Process optimization for the rapid production of adenoviral vectors for clinical trials in a disposable bioreactor system
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Recombinant adenoviral (Ad) vectors are highly efficient gene transfer vectors widely used in vaccine development and immunotherapy. To promote the industrial application of Ad vectors, studies focusing on reducing the cost of manufacturing, shortening the preclinical research period, and improving the quality of products are needed. Here, we describe a highly efficient and economical process for producing Ad vector in a novel, single-use bioreactor system suitable for clinical trials. A mini-bioreactor was used for parameter optimization and development of medium replacement protocols for Ad5-GFP production before scale-up. HEK293 cell culture and virus infection were monitored in a disposable AmProtein Current Perfusion Bioreactor and Bioflo310 bioreactor using optimized parameters and medium replacement protocols. The total cell number increased from 2.0 × 109 to 3.2 × 1010 after 6 days of culture. The total number of viral particles obtained in a single batch was 1.2 × 1015. These results demonstrate the efficiency and suitability of this system for Ad vector production for research and GMP applications.
KeywordsAdenoviral vector Disposable bioreactor HEK293 cells Process optimization
We would like to thank AmProtein (Hangzhou, China) for their assistance with ACPB photos. We also thank the native English speaking scientists of Elixigen Company (Huntington Beach, California) for editing our manuscript.
This work was supported by the State Project of Essential Drug Research and Development (grant number 2015ZX09101044) and the Science & Technology Key Program of Zhejiang, China (grant number 2014C03039).
Compliance with ethical standards
This article does not contain any studies with human participants or animals performed by any of the authors.
Conflict of interest
The authors declare that they have no conflicts of interest.
- Hui M (2007) AmProtein corporation. A method to increase dissolved oxygen in a culture vessel. In: International patent #WO2007/142664Google Scholar
- Li L, Shi M, Song Y, Bao L, Yang W, Zhang X, Ruan M, Rishton G, Joudi A, Teng Y, Xing Y, Hu F, Zhao X, Zhang X, Li H, Leng G, Yuan S, Jia Q, Hui M (2009) A single-use, scalable perfusion bioreactor system. Bioprocess Int 7(6):46–54Google Scholar
- Liu H, Liu XM, Li SC, Wu BC, Ye LL, Wang QW, Chen ZL (2009) A high-yield and scaleable adenovirus vector production process based on high density perfusion culture of HEK 293 cells as suspended aggregates. J Biosci Bioeng 107(5):524–529. https://doi.org/10.1016/j.jbiosc.2009.01.004 CrossRefPubMedGoogle Scholar
- Meuwly F, Papp F, Ruffieux PA, Bernard AR, Kadouri A, von Stockar U (2006) Use of glucose consumption rate (GCR) as a tool to monitor and control animal cell production processes in packed-bed bioreactors. J Biotechnol 122(1):122–129. https://doi.org/10.1016/j.jbiotec.2005.08.005 CrossRefPubMedGoogle Scholar
- Qian J, Li H, Hui M, Hui N, Rishton AJG, Bao L, Shi M, Zhang X, Luanfeng L, Xu J, Leng G (2008) A bioreactor system based on a novel oxygen transfer method. Bioprocess Int 6(6):66–78Google Scholar
- Schoofs G, Monica TJ, Ayala J, Horwitz J, Montgomery T, Roth G, Castillo FJ (1998) A high-yielding serum-free, suspension cell culture process to manufacture recombinant adenoviral vectors for gene therapy. Cytotechnology 28(1–3):81–89. https://doi.org/10.1023/A:1008021428969 CrossRefPubMedPubMedCentralGoogle Scholar
- Sun B, Yu XH, Kong W, Sun SY, Yang P, Zhu CL, Zhang HH, Wu YG, Chen Y, Shi YH, Zhang XZ, Jiang CL (2013) Production of influenza H1N1 vaccine from MDCK cells using a novel disposable packed-bed bioreactor. Appl Microbiol Biotechnol 97(3):1063–1070. https://doi.org/10.1007/s00253-012-4375-7 CrossRefPubMedGoogle Scholar
- Whitfield RJ, Battom SE, Barut M, Gilham DE, Ball PD (2009) Rapid high-performance liquid chromatographic analysis of adenovirus type 5 particles with a prototype anion-exchange analytical monolith column. J Chromatog A 1216(13):2725–2729. https://doi.org/10.1016/j.chroma.2008.11.010 CrossRefGoogle Scholar