A robust system for production of superabundant VP1 recombinant AAV vectors

Authors: WANG, QIZHAO - Temple University Medical School; WU, ZHONGREN - Temple University Medical School; ZHANG, JUNPING - Temple University Medical School; Firrman, Jenni; WEI, HONGYING - Temple University Medical School; ZHUANG, ZHENJING - School Of Biomedical Sciences, Huaqiao University; Liu, Linshu; MIAO, LINQING - Shriners Hospital For Children; HU, YANG - Stanford University School Of Medicine; DIAO, YONG - School Of Biomedical Sciences, Huaqiao University; XIAO, WEIDONG - Temple University Medical School

Submitted to: Molecular Therapy
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/1/2017
Publication Date: 11/7/2017
Citation: Wang, Q., Wu, Z., Zhang, J., Firrman, J., Wei, H., Zhuang, Z., Liu, L.S., Miao, L., Hu, Y., Diao, Y., Xiao, W. 2017. A robust system for production of superabundant VP1 recombinant AAV vectors. Molecular Therapy. 7:146-156. https://doi.org/10.1016/j.omtm.2017.11.002.
DOI: https://doi.org/10.1016/j.omtm.2017.11.002

Interpretive Summary: Gene therapy is a type of treatment for disease that works by delivering a gene into a human cell. Gene therapy using a small, safe virus called Adeno-Associated Virus (AAV) as a vehicle to deliver the gene into the cell has produced promising results. However, preparing AAV in large batches is a difficult process, and it is hard to make enough quality virus to use for human clinical trials. In this paper, a new method to make AAV is described. This method uses two types of viruses, a vaccinia virus and an Adenovirus/ AAV hybrid virus, to make AAV. This new method is better than the conventional method because more AAV can be made at one time, using less resources. Not only is this new method better at making AAV, but it also makes a new type of AAV, one that has changes to the exterior. The AAV with these changes are called VP1 superabundant rAAV vectors. The VP1 superabundant rAAV vectors are more infective, and therefore can improve gene therapy. In conclusion, a new method to make AAV was identified and described. This new method is not only more efficient at making AAV, but makes a new type of AAV that is more infective. This can be used to make AAV for gene therapy clinical trials.

Technical Abstract: 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 an 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 approximately 1x10(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 vs 1:1). The unique VP1 superabundant rAAV vectors produced in this new system exhibited improved transduction in vivo after intravitreal injection.