A Kinetic Model for Illustration of the Effect of Glutamine on the Apoptotic and Synthesis Pathways of Hybridoma Cells
Denizhan Yilmaz, Satish J. Parulekar, and Ali Cinar
Department of Chemical and Biological Engineering
Illinois Institute of Technology, Chicago, IL 60616
(312) 567-3044 (voice), (312) 567-8874 (fax), firstname.lastname@example.org
Monoclonal antibodies (MABs) are important reagents used in biomedical research, in diagnosis and treatment of diseases. Monoclonal antibodies (MABs) derived from hybridoma cell cultures are used extensively in diagnostic assays and have found increasing use in therapeutic applications, affinity production systems, and in vivo imaging. The cell lines, hybridoma, are produced by fusing B cells (lymphocytes) from immunized animals with myeloma cells. The simplest approach for producing a MAB in vitro is to grow the hybridoma cells in batch and fed-batch cultures and recover and purify the MAB from the culture medium. The large scale production of MABs occurs via in vitro cultivation of hybridoma cell lines in bioreactors using techniques similar to those used for microbial cultivation. A cost-effective production of MABs requires an understanding of the effects of bioreactor process variables on the physiology of hybridoma cells.
The demand for MAbs has increased substantially over the last two decades due to their therapeutic and diagnostic applications. The development of methods for optimization of cell growth and cell productivity has become a crucial issue to enhance MAb yield in vitro production. The efficiency and performance of cell cultures depend on nutrient medium optimization and process monitoring and control strategies. Creating these strategies requires understanding of process dynamics affected by cell mechanism in culture environments. Mathematical models have been significant tools for combining cell physiology and engineering to predict the behavior of cell metabolism and optimize culture conditions by identifying process parameters that significantly impact cell growth and target metabolite productivity. In this work, we develop a hybrid model for antibody synthesis in hybridoma cell cultures. The kinetic model is comprised of an unstructured kinetic representation cell growth and death, uptake of key nutrients, glucose and glutamine, and accumulation of end-products of cell metabolism, lactate and ammonia and a structured kinetic representation of MAb synthesis. The structured portion of the hybrid model describes secretion pathways throughout cell growth and product synthesis considering the effect of glutamine depletion and ammonia accumulation on the antibody synthesis rate. Glutamine provides remarkable metabolic energy for cell growth and protein synthesis and is an important precursor of proteins and peptides, as well as amino sugars, purines and pyrimidines. It has been observed that glutamine depletion in the culture greatly influences the overall antibody synthesis rate and increases the apoptotic death rates of mammalian cells. When extracellular glutamine concentration is higher than a minimum required level, the translation rates and stability of mRNAs are high, leading to maximization of MAb synthesis rate per cell. The lack of glutamine in the culture medium activates a self-digestion system called autophagy for cellular energy. After autophagy period, cells enter a highly controlled and genetically defined terminal path known as apoptosis. Glutamine exhaustion increases cell death rate as per the programmed cell death pathway. Moreover, although glutamine is an apoptosis suppressor, feeding glutamine to the culture after cells entered the apoptotic pathway does not reduce cell death rate. Performance of batch, perturbed batch (intermittent addition of glucose and/or glutamine), and fed-batch cultures is simulated. The model predictions are in good agreement with experimental data reported in the literature for Immunoglobulin G (IgG) antibodies.