Development and Optimization of a Hydrophobic Interaction Chromatography-Based Method of AAV Harvest, Capture, and Recovery
With many ongoing clinical trials utilizing adeno-associated virus (AAV) gene therapy, it is necessary to find scalable and serotype-independent primary capture and recovery methods to allow for efficient and robust manufacturing processes. Here, we demonstrate the ability of a hydrophobic interacti...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2020-12-01
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| Series: | Molecular Therapy: Methods & Clinical Development |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S232905012030200X |
| _version_ | 1819058759701364736 |
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| author | David J. McNally Bryan A. Piras Catherine M. Willis Timothy D. Lockey Michael M. Meagher |
| author_facet | David J. McNally Bryan A. Piras Catherine M. Willis Timothy D. Lockey Michael M. Meagher |
| author_sort | David J. McNally |
| collection | DOAJ |
| description | With many ongoing clinical trials utilizing adeno-associated virus (AAV) gene therapy, it is necessary to find scalable and serotype-independent primary capture and recovery methods to allow for efficient and robust manufacturing processes. Here, we demonstrate the ability of a hydrophobic interaction chromatography membrane to capture and recover AAV1, AAV5, AAV8, and AAV “Mutant C” (a novel serotype incorporating elements of AAV3B and AAV8) particles from cell culture media and cell lysate with recoveries of 76%–100% of loaded material, depending on serotype. A simple, novel technique that integrates release and recovery of cell-associated AAV capsids is demonstrated. We show that by the addition of lyotropic salts to AAV-containing cell suspensions, AAV is released at an equivalent efficiency to mechanical lysis. The addition of the lyotropic salt also promotes a phase separation, which allows physical removal of large amounts of DNA and insoluble cellular debris from the AAV-containing aqueous fraction. The AAV is then captured and eluted from a hydrophobic interaction chromatography membrane. This integrated lysis and primary capture and recovery technique facilitates substantial removal of host-cell DNA and host-cell protein impurities. |
| first_indexed | 2024-12-21T14:00:19Z |
| format | Article |
| id | doaj.art-7d9ca9ada4994606ac4ef9aba144f735 |
| institution | Directory Open Access Journal |
| issn | 2329-0501 |
| language | English |
| last_indexed | 2024-12-21T14:00:19Z |
| publishDate | 2020-12-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Molecular Therapy: Methods & Clinical Development |
| spelling | doaj.art-7d9ca9ada4994606ac4ef9aba144f7352022-12-21T19:01:23ZengElsevierMolecular Therapy: Methods & Clinical Development2329-05012020-12-0119275284Development and Optimization of a Hydrophobic Interaction Chromatography-Based Method of AAV Harvest, Capture, and RecoveryDavid J. McNally0Bryan A. Piras1Catherine M. Willis2Timothy D. Lockey3Michael M. Meagher4Department of Therapeutics Production & Quality, St. Jude Children’s Research Hospital, Memphis, TN 38105-3678, USADepartment of Therapeutics Production & Quality, St. Jude Children’s Research Hospital, Memphis, TN 38105-3678, USAChildren’s GMP, Memphis, TN, USADepartment of Therapeutics Production & Quality, St. Jude Children’s Research Hospital, Memphis, TN 38105-3678, USADepartment of Therapeutics Production & Quality, St. Jude Children’s Research Hospital, Memphis, TN 38105-3678, USA; Corresponding author: Michael Meagher, PhD, Department of Therapeutics Production & Quality, St. Jude Children’s Research Hospital, 262 Danny Thomas Place, Memphis, TN 38105-3678, USA.With many ongoing clinical trials utilizing adeno-associated virus (AAV) gene therapy, it is necessary to find scalable and serotype-independent primary capture and recovery methods to allow for efficient and robust manufacturing processes. Here, we demonstrate the ability of a hydrophobic interaction chromatography membrane to capture and recover AAV1, AAV5, AAV8, and AAV “Mutant C” (a novel serotype incorporating elements of AAV3B and AAV8) particles from cell culture media and cell lysate with recoveries of 76%–100% of loaded material, depending on serotype. A simple, novel technique that integrates release and recovery of cell-associated AAV capsids is demonstrated. We show that by the addition of lyotropic salts to AAV-containing cell suspensions, AAV is released at an equivalent efficiency to mechanical lysis. The addition of the lyotropic salt also promotes a phase separation, which allows physical removal of large amounts of DNA and insoluble cellular debris from the AAV-containing aqueous fraction. The AAV is then captured and eluted from a hydrophobic interaction chromatography membrane. This integrated lysis and primary capture and recovery technique facilitates substantial removal of host-cell DNA and host-cell protein impurities.http://www.sciencedirect.com/science/article/pii/S232905012030200Xhydrophobic interaction chromatographyAAVgene therapyprocess integrationmembrane chromatography |
| spellingShingle | David J. McNally Bryan A. Piras Catherine M. Willis Timothy D. Lockey Michael M. Meagher Development and Optimization of a Hydrophobic Interaction Chromatography-Based Method of AAV Harvest, Capture, and Recovery Molecular Therapy: Methods & Clinical Development hydrophobic interaction chromatography AAV gene therapy process integration membrane chromatography |
| title | Development and Optimization of a Hydrophobic Interaction Chromatography-Based Method of AAV Harvest, Capture, and Recovery |
| title_full | Development and Optimization of a Hydrophobic Interaction Chromatography-Based Method of AAV Harvest, Capture, and Recovery |
| title_fullStr | Development and Optimization of a Hydrophobic Interaction Chromatography-Based Method of AAV Harvest, Capture, and Recovery |
| title_full_unstemmed | Development and Optimization of a Hydrophobic Interaction Chromatography-Based Method of AAV Harvest, Capture, and Recovery |
| title_short | Development and Optimization of a Hydrophobic Interaction Chromatography-Based Method of AAV Harvest, Capture, and Recovery |
| title_sort | development and optimization of a hydrophobic interaction chromatography based method of aav harvest capture and recovery |
| topic | hydrophobic interaction chromatography AAV gene therapy process integration membrane chromatography |
| url | http://www.sciencedirect.com/science/article/pii/S232905012030200X |
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