异喹啉类化合物及其转铁蛋白靶向脂质体的抗肿瘤活性研究

发布时间：2018-05-31 22:17

本文选题：异喹啉衍生物 + 脂质体　；参考：《吉林大学》2016年硕士论文

【摘要】：恶性肿瘤作为全球公共卫生问题之一,对人类健康产生严重威胁。作为发展中国家,我国癌症患者的发病率及死亡率均逐年上升。由于已有的化疗药物毒副作用较大,新型抗肿瘤制剂的研发具有重要意义。异喹啉类化合物是一类具有抗癌活性的化合物,但具有水溶解度低,难于吸收,直接给药效果差等缺点。本研究主要对具有异喹啉结构的化合物针对抗肿瘤活性进行筛选,并开发载药系统以提高药物吸收,降低毒副作用。本研究主要包括以下几个方面:1.化合物抗肿瘤活性筛选及理化性质检测本文以11种异喹啉类化合物和α-亚甲基-γ-丁内酯类化合物为筛选对象,通过MTT细胞抑制试验筛选抗肿瘤活性强的化合物。He La、Hep G2细胞IC50实验和HEK-293T细胞抑制试验结果显示,2-[[4-溴-2-(4-溴-苯甲酰)-1-氰基-1,2-二氢-异喹啉-1-基]-(2-溴-苯基)-甲基]-丙烯酸薄荷基酯(化合物2)对肿瘤细胞的抑制作用强,对正常细胞的抑制作用弱,因此选择此化合物作为抗肿瘤药物进行研究。对该化合物的理化性质进行检测,结果表明化合物极难溶于水,在乙醇中的溶解度为1.52 mg/m L,难于直接给药,需要进行制剂的开发。2.脂质体、转铁蛋白脂质体载药系统的建立为了提高细胞对化合物2的吸收,选择了脂质体这一剂型进行制剂开发,同时对脂质体进行转铁蛋白修饰,制备得到转铁蛋白受体靶向的脂质体制剂。对脂质体、转铁蛋白脂质体进行理化性质考察和转铁蛋白浓度、载药量优化,研究结果显示优化得到的脂质体制备条件为LP:Tf=500:1,载药量10%。制备得到的脂质体粒径在77 nm左右,转铁蛋白脂质体粒径在144 nm,电位均在-7~-4 m V范围内,无明显差异。3.脂质体、转铁蛋白脂质体的体外评价本文对制备得到的脂质体和转铁蛋白脂质体载药系统进行体外评价。研究结果发现,通过血清稳定性和体外释放试验发现两种载药系统均能够在血清环境中保持较好的稳定性,且均能达到一定的缓释效果。流式细胞术和激光共聚焦结果显示在两种细胞系中,Tf-LPs的转染效率明显高于LPs,说明对脂质体进行转铁蛋白偶联能够提高肿瘤细胞对化合物的摄取效率。对细胞摄取途径进行研究,结果初步证明了Tf-LPs主要是通过Tf受体介导的内吞途径进入肿瘤细胞。MTT试验结果显示在He La和Hep G2细胞中,Tf-LPs对细胞的抑制作用强于LPs。细胞ROS水平考察和线粒体膜电位变化考察的结果也显示出Tf-LPs能够诱导细胞ROS水平提高,细胞发生凋亡。综上所述,递送新型化合物的转铁蛋白靶向脂质体是一种有潜力的肿瘤治疗药物。筛选得到的有良好抗肿瘤活性的化合物通过转铁蛋白脂质体载药系统的包载能克服原化合物溶解性差,生物利用度低的缺点,达到延长循环时间,肿瘤靶向的作用效果。本文制备的递送新型化合物的转铁蛋白脂质体,可以为今后进一步体内研究等工作打下坚实地基础。
[Abstract]:As one of the global public health problems, malignant tumors pose a serious threat to human health. As a developing country, the morbidity and mortality of cancer patients in China are increasing year by year. The research and development of new anti-tumor agents is of great significance because of the toxicity and side effects of existing chemotherapeutic drugs. Isoquinoline compounds are a class of compounds with anticancer activity, but they have the disadvantages of low solubility in water, difficulty in absorption and poor effect of direct administration. In this study, antitumor activity of compounds with isoquinoline structure was screened, and drug loading system was developed to improve drug absorption and reduce side effects. This research mainly includes the following aspects: 1. Screening of antineoplastic activity and determination of physicochemical properties 11 isoquinoline compounds and 伪 -methylene 纬 -butyrolactone compounds were selected. The results of IC50 assay and HEK-293T cell inhibition test for the screening of strong anti-tumor compound. He Lahp G2 cells by MTT cell inhibition test showed that [4- bromo-2-butadiol 4-bromo-benzoyl -1-cyano-2-dihydro-isoquinoline-1-yl] -2-bromo-phenyl-methyl]-[4- bromo-2-butadiol 4-bromo-benzoyl -1-cyanoyl] -2-bromo-phenyl-methyl]- Menthyl acrylate (compound 2) has a strong inhibitory effect on tumor cells. The inhibitory effect on normal cells was weak, so this compound was selected as an anti-tumor drug. The physical and chemical properties of the compound were determined. The results showed that the compound was extremely difficult to dissolve in water, and its solubility in ethanol was 1. 52 mg/m / L, so it was difficult to be directly administered. The establishment of liposome, transferrin liposome drug carrier system in order to improve cell absorption of compound 2, the formulation of liposome was selected to develop, and the liposome was modified with transferrin. Liposomes targeting transferrin receptors were prepared. The physicochemical properties of liposomes and transferrin liposomes were investigated and the concentration of transferrin and the amount of transferrin were optimized. The results showed that the optimized preparation conditions of liposomes were as follows: LP: TfN 500: 1, drug loading 10. 5%. The obtained liposomes were about 77 nm in diameter, while transferrin liposomes were 144 nm in size, with potentials in the range of -7 渭 -4 MV, with no significant difference of .3. Evaluation of liposomes and transferrin liposomes in vitro the preparation of liposomes and transferrin liposomes were evaluated in vitro. The results showed that the two drug delivery systems could maintain good stability in the serum environment and achieve a certain sustained release effect through the serum stability and in vitro release test. The results of flow cytometry and laser confocal focus showed that the transfection efficiency of Tf-LPs was significantly higher than that of LPswhich indicated that transferrin coupling with liposome could improve the uptake efficiency of compounds in tumor cells. The results showed that Tf-LPs entered into tumor cells mainly through TF receptor mediated endocytosis. The results showed that the inhibition of Tf-LPs on He-La and Hep G2 cells was stronger than that on LPs. The results of cell ROS and mitochondrial membrane potential changes also showed that Tf-LPs could induce the increase of ROS level and apoptosis of cells. In conclusion, transferrin-targeted liposomes for delivery of novel compounds are potential tumor drugs. The selected compounds with good antitumor activity can overcome the disadvantages of poor solubility and low bioavailability of the original compounds through the encapsulation of transferrin liposome delivery system, and achieve the effect of prolonging the circulation time and targeting the tumor. In this paper, transferrin liposomes for delivery of new compounds can lay a solid foundation for further research in vivo.
【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R943

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