Xu Zhen, Chen Feng, Zhang Lingling, Lu Jing, Xu Peng, Liu Guang, Xie Xuemin, Mu Wenli, Wang Yajun, Liu Depei
State Key Laboratory of Medical Molecular Biology, Department of Biochemistry and Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100005, China.
Sci China Life Sci. 2016 Oct;59(10):1024-1033. doi: 10.1007/s11427-016-0067-0. Epub 2016 Sep 6.
Safe and efficient gene transfer systems are the basis of gene therapy applications. Non-integrating lentiviral (NIL) vectors are among the most promising candidates for gene transfer tools, because they exhibit high transfer efficiency in both dividing and non-dividing cells and do not present a risk of insertional mutagenesis. However, non-integrating lentiviral vectors cannot introduce stable exogenous gene expression to dividing cells, thereby limiting their application. Here, we report the design of a non-integrating lentiviral vector that contains the minimal scaffold/matrix attachment region (S/MAR) sequence (SNIL), and this SNIL vector is able to retain episomal transgene expression in dividing cells. Using SNIL vectors, we detected the expression of the eGFP gene for 61 days in SNIL-transduced stable CHO cells, either with selection or not. In the NIL group without the S/MAR sequence, however, the transduced cells died under selection for the transient expression of NIL vectors. Furthermore, Southern blot assays demonstrated that the SNIL vectors were retained extrachromosomally in the CHO cells. In conclusion, the minimal S/MAR sequence retained the non-integrating lentiviral vectors in dividing cells, which indicates that SNIL vectors have the potential for use as a gene transfer tool.
安全有效的基因转移系统是基因治疗应用的基础。非整合型慢病毒（NIL）载体是基因转移工具中最具潜力的候选者之一，因为它们在分裂细胞和非分裂细胞中均表现出高转移效率，且不存在插入诱变风险。然而，非整合型慢病毒载体无法将稳定的外源基因表达引入分裂细胞，从而限制了它们的应用。在此，我们报告了一种包含最小支架/基质附着区域（S/MAR）序列（SNIL）的非整合型慢病毒载体的设计，并且这种SNIL载体能够在分裂细胞中维持附加型转基因表达。使用SNIL载体，我们在经SNIL转导的稳定CHO细胞中检测到eGFP基因表达长达61天，无论有无选择压力。然而，在没有S/MAR序列的NIL组中，转导细胞在选择压力下因NIL载体的瞬时表达而死亡。此外，Southern印迹分析表明，SNIL载体以染色体外形式保留在CHO细胞中。总之，最小S/MAR序列使非整合型慢病毒载体在分裂细胞中得以保留，这表明SNIL载体有潜力用作基因转移工具。
