Abstract：
Nucleic acid-based therapeutics encapsulated into lipid nanoparticles (LNPs) can potentially target the root cause of genetic skin diseases. Although nanoparticles are considered impermeable to skin, research and clinical studies have shown that nanoparticles can penetrate into skin with reduced skin barrier function when administered topically. Studies have shown that epidermal keratinocytes express the low-density lipoprotein receptor (LDLR) that mediates endocytosis of apolipoprotein E (apoE)-associated nanoparticles and that dermal fibroblasts express mannose receptors. Here we prepared LNPs designed to exploit these different endocytic pathways for intracellular mRNA delivery to the two most abundant skin cell types, containing: (i) labile PEG-lipids (DMG-PEG2000) prone to dissociate and facilitate apoE-binding to LNPs, enabling apoE-LDLR mediated uptake in keratinocytes, (ii) non-labile PEG-lipids (DSPE-PEG2000) to impose stealth-like properties to LNPs to enable targeting of distant cells, and (iii) mannose-conjugated PEG-lipids (DSPE-PEG2000-Mannose) to target fibroblasts or potentially immune cells containing mannose receptors. All types of LNPs were prepared by vortex mixing and formed monodisperse (PDI ∼ 0.1) LNP samples with sizes of 130 nm (±25%) and high mRNA encapsulation efficiencies (≥90%). The LNP-mediated transfection potency in keratinocytes and fibroblasts was highest for LNPs containing labile PEG-lipids, with the addition of apoE greatly enhancing transfection via LDLR. Coating LNPs with mannose did not improve transfection, and stealth-like LNPs show limited to no transfection. Taken together, our studies suggest using labile PEG-lipids and co-administration of apoE when exploring LNPs for skin delivery.
摘要：
包封到脂质纳米颗粒(LNP)中的基于核酸的治疗剂可以潜在地靶向遗传性皮肤病的根本原因。虽然纳米粒子被认为是皮肤不可渗透的，研究和临床研究表明，当局部给药时，纳米颗粒可以渗透到皮肤中，皮肤屏障功能降低。研究表明，表皮角质形成细胞表达介导载脂蛋白E（apoE）相关纳米颗粒内吞作用的低密度脂蛋白受体（LDLR），真皮成纤维细胞表达甘露糖受体。在这里，我们准备了LNP，旨在利用这些不同的内吞途径将细胞内mRNA递送到两种最丰富的皮肤细胞类型，包含：（i）不稳定的PEG-脂质（DMG-PEG2000）易于解离并促进apoE与LNP的结合，在角质形成细胞中实现apoE-LDLR介导的摄取，（ii）非不稳定的PEG-脂质（DSPE-PEG2000）对LNP施加类似隐身的特性，从而能够靶向远处的细胞，和(iii)甘露糖缀合的PEG-脂质(DSPE-PEG2000-甘露糖)靶向含有甘露糖受体的成纤维细胞或潜在免疫细胞。所有类型的LNP均通过涡旋混合制备，并形成大小为130nm（±25％）且mRNA包封效率高（≥90％）的单分散（PDI〜0.1）LNP样品。对于含有不稳定的PEG-脂质的LNP，角质形成细胞和成纤维细胞中的LNP介导的转染效力最高。随着apoE的加入，通过LDLR大大提高了转染。用甘露糖包被LNP并不能改善转染，和隐身样LNP显示限于无转染。一起来看,我们的研究建议在探索LNP用于皮肤递送时使用不稳定的PEG-脂质和apoE的共同给药。
