环状RGD多肽修饰的聚乳酸立体复合胶束对结肠癌的靶向治疗

发布时间：2018-06-23 14:55

本文选题：结肠癌 + RGD多肽　；参考：《吉林大学》2017年博士论文

【摘要】：结肠癌是常见的恶性肿瘤之一,具有较高的癌症相关性死亡率。目前,化疗仍是结肠癌术后辅助治疗及晚期结肠癌重要的治疗手段之一。传统的化疗药物因其较差的靶向性,使得药物在全身分布,肿瘤的治疗效果不理想,且常伴有严重的毒副作用。此外,由于临床应用的化疗药物本身的理化性质有一定的缺陷,如药物分子的大小、溶解性、渗透性、稳定性等,这很大程度上限制了化疗药物疗效。采用高分子聚合物纳米载体作为传统化疗药物的载体,可以有效增加治疗的靶向性及治疗效果,并可降低药物的毒副作用。而且通过包覆传统的化疗药物,可以有效的改善其溶解性、渗透性及稳定性的问题。因此,聚合物纳米载体药物传输体系为恶性肿瘤的靶向治疗提供了新的机遇与思路。目的:本研究应用纳米技术合成制备多组分聚乳酸立体复合胶束,并以环状RGD多肽作为靶向配体基团对纳米胶束进行修饰,并装载阿霉素作为抗肿瘤药物。应用鼠源结肠癌细胞C26建立小鼠结肠癌移植瘤模型。通过动物及细胞实验探讨纳米胶束载体在结肠癌治疗中的效果及靶向作用。方法:(1)通过开环聚合法合成4-arm PEG-b-PDLA、m PEG-b-PLLA及c RGD-PEG-b-PLLA。随后通过纳米沉淀法合成立体复合胶束SCM及c RGD-SCM,并将DOX装载进入胶束。通过1H NMR、TEM及DLS确定载药胶束的理化性质、并对载药胶束在体外环境中DOX的释放进行检测;(2)以鼠源结肠癌细胞C26为细胞实验对象,通过CLSM及FCM检测肿瘤细胞对药物的摄取、MTT法检测载药胶束在体外环境中的抗肿瘤效果;(3)通过高压液相法检测载药胶束在体内环境中DOX的循环时间,在C26的BALB/c小鼠移植瘤模型中,通过荧光成像法检测载药胶束在肿瘤及重要脏器中的分布,观测肿瘤生长情况及免疫组织化学法检测载药胶束在体内环境中的抗肿瘤效果。结果:(1)成功合成制备了多组分聚乳酸立体复合胶束SCM及c RGD-SCM,并成功将DOX装载其中。SCM/DOX及c RGD-SCM/DOX的载药率分别为8.9±0.37 wt.%和8.4±0.29 wt.%。在TEM下观察SCM/DOX及c RGD-SCM/DOX均为大小均一的球形结构,DLS检测结果两者的粒径大小分别为91.6±4.9nm和102.9±5.6nm。并且两种载药胶束在体外环境中(PBS溶液,p H=7.4,t=37.5℃)的DOX释放曲线均为初始快速释放而后缓慢释放的模式。(2)CLSM及FCM检测结果显示与DOX纯药相比较,SCM/DOX及c RGD-SCM/DOX均能够有效的提高肿瘤细胞对药物的摄取,且c RGD-SCM/DOX的提高效果更佳。MTT检测结果显示c RGD-SCM/DOX具有最佳的体外抗肿瘤效果。(3)药物代谢动力学检测结果显示SCM/DOX及c RGD-SCM/DOX在体内环境中呈缓慢释放的趋势,与DOX纯药相比,两种载药胶束均显著延长了DOX在体内的循环时间。荧光成像检测显示c RGD-SCM/DOX显著的提高了DOX在肿瘤部位的聚集,展现了显著的靶向性。(4)在C26小鼠移植瘤模型中,c RGD-SCM/DOX显著地抑制了肿瘤的生长,免疫组化结果显示与DOX纯药及SCM/DOX比较,c RGD-SCM/DOX在肿瘤的增殖抑制与凋亡中显示出了最佳的效果。同时,SCM/DOX及c RGD-SCM/DOX均降低了DOX纯药对小鼠主要器官(心脏、肝脏、脾脏、肺脏、肾脏)的毒副作用。结论:在本研究中,设计并制备4-arm PEG-b-PDLA、m PEG-b-PLLA和c RGD-PEG-b-PLLA,组成多元PLA SCM,并标记为c RGD-SCM/DOX,用于DOX的靶向传输。c RGD-SCM/DOX相比较于游离DOX以及无靶向性的SCM/DOX,对于αvβ3整合素阳性的C26结肠癌细胞,在体内环境中显示出了延长的循环时间、在肿瘤组织内部选择性的聚集以及强化的抗肿瘤效果。其中的靶向成分c RGD-PEG-b-PLLA可以根据不同肿瘤细胞膜表面的受体不同轻易的替换成其他靶向基团修饰的PEG-b-PLLA。当然,靶向SCM可以包埋并选择性的传输其他化学治疗药物。鉴于上述优越的性质,可切换的靶向SCM在临床针对不同的恶性肿瘤的个体化化疗中可能拥有巨大的应用前景。
[Abstract]:Colon cancer is one of the common malignant tumors and has high cancer related mortality. Chemotherapy is still one of the important treatment methods for adjuvant treatment of colon cancer and advanced colon cancer. Traditional chemotherapy drugs are distributed all over the body because of their poor targeting, and the treatment effect is not ideal and often accompanied by severe cancer. In addition, due to the physical and chemical properties of the clinical chemotherapeutic agents, such as the size, solubility, permeability, and stability of the drug molecules, this greatly restricts the efficacy of chemotherapeutic drugs. The use of polymer nanoscale as a carrier of traditional chemotherapy drugs can effectively increase the target of treatment. It can reduce the toxic and side effects of the drug, and can effectively improve its solubility, permeability and stability by wrapping traditional chemotherapeutic drugs. Therefore, the polymer nanoscale drug delivery system provides new opportunities and ideas for the targeting therapy of malignant tumors. The multicomponent polylactic acid complex micelles were synthesized and the nano micelles were modified with cyclic RGD peptide as the target ligand group, and adriamycin was loaded as an antitumor drug. The mouse colon cancer cell C26 was used to establish a mouse colon cancer transplant tumor model. The nano micellar carrier was used to treat colon cancer by animal and cell experiment. Methods: (1) 4-arm PEG-b-PDLA, m PEG-b-PLLA and C RGD-PEG-b-PLLA. were synthesized by ring opening polymerization, and then nano composite micelle SCM and C RGD-SCM were synthesized by nano precipitation method, and DOX loaded into micelle. The release of DOX in the environment was detected; (2) the mice source colon cancer cell C26 was used as the cell test object, the drug uptake by the tumor cells was detected by CLSM and FCM, and the anti tumor effect of the drug micelles in the environment was detected by MTT method. (3) the cycle time of the drug micelles in the body environment was detected by high pressure liquid chromatography, and the BALB/c mice in C26 were detected by the high pressure liquid phase method. In the transplanted tumor model, the distribution of drug loaded micelles in the tumor and important organs was detected by fluorescence imaging, the tumor growth and the antitumor effect of the drug micelles were detected by immunohistochemistry. Results: (1) the multicomponent poly milk acid stereocomplex micelle SCM and C RGD-SCM were successfully synthesized and the DOX was successfully installed. The drug loading rates of.SCM/DOX and C RGD-SCM/DOX were 8.9 + 0.37 wt.% and 8.4 + 0.29 wt.%. respectively, and SCM/DOX and C RGD-SCM/DOX were all spherical structures under TEM. The particle size of DLS detection results were 91.6 + and 102.9 + 5.6nm., and two kinds of drug bundles were in the environment. The release curves of DOX were both initial rapid release and then slow release. (2) the results of CLSM and FCM showed that SCM/DOX and C RGD-SCM/DOX could effectively improve the uptake of drugs by SCM/DOX and C RGD-SCM/DOX, and the enhanced effect of C RGD-SCM/DOX showed that C RGD-SCM/DOX was the best in vitro. (3) the results of pharmacokinetic detection showed that SCM/DOX and C RGD-SCM/DOX showed a slow release in the body environment. Compared with DOX, the two kinds of drug micelles significantly prolonged the cycle time of DOX in the body. The fluorescence imaging test showed that C RGD-SCM/DOX significantly improved the aggregation of DOX at the tumor site. Significant targeting. (4) in the C26 mouse transplanted tumor model, C RGD-SCM/DOX significantly inhibited the growth of the tumor. The immunohistochemical results showed that the C RGD-SCM/DOX showed the best effect in the proliferation inhibition and apoptosis of the tumor, and the SCM/ DOX and C RGD-SCM/DOX decreased the main organs of the DOX pure drugs to mice. Conclusion: in this study, the design and preparation of 4-arm PEG-b-PDLA, m PEG-b-PLLA and C RGD-PEG-b-PLLA, composed of multiple PLA SCM, and labeled C RGD-SCM/DOX, are used for DOX targeting transmission.C. Positive C26 colon cancer cells show an extended cycle time in the body environment, selective aggregation within the tumor tissue and enhanced anti-tumor effect. The target component C RGD-PEG-b-PLLA can easily be replaced by other target groups based on the different receptors on the membrane surface of different tumor cells. Of course, targeted SCM can be embedded and selectively transferred to other chemotherapeutic drugs. In view of the above advantages, switching target SCM may have great potential in the individualized chemotherapy for different malignant tumors.
【学位授予单位】：吉林大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R735.35

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