Enrichment of full AAV2 using multicolumn countercurrent solvent gradient purification (MCSGP)

Adeno-associated viruses (AAVs) are non-pathogenic viruses that have become promising delivery vehicles in cell and gene therapy, with several AAV-based therapeutics receiving approval in recent years. However, a critical challenge is the separation of full and empty viral capsids, as empty capsids lack therapeutic DNA and may compromise product efficacy and safety. This study describes a proof-of-concept of a twin-column multicolumn countercurrent solvent gradient purification (MCSGP) process to enhance the ratio of full to empty capsids during AAV purification. Starting from a batch method with anion exchange chromatography, the process was adapted to continuous operation. The implementation of MCSGP, with six cycles, increased full capsid content in the final product pool from 30% to 68% (ddPCR) and 27% to 61% (cryo-TEM) respectively. Moreover, MCSGP was capable of overcoming the inherent yield-purity trade-off in the polishing step by enhancing full capsid recovery. Additionally, the MCSGP process improved productivity by up to 23% and reduced buffer consumption up to 27% compared to the batch method (ddPCR). The increased productivity and reduced buffer consumption offer both economic and environmental benefits. This study demonstrates the potential of MCSGP to meet the rising demand for high-quality AAV products in gene therapy.

Schlagwörter

AAV, Adeno-associated virus, Anion exchange chromatography, Continuous purification, Full/empty separation, Multicolumn countercurrent solvent gradient purification (MCSGP)

Fachgebiet (DDC)

600 - Technik, Medizin, angewandte Wissenschaften

Projekt

Veranstaltung

Startdatum der Ausstellung

Enddatum der Ausstellung

Startdatum der Konferenz

Enddatum der Konferenz

Datum der letzten Prüfung

ISBN

ISSN

0006-3592
1097-0290

Sprache

Englisch

Während FHNW Zugehörigkeit erstellt

Ja

Zukunftsfelder FHNW

Publikationsstatus

Veröffentlicht

Begutachtung

Peer-Review der ganzen Publikation

Open Access-Status

Closed

Lizenz

Zitation

Müller, J., Müllerová, T., Tobler, D., Hauri, D., Plieninger, R., Higuchi, Y., Takahashi, R., Vogg, S., Müller‐Späth, T., & Villiger, T. (2025). Enrichment of full AAV2 using multicolumn countercurrent solvent gradient purification (MCSGP). Biotechnology & Bioengineering, 122(9), 2420–2432. https://doi.org/10.1002/bit.29036

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