CRISPR Base-Editing for Correcting Monogenic Mutations: Optimization and Off-Target Profiling

Abstract

CRISPR base-editing has now emerged as a fast, accurate, and least disruptive technology to correct monogenic mutations in animal models, demonstrating direct A-G and C-T base conversions, without formation of two-stranded breaks. This review summarizes recent advances in optimization of base-editing systems, delivery, and optimization of off-target profiling of mice, zebrafish, rabbits, and livestock. The major results are reported on the development of better versions of the EsCs: BE4max, ABE8e, and high-fidelity Cas9 enzymes can be used to obtain better on-target specificity, whereas AAV vectors, LNPs, and mRNA-based delivery are important in enabling tissue-specific editing. Species-specific variability in off-target studies due to chromatin context and deaminase behavior raises the importance of standardized profiling pipelines. Although it has not been used in therapeutic contexts, despite considerable demonstrated capabilities in metabolic, ocular, and neuromuscular disease models, there are still challenges, including limitations in the use of PAM by bystanders, mosaicism, and long-term safety. All in all, this review provides basis on the possibility of further refinement of editors, safer methods of delivery, and ethical biosafety systems to develop CRISPR base-editing into effective preclinical and translational practices.