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PMID: 9696794 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Packaging cells based on inducible gene amplification for the production of adeno-associated virus vectors.

Journal of virology ·Vol. 72 ·No. 9 ·1998-09-00 ·Pages 7024-31

Inoue N, Russell DW

Abstract

Although vectors based on adeno-associated virus (AAV) offer several unique advantages, their usage has been hampered by the difficulties encountered in vector production. In this report, we describe a new AAV packaging system based on inducible amplification of integrated helper and vector constructs containing the simian virus 40 (SV40) replication origin. The packaging and producer cell lines developed express SV40 T antigen under the control of the reverse tetracycline transactivator system, which allows inducible amplification of chromosomal loci linked to the SV40 origin. Culturing these cells in the presence of doxycycline followed by adenovirus infection resulted in helper and vector gene amplification as well as higher vector titers. Clonal producer cell lines generated vector titers that were 10 times higher than those obtained by standard methods, with approximately 10(4) vector particles produced per cell. These stocks were free of detectable replication-competent virus. The lack of a transfection step combined with the reproducibility of stable producer lines makes this packaging method ideally suited for the large-scale production of vector stocks for human gene therapy.

MeSH Terms
Cell Line, Transformed Dependovirus/genetics Gene Amplification Genetic Vectors/genetics HeLa Cells Humans Tumor Cells, Cultured
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Inoue N
Markey Molecular Medicine Center and Department of Medicine, University of Washington, Seattle, Washington 98195, USA.
Russell D W
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Article Info
Journal
Journal of virology
Abbr.
J Virol
ISSN
0022-538X
DOI
10.1128/JVI.72.9.7024-7031.1998
Published
1998-09-00
Pages
7024-31
Language
English
Region
United States
NLM ID
0113724
PMCID
PMC109922
Subset
IM
External Links
PubMed source DOI: 10.1128/JVI.72.9.7024-7031.1998 PMC Full Text View journal details
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