Hubrecht Institute, Royal Netherlands Academy of Arts and Sciences (KNAW) and University Medical Center Utrecht, Utrecht, the Netherlands.
Oncode Institute, Hubrecht Institute, Utrecht, the Netherlands.
Life Sci Alliance. 2021 Aug 9;4(10). doi: 10.26508/lsa.202000940. Print 2021 Oct.
Prime editing is a recently reported genome editing tool using a nickase-cas9 fused to a reverse transcriptase that directly synthesizes the desired edit at the target site. Here, we explore the use of prime editing in human organoids. Common TP53 mutations can be correctly modeled in human adult stem cell-derived colonic organoids with efficiencies up to 25% and up to 97% in hepatocyte organoids. Next, we functionally repaired the cystic fibrosis CFTR-F508del mutation and compared prime editing to CRISPR/Cas9-mediated homology-directed repair and adenine base editing on the CFTR-R785* mutation. Whole-genome sequencing of prime editing-repaired organoids revealed no detectable off-target effects. Despite encountering varying editing efficiencies and undesired mutations at the target site, these results underline the broad applicability of prime editing for modeling oncogenic mutations and showcase the potential clinical application of this technique, pending further optimization.
Prime editing 是一种最近报道的基因组编辑工具，它使用一种与逆转录酶融合的切口酶 cas9，直接在靶位点合成所需的编辑。在这里，我们探索了 prime editing 在人类类器官中的应用。常见的 TP53 突变可以在人类成体干细胞衍生的结肠类器官中得到正确建模，效率高达 25%，在肝细胞类器官中高达 97%。接下来，我们对囊性纤维化 CFTR-F508del 突变进行了功能修复，并将 prime editing 与 CRISPR/Cas9 介导的同源定向修复和腺嘌呤碱基编辑在 CFTR-R785*突变上进行了比较。对 prime editing 修复的类器官进行全基因组测序未发现可检测的脱靶效应。尽管在靶位点遇到了不同的编辑效率和不期望的突变，但这些结果强调了 prime editing 用于建模致癌突变的广泛适用性，并展示了该技术的潜在临床应用，有待进一步优化。
