Novel Dynamic Model to Predict the Glycosylation Pattern of Monoclonal Antibodies from Extracellular Cell Culture Conditions Ohadi Kaveh* Aghamohseni Hengameh** Gädke Johannes*** Moo-Young Murray**** Legge Raymond L.***** Scharer Jeno****** Budman Hector M.******* Chemical Engineering Department, University of Waterloo, Waterloo, Ontario, Canada * e-mail: skohadi@uwaterloo.ca, ** e-mail: h3aghamo@uwaterloo.ca, *** e-mail: j.gaedke@tu-bs.de, **** e-mail: mooyoung@uwaterloo.ca, ***** e-mail: rllegge@uwaterloo.ca ****** e-mail: jscharer @uwaterloo.ca, ******* e-mail: hbudman@uwaterloo.ca Abstract: Glycosylation is a critical protein post-translational modification with a profound impact on the therapeutic properties of Mab and research indicates that it depends on extracellular culture conditions. A novel dynamic model was developed to relate extracellular metabolites’ concentrations to a cumulative glycoprofile. The model has three components: dynamic evolution of extracellular metabolites, production of nucleotide sugars in the cytosol, and glycosylation inside the Golgi apparatus. Following comparisons with experimental data obtained from batch CHO cell cultures, the model was found capable of predicting the glycoform profile of Mab temporally, as well as the extent of galactosylation given in the form of galactosylation index. The model has the potential for use in controlling the glycoform profile by manipulating culture conditions. Keywords: Dynamic modeling, Biotechnology, Glycosylation Mammalian cell culture Monoclonal antibody 1. INTRODUCTION Monoclonal antibodies (Mabs) comprise the dominant products in the fastest growing segment of the biopharmaceutical market. Chinese hamster ovary (CHO) cells are widely used as hosts for Mab production. Several studies have reported the profound impact of glycosylation, as post-translational modification, on Mabs’ therapeutic properties. The degree and extent of glycosylation can be quantified in terms of the type and number of nucleotide sugars attached to the Mab (Durocher and Butler, 2009). N-linked glycosylation, the most common form of glycosylation in mammalian cells, is initiated in the endoplasmic reticulum (ER) by a covalent attachment of a sugar to the polypeptide chain that is followed by proper folding of the protein and removal of three glucose molecules and at least one mannose. The resulting glycoprotein is then transferred into the Golgi apparatus where the final glycosylation process takes place (Del Val et al., 2011, Hossler et al., 2007, Stanley, 2011). It has been reported (Del Val et al., 2011, Hossler et al., 2009) and experimentally observed by the authors that culture conditions can be manipulated to obtain a specific oligosaccharide (OS) structure attached to the Mab. Accordingly, a mathematical model that could describe the effect of culture conditions on glycosylation may be instrumental for manufacturing a Mab with a desired glycoprofile. Such a model should be able to relate the extracellular culture conditions to intracellular glycosylation mechanisms at the cell level. To the knowledge of the authors, no such model has yet been developed. Quality by Design (QbD) is a recent FDA initiative that promotes a systematic approach to drug development, emphasizing that product quality considerations should be introduced at the design stage of the manufacturing process. Predictive mathematical models, such as the one proposed in this study, have been identified as key for designing novel manufacturing operations within the QbD framework (Hossler et al., 2009). Fig. 1. Schematic representation of the comprehensive- model: Metabolic Flux Analysis (left), Nucleotide Sugar (centre), Glycosylation based on Golgi Maturation (right). In the current work a specifically engineered CHO cell line, provided by MabNet (National Science and Engineering Research of Canada Mab Network), has been used to produce glycosylated Mab. A comprehensive model is developed for this cell line that is composed of three main parts as shown in Fig. 1. The first part consisting of a metabolic flux analysis (MFA) based model is developed to describe changes in MFA Model Nucleotide Sugar Model Glycosylation Model Preprints of the 12th IFAC Symposium on Computer Applications in Biotechnology The International Federation of Automatic Control 16-18, 2013, December. Mumbai, India Copyright © 2013 IFAC 36