Lipid Nanoparticles for <i>In Vivo</i> Lung Delivery of CRISPR-Cas9 Ribonucleoproteins Allow Gene Editing of Clinical Targets.

Abstract

In the past 10 years, CRISPR-Cas9 has revolutionized the gene-editing field due to its modularity, simplicity, and efficacy. It has been applied for the creation of <i>in vivo</i> models, to further understand human biology, and toward the curing of genetic diseases. However, there remain significant delivery barriers for CRISPR-Cas9 application in the clinic, especially for <i>in vivo</i> and extrahepatic applications. In this work, high-throughput molecular barcoding techniques were used alongside traditional screening methodologies to simultaneously evaluate LNP formulations encapsulating ribonucleoproteins (RNPs) for <i>in vitro</i> gene-editing efficiency and <i>in vivo</i> biodistribution. This resulted in the identification of a lung-tropic LNP formulation, which shows efficient gene editing in endothelial and epithelial cells within the lung, targeting both model reporter and clinically relevant genomic targets. Further, this LNP shows no off-target indel formation in the liver, making it a highly specific extrahepatic delivery system for lung-editing applications.