Establishment of FUT8 gene knockout CHO cell line with stable expression of monoclonal antibody

Abstract

Abstract Background: Afucosylated antibodies have been become increasingly popular in clinical trials for therapeutic purposes owing to the improved efficacy compared to the fucosylated counterparts. The elimination of fucose on the heavy chain of an antibody can enhance the activity of antibody-dependent cellular cytotoxicity (ADCC). However, antibodies produced by Chinese hamster ovary (CHO) cells are heavily fucosylated, and the modification reduces the activity of ADCC. FUT8,GMD and FX encoding major fucose modification enzymes, which were knockout respectively in CHO cells using clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9). These gene knockout cell lines were evaluated for cell growth, antibody yield, N-glycan profiles and ADCC activities. Results: The results showed that the gene knockout cell lines exhibited morphology and growth kinetics similar to those of the wild CHO cells, and produced completely afucosylated recombinant antibodies. Among them, the ADCC activity of cell pool and monoclonal cell line with FUT8 gene knocked out (FUT8KO) were increased 2-2.5-fold comparing to conventional fucosylated antibodies. Conclusions: Our results indicated that FUT8KO monoclonal clone cell lines (#08E9, 05E9, 20D8, 12C4, 01E9, and 01F4) were ideal hosts for stable production of completely afucosylated high-ADCC activity antibodies, with considerable therapeutic quality and efficacy.