Efficient gene transfer into rhesus repopulating hematopoietic stem cells using a simian immunodeficiency virus-based lentiviral vector system
Efficient gene transfer into rhesus repopulating hematopoietic stem cells using a simian immunodeficiency virus-based lentiviral vector system
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DOI：
10.1182/blood-2004-01-0045
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发表时间：
2004-06-01
期刊：
影响因子：
20.3
通讯作者：
Nienhuis, AW
中科院分区：
文献类型：
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作者：
Hanawa, H;Hematti, P;Nienhuis, AW
发现高滴度的基于HIV-1的慢病毒载体颗粒非常差地（< 1%）诱导了甘氨酸动员的恒河猴CD 34（+）细胞和克隆生成祖细胞，反映了恒河猴细胞对HIV感染的感染后限制。为了克服这一障碍，我们开发了一种基于猿猴免疫缺陷病毒（SIV）的载体系统。单次暴露于低浓度的编码绿色荧光蛋白（GFP）的嗜中性假型SIV载体颗粒导致基因转移到68% +/-1%的恒河猴本体CD 34（+）细胞和75% +/-1%的克隆原祖细胞中。聚合酶链反应（PCR）分析DNA从个别造血集落证实了这些相对转导效率。为了评估SIV载体介导的干细胞基因转移在体内，3只恒河猴接受了骨髓成白细胞预处理后转导的自体精氨酸动员的外周血CD 34+细胞移植。造血重建迅速，移植后4至6个月，分别观察到平均18% +/- 8%和15% +/- 7%的GFP阳性粒细胞和单核细胞，与实时PCR测定的外周血白细胞中0.19 +/- 0.05的平均载体拷贝数一致。载体插入位点分析证明了用含有载体的细胞的多克隆重建。SIV载体似乎有希望在非人灵长类动物模型中评估基因治疗方法。(C)2004年美国血液学会
High-titer, HIV-1-based lentiviral vector particles were found to traniduce cytokine-mobilized rhesus macaque CD34(+) cells and clonogenic progenitors very poorly (< 1%), reflecting the postentry restriction in rhesus cells to HIV infection. To overcome this barrier, we developed a simian immunodeficiency virus (SIV)-based vector system. A single exposure to a low concentration of amphotropic pseudotyped SIV vector particles encoding the green fluorescent protein (GFP) resulted in gene transfer into 68% +/- 1 % of rhesus bulk CD34(+) cells and 75% +/- 1% of clonogenic progenitors. Polymerase chain reaction (PCR) analysis of DNA from individual hematopoietic colonies confirmed these relative transduction efficiencies. To evaluate SIV vector-mediated stem cell gene transfer in vivo, 3 rhesus macaques underwent transplantation with transduced, autologous cytokine-mobilized peripheral blood CD34+ cells following myeloalblative conditioning. Hematopoietic reconstitution was rapid, and an average of 18% +/- 8% and 15% +/- 7% GFP-positive granulocytes and monocyles, respectively, were observed 4 to 6 months after transplantation, consistent with the average vector copy number of 0.19 +/- 0.05 in peripheral blood leukocytes as determined by real-time PCR. Vector insertion site analysis demonstrated polyclonal reconstitution with vector-containing cells. SIV vectors appear promising for evaluating gene therapy approaches in nonhuman primate models. (C) 2004 by The American Society of Hematology