Optimized TAL effector nucleases (TALENs) for use in treatment of sickle cell disease
Optimized TAL effector nucleases (TALENs) for use in treatment of sickle cell disease
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DOI：
10.1039/c2mb05461b
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发表时间：
2012-01-01
影响因子：
--
通讯作者：
Zhao, Huimin
中科院分区：
文献类型：
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作者：
Sun, Ning;Liang, Jing;Zhao, Huimin
TAL效应物核酸酶（TALEN）代表了一类能够在体内切割长的特异性靶DNA序列的新型人工核酸酶，并且是具有潜在治疗应用的基因组编辑的有力工具。在这里，我们报告了一对定制设计的TALEN，用于人类细胞中镰状细胞病突变的靶向遗传校正，这代表了能够识别和切割人类疾病相关基因的工程TALEN的一个例子。通过使用酵母报告系统，进行了系统的研究，以优化TALEN结构，以获得最大的体内切割效率。与之前的报道相反，工程化的TALEN能够识别和切割前面有A、C或G的靶结合位点。更重要的是，优化的TALEN有效地切割了与镰状细胞病相关的人β-珠蛋白（HBB）基因内的靶序列，并将人细胞中靶向基因修复的效率提高了> 1000倍。此外，这些TALEN未显示出可检测的细胞毒性。这些结果证明了优化的TALEN作为治疗应用的强大基因组编辑工具的潜力。
TAL effector nucleases (TALENs) represent a new class of artificial nucleases capable of cleaving long, specific target DNA sequences in vivo and are powerful tools for genome editing with potential therapeutic applications. Here we report a pair of custom-designed TALENs for targeted genetic correction of the sickle cell disease mutation in human cells, which represents an example of engineered TALENs capable of recognizing and cleaving a human disease-associated gene. By using a yeast reporter system, a systematic study was carried out to optimize TALEN architecture for maximal in vivo cleavage efficiency. In contrast to the previous reports, the engineered TALENs were capable of recognizing and cleaving target binding sites preceded by A, C or G. More importantly, the optimized TALENs efficiently cleaved a target sequence within the human beta-globin (HBB) gene associated with sickle cell disease and increased the efficiency of targeted gene repair by > 1000-fold in human cells. In addition, these TALENs showed no detectable cytotoxicity. These results demonstrate the potential of optimized TALENs as a powerful genome editing tool for therapeutic applications.