Lee Seok-Hoon, Wu Jun, Im Dongjoon, Hwang Gue-Ho, Jeong You Kyeong, Jiang Hui, Lee Seok Jae, Jo Dong Hyun, Goddard William A, Kim Jeong Hun, Bae Sangsu
Department of Biomedical Sciences, Seoul National University College of Medicine, Seoul, Republic of Korea.
Fight against Angiogenesis-Related Blindness (FARB) Laboratory, Biomedical research institute, Seoul National University Hospital, Seoul 03082, Republic of Korea.
bioRxiv. 2024 Oct 25:2024.10.23.619839. doi: 10.1101/2024.10.23.619839.
Base editors (BEs) have emerged as a powerful tool for gene correction with high activity. However, bystander base editing, a byproduct of BEs, presents challenges for precise editing. Here, we investigated the effects of bystander edits on phenotypic restoration in the context of Leber congenital amaurosis (LCA), a hereditary retinal disorder, as a therapeutic model. We observed that in of LCA model mice, the highest editing activity version of an adenine base editors (ABEs), ABE8e, generated substantial bystander editing, resulting in missense mutations despite RPE65 expression, preventing restoration of visual function. Through AlphaFold-based mutational scanning and molecular dynamics simulations, we identified that the ABE8e-driven L43P mutation disrupts RPE65 structure and function. Our findings underscore the need for more stringent requirements in developing precise BEs for future clinical applications.
碱基编辑器（BEs）已成为一种具有高活性的强大基因校正工具。然而，旁观者碱基编辑作为BEs的一种副产物，给精确编辑带来了挑战。在这里，我们以遗传性视网膜疾病莱伯先天性黑蒙（LCA）作为治疗模型，研究了旁观者编辑对表型恢复的影响。我们观察到，在LCA模型小鼠中，腺嘌呤碱基编辑器（ABEs）的最高编辑活性版本ABE8e产生了大量的旁观者编辑，尽管有RPE65表达，但仍导致错义突变，从而阻止了视觉功能的恢复。通过基于AlphaFold的突变扫描和分子动力学模拟，我们确定ABE8e驱动的L43P突变破坏了RPE65的结构和功能。我们的研究结果强调了在开发用于未来临床应用的精确BEs时需要更严格的要求。
