A new PG13-based packaging cell line for stable production of clinical-grade self-inactivating γ-retroviral vectors using targeted integration

被引：43

作者：

Loew, R. ^{[1
]}

Meyer, Y. ^{[1
]}

Kuehlcke, K. ^{[1
]}

Gama-Norton, L. ^{[2
,6
]}

Wirth, D. ^{[2
]}

Hauser, H. ^{[2
]}

Stein, S. ^{[3
]}

Grez, M. ^{[3
]}

Thornhill, S. ^{[4
]}

Thrasher, A. ^{[4
]}

Baum, C. ^{[5
]}

Schambach, A. ^{[5
]}

机构：

[1] EUFETS AG, D-55743 Idar Oberstein, Germany

[2] Helmholtz Ctr Infect Res, Braunschweig, Germany

[3] GSH, Frankfurt, Germany

[4] UCL, Inst Child Hlth, Mol Immunol Unit, Ctr Immunodeficiency, London, England

[5] Hannover Med Sch, Dept Expt Hematol, D-30625 Hannover, Germany

[6] Univ Nova Lisboa, Inst Biol Expt & Tecnol, P-2780156 Oeiras, Portugal

来源：

GENE THERAPY | 2010年 / 17卷 / 02期

关键词：

SIN vector production; stable packaging lines; gamma-retroviral vectors; SEVERE COMBINED IMMUNODEFICIENCY; DEFINED CHROMOSOMAL LOCI; GENE-THERAPY; ADENOSINE-DEAMINASE; HEMATOPOIETIC-CELLS; LENTIVIRAL VECTORS; MAMMALIAN-CELLS; MOUSE MODEL; EXPRESSION; DESIGN;

D O I：

10.1038/gt.2009.134

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

The clinical application of self-inactivating (SIN) retroviral vectors has been hampered by the lack of reliable and efficient vector production technologies. To enable production of SIN gamma-retroviral vectors from stable producer clones, a new PG13-based packaging cell, known as PG368, was developed. Viral vector expression constructs can be reliably inserted at a predefined genomic locus of PG368 packaging cells by an Flp-recombinase-mediated targeted cassette exchange (RMCE) reaction. A new, carefully designed vector-targeting construct, pEMTAR-1, eliminated the co-packaging of the selectable marker gene used for the identification of successful recombination at the predefined genomic locus and thus, improved the safety of the production system. Selected clones produced vector supernatants at consistent titers. The targeted insertion of therapeutically relevant SIN vectors for chronic granulomatous disease and X-linked severe combined immunodeficiency into PG368 cells results in stable titers within the range necessary for clinical application. The production of retroviral SIN vectors from stable clinical-grade producer cells is feasible and will contribute to the safe production and application of SIN gamma-retroviral vectors for clinical trials. Gene Therapy (2010) 17, 272-280; doi: 10.1038/gt.2009.134; published online 29 October 2009

引用

页码：272 / 280

页数：9

共 43 条

[1] IDENTIFICATION OF A SIGNAL IN A MURINE RETROVIRUS THAT IS SUFFICIENT FOR PACKAGING OF NONRETROVIRAL RNA INTO VIRIONS [J].