Centrifugation is often at the centre of the clarification process occurring after mammalian cell cultures. Developing and characterising a centrifugation process is difficult because scale-down models do not reflect sufficiently the impact process conditions have on product quality. Hence, this unit operation is, perhaps, not as well characterised as others when companies map out the design space for the manufacturing process of mammalian cell-based products. During centrifugation mammalian cells are exposed to significant levels of shear: this shear increases the quantities of impurities (e.g. intracellular components) released in the cell containing fluid. These components have a significant effect on the subsequent purification steps and on the product quality as they include problematic host cell proteins such as reducing enzymes (e.g. thioredoxin reductase), and micron and submicron particles increases. The challenge is exacerbated by substantial increases in cell culture densities, a side effect of the pursuit of higher culture productivities, which centrifugation operations must contend with. The research presented here aims at developing a novel framework with a holistic view on the fermentation and centrifugation operation and their impact on filter sizing and product quality. A scale-down centrifugation model was created by adapting an existing centrifugation mimic to account for additional factors such as reduced oxygen conditions in large-scale processes. Furthermore, this model was linked to a fermentation high throughput platform - ambr®. The scale-down centrifugation model was first employed in investigating the impact of the harvest operation on the overall product quality for different products. This study was then extended to capture the effect of parameters such as centrifugation shear, cell density and viability on process performance by linking the ambr® with the scale-down centrifugation model. A Design of Experiments (DoE)-based approach was adopted in order to map the design space for the fermentation-centrifugation sequence with respect to process performance and product quality. The presentation will highlight how important trade-offs were reconciled when designing the centrifugation unit operation. Furthermore, it will illustrate how stochastic simulations were used to evaluate process robustness and identify the “sweet spot” of operation.

Integrated upstream and centrifugation scale-down framework for enhanced predictability of product heterogeneity, antibody reduction and filter sizing

2019 Spring National Meeting

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