Abstract
The terminal sugar molecules of the N-linked glycan attached to the fragment crystalizable (Fc) region is a critical quality attribute of therapeutic monoclonal antibodies (mAbs) such as immunoglobulin G (IgG). There exists naturally-occurring heterogeneity in the N-linked glycan structure of mAbs, and such heterogeneity has a significant influence on the clinical safety and efficacy of mAb drugs. We previously proposed a constraint-based modeling method called glycosylation flux analysis (GFA) to characterize the rates (fluxes) of intracellular glycosylation reactions. One contribution of this work is a significant improvement in the computational efficiency of the GFA, which is beneficial for analyzing large datasets. Another contribution of our study is the analysis of IgG glycosylation in continuous perfusion Chinese Hamster Ovary (CHO) cell cultures. The GFA of the perfusion cell culture data indicated that the dynamical changes of IgG glycan heterogeneity are mostly attributed to alterations in the galactosylation flux activity. By using a random forest regression analysis of the IgG galactosylation flux activity, we were further able to link the dynamics of galactosylation with two process parameters: cell-specific productivity of IgG and extracellular ammonia concentration. The characteristics of IgG galactosylation dynamics agree well with what we previously reported for fed-batch cultivations of the same CHO cell strain.
Highlights
Therapeutic recombinant monoclonal antibodies constitute the most important class of drugs in the biopharmaceutical industry, making up approximately half of the total revenue of biopharmaceutical products in 2013 [1]
We improved the computational efficiency of the glycosylation flux analysis (GFA) and applied the GFA to analyze the immunoglobulin G (IgG) glycosylation of four perfusion Chinese Hamster Ovary (CHO) cell culture experiments
We improved the computational efficiency of the GFA and applied the GFA to a stoichiometric model of the IgG glycosylation network to estimate IgG intracellular glycosylation analyze the IgG glycosylation of four perfusion CHO cell culture experiments
Summary
Therapeutic recombinant monoclonal antibodies constitute the most important class of drugs in the biopharmaceutical industry, making up approximately half of the total revenue of biopharmaceutical products in 2013 [1]. The state-of-the-art batch/fed-batch cell cultures are able to meet the large production volume requirement of mAbs, with reactor sizes of up to 25,000 L [2]. The application of continuous cell culture technology has far been limited to the production of unstable products that require constant recovery [7]. Whether or not product qualities and their key controlling parameters can be directly translated from the traditional batch/fed-batch cell cultures to the perfusion cell cultivations is still unresolved. Past work on converting mAb production from batch/fed-batch to perfusion cell cultures gave conflicting reports on product qualities, where a few studies demonstrated consistent product qualities [9,10], and many others showed differences between the two production modes [11,12,13,14,15]
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