Control of galactosylated glycoforms distribution in cell culture system

McCracken, Neil A.; Kowle, Ronald; Ouyang, Anli

doi:10.1002/btpr.1906

Cited by 40 publications

(23 citation statements)

References 33 publications

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“…These studies have typically demonstrated the desired effect of facilitating an increase or decrease in enzyme activity, which resulted in the targeted manipulation of the final N‐glycan oligosaccharide profile. At the substrate level, selective media supplementation of nucleotide‐sugar precursors, as well as other media supplements have showed a good deal of promise . From the literature reported results, an overall conclusion can be made that protein glycosylation profiles are fairly sensitive toward the enzymes and substrates involved in the metabolic pathway.…”

Section: Introductionmentioning

confidence: 99%

Cell culture media supplementation of uncommonly used sugars sucrose and tagatose for the targeted shifting of protein glycosylation profiles of recombinant protein therapeutics

Hossler

McDermott

Racicot

et al. 2014

Biotechnology Progress

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Protein glycosylation is an important post-translational modification toward the structure and function of recombinant therapeutics. The addition of oligosaccharides to recombinant proteins has been shown to greatly influence the overall physiochemical attributes of many proteins. It is for this reason that protein glycosylation is monitored by the developer of a recombinant protein therapeutic, and why protein glycosylation is typically considered a critical quality attribute. In this work, we highlight a systematic study toward the supplementation of sucrose and tagatose into cell culture media for the targeted modulation of protein glycosylation profiles on recombinant proteins. Both sugars were found to affect oligosaccharide maturation resulting in an increase in the percentage of high mannose N-glycan species, as well as a concomitant reduction in fucosylation. The latter effect was demonstrated to increase antibody-dependent cell-mediated cytotoxicity for a recombinant antibody. These aforementioned results were found to be reproducible at different scales, and across different Chinese hamster ovary cell lines. Through the selective supplementation of these described sugars, the targeted modulation of protein glycosylation profiles is demonstrated, as well as yet another tool in the cell culture toolbox for ensuring product comparability.

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Section: Introductionmentioning

confidence: 99%

Cell culture media supplementation of uncommonly used sugars sucrose and tagatose for the targeted shifting of protein glycosylation profiles of recombinant protein therapeutics

Hossler

McDermott

Racicot

et al. 2014

Biotechnology Progress

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“…A change in pH from 6.9 to 7.4 was shown to influence the specific growth and mAb production rates as well as the glycosylation profile of IgG3 produced in hybridoma cells (Müthing et al, 2003). Finally, a multitude of chemical supplements added in either media or feeds have been described in the literature as effective glycosylation modulators and are summarized in detail in Table 1 (Bischoff, Liscum, & Kornfeld, 1986;Bruhlmann et al, 2017;Chang et al, 2007;Chen & Harcum, 2006;Crowell, Grampp, Rogers, Miller, & Scheinman, 2007;Foddy & Hughes, 1988;Gramer et al, 2011;Gramer, Goochee, Chock, Brousseau, & Sliwkowski, 1995;Green, Adelt, Baenziger, Wilson, & Van Halbeek, 1988; Gross et al, 1986;Gu & Wang, 1998;Hollister, Conradt, & Jarvis, 2003;Hossler et al, 2014;Jing, Qian, & Li, 2010;Jones et al, 2004;Kamińska, Dzięciol, & Kościelak, 1999;Keppler, Horstkorte, Pawlita, Schmidt, & Reutter, 2001;Kuntz, Zhong, Guo, Rose, & Boons, 2009;Li, 2014;McCracken, Kowle, & Ouyang, 2014;Mitchelson, Hughes, Mondia, & Hyde-Deruyscher, (2015); Pande, Rahardjo, Livingston, & Mujacic, 2015;Rillahan et al, 2012;Rodriguez, Spearman, Huzel, & Butler, 2005;Slade, Caspary, Nargund, & Huang, 2016;Tropea et al, 1990;Yin et al, 2017;Zhang et al, 2016). Sou et al evaluated the influence of a temperature shift from 36.5 to 32°C in the late exponential phase of a fed-batch experiment on the mAb Fc-glycan profile.…”

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confidence: 99%

Impact of cell culture media additives on IgG glycosylation produced in Chinese hamster ovary cells

Ehret

Zimmermann

Eichhorn

et al. 2019

Biotech & Bioengineering

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Glycosylation is a key critical quality attribute for monoclonal antibodies and other recombinant proteins because of its impact on effector mechanisms and half‐life. In this study, a variety of compounds were evaluated for their ability to modulate glycosylation profiles of recombinant monoclonal antibodies produced in Chinese hamster ovary cells. Compounds were supplemented into the cell culture feed of fed‐batch experiments performed with a CHO K1 and a CHO DG44 cell line expressing a recombinant immunoglobulin G1 (IgG1). Experiments were performed in spin tubes or the ambr®15 controlled bioreactor system, and the impact of the compounds at various concentrations was determined by monitoring the glycosylation profile of the IgG and cell culture parameters, such as viable cell density, viability, and titer. Results indicate that the highest impact on mannosylation was achieved through 15 µM kifunensine supplementation leading to an 85.8% increase in high‐mannose containing species. Fucosylation was reduced by 76.1% through addition of 800 µM 2‐F‐peracetyl fucose. An increase of 40.9% in galactosylated species was achieved through the addition of 120 mM galactose in combination with 48 µM manganese and 24 µM uridine. Furthermore, 6.9% increased sialylation was detected through the addition of 30 µM dexamethasone in combination with the same manganese, uridine, and galactose mixture used to increase total galactosylation. Further compounds or combinations of additives were also efficient at achieving a smaller overall glycosylation modulation, required, for instance, during the development of biosimilars. To the best of our knowledge, no evaluation of the efficacy of such a variety of compounds in the same cell culture system has been described. The studied cell culture media additives are efficient modulators of glycosylation and are thus a valuable tool to produce recombinant glycoproteins.

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“…In the same study, 20 mM GlcNAc addition effected a 4% decrease of galactosylated entities. In another test, the asparagine concentration influenced the distribution of G0F, G1F and G2F, likely due to altered enzyme activity caused by increased intracellular pH, where higher G0F and lower G1F and G2F emerged with increasing asparagine levels (supplementation of 0-10 mM Asn) 125 .…”

Section: Galactosylationmentioning

confidence: 97%

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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Control of galactosylated glycoforms distribution in cell culture system

Cited by 40 publications

References 33 publications

Cell culture media supplementation of uncommonly used sugars sucrose and tagatose for the targeted shifting of protein glycosylation profiles of recombinant protein therapeutics

Cell culture media supplementation of uncommonly used sugars sucrose and tagatose for the targeted shifting of protein glycosylation profiles of recombinant protein therapeutics

Impact of cell culture media additives on IgG glycosylation produced in Chinese hamster ovary cells

Tailoring recombinant protein quality by rational media design

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