Long-term correction of hemophilia B through CRISPR/Cas9 induced homology-independent targeted integration. (December 2022)

• Record Type: Journal Article
• Authors: Chen, Xi; Niu, Xuran; Liu, Yang; Zheng, Rui; Yang, Lei; Lu, Jian; Yin, Shuming; Wei, Yu; Pan, Jiahao; Sayed, Ahmed; Ma, Xueyun; Liu, Meizhen; Jing, Fengxiang; Liu, Mingyao; Hu, Jiazhi; Wang, Liren; Li, Dali
• Is Part Of: Journal of genetics and genomics. Volume 49:Number 12(2022)
• Page Start: 1114
• Page End: 1126
• Publication Date: 2022-12
• Subjects: Hemophilia B -- Alb -- FIX -- CRISPR/Cas9-mediated HITI -- Gene therapy; Genetics -- Periodicals; Genomics -- Periodicals; 576.505
• DOI: 10.1016/j.jgg.2022.06.001
• Languages: English
• ISSNs: 1673-8527

Abstract

Abstract: CRISPR/Cas9-mediated site-specific insertion of exogenous genes holds potential for clinical applications. However, it is still infeasible because homologous recombination (HR) is inefficient, especially for non-dividing cells. To overcome the challenge, we report that a homology-independent targeted integration (HITI) strategy is used for permanent integration of high-specificity-activity Factor IX variant ( F9 Padua, R338L) at the albumin ( Alb ) locus in a novel hemophilia B (HB) rat model. The knock-in efficiency reaches 3.66%, as determined by droplet digital PCR (ddPCR). The clotting time is reduced to a normal level four weeks after treatment, and the circulating factor IX (FIX) level is gradually increased up to 52% of the normal level over nine months even after partial hepatectomy, demonstrating the amelioration of hemophilia. Through primer-extension-mediated sequencing (PEM-seq), no significant off-target effect is detected. This study not only provides a novel model for HB but also identifies a promising therapeutic approach for rare inherited diseases.