Adeno-associated virus vector enables safe and efficient Cas9 activation in neonatal and adult Cas9 knockin murine cochleae.,Gene Therapy

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Adeno-associated virus vector enables safe and efficient Cas9 activation in neonatal and adult Cas9 knockin murine cochleae.
Gene Therapy ( IF 4.6 ) Pub Date : 2020-01-31 , DOI: 10.1038/s41434-020-0124-1
Wen Kang _{1,

2,

3} , Xingle Zhao _{1,

2,

3} , Zhuoer Sun _{1,

2,

3} , Tingting Dong _{1,

2,

3} , Chenxi Jin _{1,

2,

3} , Ling Tong _{1,

2,

3} , Weidong Zhu _{1,

2,

3} , Yong Tao _{1,

2,

3} , Hao Wu _{1,

2,

3}

Affiliation

Adeno-associated virus (AAV)-mediated gene delivery systems have been shown to be effective tools for gene manipulation in the inner ear. For example, hair cells (HCs) and multiple other cell types can be transduced by the local injection of AAVs into the inner ear. However, application of the AAV-mediated CRISPR/Cas9 gene-editing approach to the inner ear in adult mice has not yet been studied. Based on our previous work, we investigated several AAV serotypes in neonatal and adult mice in parallel, and found that AAV8 had the top efficiency to transduce inner HCs. We then tested the ability of Cre-expressing AAV8 to activate Cas9 in floxed-Cas9 knockin mice, and observed significant Cas9 activation in the inner ear of both neonatal and adult animals. Neither the AAV8 virus itself nor the surgical procedures used to deliver it-cochleostomy for neonatal mice and canalostomy for adult mice-caused any damage to HCs or impaired normal hearing. Our studies indicate that the local injection of AAV8-Cre can induce Cas9 activation to perform safe and efficient gene editing in the inner ear, expanding the repertoire of gene-editing tools for regulating gene expression in the inner ear as a part of efforts to rescue genetic hearing loss, initiate regeneration of HCs, or develop gene therapy techniques.

中文翻译：

腺相关病毒载体能够在新生儿和成人 Cas9 敲入小鼠耳蜗中安全有效地激活 Cas9。

腺相关病毒 (AAV) 介导的基因传递系统已被证明是在内耳中进行基因操作的有效工具。例如，毛细胞 (HC) 和多种其他细胞类型可以通过将 AAV 局部注射到内耳中来进行转导。然而，尚未研究将 AAV 介导的 CRISPR/Cas9 基因编辑方法应用于成年小鼠的内耳。基于我们之前的工作，我们同时研究了新生小鼠和成年小鼠的几种 AAV 血清型，发现 AAV8 具有最高的转导内部 HC 的效率。然后，我们测试了表达 Cre 的 AAV8 在 floxed-Cas9 敲入小鼠中激活 Cas9 的能力，并观察到新生和成年动物内耳中的显着 Cas9 激活。AAV8 病毒本身和用于递送它的外科手术——新生小鼠的耳蜗造口术和成年小鼠的耳道造口术——都不会对 HCs 造成任何损害或正常听力受损。我们的研究表明，局部注射 AAV8-Cre 可以诱导 Cas9 激活在内耳中进行安全有效的基因编辑，从而扩展了用于调节内耳基因表达的基因编辑工具库，作为拯救工作的一部分遗传性听力损失，启动 HC 的再生，或开发基因治疗技术。

更新日期：2020-01-31

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