Downstream AAV Production: A Targeted Approach to Improve Empty | Full Capsid Separation (a Case Study with Pharmaron)

One of the greatest challenges for AAV production is the ability to purify full (product genome containing) from empty (non-product containing) viral capsids. Separation of empty from full capsids is challenging because they are similar in charge and size. The presence of empty capsids could affect the efficacy and safety of AAV vector products due to the risk of increased immunogenicity of the product. Traditionally ultracentrifugation-based purification methods have been able to provide drug substance with a high enrichment of full capsids. However, its productivity is low, the equipment is expensive, and scaling of the process is driven by a scale-out instead of a scale-up approach – essentially the need for more and more ultracentrifuges.

Although Pharmaron can deliver ultracentrifugation-based processes, Pharmaron has strategically targeted chromatography for an ‘end in mind’ approach. Chromatography based operations are scalable, thus more suitable for gene therapies and especially for those with high dose requirements. Pharmaron has a purification toolbox in place and ready to deliver multiple serotype AAV products, however its innovative approach includes evaluation of the latest advancements.

Through Pharmaron’s innovation approach, a collaboration was established with Sartorius BIA Separations to explore the potential of monoliths to separate AAV full and empty capsids. Size exclusion (SEC), cation exchange (CEX) and anion exchange (AEX) high performance liquid chromatography analytics based on Sartorius BIA Separations’ PATfix™ devices were used to estimate process recoveries with high precision and accuracy.

Working with Sartorius BIA Separations, Pharmaron was able to extend its toolbox of AAV purification solutions, providing multiple-modality options to achieve robust separation of genome-containing (full) from genome-free (empty) viral capsids with high purity and yield for multiple AAV serotypes. During this collaboration, a strong partnership was developed leading to Sartorius BIA separations presenting the work at the second webinar of the Pharmaron Cell and Gene Therapy webinar series. The webinar was entitled “Downstream AAV Production: A Targeted Approach to Optimization” and its recording is available for viewing at Pharmaron website.

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