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Optimizing and developing a scalable, chemically defined, animal component-free lentiviral vector production process in a fixed-bed bioreactor

Fiol, Carme Ripoll; Collignon, Marie-Laure; Welsh, John; Rafiq, Qasim A; (2023) Optimizing and developing a scalable, chemically defined, animal component-free lentiviral vector production process in a fixed-bed bioreactor. Molecular Therapy - Methods & Clinical Development , 30 pp. 221-234. 10.1016/j.omtm.2023.06.011. Green open access

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Abstract

Lentiviral vectors (LVVs) play a critical role in gene delivery for ex vivo gene-modified cell therapies. However, the lack of scalable LVV production methods and the high cost associated with them may limit their use. In this work, we demonstrate the optimization and development of a scalable, chemically defined, animal component-free LVV production process using adherent human embryonic kidney 293T cells in a fixed-bed bioreactor. The initial studies focused on the optimization of the culture process in 2D static cultures. Process changes such as decreasing cell seeding density on day 0 from 2.5 × 104 to 5 × 103 cells/cm2, delaying the transient transfection from 24 to 120 h post-seeding, reducing plasmid DNA to 167 ng/cm2, and adding 5 mM sodium butyrate 6 h post-transfection improved functional LVV titers by 26.9-fold. The optimized animal component-free production process was then transferred to the iCELLis Nano bioreactor, a fixed-bed bioreactor, where titers of 1.2 × 106 TU/cm2 were achieved when it was operated in perfusion. In this work, comparable functional LVV titers were obtained with FreeStyle 293 Expression medium and the conventional Dulbecco’s modified Eagle’s medium supplemented with 10% fetal bovine serum both at small and large scale.

Type: Article
Title: Optimizing and developing a scalable, chemically defined, animal component-free lentiviral vector production process in a fixed-bed bioreactor
Open access status: An open access version is available from UCL Discovery
DOI: 10.1016/j.omtm.2023.06.011
Publisher version: https://doi.org/10.1016/j.omtm.2023.06.011
Language: English
Additional information: This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
Keywords: Animal component-free, chemically defined, serum-free, virus, lentiviral vectors, iCELLis Nano bioreactor, fixed-bed bioreactor, scalability, gene therapy, immunotherapy
UCL classification: UCL
UCL > Provost and Vice Provost Offices > UCL BEAMS
UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Engineering Science
UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Engineering Science > Dept of Biochemical Engineering
URI: https://discovery.ucl.ac.uk/id/eprint/10174230
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