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Cited by 34Virus-based gene therapy researchViral Infectious Diseases and Gene Expression in InsectsRNA Interference and Gene Delivery

Scale-up and manufacturing of clinical-grade self-inactivating γ-retroviral vectors by transient transfection

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van der Loo, J C M, Swaney, W P, Grassman, E, Terwilliger, A, Higashimoto, T, Schambach, A, Baum, C, Thrasher, A J +4 more·2012·Gene Therapy
DOI 10.1038/gt.2011.102PMC PMC4167414
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Abstract

The need for γ-retroviral (gRV) vectors with a self-inactivating (SIN) design for clinical application has prompted a shift in methodology of vector manufacturing from the traditional use of stable producer lines to transient transfection-based techniques. Herein, we set out to define and optimize a scalable manufacturing process for the production of gRV vectors using transfection in a closed-system bioreactor in compliance with current good manufacturing practices (cGMP). The process was based on transient transfection of 293T cells on Fibra-Cel disks in the Wave Bioreactor. Cells were harvested from tissue culture flasks and transferred to the bioreactor containing Fibra-Cel in the presence of vector plasmid, packaging plasmids and calcium-phosphate in Dulbecco's modified Eagle's medium and 10% fetal bovine serum. Virus supernatant was harvested at 10-14 h intervals. Using optimized procedures, a total of five ecotropic cGMP-grade gRV vectors were produced (9 liters each) with titers up to 3.6 × 10(7) infectious units per milliliter on 3T3 cells. One GMP preparation of vector-like particles was also produced. These results describe an optimized process for the generation of SIN viral vectors by transfection using a disposable platform that allows for the generation of clinical-grade viral vectors without the need for cleaning validation in a cost-effective manner.

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  • Viral vectors have proven efficient in achieving stable gene transfer, but their requisite extensive safety testing adds to the cost and time to develop and manufacture CAR T cell products.

    “Although viral vectors have proven efficient in achieving stable gene transfer, their requisite extensive safety testing adds to the cost and time to develop and manufacture CAR T cell products (27,28).”

    From Scalable Manufacturing of CAR T Cells for Cancer Immunotherapy (2021) · cites this paper as [28]

    The Results (cGMP production; page unavailable) document extensive safety testing of the manufactured viral vectors. The Introduction also says that certifying stable producer cell banks is lengthy and costly, but it does not establish that safety testing itself adds cost and time to CAR T-cell product development or manufacturing. The paper concerns clinical-grade vector production, not CAR T-cell products. (LLM, conf=0.86)

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Cited by

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Authors
van der Loo, J. C. M., Swaney, W. P., Grassman, E., Terwilliger, A., Higashimoto, T., Schambach, A., Baum, C., Thrasher, A. J., Williams, D. A., Nordling, D. L., Reeves, L., & Malik, P.
Affiliations
  • Barnes-Jewish Hospital (US)
  • Cincinnati Children's Hospital Medical Center (US)
  • Medizinische Hochschule Hannover (DE)
  • Great Ormond Street Hospital (GB)
  • Boston Children's Hospital (US)
  • Indiana University School of Medicine (US)
ORCID
Venue
Gene Therapy
Publisher
Springer Science and Business Media LLC
Published
2011-07-14
Volume
19
Issue
3
Pages
246-254
ISSN
  • 0969-7128 (ISSN-L)
  • 1476-5462
DOI
10.1038/gt.2011.102
PMC
PMC4167414
OpenAlex
W2027408079
Cited by
34
References
1
Citations by year
2026: 1 · 2025: 2 · 2023: 2 · 2022: 2 · 2021: 4 · 2020: 1 · 2019: 3 · 2018: 3 · 2017: 1 · 2016: 5 · 2015: 5 · 2014: 4 · 2012: 1
FWCI
0.93
Type
journal article
Language
en
Open access
Licence
Locations
  • Gene Therapy (publishedVersion)
  • PubMed (publishedVersion)
  • PubMed Central (acceptedVersion)
  • UCL Discovery (University College London) (submittedVersion)
Funders
  • Wellcome Trust (090233)
  • Deutscher Akademischer Austauschdienst
  • National Institutes of Health (UL1TR000077)
  • Medical Research Council
  • National Institute of Allergy and Infectious Diseases (U01AI087628)
  • National Center for Advancing Translational Sciences
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Virus-based gene therapy research
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Viral Infectious Diseases and Gene Expression in Insects
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Interference and Gene Delivery
Keywords
transient transfection, vector manufacturing, scale-up, HEK293T cells, calcium phosphate transfection, cyclic GMP, ecotropic retrovirus, disposable bioreactors
MeSH terms
Animals, Biotechnology, Cell Line, Genetic Vectors, Humans, Quality Control, Transfection, Gammaretrovirus, Bioreactors, Mice, Batch Cell Culture Techniques

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