(25f) Efficient In Vivo Prime Editing with Lipid Nanoparticles

Prime editing enables a wide variety of precise genome edits for studying and treating genetic diseases. However, the in vivo delivery of prime editing components remains a challenge. Although adeno-associated viruses (AAVs) have been used to deliver prime editing components in vivo, AAVs are limited by cargo size, extended expression of the prime editor, and inability to redose patients. Transient co-delivery of prime editor mRNA and chemically synthesized prime editor guide RNA (pegRNA) via non-viral lipid nanoparticles (LNPs) is a promising approach but has previously resulted in inefficient prime editing outcomes in vivo. Here we present the systematic engineering of prime editing LNPs (PE-LNPs) to overcome bottlenecks to efficient in vivo prime editing. The optimized PE-LNPs achieve therapeutically relevant levels of prime editing in the mouse liver (up to 49% bulk liver editing following a single dose). We apply optimized PE-LNPs to treat a humanized mouse model of an inborn error of metabolism, achieving normalization of metabolite levels in the blood. The optimized PE-LNPs presented here support efficient in vivo prime editing and are a promising approach for the direct, in vivo treatment of genetic diseases.