Pilot-scale process development for recombinant adeno-associated virus (rAAV) production based on high-density Sf9 cell culture
Abstract Background In recent years, gene therapy drugs have been widely marketed, and their effectiveness and potential have been confirmed. Thus, increasing their production on an industrial scale is critical. Recombinant adeno-associated viruses (rAAVs) are optimal vectors for gene therapy applic...
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| Format: | Article |
| Language: | English |
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BMC
2024-11-01
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| Series: | Virology Journal |
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| Online Access: | https://doi.org/10.1186/s12985-024-02550-4 |
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| author | Xinran Li Jieyi Gu Haoquan Wu Yuanyuan Xie |
| author_facet | Xinran Li Jieyi Gu Haoquan Wu Yuanyuan Xie |
| author_sort | Xinran Li |
| collection | DOAJ |
| description | Abstract Background In recent years, gene therapy drugs have been widely marketed, and their effectiveness and potential have been confirmed. Thus, increasing their production on an industrial scale is critical. Recombinant adeno-associated viruses (rAAVs) are optimal vectors for gene therapy applications, and the baculovirus expression vector system (BEVS), which is based on Sf9 cell culture, is a common tool for rAAV production. Methods In this work, an Sf9 cell fed-batch process was developed using shake flasks. In the laboratory-scale bioreactor, four processes were selected as the key factors when carrying out the orthogonal experiment. On the basis of the equal P/V principle and considering the problem posed by air bubbles, a pilot-scale level bioreactor process was established. Results Here, we describe a method in which a BEVS was used to produce rAAV vectors, with the cell density increasing to 22.8 × 106 cells/mL and the rAAV titre increasing to 20 × 1011 VG/mL upon adding feed material. By resolving the problems associated with high-density cell culture and air bubbles, this process was successfully scaled to a 50 L pilot-scale level. Conclusions This successful experiment not only provides a technological basis for further scale-up but also guarantees product capacity. We hope that this development process can provide reference data for studying cell culture-based drug production. |
| format | Article |
| id | doaj-art-df07bcbad12f42859b6236a861c13e71 |
| institution | Kabale University |
| issn | 1743-422X |
| language | English |
| publishDate | 2024-11-01 |
| publisher | BMC |
| record_format | Article |
| series | Virology Journal |
| spelling | doaj-art-df07bcbad12f42859b6236a861c13e712024-11-10T12:06:38ZengBMCVirology Journal1743-422X2024-11-0121111010.1186/s12985-024-02550-4Pilot-scale process development for recombinant adeno-associated virus (rAAV) production based on high-density Sf9 cell cultureXinran Li0Jieyi Gu1Haoquan Wu2Yuanyuan Xie3College of Pharmaceutical Science, Zhejiang University of TechnologyKanglin Biotec (Hangzhou) Co., LtdKanglin Biotec (Hangzhou) Co., LtdCollege of Pharmaceutical Science, Zhejiang University of TechnologyAbstract Background In recent years, gene therapy drugs have been widely marketed, and their effectiveness and potential have been confirmed. Thus, increasing their production on an industrial scale is critical. Recombinant adeno-associated viruses (rAAVs) are optimal vectors for gene therapy applications, and the baculovirus expression vector system (BEVS), which is based on Sf9 cell culture, is a common tool for rAAV production. Methods In this work, an Sf9 cell fed-batch process was developed using shake flasks. In the laboratory-scale bioreactor, four processes were selected as the key factors when carrying out the orthogonal experiment. On the basis of the equal P/V principle and considering the problem posed by air bubbles, a pilot-scale level bioreactor process was established. Results Here, we describe a method in which a BEVS was used to produce rAAV vectors, with the cell density increasing to 22.8 × 106 cells/mL and the rAAV titre increasing to 20 × 1011 VG/mL upon adding feed material. By resolving the problems associated with high-density cell culture and air bubbles, this process was successfully scaled to a 50 L pilot-scale level. Conclusions This successful experiment not only provides a technological basis for further scale-up but also guarantees product capacity. We hope that this development process can provide reference data for studying cell culture-based drug production.https://doi.org/10.1186/s12985-024-02550-4rAAVSf9Scale-upProcess developmentGene therapy |
| spellingShingle | Xinran Li Jieyi Gu Haoquan Wu Yuanyuan Xie Pilot-scale process development for recombinant adeno-associated virus (rAAV) production based on high-density Sf9 cell culture Virology Journal rAAV Sf9 Scale-up Process development Gene therapy |
| title | Pilot-scale process development for recombinant adeno-associated virus (rAAV) production based on high-density Sf9 cell culture |
| title_full | Pilot-scale process development for recombinant adeno-associated virus (rAAV) production based on high-density Sf9 cell culture |
| title_fullStr | Pilot-scale process development for recombinant adeno-associated virus (rAAV) production based on high-density Sf9 cell culture |
| title_full_unstemmed | Pilot-scale process development for recombinant adeno-associated virus (rAAV) production based on high-density Sf9 cell culture |
| title_short | Pilot-scale process development for recombinant adeno-associated virus (rAAV) production based on high-density Sf9 cell culture |
| title_sort | pilot scale process development for recombinant adeno associated virus raav production based on high density sf9 cell culture |
| topic | rAAV Sf9 Scale-up Process development Gene therapy |
| url | https://doi.org/10.1186/s12985-024-02550-4 |
| work_keys_str_mv | AT xinranli pilotscaleprocessdevelopmentforrecombinantadenoassociatedvirusraavproductionbasedonhighdensitysf9cellculture AT jieyigu pilotscaleprocessdevelopmentforrecombinantadenoassociatedvirusraavproductionbasedonhighdensitysf9cellculture AT haoquanwu pilotscaleprocessdevelopmentforrecombinantadenoassociatedvirusraavproductionbasedonhighdensitysf9cellculture AT yuanyuanxie pilotscaleprocessdevelopmentforrecombinantadenoassociatedvirusraavproductionbasedonhighdensitysf9cellculture |