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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| Format: | Article |
| Language: | English |
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MDPI AG
2024-02-01
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| Series: | Separations |
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| Online Access: | https://www.mdpi.com/2297-8739/11/3/73 |
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| author | Sónia Mendes Tiago Q. Faria André Nascimento Marc Noverraz Franziska Bollmann Piergiuseppe Nestola António Roldão Cristina Peixoto Ricardo J. S. Silva |
| author_facet | Sónia Mendes Tiago Q. Faria André Nascimento Marc Noverraz Franziska Bollmann Piergiuseppe Nestola António Roldão Cristina Peixoto Ricardo J. S. Silva |
| author_sort | Sónia Mendes |
| collection | DOAJ |
| description | 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<sup>®</sup> 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<sup>®</sup> Smart Tangential Flow Filtration (TFF) system using a 100 kDa PES Sartocon<sup>®</sup> 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. |
| first_indexed | 2024-04-24T17:49:27Z |
| format | Article |
| id | doaj.art-6cdce523228341529b56a6e926a4ca43 |
| institution | Directory Open Access Journal |
| issn | 2297-8739 |
| language | English |
| last_indexed | 2024-04-24T17:49:27Z |
| publishDate | 2024-02-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Separations |
| spelling | doaj.art-6cdce523228341529b56a6e926a4ca432024-03-27T14:04:26ZengMDPI AGSeparations2297-87392024-02-011137310.3390/separations11030073Accelerated Development of AAV Purification Process Using a High-Throughput and Automated Crossflow SystemSónia Mendes0Tiago Q. Faria1André Nascimento2Marc Noverraz3Franziska Bollmann4Piergiuseppe Nestola5António Roldão6Cristina Peixoto7Ricardo J. S. Silva8iBET, Instituto de Biologia Experimental e Tecnológica, Apartado 12, 2780-901 Oeiras, PortugaliBET, Instituto de Biologia Experimental e Tecnológica, Apartado 12, 2780-901 Oeiras, PortugaliBET, Instituto de Biologia Experimental e Tecnológica, Apartado 12, 2780-901 Oeiras, PortugalSartorius Stedim Switzerland AG, 8317 Tagelswangen, SwitzerlandSartorius Stedim Biotech GmbH, 37079 Göttingen, GermanySartorius Stedim Switzerland AG, 8317 Tagelswangen, SwitzerlandiBET, Instituto de Biologia Experimental e Tecnológica, Apartado 12, 2780-901 Oeiras, PortugaliBET, Instituto de Biologia Experimental e Tecnológica, Apartado 12, 2780-901 Oeiras, PortugaliBET, Instituto de Biologia Experimental e Tecnológica, Apartado 12, 2780-901 Oeiras, PortugalAdeno-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<sup>®</sup> 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<sup>®</sup> Smart Tangential Flow Filtration (TFF) system using a 100 kDa PES Sartocon<sup>®</sup> 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.https://www.mdpi.com/2297-8739/11/3/73adeno-associated virusdownstream processinggene therapyprocess developmenttangential flow filtration |
| spellingShingle | Sónia Mendes Tiago Q. Faria André Nascimento Marc Noverraz Franziska Bollmann Piergiuseppe Nestola António Roldão Cristina Peixoto Ricardo J. S. Silva Accelerated Development of AAV Purification Process Using a High-Throughput and Automated Crossflow System Separations adeno-associated virus downstream processing gene therapy process development tangential flow filtration |
| title | Accelerated Development of AAV Purification Process Using a High-Throughput and Automated Crossflow System |
| title_full | Accelerated Development of AAV Purification Process Using a High-Throughput and Automated Crossflow System |
| title_fullStr | Accelerated Development of AAV Purification Process Using a High-Throughput and Automated Crossflow System |
| title_full_unstemmed | Accelerated Development of AAV Purification Process Using a High-Throughput and Automated Crossflow System |
| title_short | Accelerated Development of AAV Purification Process Using a High-Throughput and Automated Crossflow System |
| title_sort | accelerated development of aav purification process using a high throughput and automated crossflow system |
| topic | adeno-associated virus downstream processing gene therapy process development tangential flow filtration |
| url | https://www.mdpi.com/2297-8739/11/3/73 |
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