Construction of an Aptamer Modified Liposomal System Targeted to Tumor Endothelial Cells
Construction of an Aptamer Modified Liposomal System Targeted to Tumor Endothelial Cells
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DOI：
10.1248/bpb.b14-00338
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发表时间：
2014-11-01
影响因子：
2
通讯作者：
Harashima, Hideyoshi
中科院分区：
文献类型：
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作者：
Ara, Mst Naznin;Matsuda, Takashi;Harashima, Hideyoshi
在此，我们描述了一种高亲和力和特异性的DNA适配子的开发，作为一种新的配体，用于脂质体纳米粒靶向培养的小鼠肿瘤内皮细胞(MTECs)。基于主动靶向纳米技术的药物传递系统目前引起了人们的极大兴趣，因为它们具有减少副作用和促进细胞毒药物或基因以特定部位的方式传递的潜力。在这项研究中，我们报告了一个有前途的适配子候选AraHH036，它显示了对mTEC的选择性结合。该适配子不与正常细胞、正常内皮细胞或肿瘤细胞结合。为此，我们合成了适体-聚乙二醇脂结合物，并通过标准的脂质水合方法制备了适体脂质体(ALPS)。首先，我们通过孵育含有1mol%、5mol%和10mol%适体的Alps来量化Alps内化到mTECs的能力，并将结果与未修饰的聚乙二醇化脂质体(PLPs)的结果进行比较。激光共聚焦扫描显微镜(CLSM)摄取研究表明，ALPS比PLP更有效地被摄取。测得阿尔卑斯山的K-d值为142 nm。细胞内转运研究证实，大多数罗丹明标记的阿尔卑斯被摄取并与绿色溶酶追踪器共定位，从而证实它们位于溶酶体中。最后，利用基于适体的蛋白质组学方法，将适体的分子靶蛋白鉴定为热休克蛋白70(HSP70)。这些结果表明，这些阿尔卑斯有望成为一种新的载体分子，将抗血管生成药物输送到肿瘤血管系统。
We describe herein the development of a high affinity and specific DNA aptamer as a new ligand for use in liposomal nanoparticles to target cultured mouse tumor endothelial cells (mTECs). Active targeted nanotechnology based drug delivery systems are currently of great interest, due to their potential for reducing side effects and facilitating the delivery of cytotoxic drugs or genes in a site specific manner. In this study, we report on a promising aptamer candidate AraHH036 that shows selective binding towards mTECs. The aptamer does not bind to normal cells, normal endothelial cells or tumor cells. Therefore, we synthesized an aptamer-polyethylene glycol (PEG) lipid conjugate and prepared aptamer based liposomes (ALPs) by the standard lipid hydration method. First, we quantified the higher capacity of ALPs to internalize into mTECs by incubating ALPs containing 1 mol%, 5 mol% and 10 mol% aptamer of total lipids and compared the results to those for unmodified PEGylated liposomes (PLPs). A confocal laser scanning microscope (CLSM) uptake study indicated that the ALPs were taken up more efficiently than PLPs. The measured K-d value of the ALPs was 142 nM. An intracellular trafficking study confirmed that most of the rhodamine labeled ALPs were taken up and co-localized with the green lysotracker, thus confirming that they were located in lysosomes. Finally, using an aptamer based proteomics approach, the molecular target protein of the aptamer was identified as heat shock protein 70 (HSP70). The results suggest that these ALPs offer promise as a new carrier molecule for delivering anti-angiogenesis drugs to tumor vasculature.