title: A cytotoxic humanized anti-ganglioside antibody produced in a murine cell line defective of N-glycolylated-glycoconjugates.
abstract: Gangliosides containing the N-glycolyl (NGc) form of sialic acid are tumor-associated antigens and promising candidates for cancer therapy. We previously generated the murine 14F7 monoclonal antibody (mAb), specific for the N-glycolyl-GM3 ganglioside (NGcGM3), which induced an oncosis-like type of cell death on malignant cell lines expressing this antigen and recognized breast carcinoma by immunoscintigraphy in cancer patients. As humanization is expected to enhance its use for human cancer therapy, herein we describe the design and generation of two humanized versions of the 14F7 mAb by disrupting potential human T cell epitopes on its variable region. No differences in antigen reactivity or cytotoxic properties were detected among the variants tested and with respect to the chimeric counterpart. Humanized 14F7 genes were transfected into the NGcGM3-expressing NS0 cell line. Therefore, in the industrial scaling-up of the transfectoma in serum-free medium, cell viability was lost due to the cytotoxic effect of the secreted antibody. This shortcoming was solved by knocking down the CMP-N-acetylneuraminic acid hydroxylase enzyme, thus impairing the synthesis of NGc-glycoconjugates. Humanized 14F7 mAb is of potential value for the therapy of NGcGM3-expressing tumors.
authors.affiliation: Immunobiology Department, Center of Molecular Immunology, Havana, Cuba.
authors.forename: Yuniel
authors.initials: Y
authors.lastname: Fernández-Marrero
authors.forename: Lourdes
authors.initials: L
authors.lastname: Roque-Navarro
authors.forename: Tays
authors.initials: T
authors.lastname: Hernández
authors.forename: Denise
authors.initials: D
authors.lastname: Dorvignit
authors.forename: Marively
authors.initials: M
authors.lastname: Molina-Pérez
authors.forename: Addys
authors.initials: A
authors.lastname: González
authors.forename: Katya
authors.initials: K
authors.lastname: Sosa
authors.forename: Alejandro
authors.initials: A
authors.lastname: López-Requena
authors.forename: Rolando
authors.initials: R
authors.lastname: Pérez
authors.forename: Cristina Mateo
authors.initials: CM
authors.lastname: de Acosta
mesh_terms: D000230:Adenocarcinoma / Q000188:drug therapy* / Q000276:immunology / Q000473:pathology; D000818:Animals; D061067:Antibodies, Monoclonal, Humanized / Q000235:genetics / Q000378:metabolism; D000951:Antigens, Neoplasm / Q000276:immunology*; D001943:Breast Neoplasms / Q000188:drug therapy* / Q000276:immunology / Q000473:pathology; D003602:Cytotoxicity, Immunologic / Q000235:genetics; D018984:Epitopes, T-Lymphocyte / Q000235:genetics; D005260:Female; D005679:G(M3) Ganglioside / Q000031:analogs & derivatives / Q000276:immunology*; D006801:Humans; D006825:Hybridomas; D007167:Immunotherapy; D051379:Mice; D006899:Mixed Function Oxygenases / Q000235:genetics; D058990:Molecular Targeted Therapy; D016297:Mutagenesis, Site-Directed; D015202:Protein Engineering; D034741:RNA, Small Interfering / Q000235:genetics
chemical_list: D061067:Antibodies, Monoclonal, Humanized; D000951:Antigens, Neoplasm; D018984:Epitopes, T-Lymphocyte; D005679:G(M3) Ganglioside; D034741:RNA, Small Interfering; D006899:Mixed Function Oxygenases; C074383:CMPacetylneuraminate monooxygenase