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Short‐hairpin‐RNA‐mediated silencing of fucosyltransferase 8 in Chinese‐hamster ovary cells for the production of antibodies with enhanced antibody immune effector function
Author(s) -
Beuger Vincent,
Künkele KlausPeter,
Koll Hans,
Gärtner Achim,
Bähner Monika,
Burtscher Helmut,
Klein Christian
Publication year - 2009
Publication title -
biotechnology and applied biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.468
H-Index - 70
eISSN - 1470-8744
pISSN - 0885-4513
DOI - 10.1042/ba20080220
Subject(s) - chinese hamster ovary cell , microbiology and biotechnology , antibody , fucosyltransferase , biology , receptor , biochemistry , gene , immunology
Antibody‐producing Chinese‐hamster ovary cells (CHO‐DG44) were converted into cells producing antibodies with strongly enhanced ADCC (antibody‐dependent cellular cytotoxicity) by knocking down FuT8 (α‐1,6‐fucosyltransferase or fucosyltransferase 8) via constitutive expression of shRNA (short‐hairpin RNA) against FuT8. After the introduction of a FuT8 shRNA expression plasmid under the control of a U6 promoter, CHO‐DG44 clones with less than 5% residual FuT8 mRNA expression were isolated by selection for neomycin resistance, followed by low affinity nerve growth factor receptor enrichment and selection for LCA [ Lens culinaris (culinary lentil) agglutinin] resistance. The CHO‐DG44 clones identified produced highly afucosylated anti‐[IGF‐1R (insulin‐like‐growth‐factor‐1 receptor)] antibodies (up to 88%) that exhibited considerably enhanced ADCC compared with anti‐IGF‐1R wild‐type antibodies produced by parental CHO cells. At the same time, antibody productivity was not significantly decreased. Analysis of stability showed that the clones obtained may be suitable for up‐scaling, since low residual levels of FuT8 mRNA and production of afucosylated antibodies were maintained for at least 4 weeks.