纳米药物RGD-MS2-D0X的制备及在肿瘤治疗的应用

发布时间：2018-06-30 01:49

  本文选题：RGD + MS2　； 参考：《北京协和医学院》2016年博士论文

【摘要】：阿霉素(Doxorubicin, DOX)作为化疗药物在肿瘤的临床治疗中被广泛应用。但由于阿霉素水溶性差,组织选择特异性低,因此,在治疗过程中,会对患者造成严重的副作用。此外,长期使用阿霉素将会诱导肿瘤细胞产生耐药性。这些因素限制了阿霉素在肿瘤治疗中的进一步应用。纳米药物近年来不断兴起,成为肿瘤治疗的一种策略。纳米药物利用增强的药物渗透及潴留效应进入肿瘤部位发挥作用。与小分子相比,纳米药物有如下优势：具有纳米尺度的化疗药物因其分子量大而在体内具有较长的半衰期；此外,纳米载体可装载大量药物,从而使肿瘤部位的药物浓度增加；不仅如此,纳米药物具有良好的组织选择性,因此,对其它器官的副作用较小。MS2病毒样颗粒是一种无感染性,耐RNA/DNA酶,可在体外大量表达的具有纳米尺度的生物材料,因此,可作为纳米材料用于药物递送。但由于MS2缺乏靶向特异性,需要特异性的配体以提高其靶向特异性。本研究采用MS2作为载体,利用两功能交联剂SM(PEG)24将RGD和化疗药物阿霉素与MS2交联,制备出具有靶向特异性的纳米药物RGD-MS2-DOX。电镜实验和动态光散射扫描实验显示所制备的RGD-MS2-DOX形态均一,分散性良好,径粒约为30nm。为观察RGD-MS2-DOX的肿瘤杀伤功能,我们分别从细胞水平和动物水平两个方面对该药物进行研究。在细胞水平,我们采用高表达a vβ3的Hela细胞、HUVECs细胞和HepG2细胞这三种肿瘤细胞进行实验,发现RGD-MS2-DOX可通过特异性的配体-受体介导的胞吞作用进入细胞,并且进入细胞的量多于RAD-MS2-DOX和DOX。不仅如此,肿瘤细胞摄取RGD-MS2-DOX是时间依赖性的,随着药物作用时间的延长而增加药物的摄入量。电镜实验发现RGD-MS2-DOX进入细胞后可释放出DOX,进入细胞核,发挥肿瘤杀伤功能。与DOX相比,RGD-MS2-DOX对Hela细胞和HepG2细胞的增殖具有较为明显的抑制效果。为了观察RGD-MS2-DOX的安全性与体内肿瘤杀伤功能,我们选择长有人肝癌HepG2的Balb/c裸鼠作为肿瘤模型,发现RGD-MS2-DOX的毒副作用小,并且抑瘤效果明显。本研究建立了以MS2 VLPs作为纳米材料的新型广谱的主动靶向纳米药物RGD-MS2-DOX,该纳米药物可针对多种类型的肿瘤细胞发挥靶向杀伤作用,并在体内外呈现出良好的抗肿瘤作用,值得进一步深入系统的研究。
[Abstract]:Doxorubicin (DOX) is widely used as a chemotherapeutic agent in the clinical treatment of cancer. However, due to poor water solubility and low tissue selection specificity, adriamycin can cause serious side effects in the course of treatment. In addition, long-term use of adriamycin will induce drug resistance in tumor cells. These factors limit the further application of adriamycin in tumor therapy. Nanopharmaceuticals are emerging in recent years as a strategy for tumor therapy. Nanopharmaceuticals increase drug penetration and retention into the tumor site to play a role. Compared with small molecules, nanopharmaceuticals have the following advantages: nanoscale chemotherapeutic drugs have a longer half-life in vivo due to their high molecular weight, in addition, nano-carriers can load a large number of drugs, thus increasing the concentration of drugs in tumor sites; Not only that, nanopharmaceuticals have good tissue selectivity, so the side effects on other organs are small. MS2 virus-like particles are noninfectious, RNA-resistant, RNA-resistant DNAases, and can be expressed in vitro with nanoscale biomaterials, so, It can be used as nanometer material for drug delivery. However, due to the lack of targeting specificity of MS2, specific ligands are needed to improve its targeting specificity. In this study, RGD-MS2-DOX was prepared by cross-linking RGD and doxorubicin with MS2 by using two functional crosslinking agent SM (PEG) 24. The results of electron microscopy and dynamic light scattering scanning showed that the prepared RGD-MS2-DOX was homogeneous in morphology and good in dispersion. The diameter of the prepared RGD-MS2-DOX was about 30 nm. In order to observe the tumor killing function of RGD-MS2-DOX, we studied the drug from cell level and animal level. At the cell level, we used three kinds of tumor cells, HUVECs and HepG2 cells, which expressed highly a v 尾 3. We found that RGD-MS2-DOX could enter the cells through specific ligand-receptor mediated endocytosis. And the number of entering cells is more than RAD-MS2-DOX and DOX. Moreover, the uptake of RGD-MS2-DOX by tumor cells was time dependent and increased with the prolongation of drug action time. Electron microscopy showed that RGD-MS2-DOX could release DOX into the nucleus and play a role in tumor killing. Compared with DOX, RGD-MS2-DOX could inhibit the proliferation of Hela cells and HepG2 cells. In order to observe the safety of RGD-MS2-DOX and tumor killing function in vivo, Balb / c nude mice with long liver cancer HepG2 were selected as tumor model. The results showed that RGD-MS2-DOX had little toxicity and obvious tumor inhibition effect. In this study, a novel broad-spectrum active target drug RGD-MS2-DOX, using MS2VLPs as nanomaterials, was established. The drug can be targeted at various types of tumor cells and has a good anti-tumor effect in vivo and in vitro. It is worth further and systematic study.
【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R730.5
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本文编号：2084325