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

Abstract

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.

A Markov chain model for N-linked protein glycosylation – towards a low-parameter tool for model-driven glycoengineering

“…Finally, numerous previous studies have demonstrated the impact of media on glycosylation (Brühlman et al, 2015; Hossler et al, 2014; Surve and Gadgil, 2015). Thus, it will be of great value to integrate our model with models of other metabolic pathways.…”

Arabinosylation of recombinant human immunoglobulin-based protein therapeutics

“…From CHO cell line 1, mAb-1 samples were generated through cell culture media supplementation of D-arabinose, sucrose, as well as a glucose-only experimental control condition. Sucrose (70 mM) was supplemented into both the basal and feed medias to facilitate production of large quantities of high mannose N-glycans, 17 to contrast with the reduced levels of high mannose N-glycans facilitated through the use of D-arabinose. A summary of the N-glycoform profile for mAb-1 generated for the purposes of this testing is shown in Table 3.…”

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

“…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.…”