利用sf9 - rhabdovirus阴性细胞系生产重组腺相关病毒(rAAV)载体的补批工艺

IF 2.2 4区 医学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY
Xinran Li, Jieyi Gu, Haoquan Wu, Yuanyuan Xie
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引用次数: 0

摘要

背景:基因治疗已有效地应用于许多生物学研究和许多遗传或癌症疾病的治疗。目前,重组腺相关病毒(raav)是用于基因治疗的主要传递载体之一。利用Sf9细胞制备的rAAV载体具有产率高、易于规模化、成本低等优点。方法:采用Sf9横纹图病毒阴性(Sf9- rvn)细胞对rAAV的生产工艺进行验证和优化,补料分批法提高了rAAV的生产滴度。结果:在间歇培养过程中,细胞密度达到12.9×106 cells/mL,约为间歇培养过程的两倍。rAAV滴度在进料批处理过程中也提高了约8倍,达到1.5×1012 VG/mL。通过生成不同血清型和目标基因(GOI)的raav,验证了优化的工艺,所有raav的生产滴度都大于1×1012 VG/mL。结论:我们利用Sf9- rvn细胞,开发了一种替代Sf9细胞的补料批量rAAV生产工艺,达到了监管标准。该工艺适用性好,rAAV滴度至少可达1×1012 VG/mL,高于文献报道的1011 VG/mL水平。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
A Fed-batch Process for the Production of Recombinant Adeno-Associated Virus (rAAV) Vectors Using the Sf9-Rhabdovirus-Negative Cell Line.

Background: Gene therapy has been effectively applied in many biological studies and for the treatment of many genetic or cancer diseases. Currently, Recombinant Adeno- Associated Viruses (rAAVs) are one of the main types of delivery vectors used for gene therapy. rAAV vectors produced via the Sf9 cells have the advantages of high rAAV yields, easy scaleup, and low cost.

Method: Here, we used Sf9 rhabdovirus-negative (Sf9-RVN) cells to validate and optimize the rAAV production process, and the fed-batch process increased the rAAV production titre.

Results: In the fed-batch procedure, the cell density reached 12.9×106 cells/mL, which was approximately twice as high as in the batch culture process. The rAAV titre was also approximately 8-fold higher in the fed-batch process, reaching 1.5×1012 VG/mL. The optimized process was validated by generating rAAVs with various serotypes and genes of interest (GOI), all of which gave production titres greater than 1×1012 VG/mL.

Conclusion: We used Sf9-RVN cells to develop a fed-batch rAAV production process that replaces Sf9 cells to meet regulatory standards. This process has good applicability, and the rAAV titre can reach at least 1×1012 VG/mL, which is higher than the level of 1011 VG/mL reported in the literature.

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来源期刊
Current pharmaceutical biotechnology
Current pharmaceutical biotechnology 医学-生化与分子生物学
CiteScore
5.60
自引率
3.60%
发文量
203
审稿时长
6 months
期刊介绍: Current Pharmaceutical Biotechnology aims to cover all the latest and outstanding developments in Pharmaceutical Biotechnology. Each issue of the journal includes timely in-depth reviews, original research articles and letters written by leaders in the field, covering a range of current topics in scientific areas of Pharmaceutical Biotechnology. Invited and unsolicited review articles are welcome. The journal encourages contributions describing research at the interface of drug discovery and pharmacological applications, involving in vitro investigations and pre-clinical or clinical studies. Scientific areas within the scope of the journal include pharmaceutical chemistry, biochemistry and genetics, molecular and cellular biology, and polymer and materials sciences as they relate to pharmaceutical science and biotechnology. In addition, the journal also considers comprehensive studies and research advances pertaining food chemistry with pharmaceutical implication. Areas of interest include: DNA/protein engineering and processing Synthetic biotechnology Omics (genomics, proteomics, metabolomics and systems biology) Therapeutic biotechnology (gene therapy, peptide inhibitors, enzymes) Drug delivery and targeting Nanobiotechnology Molecular pharmaceutics and molecular pharmacology Analytical biotechnology (biosensing, advanced technology for detection of bioanalytes) Pharmacokinetics and pharmacodynamics Applied Microbiology Bioinformatics (computational biopharmaceutics and modeling) Environmental biotechnology Regenerative medicine (stem cells, tissue engineering and biomaterials) Translational immunology (cell therapies, antibody engineering, xenotransplantation) Industrial bioprocesses for drug production and development Biosafety Biotech ethics Special Issues devoted to crucial topics, providing the latest comprehensive information on cutting-edge areas of research and technological advances, are welcome. Current Pharmaceutical Biotechnology is an essential journal for academic, clinical, government and pharmaceutical scientists who wish to be kept informed and up-to-date with the latest and most important developments.
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