Specific transgene expression in human and mouse CD4+ cells using lentiviral vectors with regulatory sequences from the CD4 gene
Specific transgene expression in human and mouse CD4+ cells using lentiviral vectors with regulatory sequences from the CD4 gene
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DOI：
10.1182/blood-2002-02-0578
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发表时间：
2003-05-01
期刊：
影响因子：
20.3
通讯作者：
Klatzmann, D
中科院分区：
文献类型：
--
作者：
Marodon, G;Mouly, E;Klatzmann, D
通过基因转移载体实现治疗性转基因的细胞特异性表达代表基因治疗的主要目标。为了实现转基因在CD 4（+）细胞中的特异性表达，我们已经产生了表达增强型绿色荧光蛋白（eGFP）报告基因的慢病毒载体，该报告基因在来自CD 4基因的调控序列的控制下-最小启动子和近端增强子，有或没有沉默子。两种慢病毒载体都可以以高滴度（每毫升超过10（7）个感染性颗粒）产生，并用于培养健康的鼠造血干细胞（HSC）。在用转导的HSC重建RAG-2缺陷小鼠时，特异性载体在T细胞中有效表达，在B细胞中最低限度地表达，并且在骨髓的未成熟细胞中不表达。在载体调控序列中添加CD 4基因沉默元件导致重组小鼠和体外转导的人T细胞中eGFP表达进一步限制到CD 4（+）T细胞中。来自人CD 34（+）细胞的非T CD 4（+）树突状细胞和巨噬细胞在体外表达特定载体的转基因，尽管水平低于CD 4（+）T细胞。总之，我们已经产生了慢病毒载体，其允许在转导的小鼠HSC和人成熟T细胞分化后特异性靶向转基因表达至CD 4（+）细胞。最终，这些载体可能被证明可用于原位注射用于HIV感染或遗传性免疫缺陷的体内基因治疗。(C)2003年，美国血液学会。
Achieving cell-specific expression of a therapeutic transgene by gene transfer vectors represents a major goal for gene therapy. To achieve specific expression of a transgene in CD4(+) cells, we have generated lentiviral vectors expressing the enhanced green fluorescent protein (eGFP) reporter gene under the control of regulatory sequences derived from the CD4 gene-a minimal promoter and the proximal enhancer, with or without the silencer. Both lentiviral vectors could be produced at high titers (more than 10(7) infectious particles per milliliter) and were used to transduce healthy murine hematopoietic stem cells (HSCs). On reconstitution of RAG-2-deficient mice with transduced HSCs, the specific vectors were efficiently expressed in T cells, minimally expressed in B cells, and not expressed in immature cells of the bone marrow. Addition of the CD4 gene-silencing element in the vector regulatory sequences led to further restriction of eGFP expression into CD4(+) T cells in reconstituted mice and in ex vivo-transduced human T cells. Non-T CD4(+) dendritic and macrophage cells derived from human CD34(+) cells in vitro expressed the transgene of the specific vectors, albeit at lower levels than CD4(+) T cells. Altogether, we have generated lentiviral vectors that allow specific targeting of transgene expression to CD4(+) cells after differentiation of transduced mice HSCs and human mature T cells. Ultimately, these vectors may prove useful for in situ injections for in vivo gene therapy of HIV infection or genetic immunodeficiencies. (C) 2003 by The American Society of Hematology.