Volodina Olga V, Demchenko Anna G, Anuchina Arina A, Ryzhkova Oxana P, Kovalskaya Valeriia A, Kondrateva Ekaterina V, Tabakov Vyacheslav Y, Lavrov Alexander V, Smirnikhina Svetlana A
DIV, Research Centre for Medical Genetics, Moscow, 115522, Russia.
Biochemistry (Mosc). 2025 Jun;90(6):773-785. doi: 10.1134/S0006297924604672.
Prime editing is a highly promising strategy for treating hereditary disorders due to its superior efficiency and safety profile compared to the conventional CRISPR-Cas9 systems. This study is dedicated to development of a causal therapy for cystic fibrosis by targeting the F508del variant of the gene using prime editing, as this specific deletion accounts for a substantial proportion of cystic fibrosis cases. While prime editing has shown remarkable precision in introducing targeted genetic modifications, its application in AT-rich genomic regions, such as the one containing the F508del variant, remains challenging. To overcome this limitation, we systematically evaluated 24 pegRNAs designed for two distinct prime editing systems, PEmax and PE2-NG. Efficiency of the F508del variant correction reached 2.81% (without normalization for transfection efficiency) in the airway basal cells from the patients with homozygous F508del mutation. However, the average transfection efficiency was only 11.9%, emphasizing critical need for the advancements in delivery methodologies. These findings highlight potential of prime editing as an approach for treating cystic fibrosis, while also underscoring necessity for further optimization of both editing constructs and delivery vectors to achieve clinically relevant correction levels.
与传统的CRISPR-Cas9系统相比，碱基编辑因其更高的效率和安全性，是一种治疗遗传性疾病非常有前景的策略。本研究致力于通过碱基编辑靶向该基因的F508del变体，开发一种针对囊性纤维化的病因疗法，因为这种特定的缺失在囊性纤维化病例中占相当大的比例。虽然碱基编辑在引入靶向基因修饰方面已显示出显著的精确性，但其在富含AT的基因组区域（如包含F508del变体的区域）的应用仍然具有挑战性。为克服这一限制，我们系统评估了为两种不同的碱基编辑系统PEmax和PE2-NG设计的24种pegRNA。在纯合F508del突变患者的气道基底细胞中，F508del变体校正效率达到2.81%（未对转染效率进行标准化）。然而，平均转染效率仅为11.9%，这突出表明迫切需要改进递送方法。这些发现凸显了碱基编辑作为治疗囊性纤维化方法的潜力，同时也强调了进一步优化编辑构建体和递送载体以达到临床相关校正水平的必要性。
