Biomedical Pioneering Innovation Center, Beijing Advanced Innovation Center for Genomics, Peking-Tsinghua Center for Life Sciences, Peking University Genome Editing Research Center, State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, Peking University, Beijing, China.
Peking University-Tsinghua University-National Institute of Biological Sciences Joint Graduate Program, Peking University, Beijing, China.
Nat Biotechnol. 2019 Sep;37(9):1059-1069. doi: 10.1038/s41587-019-0178-z. Epub 2019 Jul 15.
Current tools for targeted RNA editing rely on the delivery of exogenous proteins or chemically modified guide RNAs, which may lead to aberrant effector activity, delivery barrier or immunogenicity. Here, we present an approach, called leveraging endogenous ADAR for programmable editing of RNA (LEAPER), that employs short engineered ADAR-recruiting RNAs (arRNAs) to recruit native ADAR1 or ADAR2 enzymes to change a specific adenosine to inosine. We show that arRNA, delivered by a plasmid or viral vector or as a synthetic oligonucleotide, achieves editing efficiencies of up to 80%. LEAPER is highly specific, with rare global off-targets and limited editing of non-target adenosines in the target region. It is active in a broad spectrum of cell types, including multiple human primary cell types, and can restore α-L-iduronidase catalytic activity in Hurler syndrome patient-derived primary fibroblasts without evoking innate immune responses. As a single-molecule system, LEAPER enables precise, efficient RNA editing with broad applicability for therapy and basic research.
目前用于靶向 RNA 编辑的工具依赖于外源性蛋白质或化学修饰的向导 RNA 的递送，这可能导致效应物活性异常、递送障碍或免疫原性。在这里，我们提出了一种称为利用内源性 ADAR 进行可编程 RNA 编辑（LEAPER）的方法，该方法使用短的工程 ADAR 募集 RNA（arRNA）募集天然 ADAR1 或 ADAR2 酶将特定的腺苷转变为肌苷。我们表明，通过质粒或病毒载体或合成寡核苷酸递送的 arRNA 可实现高达 80%的编辑效率。LEAPER 具有高度特异性，罕见的全局脱靶和靶区中非靶腺苷的有限编辑。它在广泛的细胞类型中具有活性，包括多种人类原代细胞类型，并且可以在不引发先天免疫反应的情况下恢复 Hurler 综合征患者来源的原代成纤维细胞中的α-L-艾杜糖苷酸酶催化活性。作为一种单分子系统，LEAPER 能够实现精确、高效的 RNA 编辑，具有广泛的治疗和基础研究适用性。
