Adenine Base Editing In Vivo with a Single Adeno-Associated Virus Vector.
Adenine Base Editing In Vivo with a Single Adeno-Associated Virus Vector.
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DOI：
10.1089/genbio.2022.0015
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发表时间：
2022-06
期刊：
影响因子：
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通讯作者：
Han Zhang;Nathan Bamidele;Pengpeng Liu;O. Ojelabi;Xin D Gao;Tom S Rodriguez;Hao-Feng Cheng;
中科院分区：
文献类型：
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作者：
Han Zhang;Nathan Bamidele;Pengpeng Liu;O. Ojelabi;Xin D Gao;Tom S Rodriguez;Hao-Feng Cheng;
碱基编辑器为遗传疾病的治疗开辟了新的途径。然而，需要在递送方法方面取得进展，以使疾病靶向范围广泛的组织和细胞类型。腺相关病毒（AAV）载体仍然是基因治疗中最有前途的递送载体之一。目前，大多数BE/guide组合及其启动子超过了aav的包装极限（~ 5kb）。双重aav递送策略通常需要高病毒剂量，这会带来安全问题。在这项研究中，我们利用来自脑膜炎奈瑟菌的紧凑Cas9 （Nme2Cas9）设计了腺嘌呤碱基编辑器（ABE）。与已被充分表征的含化脓链球菌cas9的ABEs相比，使用Nme2Cas9 （Nme2-ABE）的ABEs具有独特的原间隔器邻近基序（N4CC）和编辑窗口，表现出更少的脱靶效应，并且可以在人类和小鼠基因组中有效地安装治疗相关突变。重要的是，我们发现通过单个AAV载体在体内递送Nme2-ABE及其引导RNA可以有效地编辑小鼠基因组位点，并在成年小鼠酪氨酸血症模型中恢复疾病突变和表型。我们预计Nme2-ABE凭借其紧凑的尺寸和广泛的靶向范围，将使一系列治疗应用具有更高的安全性和有效性，部分原因是在单载体系统中包装。
Base editors (BEs) have opened new avenues for the treatment of genetic diseases. However, advances in delivery approaches are needed to enable disease targeting of a broad range of tissues and cell types. Adeno-associated virus (AAV) vectors remain one of the most promising delivery vehicles for gene therapies. Currently, most BE/guide combinations and their promoters exceed the packaging limit (∼5 kb) of AAVs. Dual-AAV delivery strategies often require high viral doses that impose safety concerns. In this study, we engineered an adenine base editor (ABE) using a compact Cas9 from Neisseria meningitidis (Nme2Cas9). Compared with the well-characterized Streptococcus pyogenes Cas9-containing ABEs, ABEs using Nme2Cas9 (Nme2-ABE) possess a distinct protospacer adjacent motif (N4CC) and editing window, exhibit fewer off-target effects, and can efficiently install therapeutically relevant mutations in both human and mouse genomes. Importantly, we show that in vivo delivery of Nme2-ABE and its guide RNA by a single AAV vector can efficiently edit mouse genomic loci and revert the disease mutation and phenotype in an adult mouse model of tyrosinemia. We anticipate that Nme2-ABE, by virtue of its compact size and broad targeting range, will enable a range of therapeutic applications with improved safety and efficacy due in part to packaging in a single-vector system.