title: Antitumor Efficacy of Anti-GD2 IgG1 Is Enhanced by Fc Glyco-Engineering.
abstract: The affinity of therapeutic antibodies for Fcγ receptors (FcγRs) strongly influences their antitumor potency. To generate antibodies with optimal binding and immunologic efficacy, we compared the affinities of different versions of an IgG1 Fc region that had an altered peptide backbone, altered glycans, or both. To produce IgG1 with glycans that lacked α1,6-fucose, we used CHO cells that were deficient in the enzyme UDP-N-acetylglucosamine: α-3-d-mannoside-β-1,2-N-acetylglucosaminyltransferase I (GnT1), encoded by the MGAT1 gene. Mature N-linked glycans require this enzyme, and without it, CHO cells synthesize antibodies carrying only Man5-GlcNAc2, which were more effective in antibody-dependent cell-mediated cytotoxicity (ADCC). Our engineered IgG1, hu3F8-IgG1, is specific for GD2, a neuroendocrine tumor ganglioside. Its peptide mutant is IgG1-DEL (S239D/I332E/A330L), both produced in wild-type CHO cells. When produced in GnT1-deficient CHO cells, we refer to them as IgG1n and IgG1n-DEL, respectively. Affinities for human FcγRs were measured using Biacore T-100 (on CD16 and CD32 polymorphic alleles), their immunologic properties compared for ADCC and complement-mediated cytotoxicity (CMC) in vitro, and pharmacokinetics and antitumor effects were compared in vivo in humanized mice. IgG1n and IgG1n-DEL contained only mannose and acetylglucosamine and had preferential affinity for activating CD16s, over inhibitory CD32B, receptors. In vivo, the antitumor effects of IgG1, IgG1-DEL, and IgG1n-DEL were similar but modest, whereas IgG1n was significantly more effective (P < 0.05). Thus, IgG1n antibodies produced in GnT1-deficient CHO cells may have potential as improved anticancer therapeutics. Cancer Immunol Res; 4(7); 631-8. ©2016 AACR.
authors.affiliation: Department of Pediatrics, Memorial Sloan Kettering Cancer Center, New York, New York.
authors.forename: Hong
authors.initials: H
authors.lastname: Xu
authors.affiliation: Department of Pediatrics, Memorial Sloan Kettering Cancer Center, New York, New York.
authors.forename: Hongfen
authors.initials: H
authors.lastname: Guo
authors.affiliation: Department of Pediatrics, Memorial Sloan Kettering Cancer Center, New York, New York.
authors.forename: Irene Y
authors.initials: IY
authors.lastname: Cheung
authors.affiliation: Department of Pediatrics, Memorial Sloan Kettering Cancer Center, New York, New York. cheungn@mskcc.org.
authors.forename: Nai-Kong V
authors.initials: NK
authors.lastname: Cheung
mesh_terms: D000818:Animals; D000915:Antibody Affinity / Q000276:immunology; D000920:Antibody-Dependent Cell Cytotoxicity / Q000276:immunology; D000074322:Antineoplastic Agents, Immunological / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology*; D016466:CHO Cells; D045744:Cell Line, Tumor; D003412:Cricetulus; D004195:Disease Models, Animal; D005732:Gangliosides / Q000037:antagonists & inhibitors* / Q000276:immunology / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000276:immunology* / Q000378:metabolism; D007074:Immunoglobulin G / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology*; D007697:Kinesics; D051379:Mice; D018345:Mice, Knockout; D009369:Neoplasms / Q000188:drug therapy / Q000276:immunology* / Q000473:pathology; D011485:Protein Binding; D023041:Xenograft Model Antitumor Assays
chemical_list: D000074322:Antineoplastic Agents, Immunological; D005732:Gangliosides; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; C019403:ganglioside, GD2