Inhibition of HIV-1 infection by lentiviral vectors expressing pol III-promoted anti-HIV RNAs
Inhibition of HIV-1 infection by lentiviral vectors expressing pol III-promoted anti-HIV RNAs
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DOI：
10.1016/s1525-0016(03)00165-5
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发表时间：
2003-08-01
影响因子：
12.4
通讯作者：
Rossi, JJ
中科院分区：
文献类型：
--
作者：
Li, MJ;Bauer, G;Rossi, JJ
慢病毒载体的一个主要优势是它们能够穿过核膜进入细胞核，从而允许转导未分裂的细胞。利用基于HIV的慢病毒载体，我们将抗CCR5核酶(CCR5RZ)，一种核仁定位的TAR RNA诱饵，或Pol III表达的siRNA基因导入培养细胞和原代细胞。CCR5RZ由腺病毒VA1 Pol III启动子驱动，而人U6 SnRNA Pol III转录的TAR诱骗物嵌入U16 snoRNA(命名为U16TAR)，siRNAs来自人U6 Pol III启动子。这些载体在293细胞中的转导效率为96-98%，在原代PBMC中的转导效率为15-20%。CCR5RZ和U16TAR诱骗在单个载体骨架中的组合在原代T细胞和CD34(+)来源的单核细胞中的选择性存活试验中对HIV-1攻击具有增强的保护作用。慢病毒载体骨架表达的siRNAs也显示出对HIV-1攻击的PBMC中p24表达的有效抑制。总之，我们的结果表明，慢病毒载体可以有效地将Pol III治疗性表达单位的单个构建和组合输送到原代造血细胞中进行抗HIV基因治疗，并有望用于基于干细胞或T细胞的HIV-1感染的基因治疗。
A primary advantage of lentiviral vectors is their ability to pass through the nuclear envelope into the cell nucleus thereby allowing transduction of nondividing cells. Using HIV-based lentiviral vectors, we delivered an anti-CCR5 ribozyme (CCR5RZ), a nucleolar localizing TAR RNA decoy, or Pol III-expressed siRNA genes into cultured and primary cells. The CCR5RZ is driven by the adenoviral VA1 Pol III promoter, while the human U6 snRNA Pol III-transcribed TAR decoy is embedded in a U16 snoRNA (designated U16TAR), and the siRNAs were expressed from the human U6 Pol III promoter. The transduction efficiencies of these vectors ranged from 96-98% in 293 cells to 15-20% in primary PBMCs. A combination of the CCR5RZ and U16TAR decoy in a single vector backbone gave enhanced protection against HIV-1 challenge in a selective survival assay in both primary T cells and CD34(+)-derived monocytes. The lentiviral vector backbone-expressed siRNAs also showed potent inhibition of p24 expression in PBMCs challenged with HIV-1. Overall our results demonstrate that the lentiviral-based vectors can efficiently deliver single constructs as well as combinations of Pol III therapeutic expression units into primary hematopoietic cells for anti-HIV gene therapy and hold promise for stem or T-cell-based gene therapy for HIV-1 infection.