in the cell culture process to generate antibodies with a distribution of high mannose glycoforms. Antibodies were then purified and treated with a mannosidase for trimming to Man5 in vitro. This 2-step approach can consistently generate antibodies with > 99% Man5 glycan. Antibodies bearing varying levels of Man5 were studied to compare ADCC and Fcγ receptor binding, and they showed enhanced ADCC activity and increased binding affinity to the FcγRIIIA. In addition, the clearance rate of antibodies bearing Man8/9 and Man5 glycans was determined in a pharmacokinetics study in mice. When compared with historical data, the antibodies bearing the high mannose glycoform exhibited faster clearance rate compared with antibodies bearing the fucosylated complex glycoform, while the pharmacokinetic properties of antibodies with Man8/9 and Man5 glycoforms appeared similar. In addition, we identified the presence of a mannosidase in mouse serum that converted most Man8/9 to Man6 after 24 h. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|