Tregidgo, Molly Beth; (2022) Scale-down Technologies for Perfusion Culture for Rapid Biopharmaceutical Process Development. Doctoral thesis (Eng.D), UCL (University College London).

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Abstract

Perfusion culture is becoming an increasingly popular choice for the production of therapeutic proteins, however few scale-down devices capable of small scale and/or high throughput optimisation of high cell density perfusion cultures have been published. To address this technology gap, this thesis describes the development, implementation, and engineering characterisation of two scale-down technologies; (i) a quasi-perfusion method at mL-scale in microwell plates (MWPs) and (ii) a purposely designed novel bioreactor (BR) at 250mL scale. Quasi-perfusion in MWP was developed at mL-scale and is capable of achieving many of the specific characteristics of perfusion culture, namely elevated cell density, cell retention and good productivity. The quasi-perfusion methodology was implemented to screen a range of process conditions, exchanging at a constant vessel volume per day (VVD) between 0.5-1.8, or at constant cell specific perfusion rate (CSPR) with a range of media, with cell retention achieved via sedimentation or centrifugation. Viable cell densities (VCDs) of 42 × 10^{6} cells mL-1and volumetric productivities up to 2 fold greater than fed-batch were achieved. Design and engineering characterisation of the novel 250mL BR ensured favourable hydrodynamics, with a dual impeller system selected to maximise mass transfer. Cell retention was achieved via a tangential flow filter (TFF) at perfusion rates between 0.5- 1.8 VVD, maintaining maximum VCDs of 92 × 10^{6} cells mL-1 at >95% viability. Good scalability was demonstrated for a range of performance metrics, including µmax, qAb and biomass production, between each system when compared to a bench scale 5L BR, while scaling was based on constant volumetric power input. The combined use of quasi-perfusion methodologies, shown to be sensitive to changes in perfusion rate and media composition, for high throughput screening studies, followed by the 250mL BR for in-depth study of a small range of conditions, is shown to be a powerful tool for the development and optimisation of perfusion processes.

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