De Villiers, Ann-Marie; (2018) High cell density purification strategies for application to antibody and virus-vectored biopharmaceutical products. Doctoral thesis (Ph.D), UCL (University College London).

Abstract

Record high titres of high value products have resulted from recent developments in cell culture production methods that are capable of producing high cell densities. These enhanced-titre, high cell density processes are thought to be a bottleneck for downstream processing due to the excessive solids content, product titre and impurity profile. This thesis explored these challenges and endeavours to find ways to overcome them. A mAb and an adenovirus have been used to represent two common biotech products, which represents the size range and complexity of biotech products used in these processes. High cell densities are challenging to clarify; however, this thesis shows that contrary to expectations from literature tangential flow microfiltration could be used to successfully clarify mAb harvests with cell densities up to 200 x 106 cell/ml, and could be successfully used for both mAbs harvest and more complex adenovirus harvests which contained lysed mammalian cells. In adenovirus harvests, high impurity concentrations are a particular challenge. This thesis shows that when dealing with high cell density harvests, using a DNA precipitation step with a variable detergent concentration linked to cell densities allows for predictable process performance. By being able to predict the range of the concentration of HC-DNA remaining after DNA precipitation, it is then also possible to design an AEX step which is capable of delivering robust performance. The economics of high-cell density processes was also investigated to better understand its impact on purification. High product concentrations present their own challenges, but increases in titre usually bring substantial reduction in production costs, sometimes is unexpected ways. Here we show for mAbs, increasing the titre also lead to increases in the dynamic binding capacity of both Protein A and cation exchange resins, which reduced the cost of goods.

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