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A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors

Recombinant adeno-associated viral (rAAV) vectors have been widely used in human gene therapy. One major impediment to its broad application is the inability to produce high-quality vectors in mass quantity. Here, an efficient and scalable suspension cell culture system for the production of rAAV ve...

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Autores principales: Wang, Qizhao, Wu, Zhongren, Zhang, Junping, Firrman, Jenni, Wei, Hongying, Zhuang, Zhengjing, Liu, LinShu, Miao, Linqing, Hu, Yang, Li, Dong, Diao, Yong, Xiao, Weidong
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Society of Gene & Cell Therapy 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5721209/
https://www.ncbi.nlm.nih.gov/pubmed/29255740
http://dx.doi.org/10.1016/j.omtm.2017.11.002
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author Wang, Qizhao
Wu, Zhongren
Zhang, Junping
Firrman, Jenni
Wei, Hongying
Zhuang, Zhengjing
Liu, LinShu
Miao, Linqing
Hu, Yang
Li, Dong
Diao, Yong
Xiao, Weidong
author_facet Wang, Qizhao
Wu, Zhongren
Zhang, Junping
Firrman, Jenni
Wei, Hongying
Zhuang, Zhengjing
Liu, LinShu
Miao, Linqing
Hu, Yang
Li, Dong
Diao, Yong
Xiao, Weidong
author_sort Wang, Qizhao
collection PubMed
description Recombinant adeno-associated viral (rAAV) vectors have been widely used in human gene therapy. One major impediment to its broad application is the inability to produce high-quality vectors in mass quantity. Here, an efficient and scalable suspension cell culture system for the production of rAAV vectors is described. In this system, the AAV trans factors, Rep78, Rep52, VP1, VP2, and VP3, were stably integrated into a single vaccinia virus carrier by maximizing the use of alternative codons between genes with identical amino acids, and the cis rAAV genome was carried by an E1/E3 gene-deleted adenovirus. Infection of improved, E1 integrated, suspension-cultured cells with these two viral vectors resulted in the robust production of rAAV vectors. The newly enhanced system can consistently produce ∼1 × 10(15) genome containing rAAV vectors per liter of suspension cells. Moreover, the capsid composition of rAAV vectors produced by this system is markedly different from those produced using the traditional system in that the VP1 protein is more abundant than the VP2 protein (19:1 versus 1:1). The unique VP1 superabundant rAAV vectors produced in this new system exhibited improved transduction in vivo after intravitreal injection.
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spelling pubmed-57212092017-12-18 A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors Wang, Qizhao Wu, Zhongren Zhang, Junping Firrman, Jenni Wei, Hongying Zhuang, Zhengjing Liu, LinShu Miao, Linqing Hu, Yang Li, Dong Diao, Yong Xiao, Weidong Mol Ther Methods Clin Dev Article Recombinant adeno-associated viral (rAAV) vectors have been widely used in human gene therapy. One major impediment to its broad application is the inability to produce high-quality vectors in mass quantity. Here, an efficient and scalable suspension cell culture system for the production of rAAV vectors is described. In this system, the AAV trans factors, Rep78, Rep52, VP1, VP2, and VP3, were stably integrated into a single vaccinia virus carrier by maximizing the use of alternative codons between genes with identical amino acids, and the cis rAAV genome was carried by an E1/E3 gene-deleted adenovirus. Infection of improved, E1 integrated, suspension-cultured cells with these two viral vectors resulted in the robust production of rAAV vectors. The newly enhanced system can consistently produce ∼1 × 10(15) genome containing rAAV vectors per liter of suspension cells. Moreover, the capsid composition of rAAV vectors produced by this system is markedly different from those produced using the traditional system in that the VP1 protein is more abundant than the VP2 protein (19:1 versus 1:1). The unique VP1 superabundant rAAV vectors produced in this new system exhibited improved transduction in vivo after intravitreal injection. American Society of Gene & Cell Therapy 2017-11-07 /pmc/articles/PMC5721209/ /pubmed/29255740 http://dx.doi.org/10.1016/j.omtm.2017.11.002 Text en © 2017 The Author(s) http://creativecommons.org/licenses/by-nc-nd/4.0/ This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
spellingShingle Article
Wang, Qizhao
Wu, Zhongren
Zhang, Junping
Firrman, Jenni
Wei, Hongying
Zhuang, Zhengjing
Liu, LinShu
Miao, Linqing
Hu, Yang
Li, Dong
Diao, Yong
Xiao, Weidong
A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors
title A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors
title_full A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors
title_fullStr A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors
title_full_unstemmed A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors
title_short A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors
title_sort robust system for production of superabundant vp1 recombinant aav vectors
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5721209/
https://www.ncbi.nlm.nih.gov/pubmed/29255740
http://dx.doi.org/10.1016/j.omtm.2017.11.002
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