Spatial profiling of gene editing by in situ sequencing in mice and macaques.

Abstract

Base and prime editing technologies precisely install defined nucleotide edits in both dividing and non-dividing cells, offering potential for correcting pathogenic mutations directly in organisms. However, to fully leverage their therapeutic potential, accurately measuring editing rates with high spatial resolution is crucial. Here we use imaging-based in situ sequencing (ISS) to map base and prime editing events within native tissues. We establish and validate this technology in mouse brains treated with intein-split adenine base editors or prime editors delivered via adeno-associated viral vectors. We next apply ISS in the liver of mice and macaques treated with adenine base editors encoded on lipid nanoparticle-encapsulated mRNA and guide RNA (RNA-LNP). Effective editing was observed across all metabolic zones of liver lobules. Moreover, in experiments where repeated doses of RNA-LNP are administered, the initial dose does not affect the editing efficiency and distribution of the subsequent dose. Our results demonstrate how ISS can visualize gene editing events in vivo and suggest that base editor delivery using RNA-LNP could be used to address a wide spectrum of metabolic liver diseases.