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Article

Comparison of Different Liquid Chromatography-Based Purification Strategies for Adeno-Associated Virus Vectors

1
Department of Pharmacy—Center for Drug Research, Ludwig-Maximilians-Universität München, Butenandtstr. 5-13, 81377 Munich, Germany
2
Coriolis Pharma, Fraunhoferstr. 18 b, 82152 Martinsried, Germany
3
Department of Ophthalmology, University Hospital, LMU Munich, Mathildenstr. 8, 80336 Munich, Germany
*
Authors to whom correspondence should be addressed.
Academic Editor: Gabriele Candiani
Pharmaceutics 2021, 13(5), 748; https://doi.org/10.3390/pharmaceutics13050748
Received: 29 March 2021 / Revised: 10 May 2021 / Accepted: 11 May 2021 / Published: 18 May 2021
Recombinant adeno-associated virus (rAAV) vectors have evolved as one of the most promising technologies for gene therapy due to their good safety profile, high transduction efficacy, and long-term gene expression in nondividing cells. rAAV-based gene therapy holds great promise for treating genetic disorders like inherited blindness, muscular atrophy, or bleeding disorders. There is a high demand for efficient and scalable production and purification methods for rAAVs. This is particularly true for the downstream purification methods. The current standard methods are based on multiple steps of gradient ultracentrifugation, which allow for the purification and enrichment of full rAAV particles, but the scale up of this method is challenging. Here, we explored fast, scalable, and universal liquid chromatography-based strategies for the purification of rAAVs. In contrast to the hydrophobic interaction chromatography (HIC), where a substantial amount of AAV was lost, the cation exchange chromatography (CEX) was performed robustly for multiple tested serotypes and resulted in a mixture of full and empty rAAVs with a good purity profile. For the used affinity chromatography (AC), a serotype dependence was observed. Anion exchange chromatography (AEX) worked well for the AAV8 serotype and achieved high levels of purification and a baseline separation of full and empty rAAVs. Depending on the AAV serotype, a combination of CEX and AEX or AC and AEX is recommended and holds promise for future translational projects that require highly pure and full particle-enriched rAAVs. View Full-Text
Keywords: adeno-associated virus; rAAV vector; column purification; empty capsids; ion exchange chromatography; affinity chromatography adeno-associated virus; rAAV vector; column purification; empty capsids; ion exchange chromatography; affinity chromatography
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MDPI and ACS Style

Rieser, R.; Koch, J.; Faccioli, G.; Richter, K.; Menzen, T.; Biel, M.; Winter, G.; Michalakis, S. Comparison of Different Liquid Chromatography-Based Purification Strategies for Adeno-Associated Virus Vectors. Pharmaceutics 2021, 13, 748. https://doi.org/10.3390/pharmaceutics13050748

AMA Style

Rieser R, Koch J, Faccioli G, Richter K, Menzen T, Biel M, Winter G, Michalakis S. Comparison of Different Liquid Chromatography-Based Purification Strategies for Adeno-Associated Virus Vectors. Pharmaceutics. 2021; 13(5):748. https://doi.org/10.3390/pharmaceutics13050748

Chicago/Turabian Style

Rieser, Ruth, Johanna Koch, Greta Faccioli, Klaus Richter, Tim Menzen, Martin Biel, Gerhard Winter, and Stylianos Michalakis. 2021. "Comparison of Different Liquid Chromatography-Based Purification Strategies for Adeno-Associated Virus Vectors" Pharmaceutics 13, no. 5: 748. https://doi.org/10.3390/pharmaceutics13050748

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