Accelerated Development of AAV Purification Process Using a High-Throughput and Automated Crossflow System

Adeno-associated viruses (AAV) are currently predominant viral transfer tools for gene therapy, and efforts are being made to design faster and more efficient methods and technologies for their manufacturing. The early selection of high-performing filters is essential for developing an ultrafiltrati...

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Bibliographic Details
Published in:Separations 2024-03, Vol.11 (3), p.73
Main Authors: Mendes, Sónia, Faria, Tiago Q., Nascimento, André, Noverraz, Marc, Bollmann, Franziska, Nestola, Piergiuseppe, Roldão, António, Peixoto, Cristina, Silva, Ricardo J. S.
Format: Article
Language:English
Subjects:
adeno-associated virus
Automation
Cassettes
Cell culture
Cross flow
Design of experiments
downstream processing
Gene therapy
Genetic vectors
process development
Process parameters
Sensors
tangential flow filtration
Ultrafiltration
Viruses
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Summary:Adeno-associated viruses (AAV) are currently predominant viral transfer tools for gene therapy, and efforts are being made to design faster and more efficient methods and technologies for their manufacturing. The early selection of high-performing filters is essential for developing an ultrafiltration and diafiltration (UF/DF) process, especially when feed material is scarce, and timelines are short. However, few methods and technologies exist to enable process optimization with multiple variations in a single run. In this study, we explored the potential of Ambr® Crossflow for high-throughput, automated screening of different membrane materials, pore sizes and different process conditions for the UF/DF step of AAV8. The best overall performance was obtained with a 100 kDa PES flat sheet cassette. The UF/DF process was further transferred to a larger scale to the Sartoflow® Smart Tangential Flow Filtration (TFF) system using a 100 kDa PES Sartocon® Slice 200 cassette and compared to a 100 kDa PES hollow fiber. Virus recovery, permeate flux and total protein removal values of the flat sheet cassette were similar to those achieved in small-scale devices, and higher than those of the hollow fiber, thus demonstrating similar performance at a larger process scale. The high-throughput, automated method described herein allowed to screen multiple materials and process parameters of a UF/DF process in a time- and resource-efficient way, making it a useful tool to accelerate early-stage downstream process development of AAV.
ISSN:2297-8739
2297-8739
DOI:10.3390/separations11030073