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Implementation and Automation of Downstream UF/DF of AAV Using Flatsheet Membrane

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The use of adeno-associated viruses (AAVs) is common in gene therapy technology to deliver the gene of interest in a viral capsid.  The upstream process for AAV includes cell expansion, plasmid transfection, and viral vector production.  The cell culture material is then harvested and purified to prepare for formulation and fill/finish.  After the harvest step, an ultrafiltration/diafiltration step is performed using a flatsheet membrane to concentrate the product 10-20x and exchange the buffer in preparation for further chromatographic separation.  As the Takeda Cambridge pilot team transferred gene therapy capabilities into the facility, the group transitioned from stainless-steel equipment to single-use equipment, with a focus on data collection and automation.  The initial implementation of the system included controlling retentate flow via the Levitronix® controller with a manual TMP-control clamp which requires adjustments during the run.  To further automate the process, the manual TMP-control clamp was replaced with a second Levitronix pump on the retentate line in reverse flow direction, enabling automatic TMP control during operations.  The use of the levitronix pump resulted in a faster response from the controller, constant control at the set point compared to a manual clamp or an automated valve, and reduced shear on the product during recirculation and processing.  Recipes were developed on the controller with support from Levitronix® to perform load and ultrafiltration 1, diafiltration, and ultrafiltration 2 with minimal manual intervention, and the system was integrated with DeltaV for further control capabilities.  The process was successfully scaled from 10L to up to 50L bioreactor scale using different sized Levitronix® single-use pumps, and the data capture provided an easy solution for trending and analyzing the performance across the TFF step.

Author James Hilton
Company Takeda