Case Study: Can Cell-Ess® be used to increase consistency of protein quality in
CHO cell monoclonal antibody production?
Adam Elhofy, PhD, Chief Science Officer, Essential Pharmaceuticals, LLC
Introduction
The most common method to extend culture times and increase titer during bioproduction is to employ
a feed strategy during the batch run. The method, called fed-batch, can employ a variety of feeds to add
back nutrients that were utilized by the cells in the bioreactor. Including lipids as part of a feed regimen
can be beneficial if supplied throughout the bioproduction run. Cells need lipids for membrane
formation (for example, cellular membrane, Golgi apparatus, and endoplasmic reticulum (ER). Healthy,
functional membranes are critical for protein synthesis and post-translational modification. Posttranslational modifications such as glycosylation have a direct effect on protein quality and therefore
therapeutic effectiveness. The Golgi apparatus is the primary location for post-translational
modification, while the ER is where synthesis and lower forms of glycosylation occur. The Golgi
apparatus has a complex membrane with a high amount of cholesterol. Previously, fetal bovine serum
added to media acted as a source and delivery mechanism of lipids to cells. However, the shift of the
bioproduction industry toward serum-free media has removed that source of lipids. Cell-Ess® universal
titer boost and optimizer provides key nutrients and lipids to media enabling a greater level of control
over post-translation modification and consistency while simultaneously increasing titer.
The Challenge
Increase reproducibility of glycosylation patterns and quality features for CHO cell monoclonal antibody
production.
ambr systems (sold by Sartorius Stedim Biotech) can be used as a model for large-scale bioreactors. At
the small scale, feeds can be tested economically at various concentrations and feeding schedules to
determine the optimum feed strategy for individual clones before they are scaled up to a bioreactor for
production. In previous case studies, Cell-Ess was shown to increase productivity in cells and overall
titer when added as an initial supplement (1).
In this case study, we investigate whether Cell-Ess can be added as a feed to improve reproducibility of
protein quality measure through glycosylation patterns.
Method
To test our hypothesis, we modeled a fed-batch system with the ambr system. We used 5 different feed
strategies in duplicate with Cell-Ess. The control vessel did not receive any Cell-Ess addition. All groups
fed with Cell-Ess had an increase in monoclonal antibody yield compared to control. The monoclonal
antibody from each Cell-Ess group and the control group was purified and then analyzed for the glycan
profile. The glycan profile was then analyzed by Prism (sold by GraphPad Software, Inc.) for standard
error around the mean looking for variability.
Results
There was a significant increase in consistency, demonstrated by less variability, with the use of Cell-Ess
regardless of the feed strategy tested (Figure 1). In addition, for all feeds tested, there was an increase
in the yield of monoclonal antibody with the greatest yield increase occurring with 5% of Cell-Ess fed 3
times. These results suggest that Cell-Ess can improve reproducibility in product quality in addition to
increasing titer. One of the important features companies are looking for is the level of more complex
glycosylation patterns. The group with more complex glycoforms is represented as galactose in Figure 1.
There is significantly less variation in the higher order glycoforms when using Cell-Ess as a feed.
SEM
0.6
Control
Cell-Ess
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Figure 1. Glycan profiles were measured for their variability with and without treatment with Cell-Ess.
There is less variability as measured by the SEM (standard error of mean) in 4 out of the 6 groups with
variability of approximately 0.1.
Conclusions
Cell-Ess has been used to increase titer and yield when used as a feed. Possible reasons for increased
productivity are: (1) there are better energetics in the systems, allowing for more energy for protein
synthesis, and (2) there is improved membrane function, including in the ER and Golgi apparatus, where
synthesis and post-translational modification occur. In this study, we demonstrate that there is more
consistent protein quality as shown by less variability in the glycolytic patterns from monoclonal
antibodies, demonstrating the membrane complexity of the Golgi apparatus was improved. A higher
functioning Golgi apparatus would lead to more consistent quality attributes from run to run. This study
demonstrated the utility of having a tool such as Cell-Ess to reduce protein quality variability while
increasing monoclonal antibody titer in a fed-batch system. In further studies, we will investigate the
use of Cell-Ess as a feed to increase higher order glycoforms measured by increase galactosylation.
Reference
1
Poster: Novel Lipid Supplement Enhances Protein Production in Small Bioreactors. Adam Elhofy, Ph.D.,
Essential Pharmaceuticals, Ewing, NJ.
Phone: (844) Ess Prod (377-7763)
Email: [email protected]