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High-cell-density production of adeno-associated viral vector serotype 6 by triple transfection in suspension HEK293 cell cultures
  • Pablo Moço,
  • Xingge Xu,
  • Amine Kamen
Pablo Moço
McGill University
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Xingge Xu
McGill University
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Amine Kamen
McGill University

Corresponding Author:[email protected]

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The use of adeno-associated viruses (AAV) as vectors for gene and cell therapy has risen considerably in recent years. Consequently, the amount of AAV vectors required during the validation and clinical trials has also increased. AAV serotype 6 (AAV6) is well-documented for its efficiency in transducing different cell types and has been successfully used in gene and cell therapy protocols. However, the number of vectors required to effectively deliver the transgene to one single cell has been estimated at 106 viral genomes (VG). Overall, this means that large-scale production of AAV6 is needed. Suspension cell-based platforms are currently limited to low-cell-density productions, hindering the potential of this production process to increase yields. Here, we investigate the improvement of the production of AAV6 at higher cell densities. The production was performed by transient transfection of HEK293SF cells. When the plasmid DNA is provided on a cell basis, the production can be carried out at medium cell density without effects on cell-specific titer or particle functionality, resulting in titers above 1010 VG/mL. Medium supplementation alleviated the cell density effect, in terms of VG/cell, at high-cell-density productions. On the other hand, the cell-specific functional titer was not maintained, and further studies are necessary to understand the observed limitations. The medium-cell-density production method reported here lays the foundation for large-scale process operations, potentially solving the current vector shortage in AAV manufacturing.
01 Feb 2023Submitted to Biotechnology Journal
02 Feb 2023Submission Checks Completed
02 Feb 2023Assigned to Editor
21 Feb 2023Reviewer(s) Assigned
05 Mar 2023Review(s) Completed, Editorial Evaluation Pending
09 Mar 2023Editorial Decision: Revise Major
13 May 20231st Revision Received
17 May 2023Submission Checks Completed
17 May 2023Assigned to Editor
17 May 2023Reviewer(s) Assigned
26 May 2023Review(s) Completed, Editorial Evaluation Pending
30 May 2023Editorial Decision: Accept