Precise Correction of the Pde6b-L659P Mutation Causing Retinal Degeneration with Minimum Bystander Editing by...

Area of Science:

• Genetics
• Molecular Biology
• Ophthalmology

Background:

• Novel genome editing technologies like base editing (BE), prime editing (PE), and click editing (CE) offer precise gene modification with reduced risks.
• Systematic comparisons of these advanced editing tools for correcting disease-causing mutations are lacking.

Purpose of the Study:

• To evaluate and compare the efficacy and precision of BE, PE, and CE in correcting the Pde6b (c.1976T>C, p.L659P) mutation responsible for retinal degeneration.
• To assess the potential of these technologies for therapeutic applications in a mouse model of inherited retinal disease.

Main Methods:

• In vitro optimization of BE, PE, and CE systems using Pde6b-L659P cell models.
• Deep sequencing analysis to compare editing efficiency, precision, and bystander effects across the three technologies.
• In vivo electroporation of the optimized prime editing system into Pde6b-L659P mice.

Main Results:

• Prime editing demonstrated the highest precision, significantly minimizing bystander edits compared to base editing and click editing.
• Click editing exhibited lower efficiency and higher indel rates, indicating a need for further optimization.
• In vivo application of prime editing in mice achieved 12.4% targeted repair, leading to partial rescue of retinal degeneration.

Conclusions:

• Prime editing is a highly precise genome editing tool suitable for correcting the Pde6b mutation causing retinal degeneration.
• This study provides proof of concept for using BE, PE, and CE technologies to address genetic mutations underlying retinal diseases.
• The findings highlight the potential of precision gene editing for future therapeutic strategies in inherited retinal disorders.