Lipid Nanoparticle-Mediated Hit-and-Run Approaches Yield Efficient and Safe In Situ Gene Editing in Human Skin

作者：Juliana Bolsoni, Danny Liu, Fatemeh Mohabatpour, R. Ebner, Gaurav Sadhnani, Belal Tafech, Jerry Leung, Selina Rahman Shanta, Kevin An, Tessa Morin, Yihang Chen, A. Emilia Arguello, Keith A. Choate, Eric Jan, Colin J.D. Ross, Davide Brambilla, Dominik Witzigmann, Jayesh A. Kulkarni, Pieter R. Cullis, Sarah Hedtrich · 发表于：ACS Nano · 年份：2023 · DOI：10.1021/acsnano.3c08644 · 被引用次数：37 · 研究领域：RNA Interference and Gene Delivery、CRISPR and Genetic Engineering、Virus-based gene therapy research

High Resolution Image Download MS PowerPoint Slide Despite exciting advances in gene editing, the efficient delivery of genetic tools to extrahepatic tissues remains challenging. This holds particularly true for the skin, which poses a highly restrictive delivery barrier. In this study, we ran a head-to-head comparison between Cas9 mRNA or ribonucleoprotein (RNP)-loaded lipid nanoparticles (LNPs) to deliver gene editing tools into epidermal layers of human skin, aiming for in situ gene editing. We observed distinct LNP composition and cell-specific effects such as an extended presence of RNP in slow-cycling epithelial cells for up to 72 h. While obtaining similar gene editing rates using Cas9 RNP and mRNA with MC3-based LNPs (10–16%), mRNA-loaded LNPs proved to be more cytotoxic. Interestingly, ionizable lipids with a p K a ∼ 7.1 yielded superior gene editing rates (55%–72%) in two-dimensional (2D) epithelial cells while no single guide RNA-dependent off-target effects were detectable. Unexpectedly, these high 2D editing efficacies did not translate to actual skin tissue where overall gene editing rates between 5%–12% were achieved after a single application and irrespective of the LNP composition. Finally, we successfully base-corrected a disease-causing mutation with an efficacy of ∼5% in autosomal recessive congenital ichthyosis patient cells, showcasing the potential of this strategy for the treatment of monogenic skin diseases. Taken together, this study demonstrates the...