A new generation of retroviral producer cells: predictable and stable virus production by Flp-mediated site-specific integration of retroviral vectors.

2.50
Hdl Handle:
http://hdl.handle.net/10033/14594
Title:
A new generation of retroviral producer cells: predictable and stable virus production by Flp-mediated site-specific integration of retroviral vectors.
Authors:
Schucht, R; Coroadinha, A S; Zanta-Boussif, M A; Verhoeyen, E; Carrondo, M J T; Hauser, H; Wirth, Dagmar
Abstract:
We developed a new strategy that provides well-defined high-titer producer cells for recombinant retroviruses in a minimum amount of time. The strategy involves the targeted integration of the retroviral vector into a chromosomal locus with favorable properties. For proof of concept we established a novel HEK293-based retroviral producer cell line, called Flp293A, with a single-copy retroviral vector integrated at a selected chromosomal locus. The vector was flanked by noninteracting Flp-recombinase recognition sites and was exchanged for different retroviral vectors via Flp-mediated cassette exchange. All analyzed cell clones showed correct integration and identical titers for each of the vectors, confirming that the expression characteristics from the parental cell were preserved. Titers up to 2.5 x 10(7) infectious particles/10(6) cells were obtained. Also, high-titer producer cells for a therapeutic vector that encodes the 8.9-kb collagen VII cDNA in a marker-free cassette were obtained within 3 weeks without screening. Thus, we provide evidence that the precise integration of viral vectors into a favorable chromosomal locus leads to high and predictable virus production. This method is compatible with other retroviral vectors, including self-inactivating vectors and marker-free vectors. Further, it provides a tool for evaluation of different retroviral vector designs.
Citation:
Mol. Ther. 2006, 14(2):285-92
Issue Date:
1-Aug-2006
URI:
http://hdl.handle.net/10033/14594
DOI:
10.1016/j.ymthe.2005.12.003
PubMed ID:
16697259
Type:
Article
Language:
en
ISSN:
1525-0016
Appears in Collections:
Publications of Dept. Gene Regulation and Differentiation (RDIF)

Full metadata record

DC FieldValue Language
dc.contributor.authorSchucht, R-
dc.contributor.authorCoroadinha, A S-
dc.contributor.authorZanta-Boussif, M A-
dc.contributor.authorVerhoeyen, E-
dc.contributor.authorCarrondo, M J T-
dc.contributor.authorHauser, H-
dc.contributor.authorWirth, Dagmar-
dc.date.accessioned2007-11-16T14:34:49Z-
dc.date.available2007-11-16T14:34:49Z-
dc.date.issued2006-08-01-
dc.identifier.citationMol. Ther. 2006, 14(2):285-92en
dc.identifier.issn1525-0016-
dc.identifier.pmid16697259-
dc.identifier.doi10.1016/j.ymthe.2005.12.003-
dc.identifier.urihttp://hdl.handle.net/10033/14594-
dc.description.abstractWe developed a new strategy that provides well-defined high-titer producer cells for recombinant retroviruses in a minimum amount of time. The strategy involves the targeted integration of the retroviral vector into a chromosomal locus with favorable properties. For proof of concept we established a novel HEK293-based retroviral producer cell line, called Flp293A, with a single-copy retroviral vector integrated at a selected chromosomal locus. The vector was flanked by noninteracting Flp-recombinase recognition sites and was exchanged for different retroviral vectors via Flp-mediated cassette exchange. All analyzed cell clones showed correct integration and identical titers for each of the vectors, confirming that the expression characteristics from the parental cell were preserved. Titers up to 2.5 x 10(7) infectious particles/10(6) cells were obtained. Also, high-titer producer cells for a therapeutic vector that encodes the 8.9-kb collagen VII cDNA in a marker-free cassette were obtained within 3 weeks without screening. Thus, we provide evidence that the precise integration of viral vectors into a favorable chromosomal locus leads to high and predictable virus production. This method is compatible with other retroviral vectors, including self-inactivating vectors and marker-free vectors. Further, it provides a tool for evaluation of different retroviral vector designs.en
dc.format.extent387713 bytes-
dc.format.mimetypeapplication/pdf-
dc.language.isoenen
dc.titleA new generation of retroviral producer cells: predictable and stable virus production by Flp-mediated site-specific integration of retroviral vectors.en
dc.typeArticleen
dc.format.digYES-

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