小柯机器人

美国哈佛医学院刘如谦等研究人员合作发现，造血干细胞的碱基编辑可治疗小鼠的镰状细胞病。相关论文于2021年6月2日在线发表于国际学术期刊《自然》。

研究人员使用一种定制的腺嘌呤碱基编辑器（ABE8e-NRCH）将镰状细胞病（SCD）等位基因（HBB^S）改造为Makassar β-珠蛋白 (HBB^G)，这是一种非致病性变体。将编码碱基编辑器和靶向向导RNA的mRNA离体递送到SCD患者的造血干细胞和祖细胞 (HSPC) 中，导致80%的HBB^S转化为HBB^G。将编辑过的人类HSPC移植到免疫缺陷小鼠中16周后，HBB^G的频率为68%，缺氧诱导的骨髓网织红细胞镰状细胞减少了5倍，表明基因编辑是持久的。

为了评估HBB^S碱基编辑的生理效应，研究人员提供了ABE8e-NRCH并将RNA从人源化SCD小鼠导入HSPC，然后将这些细胞移植到辐射小鼠中。十六周后，Makassar β-珠蛋白占血液中79%的β-珠蛋白，缺氧引起的镰状细胞减少了三倍。与接受未编辑细胞的小鼠相比，接受碱基编辑的HSPC的小鼠显示出接近正常的血液学参数和脾脏病理的减少。编辑骨髓的二次移植证实了基因编辑在长期造血干细胞中是持久的，并表明20%或更多的HBB^S-to-HBB^G编辑足以进行表型挽救。人类HSPC的碱基编辑避免了在Cas9核酸酶处理后观察到的p53激活和更大的缺失。这些发现表明SCD的一次性自体治疗可以消除致病性HBB^S，产生良性HBB^G，并最大限度地减少双链DNA断裂的不良后果。

据悉，SCD是由 β-珠蛋白基因HBB的突变引起的。

附：英文原文

Title: Base editing of haematopoietic stem cells rescues sickle cell disease in mice

Author: Gregory A. Newby, Jonathan S. Yen, Kaitly J. Woodard, Thiyagaraj Mayuranathan, Cicera R. Lazzarotto, Yichao Li, Heather Sheppard-Tillman, Shaina N. Porter, Yu Yao, Kalin Mayberry, Kelcee A. Everette, Yoonjeong Jang, Christopher J. Podracky, Elizabeth Thaman, Christophe Lechauve, Akshay Sharma, Jordana M. Henderson, Michelle F. Richter, Kevin T. Zhao, Shannon M. Miller, Tina Wang, Luke W. Koblan, Anton P. McCaffrey, John F. Tisdale, Theodosia A. Kalfa, Shondra M. Pruett-Miller, Shengdar Q. Tsai, Mitchell J. Weiss, David R. Liu

Issue&Volume: 2021-06-02

Abstract: Sickle cell disease (SCD) is caused by a mutation in the β-globin gene HBB1. We used a custom adenine base editor (ABE8e-NRCH)2,3 to convert the SCD allele (HBBS) into Makassar β-globin (HBBG), a non-pathogenic variant4,5. Ex vivo delivery of mRNA encoding the base editor with a targeting guide RNA into haematopoietic stem and progenitor cells (HSPCs) from patients with SCD resulted in 80% conversion of HBBS to HBBG. Sixteen weeks after transplantation of edited human HSPCs into immunodeficient mice, the frequency of HBBG was 68% and hypoxia-induced sickling of bone marrow reticulocytes had decreased fivefold, indicating durable gene editing. To assess the physiological effects of HBBS base editing, we delivered ABE8e-NRCH and guide RNA into HSPCs from a humanized SCD mouse6 and then transplanted these cells into irradiated mice. After sixteen weeks, Makassar β-globin represented 79% of β-globin protein in blood, and hypoxia-induced sickling was reduced threefold. Mice that received base-edited HSPCs showed near-normal haematological parameters and reduced splenic pathology compared to mice that received unedited cells. Secondary transplantation of edited bone marrow confirmed that the gene editing was durable in long-term haematopoietic stem cells and showed that HBBS-to-HBBG editing of 20% or more is sufficient for phenotypic rescue. Base editing of human HSPCs avoided the p53 activation and larger deletions that have been observed following Cas9 nuclease treatment. These findings point towards a one-time autologous treatment for SCD that eliminates pathogenic HBBS, generates benign HBBG, and minimizes the undesired consequences of double-strand DNA breaks.

DOI: 10.1038/s41586-021-03609-w

Source: https://www.nature.com/articles/s41586-021-03609-w

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