羟基喜树碱论文：携载羟基喜树碱酸度敏感聚合物微球的研究.doc

羟基喜树碱论文：携载羟基喜树碱酸度敏感聚合物微球的研究【中文摘要】羟基喜树碱(HCPT)具有较广抗肿瘤谱,但因其不溶于水及生理上可接受的有机溶剂,并易转变为抗肿瘤活性低且有严重副作用的羧酸盐形式,从而限制了HCPT的临床应用。利用肿瘤部位的微酸性环境,使用酸度敏感聚合物作为抗癌药物制剂的载体材料,同时利用肝癌细胞表面含有大量的去唾液酸糖蛋白受体构建含靶向基团的纳米级微球制剂,将药物选择性富集于肿瘤部位,以提高药物的生物利用度和抑制肿瘤生长的效果。在静电喷射技术制备载药微球中,运用正交实验法,获得了影响载药微球的粒径和HCPT包裹效率的主次因素,建立了纳米微球粒径和HCPT包裹效率与主要因素的回归方程,验证实验表明计算值与实验结果有较好的一致性。采用采用优化的工艺条件,以聚乙二醇与聚乳酸共聚物(PELA)、含缩醛结构的酸度敏感聚合物(PBELA)和含半乳糖基的酸度敏感聚合物(PGBELA)为载体材料；制备了载HCPT纳米微球。结果表明微球形态规整,大小均一,平均粒径约为250 nm。荧光显微镜图片显示大部分药物包裹到微球当中,微球的载药量在1.3%左右,药物包裹效率达到40%以上。分别在pH 7.4,6.0,5.0的缓冲液中,从微球失重、分子量降低和分子量分布变宽等几方面考察三种聚合物微球的降解行为和药物释放情况。结果表明PBELA和PGBELA微球在酸性条件下降解速度要远高于中性条件下的降解,而PELA微球在酸性条件下则显得相对稳定,体现出PBELA和PGBELA微球的酸度敏感性,且PGBELA微球的酸度敏感性更为明显。三种聚合物载药微球的释放行为都表现为前期的突释和后期的缓慢释放,同时基于PBELA和PGBELA微球的酸度敏感性,其突释量和最终的累计释放量都大幅增加。在中性和酸性条件下考察三种聚合物载药微球的细胞毒性和细胞吞噬情况。在pH6.8环境下HCPT/PBELA和HCPT/PGBELA微球表现出较高的抑制肿瘤细胞生长的效果,IC50接近中性环境下的50%,而HCPT/PELA微球则无明显差别。细胞吞噬实验结果表明,HepG-2细胞对HCPT/PGBELA微球的吞噬要远高于其他两种载药微球。通过瘤内注射载药聚合物微球,基于肿瘤生长曲线、动物死亡曲线和肿瘤的组织学分析等结果,表明三种聚合物空白微球和生理盐水都没有对肿瘤产生抑制效果,而三种聚合物的载药微球则表现出良好的肿瘤抑制效果,且抑制效果都优于自由HCPT。【英文摘要】Hydroxycamptothecin (HCPT) has wide antitumor spectrum, but it is insoluble in both water and physiological acceptable organic solvents and tends to change into inactive carboxylate form, which restricted the clinical application. Currently, systematical administration of the antineoplasma agent is one of the main strategies for chemotherapy. But it can be hardly delivered to target region, leading to strong side effects and low compliance and therapeutic efficacy. Acid-labile polymers have been developed as carriers of anticancer drugs through the utilization of the pH differences between the tumor and the normal tissues. There are various receptors on the surface of macrophages. and the galactosylation shows the potential strategy for hepatic cells targeting. Acid-labile nanospheres with galactose groups were proposed to target delivery of HCPT to hepatocarcinoma, thereby enhancing the availability of drug material and inhibiting effect on tumor growth.Electrosprayed nanoparticles is a novel processing technique for the production of polymer nanoparticles with diameter of hundreds nanometers and tens micrometers from electrically charged liquid jets of polymer solutions or melts under static electric field. Orthogonal experimental method was firstly used to investigate process parameters of the electrospraying system. Results of statistical analysis showed that significant influences were observed for polymer solution concentration, jet speed and needle size on nanoparticle size and encapsulation efficiency of anti-cancer drugs. Validation test showed that the experimental values of nanoparticle size and encapsulation efficiency of anti-cancer drugs were in good agreement with the calculated ones. Poly(DL-lactide)-poly(ethylene glycol) (PELA), acetal groups containing PELA (PBELA) and galactose modified PBELA (PGBELA) were introduced as nanosphere carriers for HCPT by electrospraying method. Based on the optimized process parameters. HCPT encapsulated nanospheres possessed uniformly spherical and smooth features with the average size of around 250 nm. The loading efficiency of HCPT in nanospheres was about 1.5%, and up to 50% of loading efficiency was detected.HCPT loaded PELA, PBELA and PGBELA microspheres were incubated in buffer solution of pH 7.4, pH 6.0 and pH 5.0, and the matrix degradation and drug release profiles were investigated. HCPT loaded PELA microspheres showed similar profiles of matrix degradation and drug release in the different buffer solutions, which were significantly enhanced for HCPT loaded PBELA and PGBELA microspheres after incubated in pH 6.0 and pH 5.0 buffer solutions. In vitro cell experiments indicated that polymer nanospheres without HCPT had no significant influence on cell viability, while HCPT-loaded nanospheres exhibited significant cytotoxicity. Compared with PELA and PBELA nanospheres, the cellular uptake efficiency of HepG2 cells for PGBELA nanospheres was significantly greater. In vivo study was performed through intra-tumor injection of HCPT-loaded nanospheres with respect to the tumor growth rate, animal survival rate and histological observation. all the drug treatment groups showed significant inhibition of tumor growth, and HCPT-loaded nanoparticles groups indicated the most remarkable inhibition effect.【关键词】羟基喜树碱 静电喷射 酸度敏感 控制释放 抗肿瘤活性【英文关键词】Electrospraying Acid-labile polymer Targeting drug delivery system Anti-tumor activity【目录】携载羟基喜树碱酸度敏感聚合物微球的研究摘要7-9Abstract9-10目录11-14第1章 绪论14-261.1 癌症及癌症治疗14-171.1.1 癌症的现状14-151.1.2 癌症的治疗方法15-161.1.3 癌症化疗的研究现状16-171.2 药物控制释放体系17-201.2.1 药物控释体系的释放机制181.2.2 药物控释体系的靶向机制18-191.2.3 抗癌药物控释体系的研究现状19-201.3 酸度敏感聚合物20-211.3.1 酸度敏感聚合物的特点及种类201.3.2 酸度敏感聚合物在药物控释体系中的应用20-211.4 喜树碱类药物及其剂型研究21-221.4.1 喜树碱类药物的特点211.4.2 喜树碱类药物控释剂型的研究现状21-221.5 纳米微球药物制剂22-241.5.1 纳米微球制剂的特点221.5.2 纳米微球制剂的制备方法22-231.5.3 静电喷射法制备纳米微球的研究现状23-241.6 课题的研究意义、内容及创新24-261.6.1 课题的研究目的和意义241.6.2 课题的研究内容24-251.6.3 课题的创新点25-26第2章 正交设计法研究静电喷射微球的工艺参数26-352.1 实验部分26-282.1.1 实验材料26-272.1.2 交试验设计272.1.3 微球的制备27-282.1.4 微球的表征282.2 结果与讨论28-342.2.1 微球的表征28-292.2.2 极差分析29-302.2.3 方差分析30-322.2.4 交互作用分析322.2.5 回归分析32-332.2.6 验证实验33-342.3 本章小结34-35第3章 携载羟基喜树碱微球制剂的研究35-473.1 实验部分35-393.1.1 实验材料35-363.1.2 酸度敏感聚合物的制备及表征363.1.3 载药微球的制备36-373.1.4 载药微球的表征373.1.5 微球中羟基喜树碱的包裹效率及载药量37-383.1.6 微球的体外降解383.1.7 微球降解的表征38-393.1.8 载药微球的体外释放393.1.9 统计学分析393.2 结果与讨论39-453.2.1 酸度敏感聚合物的表征39-403.2.2 载药微球的表征40-413.2.3 载药微球的包裹效率及载药量41-423.2.4 微球的体外降解行为42-443.2.5 载药微球的体外释放行为44-453.3 本章小结45-47第4章 携载羟基喜树碱微球制剂的抗肿瘤活性研究47-594.1 实验部分47-5