Bystander base editing interferes with visual function restoration in Leber congenital amaurosis

Area of Science:

• Molecular Biology
• Gene Therapy
• Ophthalmology

Background:

• Base editors (BEs) offer high-efficiency gene correction but produce bystander edits.
• Bystander editing poses a challenge for precise therapeutic outcomes.
• Leber congenital amaurosis (LCA) is a hereditary retinal disorder impacting vision.

Purpose of the Study:

• To investigate the impact of bystander edits on phenotypic restoration in an LCA mouse model.
• To evaluate the efficacy of adenine base editors (ABEs) in correcting LCA-associated mutations.
• To understand the molecular mechanisms underlying editing-induced dysfunction.

Main Methods:

• Utilized LCA model mice (rd12) with adenine base editors (ABEs), including ABE8e.
• Performed AlphaFold-based mutational scanning and molecular dynamics simulations.
• Assessed RPE65 expression and visual function restoration.

Main Results:

• ABE8e generated significant bystander editing in LCA model mice.
• Bystander mutations, including L43P in RPE65, prevented visual function restoration despite RPE65 expression.
• The L43P mutation was identified to disrupt RPE65 structure and function.

Conclusions:

• Bystander editing by highly active ABEs can impede therapeutic gene correction.
• Precise base editing strategies are essential for successful gene therapy in LCA.
• Development of base editors with reduced bystander activity is critical for clinical translation.