Liter-scale manufacturing of shelf-stable plasmid DNA/PEI transfection particles for viral vector production
© 2024 The Authors..
The transfection efficiency and stability of the delivery vehicles of plasmid DNA (pDNA) are critical metrics to ensure high-quality and high-yield production of viral vectors. We previously identified that the optimal size of pDNA/poly(ethylenimine) (PEI) transfection particles is 400-500 nm and developed a bottom-up assembly method to construct stable 400-nm pDNA/PEI particles and benchmarked their transfection efficiency in producing lentiviral vectors (LVVs). Here, we report scale-up production protocols for such transfection particles. Using a two-inlet confined impinging jet (CIJ) mixer with a dual syringe pump set-up, we produced a 1-L batch at a flow rate of 100 mL/min, and further scaled up this process with a larger CIJ mixer and a dual peristaltic pump array, allowing for continuous production at a flow rate of 1 L/min without a lot size limit. We demonstrated the scalability of this process with a 5-L lot and validated the quality of these 400-nm transfection particles against the target product profile, including physical properties, shelf and on-bench stability, transfection efficiency, and LVV production yield in both 15-mL bench culture and 2-L bioreactor runs. These results confirm the potential of this particle assembly process as a scalable manufacturing platform for viral vector production.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2024 |
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| Erschienen: |
2024 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:32 |
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| Enthalten in: |
Molecular therapy. Methods & clinical development - 32(2024), 1 vom: 14. Feb., Seite 101194 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Hu, Yizong [VerfasserIn] |
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| Links: |
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| Themen: |
Endosomal escape |
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| Anmerkungen: |
Date Revised 15.02.2024 published: Electronic-eCollection Citation Status PubMed-not-MEDLINE |
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| doi: |
10.1016/j.omtm.2024.101194 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
NLM368423816 |
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| 520 | |a © 2024 The Authors. | ||
| 520 | |a The transfection efficiency and stability of the delivery vehicles of plasmid DNA (pDNA) are critical metrics to ensure high-quality and high-yield production of viral vectors. We previously identified that the optimal size of pDNA/poly(ethylenimine) (PEI) transfection particles is 400-500 nm and developed a bottom-up assembly method to construct stable 400-nm pDNA/PEI particles and benchmarked their transfection efficiency in producing lentiviral vectors (LVVs). Here, we report scale-up production protocols for such transfection particles. Using a two-inlet confined impinging jet (CIJ) mixer with a dual syringe pump set-up, we produced a 1-L batch at a flow rate of 100 mL/min, and further scaled up this process with a larger CIJ mixer and a dual peristaltic pump array, allowing for continuous production at a flow rate of 1 L/min without a lot size limit. We demonstrated the scalability of this process with a 5-L lot and validated the quality of these 400-nm transfection particles against the target product profile, including physical properties, shelf and on-bench stability, transfection efficiency, and LVV production yield in both 15-mL bench culture and 2-L bioreactor runs. These results confirm the potential of this particle assembly process as a scalable manufacturing platform for viral vector production | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a HEK293 cells | |
| 650 | 4 | |a PEI | |
| 650 | 4 | |a endosomal escape | |
| 650 | 4 | |a nanoparticles | |
| 650 | 4 | |a plasmid DNA | |
| 650 | 4 | |a poly(ethylenimine) | |
| 650 | 4 | |a scale-up production | |
| 650 | 4 | |a transient transfection | |
| 700 | 1 | |a Eder, Brendan A |e verfasserin |4 aut | |
| 700 | 1 | |a Lin, Jinghan |e verfasserin |4 aut | |
| 700 | 1 | |a Li, Sixuan |e verfasserin |4 aut | |
| 700 | 1 | |a Zhu, Yining |e verfasserin |4 aut | |
| 700 | 1 | |a Wang, Tza-Huei |e verfasserin |4 aut | |
| 700 | 1 | |a Guo, Ting |e verfasserin |4 aut | |
| 700 | 1 | |a Mao, Hai-Quan |e verfasserin |4 aut | |
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