Safer and efficient base editing and prime editing via ribonucleoproteins delivered through optimized lipid-nanoparticle formulations

Delivering ribonucleoproteins (RNPs) for in vivo genome editing is safer
than using viruses encoding for Cas9 and its respective guide RNA.
However, transient RNP activity does not typically lead to optimal editing
outcomes. Here, we show that the efficiency of delivering RNPs can be
enhanced by cell-penetrating peptides (covalently fused to the protein or
as excipients) and that lipid nanoparticles (LNPs) encapsulating RNPs can
be optimized for enhanced RNP stability, delivery efficiency, and editing
potency. Specifically, after screening for suitable ionizable cationic
lipids and by optimizing the concentration of the synthetic lipid DMG-PEG
2000, we show that the encapsulation, via microfluidic mixing, of adenine
base editor and prime editor RNPs within LNPs using the ionizable lipid
SM102 can result in in vivo editing-efficiency enhancements larger than
300-fold (with respect to the delivery of the naked RNP) without
detectable off-target edits. We believe that chemically defined LNP
formulations optimized for RNP-encapsulation stability and delivery
efficiency will lead to safer genome editing.