Institut de Pharmacologie de Sherbrooke, Department of Pharmacology and Physiology, Faculty of Medicine and Health Sciences, Université de Sherbrooke, Sherbrooke, Quebec J1H 5N4, Canada.
Mol Pharm. 2023 Mar 6;20(3):1577-1590. doi: 10.1021/acs.molpharmaceut.2c00783. Epub 2023 Feb 13.
To investigate the structure-cellular penetration relationship of guanidinium-rich transporters (GRTs), we previously designed PGua, a five-amino acid peptoid containing a conformationally restricted pattern of eight guanidines, which showed high cell-penetrating abilities and low cell toxicity. Herein, we characterized the cellular uptake selectivity, internalization pathway, and intracellular distribution of PGua, as well as its capacity to deliver cargo. PGua exhibits higher penetration efficiency in HeLa cells than in six other cell lines (A549, Caco-2, fibroblast, HEK293, Mia-PaCa2, and MCF7) and is mainly internalized by clathrin-mediated endocytosis and macropinocytosis. Confocal microscopy showed that it remained trapped in endosomes at low concentrations but induced pH-dependent endosomal membrane destabilization at concentrations ≥10 μM, allowing its diffusion into the cytoplasm. Importantly, PGua significantly enhanced macropinocytosis and the cellular uptake and cytosolic delivery of large IgGs following noncovalent complexation. Therefore, in addition to its peptoid nature conferring high resistance to proteolysis, PGua presents characteristics of a promising tool for IgG delivery and therapeutic applications.
为了研究富含胍基的转运体（GRTs）的结构-细胞穿透关系，我们之前设计了 PGua，这是一种含有八个胍基的受限构象模式的五肽缩氨酸，具有很高的细胞穿透能力和低细胞毒性。在这里，我们描述了 PGua 的细胞摄取选择性、内化途径和细胞内分布，以及其传递货物的能力。PGua 在 HeLa 细胞中的穿透效率高于其他六种细胞系（A549、Caco-2、成纤维细胞、HEK293、Mia-PaCa2 和 MCF7），主要通过网格蛋白介导的内吞作用和巨胞饮作用内化。共聚焦显微镜显示，在低浓度下，它仍被困在内体中，但在浓度≥10 μM 时诱导 pH 依赖性内体膜不稳定，允许其扩散到细胞质中。重要的是，PGua 显著增强了大 IgG 的巨胞饮作用以及非共价复合物化后的细胞摄取和细胞质传递。因此，除了其肽性质赋予高抗蛋白水解性外，PGua 还具有作为 IgG 传递和治疗应用的有前途工具的特征。
