The availability of glucose to CHO cells affects the intracellular lipid-linked oligosaccharide distribution, site occupancy and the N-glycosylation profile of a monoclonal antibody

Liu, Bo; Spearman, Maureen; Doering, J. P.; Lattová, Erica; Perreault, Hélène; Butler, Michael

doi:10.1016/j.jbiotec.2013.11.007

Cited by 100 publications

(94 citation statements)

References 121 publications

(147 reference statements)

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“…[33] The branching and terminal residue of the N-glycan is influenced by cell culture conditions such as glucose limitation, dissolved oxygen and pH, with the exact effect being dependent on cell line, and glycoprotein. [34,35] The osmotic conditions and mechanical stress in cell culture do not seem to affect the N-glycan structure. [35] For a detailed review on approaches on how to control antibody glycosylation, the reader is encouraged to read the recent review by Sha et al [36] While N-glycosylation is by far the predominant form of glycosylation present in IgG, O-linked glycans can occasionally be found in the hinge region of IgG3.…”

Section: N-glycosylationmentioning

confidence: 99%

Microheterogeneity of Recombinant Antibodies: Analytics and Functional Impact

Beyer

Schuster

Jungbauer

et al. 2017

Biotechnology Journal

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Antibodies are typical examples of biopharmaceuticals which are composed of numerous, almost infinite numbers of potential molecular entities called variants or isoforms, which constitute the microheterogeneity of these molecules. These variants are generated during biosynthesis by so‐called posttranslational modification, during purification or upon storage. The variants differ in biological properties such as pharmacodynamic properties, for example, Antibody Dependent Cellular Cytotoxicity, complement activation, and pharmacokinetic properties, for example, serum half‐life and safety. Recent progress in analytical technologies such as various modes of liquid chromatography and mass spectrometry has helped to elucidate the structure of a lot of these variants and their biological properties. In this review the most important modifications (glycosylation, terminal modifications, amino acid side chain modifications, glycation, disulfide bond variants and aggregation) are reviewed and an attempt is made to give an overview on the biological properties, for which the reports are often contradictory. Even though there is a deep understanding of cellular and molecular mechanism of antibody modification and their consequences, the clinical proof of the effects observed in vitro and in vivo is still not fully rendered. For some modifications such as core‐fucosylation of the N‐glycan and aggregation the effects are clear and should be monitored, but with others such as C‐terminal lysine clipping the reports are contradictory. As a consequence it seems too early to tell if any modification can be safely ignored.

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Section: N-glycosylationmentioning

confidence: 99%

Microheterogeneity of Recombinant Antibodies: Analytics and Functional Impact

Beyer

Schuster

Jungbauer

et al. 2017

Biotechnology Journal

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“…Glucose limitations in a chemostat culture produced an increase in non-glycosylated gamma-interferon, which could be rapidly restored with pulsed glucose additions (Hayter et al 1992). Both reduced site-occupancy and truncation of N-glycans under low glucose conditions have been shown in several other recombinant glycoproteins and viral proteins (Davidson and Hunt 1985;Liu et al 2014;Seo et al 2014;Tachibana et al 1996).…”

Section: Culture Parameters Affecting Glycosylationmentioning

confidence: 95%

Glycosylation in Cell Culture

Spearman

Butler

2014

Cell Engineering

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Glycosylation affects many general functional factors of a glycoprotein, such as stability, inhibition of proteolysis, solubility, aggregation, as well as other attributes critical to its use as a biotherapeutic. Therefore our understanding of the glycosylation pathway, factors that affect it, and our ability to manipulate glycosylation are essential in producing superior biotherapeutics. The complex synthetic pathway of N-linked glycosylation occurs in both the endoplasmic reticulum and Golgi and involves a large number of precursors and enzymes, leading to a large array of possible glycan structures. Thus, variability in glycosylation may be influenced by numerous factors that affect this pathway, such as host cell type, nutrient levels and supplements, dissolved oxygen, pH, temperature, and by-product accumulation, and thus must be closely monitored. Better analytical techniques have allowed linking specific glycan structures to functionality of glycoproteins, which have led to efforts to modify glycosylation through genetic engineering, sequence-interfering RNA (siRNA), glycosylation inhibitors or chemoenzymatic modification.

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“…The absence of glucose affects the synthesis of oligosaccharide precursors, as glucose-starved cells alter its synthesis process 80 . In a CHO cell culture test it was shown that the proportion of nonglycosylated antibody was correlated to the duration the cells were deprived of glucose: the absence of glucose during 24 hours led to 45% of non-glycosylated mAb 81 . In another study with the human cell line rF2N78 in fed-batch culture it was observed that in the absence of glucose in the feed about 44% of the product was aglycosylated, while no aglycosylated antibody was expressed when feeding glucose throughout the culture 82 .…”

Section: Non Specific Impact Of Media Componentsmentioning

confidence: 99%

Tailoring recombinant protein quality by rational media design

Brühlmann

Jordan

Hemberger³

et al. 2015

Biotechnology Progress

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Clinical efficacy and safety of recombinant proteins are closely associated with their structural characteristics. The major quality attributes comprise glycosylation, charge variants (oxidation, deamidation, and C‐ & N‐terminal modifications), aggregates, low‐molecular‐weight species (LMW), and misincorporation of amino acids in the protein backbone. Cell culture media design has a great potential to modulate these quality attributes due to the vital role of medium in mammalian cell culture. The purpose of this review is to provide an overview of the way both classical cell culture medium components and novel supplements affect the quality attributes of recombinant therapeutic proteins expressed in mammalian hosts, allowing rational and high‐throughput optimization of mammalian cell culture media. A selection of specific and/or potent inhibitors and activators of oligosaccharide processing as well as components affecting multiple quality attributes are presented. Extensive research efforts in this field show the feasibility of quality engineering through media design, allowing to significantly modulate the protein function. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:615–629, 2015

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The availability of glucose to CHO cells affects the intracellular lipid-linked oligosaccharide distribution, site occupancy and the N-glycosylation profile of a monoclonal antibody

Cited by 100 publications

References 121 publications

Microheterogeneity of Recombinant Antibodies: Analytics and Functional Impact

Microheterogeneity of Recombinant Antibodies: Analytics and Functional Impact

Glycosylation in Cell Culture

Tailoring recombinant protein quality by rational media design

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