A Robust System for Production of Superabundant VP1 Recombinant AAV Vectors

• Published in: Molecular therapy. Methods & clinical development Vol. 7; no. C; pp. 146 - 156
• Main Authors: Wang, Qizhao, Wu, Zhongren, Zhang, Junping, Firrman, Jenni, Wei, Hongying, Zhuang, Zhengjing, Liu, LinShu, Miao, Linqing, Hu, Yang, Li, Dong, Diao, Yong, Xiao, Weidong
• Format: Journal Article
• Language: English
• Published: United States Elsevier Limited 15.12.2017; American Society of Gene & Cell Therapy; Elsevier
• Subjects: AAV; Cell culture; Codons; Cytomegalovirus; E3 gene; Experiments; Expression vectors; Gene therapy; Genes; Genomes; Immunoglobulins; Infections; Proteins; vaccinia virus; vector production; VP1; VP1 protein; VP2 protein; VP3 protein; gene therapy; vaccinia virus; AAV; VP1; vector production
• ISSN: 2329-0501; 2329-0501
• DOI: 10.1016/j.omtm.2017.11.002

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 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 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 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 after intravitreal injection.