2.50
Hdl Handle:
http://hdl.handle.net/10033/129836
Title:
Retroviral vector performance in defined chromosomal Loci of modular packaging cell lines.
Authors:
Gama-Norton, L; Herrmann, S; Schucht, R; Coroadinha, A S; Löw, R; Alves, P M; Bartholomae, C C; Schmidt, M; Baum, C; Schambach, A; Hauser, H; Wirth, D
Abstract:
The improvement of safety and titer of retroviral vectors produced in standard retroviral packaging cell lines is hampered because production relies on uncontrollable vector integration events. The influences of chromosomal surroundings make it difficult to dissect the performance of a specific vector from the chromosomal surroundings of the respective integration site. Taking advantage of a technology that relies on the use of packaging cell lines with predefined integration sites, we have systematically evaluated the performance of several retroviral vectors. In two previously established modular packaging cell lines (Flp293A and 293 FLEX) with single, defined chromosomal integration sites, retroviral vectors were integrated by means of Flp-mediated site-specific recombination. Vectors that are distinguished by different long terminal repeat promoters were introduced in either the sense or reverse orientation. The results show that the promoter, viral vector orientation, and integration site are the main determinants of the titer. Furthermore, we exploited the viral production systems to evaluate read-through activity. Read-through is thought to be caused by inefficient termination of vector transcription and is inherent to the nature of retroviral vectors. We assessed the frequency of transduction of sequences flanking the retroviral vectors from both integration sites. The approach presented here provides a platform for systematic design and evaluation of the efficiency and safety of retroviral vectors optimized for a given producer cell line.
Affiliation:
Helmholtz Center for Infection Research (HZI), 38124 Braunschweig, Germany.
Citation:
Retroviral vector performance in defined chromosomal Loci of modular packaging cell lines. 2010, 21 (8):979-91 Hum. Gene Ther.
Journal:
Human gene therapy
Issue Date:
Aug-2010
URI:
http://hdl.handle.net/10033/129836
DOI:
10.1089/hum.2009.089
PubMed ID:
20222806
Type:
Article
Language:
en
ISSN:
1557-7422
Appears in Collections:
Publications of Dept. Gene Regulation and Differentiation (RDIF)

Full metadata record

DC FieldValue Language
dc.contributor.authorGama-Norton, Len
dc.contributor.authorHerrmann, Sen
dc.contributor.authorSchucht, Ren
dc.contributor.authorCoroadinha, A Sen
dc.contributor.authorLöw, Ren
dc.contributor.authorAlves, P Men
dc.contributor.authorBartholomae, C Cen
dc.contributor.authorSchmidt, Men
dc.contributor.authorBaum, Cen
dc.contributor.authorSchambach, Aen
dc.contributor.authorHauser, Hen
dc.contributor.authorWirth, Den
dc.date.accessioned2011-05-20T10:42:53Z-
dc.date.available2011-05-20T10:42:53Z-
dc.date.issued2010-08-
dc.identifier.citationRetroviral vector performance in defined chromosomal Loci of modular packaging cell lines. 2010, 21 (8):979-91 Hum. Gene Ther.en
dc.identifier.issn1557-7422-
dc.identifier.pmid20222806-
dc.identifier.doi10.1089/hum.2009.089-
dc.identifier.urihttp://hdl.handle.net/10033/129836-
dc.description.abstractThe improvement of safety and titer of retroviral vectors produced in standard retroviral packaging cell lines is hampered because production relies on uncontrollable vector integration events. The influences of chromosomal surroundings make it difficult to dissect the performance of a specific vector from the chromosomal surroundings of the respective integration site. Taking advantage of a technology that relies on the use of packaging cell lines with predefined integration sites, we have systematically evaluated the performance of several retroviral vectors. In two previously established modular packaging cell lines (Flp293A and 293 FLEX) with single, defined chromosomal integration sites, retroviral vectors were integrated by means of Flp-mediated site-specific recombination. Vectors that are distinguished by different long terminal repeat promoters were introduced in either the sense or reverse orientation. The results show that the promoter, viral vector orientation, and integration site are the main determinants of the titer. Furthermore, we exploited the viral production systems to evaluate read-through activity. Read-through is thought to be caused by inefficient termination of vector transcription and is inherent to the nature of retroviral vectors. We assessed the frequency of transduction of sequences flanking the retroviral vectors from both integration sites. The approach presented here provides a platform for systematic design and evaluation of the efficiency and safety of retroviral vectors optimized for a given producer cell line.en
dc.language.isoenen
dc.subject.meshCell Lineen
dc.subject.meshChromosomesen
dc.subject.meshGene Targetingen
dc.subject.meshGene Therapyen
dc.subject.meshGenetic Locien
dc.subject.meshGenetic Vectorsen
dc.subject.meshPromoter Regions, Geneticen
dc.subject.meshRetroviridaeen
dc.subject.meshTransduction, Geneticen
dc.subject.meshVirus Assemblyen
dc.subject.meshVirus Integrationen
dc.titleRetroviral vector performance in defined chromosomal Loci of modular packaging cell lines.en
dc.typeArticleen
dc.contributor.departmentHelmholtz Center for Infection Research (HZI), 38124 Braunschweig, Germany.en
dc.identifier.journalHuman gene therapyen

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