The Evolution of Protein Expression and Cell Culture - - BioPharm International


The Evolution of Protein Expression and Cell Culture

BioPharm International
Volume 20, Issue 10

The tedious task of medium development is typically accomplished in shake flasks and small-scale reactors. Two- to three- liter bioreactors are most commonly used for this purpose. The key to speedy medium development is the use of statistical designs that help screen and optimize the concentrations of critical components. For this reason, it is highly desirable to use a high throughput system. The implementation of such systems for mammalian cells has been successfully demonstrated.34 These developments are occurring in conjunction with promising developments in the area of high throughput systems for analytical measurements, e.g., systems for measuring antibody yield in 96-well format.

Although limited, some reports in the literature describe how microarray analysis can help assist with the medium design. Allison et al. designed a serum-free formulation for growing the normal human fibroblast, WI-38, which normally requires the presence of FBS in the growth medium. The microarray analysis for the cultures grown in serum-containing medium indicated the expression of 17 receptors. The inclusion of ligands of four of these 17 receptors resulted in a formulation that could support the growth of the cells without serum.35 The future belongs to the combined approaches of genomics, proteomics, and metabolomics for the design of media for mammalian cells. Such an approach, where the combined analysis of transcriptomic, proteomic, and protein interaction data was used, has been explored to better understand the galactose metabolism of S. cerevisae.36


The demand of therapeutic proteins and antibodies will keep increasing the scale at which these molecules have to be manufactured. A number of industrial biologics are currently manufactured by cell culture at the 10,000–20,000 L scale. The scale-up of cell culture processes still remains poorly understood, however, and it often leads to altered yields and altered product quality attributes. Although a number of empirical approaches are used for scale-up, the exact scale-up criterion is still unknown. The scale-up of processes in the past often was constrained because the mammalian cells were thought to be highly sensitive to shear. The notion of shear sensitivity of mammalian cells, particularly the ones used in industry, is slowly fading away and this certainly gives scientists more flexibility in scale-up. With no gas–liquid interfaces present, the local energy dissipation rate that animal cells would encounter in typical bioprocessing situations typically is orders of magnitude lower than the rate that has been experimentally demonstrated to catastrophically damage mammalian cells.37 Nonetheless, the non-lethal effects to cells of high energy dissipation rates still need to be fully established.

The likely reasons for imprecise scale-up and scale-down may lie in either raw material differences or dissimilar bioreactor hydrodynamics at different scales. Variation caused by raw materials might result from differences in manufacturing processes used to produce the raw materials for large scale, lot-to-lot variability, trace impurities present in the raw materials, or media preparation issues at large scale. Understanding variation caused by raw materials is challenging because of limited analytical capabilities, resources, and knowledge of process sensitivity to different analytes. Equally challenging is to understand how different fluid microenvironments affect cellular behavior. Computational fluid dynamics is an exciting technology that can be effectively used to understand bioreactor hydrodynamics.38 It should be borne in mind, however, that such technology must be used along with experimental data generated at different scales or in different bioreactor configurations at a given scale. The combination of computational and experimental data holds the key to precisely define a scale-up criterion.

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