Cargando…
High-efficiency purification of divergent AAV serotypes using AAVX affinity chromatography
The adeno-associated viral vector (AAV) provides a safe and efficient gene therapy platform with several approved products that have marked therapeutic impact for patients. However, a major bottleneck in the development and commercialization of AAV remains the efficiency, cost, and scalability of AA...
Autores principales: | , , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Society of Gene & Cell Therapy
2022
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9823220/ https://www.ncbi.nlm.nih.gov/pubmed/36654797 http://dx.doi.org/10.1016/j.omtm.2022.12.009 |
_version_ | 1784866107612463104 |
---|---|
author | Florea, Michael Nicolaou, Fotini Pacouret, Simon Zinn, Eric M. Sanmiguel, Julio Andres-Mateos, Eva Unzu, Carmen Wagers, Amy J. Vandenberghe, Luk H. |
author_facet | Florea, Michael Nicolaou, Fotini Pacouret, Simon Zinn, Eric M. Sanmiguel, Julio Andres-Mateos, Eva Unzu, Carmen Wagers, Amy J. Vandenberghe, Luk H. |
author_sort | Florea, Michael |
collection | PubMed |
description | The adeno-associated viral vector (AAV) provides a safe and efficient gene therapy platform with several approved products that have marked therapeutic impact for patients. However, a major bottleneck in the development and commercialization of AAV remains the efficiency, cost, and scalability of AAV production. Chromatographic methods have the potential to allow purification at increased scales and lower cost but often require optimization specific to each serotype. Here, we demonstrate that the POROS CaptureSelect AAVX affinity resin efficiently captures a panel of 15 divergent AAV serotypes, including the commonly used AAV2, AAV8, AAV9, PHP.B, and Anc80. We also find that AAVX resin can be regenerated repeatedly without loss of efficiency or carry-over contamination. While AAV preps purified with AAVX showed a higher fraction of empty capsids than preps purified using iodixanol ultracentrifugation, the potency of the AAVX purified vectors was comparable with that of iodixanol purified vectors both in vitro and in vivo. Finally, optimization of the purification protocol resulted in a process with an overall efficiency of 65%–80% across all scales and AAV serotypes tested. These data establish AAVX affinity chromatography as a versatile and efficient method for purification of a broad range of AAV serotypes. |
format | Online Article Text |
id | pubmed-9823220 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | American Society of Gene & Cell Therapy |
record_format | MEDLINE/PubMed |
spelling | pubmed-98232202023-01-17 High-efficiency purification of divergent AAV serotypes using AAVX affinity chromatography Florea, Michael Nicolaou, Fotini Pacouret, Simon Zinn, Eric M. Sanmiguel, Julio Andres-Mateos, Eva Unzu, Carmen Wagers, Amy J. Vandenberghe, Luk H. Mol Ther Methods Clin Dev Original Article The adeno-associated viral vector (AAV) provides a safe and efficient gene therapy platform with several approved products that have marked therapeutic impact for patients. However, a major bottleneck in the development and commercialization of AAV remains the efficiency, cost, and scalability of AAV production. Chromatographic methods have the potential to allow purification at increased scales and lower cost but often require optimization specific to each serotype. Here, we demonstrate that the POROS CaptureSelect AAVX affinity resin efficiently captures a panel of 15 divergent AAV serotypes, including the commonly used AAV2, AAV8, AAV9, PHP.B, and Anc80. We also find that AAVX resin can be regenerated repeatedly without loss of efficiency or carry-over contamination. While AAV preps purified with AAVX showed a higher fraction of empty capsids than preps purified using iodixanol ultracentrifugation, the potency of the AAVX purified vectors was comparable with that of iodixanol purified vectors both in vitro and in vivo. Finally, optimization of the purification protocol resulted in a process with an overall efficiency of 65%–80% across all scales and AAV serotypes tested. These data establish AAVX affinity chromatography as a versatile and efficient method for purification of a broad range of AAV serotypes. American Society of Gene & Cell Therapy 2022-12-16 /pmc/articles/PMC9823220/ /pubmed/36654797 http://dx.doi.org/10.1016/j.omtm.2022.12.009 Text en © 2022 The Authors https://creativecommons.org/licenses/by/4.0/This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Original Article Florea, Michael Nicolaou, Fotini Pacouret, Simon Zinn, Eric M. Sanmiguel, Julio Andres-Mateos, Eva Unzu, Carmen Wagers, Amy J. Vandenberghe, Luk H. High-efficiency purification of divergent AAV serotypes using AAVX affinity chromatography |
title | High-efficiency purification of divergent AAV serotypes using AAVX affinity chromatography |
title_full | High-efficiency purification of divergent AAV serotypes using AAVX affinity chromatography |
title_fullStr | High-efficiency purification of divergent AAV serotypes using AAVX affinity chromatography |
title_full_unstemmed | High-efficiency purification of divergent AAV serotypes using AAVX affinity chromatography |
title_short | High-efficiency purification of divergent AAV serotypes using AAVX affinity chromatography |
title_sort | high-efficiency purification of divergent aav serotypes using aavx affinity chromatography |
topic | Original Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9823220/ https://www.ncbi.nlm.nih.gov/pubmed/36654797 http://dx.doi.org/10.1016/j.omtm.2022.12.009 |
work_keys_str_mv | AT floreamichael highefficiencypurificationofdivergentaavserotypesusingaavxaffinitychromatography AT nicolaoufotini highefficiencypurificationofdivergentaavserotypesusingaavxaffinitychromatography AT pacouretsimon highefficiencypurificationofdivergentaavserotypesusingaavxaffinitychromatography AT zinnericm highefficiencypurificationofdivergentaavserotypesusingaavxaffinitychromatography AT sanmigueljulio highefficiencypurificationofdivergentaavserotypesusingaavxaffinitychromatography AT andresmateoseva highefficiencypurificationofdivergentaavserotypesusingaavxaffinitychromatography AT unzucarmen highefficiencypurificationofdivergentaavserotypesusingaavxaffinitychromatography AT wagersamyj highefficiencypurificationofdivergentaavserotypesusingaavxaffinitychromatography AT vandenberghelukh highefficiencypurificationofdivergentaavserotypesusingaavxaffinitychromatography |