Arbor Biotechnologies, 20 Acorn Park Drive, Tower 500, Cambridge, MA, USA.
Nat Commun. 2022 May 20;13(1):2833. doi: 10.1038/s41467-022-30465-7.
The CRISPR-Cas type V-I is a family of Cas12i-containing programmable nuclease systems guided by a short crRNA without requirement for a tracrRNA. Here we present an engineered Type V-I CRISPR system (Cas12i), ABR-001, which utilizes a tracr-less guide RNA. The compact Cas12i effector is capable of self-processing pre-crRNA and cleaving dsDNA targets, which facilitates versatile delivery options and multiplexing, respectively. We apply an unbiased mutational scanning approach to enhance initially low editing activity of Cas12i2. The engineered variant, ABR-001, exhibits broad genome editing capability in human cell lines, primary T cells, and CD34+ hematopoietic stem and progenitor cells, with both robust efficiency and high specificity. In addition, ABR-001 achieves a high level of genome editing when delivered via AAV vector to HEK293T cells. This work establishes ABR-001 as a versatile, specific, and high-performance platform for ex vivo and in vivo gene therapy.
CRISPR-Cas 类型 V-I 是一类 Cas12i 包含的可编程核酸酶系统，由一条短的 crRNA 引导，而不需要 tracrRNA。在这里，我们展示了一种经过工程改造的 V-I 型 CRISPR 系统（Cas12i），ABR-001，它利用了一种无 tracr 的向导 RNA。紧凑型 Cas12i 效应子能够自我加工预 crRNA 并切割 dsDNA 靶标，分别促进了多样化的递送选择和多重操作。我们应用了一种无偏的突变扫描方法来提高 Cas12i2 最初的低编辑活性。经过工程改造的变体 ABR-001 在人细胞系、原代 T 细胞和 CD34+造血干细胞和祖细胞中表现出广泛的基因组编辑能力，具有强大的效率和高度的特异性。此外，ABR-001 通过 AAV 载体递送到 HEK293T 细胞时，可实现高水平的基因组编辑。这项工作确立了 ABR-001 作为一种用于体外和体内基因治疗的通用、特异和高性能平台。
