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. 2023 Jul;120(7):1953-1960.
doi: 10.1002/bit.28414. Epub 2023 May 26.

A new T-antigen negative HEK293 cell line with improved AAV productivity

Coralie Croissant  1 , Joshua Armitano  1 , Bertrand Lazuech  1 , Danijel Švec  1 , Cyril Pugin  1 , Anaïs Guesdon  1 , Louise Bryan  1 , Antonio Castro  1 , Léa Neuhaus  1 , Giulia Fonti  1 , Jacopo Martinis  1 , Maria J Wurm  1 , Florian M Wurm  1 , Paco Pino  1
Affiliations

A new T-antigen negative HEK293 cell line with improved AAV productivity

Coralie Croissant et al. Biotechnol Bioeng. 2023 Jul.

Abstract

Viral vectors for gene therapy, such as recombinant adeno-associated viruses, are produced in human embryonic kidney (HEK) 293 cells. However, the presence of the SV40 T-antigen-encoding CDS SV40GP6 and SV40GP7 in the HEK293T genome raises safety issues when these cells are used in manufacturing for clinical purposes. We developed a new T-antigen-negative HEK cell line from ExcellGene's proprietary HEKExpress,® using the CRISPR-Cas9 strategy. We obtained a high number of clonally-derived cell populations and all of them were demonstrated T-antigen negative. Stability study and AAV production evaluation showed that the deletion of the T-antigen-encoding locus did not impact neither cell growth nor viability nor productivity. The resulting CMC-compliant cell line, named HEKzeroT,® is able to produce high AAV titers, from small to large scale.

Keywords: AAV; HEK293; T-antigen; gene therapy; manufacturing.

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References

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