A comparative study of cell growth and adenovirus production using suspension and stationary cell culture

Abstract

The growing demand of adenoviral vectors for gene therapy, especially for cancer treatment, creates the need for large scale production of these vectors [2, 4]. In order to do this, characterization and optimization of culture conditions for HEK293 cells is essential to improve productivity. This study has two main objectives: characterizing growth and adenovirus productivity of HEK293 cells in three different culture systems: stationary, microcarriers and suspension; and determining media composition that is best suited for cell growth and adenovirus production under low serum conditions in terms of glucose and glutamine concentrations. Culture parameters were determined for HEK293 cells growing attached in basal media supplemented with 10% FBS. HEK293 cells were gradually adapted to grow in suspension in the absence of serum in supplemented media. We compared cell growth in stationary culture with 10% serum, Cytodex 3 with 10% serum and suspension culture without serum. The infectivity of adenovirus and final viral titer were also assessed in all systems. We applied metabolic flux analysis to all culture systems pre and post infection, considering the main carbon metabolism pathways, a biomass production term based on the reported stoichiometric composition of mammalian cells and a virus production equation based on protein and nucleic material content of the virus [1, 3]. This allows us to characterize and compare the metabolic state of cells and resource distribution during cell growth and virus production. Based on metabolic demand, media composition that was best suited for cell growth and adenovirus production in terms of glucose and glutamine concentrations was determined. In microcarriers we obtained a higher cell density than in stationary and suspension cultures. However, a different metabolic flux distribution, lower specific lactate production and a lower lactate produced to glucose consumed ratio was observed in suspension cultures. Results obtained can provide further insight into the metabolism of HEK293 cells during growth and virus production stages and evaluate the use of suspension and microcarrier cell culture technology for adenoviral vector production.