title: A multi-pronged investigation into the effect of glucose starvation and culture duration on fed-batch CHO cell culture.
abstract: In this study, omics-based analysis tools were used to explore the effect of glucose starvation and culture duration on monoclonal antibody (mAb) production in fed-batch CHO cell culture to gain better insight into how these parameters can be controlled to ensure optimal mAb productivity and quality. Titer and N-glycosylation of mAbs, as well as proteomic signature and metabolic status of the production cells in the culture were assessed. We found that the impact of glucose starvation on the titer and N-glycosylation of mAbs was dependent on the degree of starvation during early stationary phase of the fed-batch culture. Higher degree of glucose starvation reduced intracellular concentrations of UDP-GlcNAc and UDP-GalNAc, but increased the levels of UDP-Glc and UDP-Gal. Increased GlcNAc and Gal occupancy correlated well with increased degree of glucose starvation, which can be attributed to the interplay between the dilution effect associated with change in specific productivity of mAbs and the changed nucleotide sugar metabolism. Herein, we also show and discuss that increased cell culture duration negatively affect the maturation of glycans. In addition, comparative proteomics analysis of cells was conducted to observe differences in protein abundance between early growth and early stationary phases. Generally higher expression of proteins involved in regulating cellular metabolism, extracellular matrix, apoptosis, protein secretion and glycosylation was found in early stationary phase. These analyses offered a systematic view of the intrinsic properties of these cells and allowed us to explore the root causes correlating culture duration with variations in the productivity and glycosylation quality of monoclonal antibodies produced with CHO cells.
authors.affiliation: Network Engineering of Eukaryotic Cell Factories, Department of Systems Biology, Technical University of Denmark, Building 223, 2800 Kgs. Lyngby, Denmark.|Symphogen A/S, Pederstrupvej, 93, 2750, Ballerup, Denmark.
authors.forename: Yuzhou
authors.initials: Y
authors.lastname: Fan
authors.affiliation: Center for Process Systems Engineering, Department of Chemical Engineering, Imperial College London, South Kensington Campus, London, SW7 2AZ, UK.
authors.forename: Ioscani
authors.initials: I
authors.lastname: Jimenez Del Val
authors.affiliation: Symphogen A/S, Pederstrupvej, 93, 2750, Ballerup, Denmark.
authors.forename: Christian
authors.initials: C
authors.lastname: Müller
authors.affiliation: Network Engineering of Eukaryotic Cell Factories, Department of Systems Biology, Technical University of Denmark, Building 223, 2800 Kgs. Lyngby, Denmark.
authors.forename: Anne Mathilde
authors.initials: AM
authors.lastname: Lund
authors.affiliation: Symphogen A/S, Pederstrupvej, 93, 2750, Ballerup, Denmark.
authors.forename: Jette Wagtberg
authors.initials: JW
authors.lastname: Sen
authors.affiliation: Symphogen A/S, Pederstrupvej, 93, 2750, Ballerup, Denmark.
authors.forename: Søren Kofoed
authors.initials: SK
authors.lastname: Rasmussen
authors.affiliation: Center for Process Systems Engineering, Department of Chemical Engineering, Imperial College London, South Kensington Campus, London, SW7 2AZ, UK.
authors.forename: Cleo
authors.initials: C
authors.lastname: Kontoravdi
authors.affiliation: Department of Chemical and Biomolecular Engineering, Johns Hopkins University, Baltimore, Maryland 21218.
authors.forename: Deniz
authors.initials: D
authors.lastname: Baycin-Hizal
authors.affiliation: Department of Chemical and Biomolecular Engineering, Johns Hopkins University, Baltimore, Maryland 21218.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Betenbaugh
authors.affiliation: Symphogen A/S, Pederstrupvej, 93, 2750, Ballerup, Denmark. dw@symphogen.com.
authors.forename: Dietmar
authors.initials: D
authors.lastname: Weilguny
authors.affiliation: Network Engineering of Eukaryotic Cell Factories, Department of Systems Biology, Technical University of Denmark, Building 223, 2800 Kgs. Lyngby, Denmark. mr@bio.dtu.dk.
authors.forename: Mikael Rørdam
authors.initials: MR
authors.lastname: Andersen
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000235:genetics; D016466:CHO Cells; D018929:Cell Culture Techniques / Q000379:methods*; D003412:Cricetulus; D003470:Culture Media / Q000737:chemistry*; D005947:Glucose / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D003470:Culture Media; D011994:Recombinant Proteins; D005947:Glucose
keywords: chinese hamster ovary cells; culture duration; fed-batch; glucose starvation; glycosylation; monoclonal antibody; proteomics