期刊
MOLECULAR THERAPY
卷 21, 期 6, 页码 1259-1269出版社
NATURE PUBLISHING GROUP
DOI: 10.1038/mt.2013.65
关键词
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资金
- California Institute for Regenerative Medicine (CIRM) [DR1-01490]
- Swiss National Science Foundation [PBZHP3-133452]
- Swiss National Science Foundation (SNF) [PBZHP3-133452] Funding Source: Swiss National Science Foundation (SNF)
The HIV-1 coreceptor CCR5 is a validated target for HIV/AIDS therapy. The apparent elimination of HIV-1 in a patient treated with an allogeneic stem cell transplant homozygous for a naturally occurring CCR5 deletion mutation (CCR5(Delta 32/Delta 32)) supports the concept that a single dose of HIV-resistant hematopoietic stem cells can provide disease protection. Given the low frequency of naturally occurring CCR5(Delta 32/Delta 32) donors, we reasoned that engineered autologous CD34(+) hematopoietic stem/progenitor cells (HSPCs) could be used for AIDS therapy. We evaluated disruption of CCR5 gene expression in HSPCs isolated from granulocyte colony-stimulating factor (CSF)-mobilized adult blood using a recombinant adenoviral vector encoding a CCR5-specific pair of zinc finger nucleases (CCR5-ZFN). Our results demonstrate that CCR5-ZFN RNA and protein expression from the adenoviral vector is enhanced by pretreatment of HSPC with protein kinase C (PKC) activators resulting in >25% CCR5 gene disruption and that activation of the mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) signaling pathway is responsible for this activity. Importantly, using an optimized dose of PKC activator and adenoviral vector we could generate CCR5-modified HSPCs which engraft in a humanized mouse model (albeit at a reduced level) and support multilineage differentiation in vitro and in vivo. Together, these data establish the basis for improved approaches exploiting adenoviral vector delivery in the modification of HSPCs.
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