美国哈佛大学刘如谦小组发现，噬菌体辅助演化和蛋白质工程产生紧凑高效的先导编辑器。相关论文于2023年8月31日发表在《细胞》杂志上。
研究人员利用蛋白质演化和工程技术生成了体积更小、效率更高的先导编辑器。利用噬菌体辅助演化，研究人员将紧凑型反转录酶的编辑效率提高了22倍，并生成了比当前一代编辑器PEmax小516-810个碱基对的先导编辑器。研究人员发现不同的反转录酶擅长不同类型的编辑，并利用这一洞察力生成了性能优于PEmax和PEmaxΔRNaseH的反转录酶，PEmaxΔRNaseH是双AAV运送系统中使用的截短编辑器。
最后，研究人员生成的Cas9结构域提高了先导编辑能力。这些生成的编辑器（PE6a-g）提高了患者来源成纤维细胞和原代人类T细胞的治疗相关编辑能力。PE6变体还能在体内通过双AAV传播实现更长的插入，在小鼠大脑皮层中实现了40%的loxP插入，比以前最先进的先导编辑器提高了24倍。
据悉，先导编辑能在活细胞中进行各种精确的基因组编辑。
附：英文原文
Title: Phage-assisted evolution and protein engineering yield compact, efficient prime editors
Author: Jordan L. Doman, Smriti Pandey, Monica E. Neugebauer, Meirui An, Jessie R. Davis, Peyton B. Randolph, Amber McElroy, Xin D. Gao, Aditya Raguram, Michelle F. Richter, Kelcee A. Everette, Samagya Banskota, Kathryn Tian, Y. Allen Tao, Jakub Tolar, Mark J. Osborn, David R. Liu
Issue&Volume: 2023/08/31
Abstract: Prime editing enables a wide variety of precise genome edits in living cells. Here we use protein evolution and engineering to generate prime editors with reduced size and improved efficiency. Using phage-assisted evolution, we improved editing efficiencies of compact reverse transcriptases by up to 22-fold and generated prime editors that are 516–810 base pairs smaller than the current-generation editor PEmax. We discovered that different reverse transcriptases specialize in different types of edits and used this insight to generate reverse transcriptases that outperform PEmax and PEmaxΔRNaseH, the truncated editor used in dual-AAV delivery systems. Finally, we generated Cas9 domains that improve prime editing. These resulting editors (PE6a-g) enhance therapeutically relevant editing in patient-derived fibroblasts and primary human T-cells. PE6 variants also enable longer insertions to be installed in vivo following dual-AAV delivery, achieving 40% loxP insertion in the cortex of the murine brain, a 24-fold improvement compared to previous state-of-the-art prime editors.
DOI: 10.1016/j.cell.2023.07.039
Source: https://www.cell.com/cell/fulltext/S0092-8674(23)00854-1