(157o) A Dynamic Elementary Flux Mode-Based Model for Antibody Producing Cell Lines and Model Based Evaluation of Fed-Batch Cultures

Monoclonal antibodies (MAbs) are significant reagents used extensively in diagnostic assays, therapeutic applications, and in vivo imaging. Mammalian cell cultures have become the favored production hosts for MAbs, since microbial systems are not able to carry out the complex post-translational and functional modifications of these proteins, such as glycosylation. Chinese Hamster Ovary (CHO) cells and hybridoma cells, which share similar metabolic characteristics, have been popular cell types for production of MAbs. The efficient performance of these cell cultures requires highly specialized culture media to enhance MAb yield for in vitro production in view of substantial cell death and reduced MAb productivity due to the variations in culture conditions. Although production practices have been employed for decades, cell kinetics is still under investigation to obtain quantitatively as well as qualitatively cost-effective production strategies. Creating these strategies requires understanding of cell metabolism affected by process dynamics in culture environments. Kinetic models empower us to illustrate quantitative cell growth and metabolic activity, which allows prediction of different cell phenotypes and provides better understanding of cell physiology, which is important in optimization of MAb production in animal cell cultures.

Based on elementary flux mode (EFM) analysis, a novel approach is presented for monoclonal antibody (MAb) production by GS-CHO cells. An EFM-based kinetic model is developed on the basis of a set of macro-reactions, which can predict the time-dependent concentrations of metabolites, cell growth, and MAb productivity over a range of culture conditions. The rate of each macro-reaction through the central carbon metabolism contributes to ATP production. Energy metabolism and availability of substrates determine the specific biomass and MAb synthesis rates. The model provides a framework to predict different cell phenotypes under different culture conditions with a reduced metabolic network comprised of 23 macro-reactions. The model incorporates energy metabolism with biomass and MAb formation, with the specific ATP production rate being decided by the central carbon metabolism and used for estimation of biomass and MAb synthesis rates. The reaction rate expressions are represented by Michaelis-Menten kinetics based on extracellular metabolite concentrations, which determine ATP production by glycolysis and respiratory chain. Glutamine and asparagine are considered as regulatory metabolites for GS-CHO cells. Glutamine determines asparagine utilization route and energetic state of cells, while asparagine regulates the uptake rates of aspartate and glutamate. Glutamine is considered as a regulatory metabolite for GS-CHO cell lines - its variation in the culture influences cell metabolism and growth. With glutamine-free media, asparagine is the best alternative amino acid for GS-CHO cell lines. The model was calibrated for glutamine-free and glutamine-available cases and validated for fed-batch cultures supplied with glutamine. Un-optimized fed-batch cultures have been simulated considering a constant feeding strategy and a daily feeding strategy. Higher viable cell accumulation was observed in constant fed-batch operation, due to suppression of lactate formation. The model predictions are in good agreement with the experimental data reported in literature. The proposed model can be used for design of optimal nutrient media and optimization and control of fed-batch cultures.