Lentiviral delivery of co-packaged Cas9 mRNA and a Vegfa-targeting guide RNA prevents wet age-related macular degeneration in mice
Lentiviral delivery of co-packaged Cas9 mRNA and a Vegfa-targeting guide RNA prevents wet age-related macular degeneration in mice
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DOI：
10.1038/s41551-020-00656-y
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发表时间：
2021-01-04
影响因子：
28.1
通讯作者：
Cai, Yujia
中科院分区：
文献类型：
--
作者：
Ling, Sikai;Yang, Shiqi;Cai, Yujia
治疗性基因组编辑需要有效和有针对性的传递方法。用腺相关病毒携带Cas9基因可以产生很强的体内治疗效果，但也可能导致持续的Cas9表达、抗Cas9免疫反应和非靶点编辑。慢病毒载体已经被设计成运送瞬时表达的核酸酶，但在体内缺乏其生物医学效果的证据。在这里，我们证明了慢病毒共传递化脓性链球菌Cas9 mRNA和编码针对血管内皮生长因子A(Vegfa)的引导RNA的表达盒在由Vegfa诱导的湿性年龄相关性黄斑变性小鼠模型中是有效的。一次视网膜下注射工程慢病毒，在视网膜色素上皮中杀死44%的Vegfa，并将脉络膜新生血管的面积减少63%，而不会引发偏离目标的编辑或抗Cas9免疫反应。针对核酸酶瞬时表达的工程慢病毒可能形成治疗视网膜新生血管疾病的新方法的基础。慢病毒共包装编码SpCas9 mRNA的表达盒和针对Vegfa基因的引导RNA，可防止小鼠患上Vegfa诱导的湿性老年性黄斑变性。
Therapeutic genome editing requires effective and targeted delivery methods. The delivery of Cas9 mRNA using adeno-associated viruses has led to potent in vivo therapeutic efficacy, but can cause sustained Cas9 expression, anti-Cas9 immune responses and off-target edits. Lentiviral vectors have been engineered to deliver nucleases that are expressed transiently, but in vivo evidence of their biomedical efficacy is lacking. Here, we show that the lentiviral codelivery of Streptococcus pyogenesCas9 mRNA and expression cassettes that encode a guide RNA that targets vascular endothelial growth factor A (Vegfa) is efficacious in a mouse model of wet age-related macular degeneration induced by Vegfa. A single subretinal injection of engineered lentiviruses knocked out 44% of Vegfa in retinal pigment epithelium and reduced the area of choroidal neovascularization by 63% without inducing off-target edits or anti-Cas9 immune responses. Engineered lentiviruses for the transient expression of nucleases may form the basis of new treatments for retinal neovascular diseases.Lentiviruses co-packaging expression cassettes that encode SpCas9 mRNA and a guide RNA targeting the Vegfa gene prevent mice from developing wet age-related macular degeneration induced by Vegfa.