Viral vectors for the treatment of alcoholism: Use of metabolic flux analysis for cell cultivation and vector production

Centre for Biochemical Engineering and Biotechnology, Department of Chemical Engineering and Biotechnology, Institute for Cell Dynamics and Biotechnology (ICDB): a Centre for Systems Biology, University of Chile, Beauchef 861, Santiago, Chile.
Metabolic Engineering (Impact Factor: 6.77). 10/2009; 12(2):129-37. DOI: 10.1016/j.ymben.2009.09.003
Source: PubMed


The HEK293 cell line has been used for the production of adenovirus vectors to be used in the potential treatment of alcoholism using a gene therapy strategy. Culture optimization and scale-up has been achieved by first adapting the cells to serum-free media and secondly by growing them in suspension. Adenovirus production after infection was increased, resulting in higher specific glucose consumption and lactate accumulation rates compared to the growth phase. We applied media design tools and Metabolic Flux Analysis (MFA) to compare the metabolic states of cells during growth and adenovirus production and to optimize culture media according to the metabolic demand of the cells in terms of glucose and glutamine concentrations. This allowed obtaining a higher maximum cell concentration and increased adenovirus production by minimizing the production of metabolites that can have an inhibitory effect on cell growth. We have proposed a stoichiometric equation for adenovirus synthesis. MFA results allowed determination of how these changes in composition affected the way cells distribute their nutrient resources during cell growth and virus production. Virus purification was successfully achieved using chromatography and Aqueous Two-Phase Systems (ATPS).

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Available from: Veronica Sofia Martinez, Jul 18, 2014
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    • "These studies boosted our fundamental understanding on the physiological changes following the development, adaptation, selection and engineering of cell hosts opening the door for a variety of metabolic interventions towards improved productivities (Lim et al. 2010; Seth et al. 2006). In the case of recombinant virus production, however, the translation of gathered knowledge for the rational design of process and cell engineering is still scarce, with some notable exceptions (Hansen et al. 2005; Martinez et al. 2009; Mitta et al. 2005). Although viruses induce appreciable changes to cell metabolism as part of their replication strategy, the majority of reported work has been conducted from a basic virology point of view aiming at inhibiting viral replication. "
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