Nonhuman Primate Genomes Have Limited Relevance for Evaluating Human CRISPR Off-Target Effects

Area of Science:

• Genomic Medicine
• Biotechnology
• Bioinformatics

Background:

• CRISPR technology enables precise genome editing for therapeutic applications.
• Off-target mutations caused by CRISPR nucleases pose risks for clinical development.
• Nonhuman primates (NHPs) are frequently used to assess therapeutic safety, but their utility for evaluating human CRISPR off-target activity is uncertain.

Purpose of the Study:

• To computationally assess the recapitulation of human CRISPR off-target sites in commonly used NHP species.
• To evaluate the relevance of NHP models for predicting human CRISPR off-target activity in preclinical safety assessments.

Main Methods:

• Designed millions of Cas12a and Cas9 spacer sequences targeting human genes.
• Computationally predicted off-target editing sites in humans and five NHP species.
• Analyzed the overlap between human and NHP off-target sites based on defined inclusion criteria.

Main Results:

• A significant proportion of human CRISPR off-target sites were not found in NHP genomes.
• Only 14-21% of Cas12a and 7-15% of Cas9 human off-targets were recapitulated in commonly used NHPs.
• This indicates a limited ability of NHPs to model human CRISPR off-target effects.

Conclusions:

• NHP models have limitations in accurately reflecting human CRISPR spacer specificity.
• Preclinical safety assessments in NHPs may not fully capture the risks of human off-target editing.
• Results contextualize human risk assessment for CRISPR-based therapies.