Establishment of a GDP-mannose 4,6-dehydratase (GMD) knockout host cell line: A new strategy for generating completely non-fucosylated recombinant therapeutics

Roles of Pofut1 and O-Fucose in Mammalian Notch Signaling

“…Although it is clear that O-fucose on Notch is an essential substrate of Fringe (8,9), Fringe-independent Notch signaling defects are observed in Drosophila Ofut1 mutants (12,13), indicative of signaling functions that are regulated solely by O-fucose on Notch. This is consistent with the reduced Jagged1-induced Notch signaling observed in Lec13 CHO cells that have very low GDP-fucose (32)(33)(34)(35) and little Fringe activity (8,15) and with rescue of Notch signaling by exogenous fucose or genetic complementation of the Lec13 fucosylation defect (8,15).…”

“…Pro Ϫ 5Lec1.3C CHO cells (42) with the vector pMIRB are equivalent to parent CHO in Notch signaling activity (8,15) and are referred to as Lec1. Lec13.6A CHO cells were previously characterized as fucosylation-defective (32)(33)(34)(35) and are referred to as Lec13. For maximum expression of the Lec13 phenotype, Lec13 cells were cultured in ␣-modified minimal essential medium and 10% dialyzed fetal bovine serum (Gemini).…”

“…These papers reported no detectable Gmd transcripts or GDP-fucose in Lec13 cells, but a recent analysis found that Lec13 has about 3% of the GDP-fucose levels in CHO cells (35). Since Lec13 cells are rescued by exogenous fucose added to the culture medium (32), culture conditions affect the level of fucosylation.…”

EB66 cell line, a duck embryonic stem cell-derived substrate for the industrial production of therapeutic monoclonal antibodies with enhanced ADCC activity

“…[22][23][24] An alternative strategy is the experimental knockdown in CHO cells of two key genes involved in oligosaccharide fucose modification, a1,6-fucosyltransferase (FUT8) and GDP-mannose 4,6-dehydratase (GMD). [25][26][27] Several preclinical and clinical trials using nonfucosylated antibodies generated with such engineered CHO cell lines are currently underway.…”

Cell Culture Processes in Monoclonal Antibody Production

“…RNAi and gene deletion technologies have also been used to decrease or eliminate the fucose on antibodies to dramatically increase ADCC activity. 27,28 In general, glycosylation is difficult to control precisely in mammalian cells as it is dependent on a variety of factors such as clonal variations, media, as well as culture conditions. CHO stable cell lines have often been selected using metabolic selective markers including methotrexate (MTX) (dihydrofolate reductase gene mediated) and methionine sulphoximine (MSX) (glutamine synthetase gene mediated).…”