Details

Autor(en) / Beteiligte

• Cai, Yujia
• Laustsen, Anders
• Zhou, Yan
• Sun, Chenglong
• Anderson, Mads Valdemar
• Li, Shengting
• Uldbjerg, Niels
• Luo, Yonglun
• Jakobsen, Martin R
• Mikkelsen, Jacob Giehm

Titel

Targeted, homology-driven gene insertion in stem cells by ZFN-loaded 'all-in-one' lentiviral vectors

Ist Teil von

• eLife, 2016-06, Vol.5

Ort / Verlag

England: eLife Science Publications, Ltd

Erscheinungsjahr

2016

Link zum Volltext

Quelle

MEDLINE

Beschreibungen/Notizen

• Biased integration remains a key challenge for gene therapy based on lentiviral vector technologies. Engineering of next-generation lentiviral vectors targeting safe genomic harbors for insertion is therefore of high relevance. In a previous paper (Cai et al., 2014a), we showed the use of integrase-defective lentiviral vectors (IDLVs) as carriers of complete gene repair kits consisting of zinc-finger nuclease (ZFN) proteins and repair sequences, allowing gene correction by homologous recombination (HR). Here, we follow this strategy to engineer ZFN-loaded IDLVs that insert transgenes by a homology-driven mechanism into safe loci. This insertion mechanism is driven by time-restricted exposure of treated cells to ZFNs. We show targeted gene integration in human stem cells, including CD34(+) hematopoietic progenitors and induced pluripotent stem cells (iPSCs). Notably, targeted insertions are identified in 89% of transduced iPSCs. Our findings demonstrate the applicability of nuclease-loaded 'all-in-one' IDLVs for site-directed gene insertion in stem cell-based gene therapies.