先导编辑修复患者来源诱导多能干细胞中的 HBB 密码子 8/9（+G）突变，并在红系细胞中恢复 β 珠蛋白表达
Prime Editing Corrects the HBB Codon 8/9 (+G) Mutation in Patient-Derived Induced Pluripotent Stem Cells and Restores β-Globin Expression in iPSC-Derived Erythroid Cells
HBB 基因密码子 8/9 的 +G 插入突变会导致输血依赖型 β 地中海贫血，在南亚地区较为常见。先导编辑（prime editing）是一种无需造成 DNA 双链断裂、也无需外源供体 DNA 的精准基因组编辑技术，有望直接纠正这一插入突变，但效率取决于引导 RNA 的设计。研究者从一名纯合患者身上制备诱导多能干细胞（iPSC），优化了编辑条件，并将患者、编辑后和对照的 iPSC 分化为红系细胞进行比较。编辑后的细胞成熟度和集落形成更接近对照，HBB 转录本恢复到约为对照的四倍水平，检测到 β 珠蛋白蛋白，并出现与成人血红蛋白（HbA）一致的色谱峰，而胎儿和胚胎型珠蛋白仍占主导。作者指出，还需在更多供者、造血干细胞和体内验证。该研究尚未经同行评审。
为什么推荐给您：先导编辑在患者 iPSC 中修复致病突变并恢复功能蛋白，属有转化前景的基因治疗早期验证。
不需要生物学背景，多打比方
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摘要Abstract
Homozygosity for the HBB codon 8/9 (+G) frameshift (c.27dup (p.Ser10ValfsTer14)) causes transfusion-dependent {beta}-thalassaemia and is common in South Asia. Prime editing can reverse this insertion without double-strand breaks or donor DNA, but its efficiency depends on pegRNA design. We derived Sendai-reprogrammed iPSCs from a homozygous patient, optimised PEmax editing (spacer, pegRNA extension, secondary nick, MLH1dn) and compared patient, PEmax-treated and control iPSC-derived erythroid cells by flow cytometry, colony assays, RT-qPCR, western blotting and cation-exchange HPLC. Patient iPSCs had a normal 46,XY karyotype, expressed pluripotency markers, formed all three germ layers and were Sendai-free by passage 15. A PAM-disrupting spacer with a +68 nicking sgRNA gave the highest intended-edit frequency 8.4%; MLH1dn added little. PEmax-treated cultures matured and formed colonies more like control than patient cultures, restored HBB transcript (about 4-fold control iPSC-derived cells) and detectable {beta}-globin protein, and contained an HPLC fraction consistent with HbA ( undetectable in patient cells). Fetal haemoglobin (about 80%) and embryonic globin remained predominant. Prime editing corrects HBB c.27dupG in patient iPSCs and restores {beta}-globin expression, with partial restoration of adult haemoglobin in a fetal/embryonic-type erythroid background. Validation in additional donors, haematopoietic stem cells and in vivo is required.