PO.ET02.08 · 实验与分子治疗
研究常规及新型阳离子纳米脂质体中的磷脂酰胆碱和磷脂酰乙醇胺磷脂与慢性髓性白血病细胞体外相互作用
Investigating phosphatidylcholine and phosphoethanolamine phospholipids in conventional and novel cationic nanoliposomes for interactions with chronic myeloid leukemia cells in vitro
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
背景：脂质纳米颗粒（LNPs）已被确立为慢性髓性白血病（CML）药物/核苷酸递送的优良载体。阳离子脂质和辅助脂质（如基于磷脂酰胆碱（PC）和磷脂酰乙醇胺（PE）的脂质）比例的优化决定了LNPs的选择性。此外，纳入源自靶细胞膜的LE（脂质提取物）已被用于开发CLENs（细胞膜脂质提取纳米脂质体）以实现更优的靶向。然而，PE和PC基脂质在LE存在下的行为尚不清楚。在本研究中，我们使用模型CML（K562-GFP）细胞系，研究了由PE或PC磷脂以不同阳离子脂质和/或脂质提取物成分组合构成的LNPs的细胞相互作用。
方法：K562-GFP人类CML细胞系（CCL-243-GFP）培养于补充10%胎牛血清（FBS）的RPMI-1640培养基中。LNPs采用薄膜水化法制备，并对粒径和zeta电位进行表征。研究的第I阶段比较了LNPs的细胞摄取，以不同浓度（0-50 mol%）的四种阳离子脂质（DOTAP、EPC、DOTMA、DODMA）替代辅助脂质DOPC。第II阶段涉及在第I阶段确定的固定阳离子脂质浓度下比较PC与PE辅助脂质。研究的第III阶段旨在研究在不同浓度下纳入LE后靶细胞系和非靶细胞系对LNPs的细胞摄取。
结果：从第I阶段我们确定，与对照（DOPC 100 mol%）相比，含25 mol% DOTAP或EPC的LNPs的细胞摄取高约250%。第II阶段揭示，含辅助脂质DOPC的LNPs在所有时间点（0至120分钟）表现显著更佳，与辅助脂质DOPE相比细胞摄取增加。在第III阶段，在阳离子LNPs中纳入源自靶细胞的LE导致靶细胞系的选择性摄取增加，非靶细胞系的摄取减少。
结论：迄今为止的结果揭示，LNPs的细胞摄取在很大程度上取决于纳米颗粒开发中所用阳离子脂质与辅助脂质类型的最佳比例，并且纳入LE可实现更优的靶向。
查看英文原文 English abstract
Background: Lipid Nanoparticles (LNPs) have been established as excellent vehicles for drug/nucleotide delivery for Chronic Myeloid Leukemia (CML). The optimization of lipid ratios of cationic and helper lipids, such as phosphatidylcholine (PC) and phosphoethanolamine (PE)- based lipids, governs the selectivity of LNPs. Furthermore, the incorporation of LE (lipid extracts) derived from target cell membranes has been used to develop CLENs (cell membrane lipid-extracted nanoliposomes) for superior targeting. However, the behavior of PE- and PC-based lipids in the presence of LE is not well understood. In this study, we investigate the cellular interactions of LNPs consisting of PE or PC phospholipids employed in different combinations of cationic lipids and/or lipid extract ingredients using a model CML (K562-GFP) cell line.
Methods: The K562-GFP human CML cell line (CCL-243-GFP) was cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum (FBS). LNPs were prepared using the thin film hydration method and characterized for size and zeta potential. Phase I of the study compared the cellular uptake of LNPs, substituting four cationic lipids (DOTAP, EPC, DOTMA, DODMA) at varying concentrations (0-50 mol%) with the helper lipid DOPC. Phase II involved comparing the PC versus PE helper lipid at a fixed cationic lipid concentration determined in Phase I. Phase III of the study aims to investigate the cellular uptake of LNPs after the inclusion of LE at various concentrations by target and non-target cell lines.
Results: We identified the cellular uptake to be about 250% higher for LNPs containing 25 mol% of DOTAP or EPC compared to the control (DOPC 100 mol%) from Phase I. Phase II revealed that LNPs with helper lipid DOPC performed significantly better at all time points (0 to 120 min) with an increase in cellular uptake compared to helper lipid DOPE. In phase III, the inclusion of LE derived from the target cell in the cationic LNPs led to a selective uptake by the target cell line and decreased uptake by the non-target cell lines.
Conclusion: Results to date revealed that the cellular uptake of LNPs depends largely on the optimum ratio of the cationic lipid to the helper lipid type employed in the development of the nanoparticles, and that inclusion of LE leads to superior targeting.
利益披露 Disclosure
S. Y. Barve, None..
R. B. Campbell, None.