Production of Lentiviral Vectors Using Suspension Cells Grown in Serum-free Media.
HIV
gene and cell therapy
immunotherapy
lentiviral
lentivirus
production
pseudotype
scalable
suspension
vector
Journal
Molecular therapy. Methods & clinical development
ISSN: 2329-0501
Titre abrégé: Mol Ther Methods Clin Dev
Pays: United States
ID NLM: 101624857
Informations de publication
Date de publication:
12 Jun 2020
12 Jun 2020
Historique:
received:
31
08
2019
accepted:
15
11
2019
entrez:
1
1
2020
pubmed:
1
1
2020
medline:
1
1
2020
Statut:
epublish
Résumé
Lentiviral vectors are increasingly utilized in cell and gene therapy applications because they efficiently transduce target cells such as hematopoietic stem cells and T cells. Large-scale production of current Good Manufacturing Practices-grade lentiviral vectors is limited because of the adherent, serum-dependent nature of HEK293T cells used in the manufacturing process. To optimize large-scale clinical-grade lentiviral vector production, we developed an improved production scheme by adapting HEK293T cells to grow in suspension using commercially available and chemically defined serum-free media. Lentiviral vectors with titers equivalent to those of HEK293T cells were produced from SJ293TS cells using optimized transfection conditions that reduced the required amount of plasmid DNA by 50%. Furthermore, purification of SJ293TS-derived lentiviral vectors at 1 L yielded a recovery of 55% ± 14% (n = 138) of transducing units in the starting material, more than a 2-fold increase over historical yields from adherent HEK293T serum-dependent lentiviral vector preparations. SJ293TS cells were stable to produce lentiviral vectors over 4 months of continuous culture. SJ293TS-derived lentiviral vectors efficiently transduced primary hematopoietic stem cells and T cells from healthy donors. Overall, our SJ293TS cell line enables high-titer vector production in serum-free conditions while reducing the amount of input DNA required, resulting in a highly efficient manufacturing option.
Identifiants
pubmed: 31890741
doi: 10.1016/j.omtm.2019.11.011
pii: S2329-0501(19)30136-6
pmc: PMC6931067
doi:
Types de publication
Journal Article
Langues
eng
Pagination
58-68Subventions
Organisme : NHLBI NIH HHS
ID : P01 HL053749
Pays : United States
Informations de copyright
© 2019 The Author(s).
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