Department of Pharmaceutical Sciences, College of Pharmacy, Oregon State University, Robertson Life Sciences Building, 2730 Southwest Moody Avenue, Portland, OR, 97201, USA.
Moderna Therapeutics, 200 Technology Square, Cambridge, MA, 02139, USA.
Nat Commun. 2020 Feb 20;11(1):983. doi: 10.1038/s41467-020-14527-2.
Endosomal sequestration of lipid-based nanoparticles (LNPs) remains a formidable barrier to delivery. Herein, structure-activity analysis of cholesterol analogues reveals that incorporation of C-24 alkyl phytosterols into LNPs (eLNPs) enhances gene transfection and the length of alkyl tail, flexibility of sterol ring and polarity due to -OH group is required to maintain high transfection. Cryo-TEM displays a polyhedral shape for eLNPs compared to spherical LNPs, while x-ray scattering shows little disparity in internal structure. eLNPs exhibit higher cellular uptake and retention, potentially leading to a steady release from the endosomes over time. 3D single-particle tracking shows enhanced intracellular diffusivity of eLNPs relative to LNPs, suggesting eLNP traffic to productive pathways for escape. Our findings show the importance of cholesterol in subcellular transport of LNPs carrying mRNA and emphasize the need for greater insights into surface composition and structural properties of nanoparticles, and their subcellular interactions which enable designs to improve endosomal escape.
内体隔离仍然是脂质纳米颗粒 (LNPs) 递药的一个巨大障碍。本文通过对胆固醇类似物的构效分析表明，将 C-24 烷基植物甾醇掺入 LNPs (eLNPs) 中可以增强基因转染，并且由于 -OH 基团的存在，烷基链的长度、甾醇环的柔性和极性都需要保持高转染效率。与球形 LNPs 相比，冷冻透射电镜显示 eLNPs 呈多面体形，而 X 射线散射显示内部结构几乎没有差异。eLNPs 表现出更高的细胞摄取和保留能力，可能会导致内体中的物质随着时间的推移而稳定释放。3D 单颗粒跟踪显示，与 LNPs 相比，eLNPs 的细胞内扩散性增强，表明 eLNP 能够向有利于逃逸的有效途径进行运输。我们的研究结果表明了胆固醇在携带 mRNA 的 LNPs 亚细胞转运中的重要性，并强调了需要更深入地了解纳米颗粒的表面组成和结构特性，以及它们的亚细胞相互作用，这可以促进设计来改善内体逃逸。
