Davidson, Alexander James; (2024) Investigating the optimal expansion platform and bioprocessing parameters for the improved production of autologous chimeric antigen receptor T-cell immunotherapies. Doctoral thesis (Eng.D), UCL (University College London).

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Abstract

Novel autologous chimeric antigen receptor (CAR) T cell immunotherapies, which have demonstrated significant clinical success in the treatment of various haematological malignancies, are undergoing process related bottlenecks during the manufacture of these drug products. Although trends toward automation and encapsulating the process into single solution technologies such as the CliniMACS Prodigy® system have enabled a much needed shift away from the static culture flask and bag manual-processing approach developed in academic laboratories, there remain concerns regarding reduced modular flexibility, scalability and limitations in process analytical technologies (PATs). This doctoral thesis aimed to demonstrate that modular semi-automated bioreactor systems for autologous and allogeneic CAR T cell expansion can potentially alleviate the processing bottlenecks evident in commercial scale-out manufacture when using static culture platforms and when using single-solution technologies. Initially, static culture platforms that remain most prevalent in clinical and commercial manufacture, along with respective culture medium conditions frequently adopted were systematically characterised. Outcomes from this study demonstrated improved growth kinetics and desirable drug product quality attributes over a 7 day culture period when expanding cells using G-Rex® systems (56.75 ± 6.41 mean fold expansion) compared to the commonly used static culture bags (8.39 ± 3.47 mean fold expansion). These findings were further translated to demonstrate the cultivation of CAR T cells using the FACER Cell Culture Bioreactor (Aglaris Ltd) that implements similar culture mechanisms to the G-Rex® but with enhanced automation, modular flexibility and PATs. This novel semi-automated bioreactor displayed significantly higher specific growth rates (µ = 0.0259 ± 0.0018 h-1 , td = 26.83 ± 1.85 h) and equivalent cell quality and cytolytic potency when compared to both static culture platforms and single-solution automated technologies such as the CliniMACS Prodigy® (µ = 0.0167 ± 0.0003 h-1 , td = 41.44 ± 0.77 h), potentially enabling a therapeutic batch to be manufactured in a shorter expansion time frame. Aspects of PATs, improved monitoring and feedback control were developed further in CAR T cell expansion studies using the Applikon® MiniBio stirred-tank bioreactor (Getinge AB), specifically investigating an allogeneic approach to the scale-up of manufacture. In allogeneic manufacturing cases, high cell densities are required during culture. Using the Applikon® system, feedback control of both pH and dissolved oxygen enabled an improved cell culture environment to be maintained during expansion to achieve CAR T cell densities of 8.11 × 106 cells ml-1 in 7 days of culture that still exhibited high cell viability (96%).

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