Zhang Zhenwu, Bao Xinyu, Lin Chao-Po
School of Life Science and Technology, ShanghaiTech University, Shanghai 201210, China.
Biomedicines. 2023 Aug 1;11(8):2168. doi: 10.3390/biomedicines11082168.
Applying programmable nucleases in gene editing has greatly shaped current research in basic biology and clinical translation. Gene editing in human pluripotent stem cells (PSCs), including embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), is highly relevant to clinical cell therapy and thus should be examined with particular caution. First, since all mutations in PSCs will be carried to all their progenies, off-target edits of editors will be amplified. Second, due to the hypersensitivity of PSCs to DNA damage, double-strand breaks (DSBs) made by gene editing could lead to low editing efficiency and the enrichment of cell populations with defective genomic safeguards. In this regard, DSB-independent gene editing tools, such as base editors and prime editors, are favored due to their nature to avoid these consequences. With more understanding of the microbial world, new systems, such as Cas-related nucleases, transposons, and recombinases, are also expanding the toolbox for gene editing. In this review, we discuss current applications of programmable nucleases in PSCs for gene editing, the efforts researchers have made to optimize these systems, as well as new tools that can be potentially employed for differentiation modeling and therapeutic applications.
在基因编辑中应用可编程核酸酶极大地塑造了当前基础生物学研究和临床转化的格局。人类多能干细胞（PSCs）中的基因编辑，包括胚胎干细胞（ESCs）和诱导多能干细胞（iPSCs），与临床细胞治疗高度相关，因此应格外谨慎地进行研究。首先，由于PSCs中的所有突变都会传递给其所有后代，编辑工具的脱靶编辑会被放大。其次，由于PSCs对DNA损伤高度敏感，基因编辑产生的双链断裂（DSBs）可能导致编辑效率低下，并使具有缺陷基因组保护机制的细胞群体富集。在这方面，不依赖DSB的基因编辑工具，如碱基编辑器和引导编辑器，因其能够避免这些后果的特性而受到青睐。随着对微生物世界的更多了解，新的系统，如与Cas相关的核酸酶、转座子和重组酶，也在不断扩展基因编辑的工具库。在这篇综述中，我们讨论了可编程核酸酶在PSCs基因编辑中的当前应用、研究人员为优化这些系统所做的努力，以及可潜在用于分化建模和治疗应用的新工具。
