用于肿瘤诊断治疗的仿细胞膜磷酸胆碱胶束的研究

发布时间：2018-02-21 01:48

本文关键词： 纳米医学磷酸胆碱两性离子聚合物胶束 pH响应诊疗一体化　出处：《浙江大学》2016年博士论文　论文类型：学位论文

【摘要】：聚合物纳米胶束,由于可以改善药物溶解度、增加在肿瘤部位靶向富集并提高生物利用度,已成为提高癌症诊断与治疗效果并减少毒副作用的重要纳米载体之一。本文依据磷酸胆碱两性离子的细胞膜仿生特性,致力于设计合成磷酸胆碱基载体材料,构建以两性离子磷酸胆碱为亲水壳层的聚合物纳米胶束,并通过对胶束内核的不同设计实现其在癌症诊断与治疗上的多功能化作用,主体包含以下四个方面:1.研究采用RAFT聚合制备了磷酸胆碱基嵌段共聚物PMPC-b-PDDMMA。该共聚物能自组装形成单分散的纳米胶束,并能在蛋白质和培养基溶液中保持粒径稳定。该共聚物疏水嵌段中的环状缩酮基团,可在酸性条件下实现断裂。体外药物释放结果显示,相比于正常生理环境,酸性环境下阿霉素的释放速度显著增强。细胞实验也表明该载药胶束能有效实现细胞内的阿霉素释放并抑制HepG2肝癌细胞的增殖。同时,包载IR-780近红外染料胶束的活体成像结果显示,磷酸胆碱化胶束能有效富集于肿瘤组织。2.研究进一步基于非共价键的主客体作用,设计合成了酸性敏感的磷酸胆碱化聚合物前药胶束。通过RAFT聚合制备了含有金刚烷的无规共聚物poly(MPC-co-Ada),并用2DNOESYNMR证实了金刚烷与阿霉素修饰的β-环糊精(DOX-hyd-CD)之间的主客体络合作用。体外药物释放结果表明,所制备的前药胶束可在模拟溶酶体/内涵体的酸性条件下通过腙键的断裂实现阿霉素的快速释放。细胞实验结果证实了前药胶束的有效细胞内吞以及对HepG2肝癌细胞增殖的抑制作用。得益于“即插即用”式的主客体作用,该前药胶束还有望进一步实现多功能化的肿瘤治疗效果。3.研究进一步通过RAFT聚合制备了含苯甲醛基团的磷酸胆碱基共聚物,并通过腙键接枝了 AIE分子四苯基乙烯(TPE)。所得到的共聚物PMPC-hyd-TPE能自组装形成以磷酸胆碱为壳、以TPE为核的纳米胶束。疏水的TPE分子聚集成核,不但可以激发AIE效应还能通过疏水相互作用包载阿霉素。体外药物释放实验表明,在模拟细胞内酸性环境下,腙键键接TPE的断裂使得胶束溶胀并快速释放出阿霉素。细胞实验显示,该聚合物胶束具备良好的AIE成像效果并可有效抑制HepG2肝癌细胞的增殖。同时,DLS结果表明磷酸胆碱化胶束在含FBS的DMEM培养基里能保持长效稳定,且与血浆蛋白BSA和FBG共孵育24h后,胶束吸附蛋白质的含量维持在较低水平。进一步动物实验显示,磷酸胆碱化载药胶束能靶向富集并传递阿霉素至肿瘤组织。4.研究进一步探索了通过简便的一步聚合以及单一近红外染料负载制备诊疗一体化的磷酸胆碱均聚物胶束的可行性。通过增长磷酸胆碱单体MPC的烷基链长度得到了两亲性单体MDPC,经一步RAFT聚合便可得到两亲性均聚物PMDPC。该两亲性均聚物PMDPC同样能自组装形成稳定的胶束,并实现对近红外染料IR-780的有效负载。经均聚物胶束负载后,近红外染料IR-780的光稳定性显著增强,并具备高效的光热转换效果,能有效杀伤BxPC-3胰腺癌细胞。进一步的动物实验表明,IR-780包载的磷酸胆碱化均聚物胶束能有效富集于肿瘤区域,具有高分辨且长效的近红外成像效果。
[Abstract]:The polymer micelles, which can improve drug solubility and bioavailability to increase and enriched in tumor target, improve the diagnosis and treatment of cancer has become one of the important carrier of nano effect and reduce toxic side effects. Based on the characteristics of cell membrane biomimetic phosphorylcholine zwitterionic, committed to the synthesis of choline phosphate based carrier material design, to construct zwitterionic phosphorylcholine polymer micelles as hydrophilic shell, and through the different design of the micelle core realization of multiple functions in cancer diagnosis and treatment of the main effect, including the following four aspects: 1. study by RAFT polymerization preparation of phosphorylcholine based block copolymers PMPC-b-PDDMMA. the copolymer can self assemble to form micelles dispersed, and can keep the culture medium solution in stable particle size. The copolymer protein and hydrophobic block in cyclic ketal Groups can realize fracture under acidic conditions. The in vitro drug release results showed that, compared to the normal physiological environment, the release rate of adriamycin in acidic environment was significantly enhanced. Cell experiments showed that the micelle can effectively achieve the intracellular release of doxorubicin and inhibit HepG2 liver cancer cell proliferation. At the same time, the results of in vivo imaging encapsulation IR-780 near infrared dye micelles showed that the micelles could effectively enrich bile alkaline phosphate on.2. tumor tissue further based on object function of non covalent bonds, bile acid sensitive polymer alkaline phosphate prodrug micelles were designed and synthesized. The preparation of Copolymer Poly containing adamantane via RAFT polymerization (MPC-co-Ada), and Adamantane and adriamycin modified beta cyclodextrin confirmed by 2DNOESYNMR (DOX-hyd-CD) complex interaction between subject and object. The in vitro drug release results showed that the micelles prepared before Quick release can be realized by adriamycin in acidic conditions of hydrazone bond fracture simulation under the lysosomal / endosomal cells. Experimental results confirm the effective cell prodrug micelles endocytosis and inhibit the proliferation of HepG2 cells. Thanks to the "plug and play" object type for.3. of tumor therapy the prodrug micelles is also expected to further realize multiple functions of the preparation of phosphorylcholine based copolymers containing benzaldehyde groups further RAFT polymerization, and the hydrazone bond grafted onto the AIE molecule four phenyl ethylene (TPE) copolymer. The PMPC-hyd-TPE can self assemble to form with phosphorylcholine as shell with TPE micelles nuclear. TPE hydrophobic aggregation nucleation, but also can not only stimulate the AIE effect of doxorubicin loaded by hydrophobic interaction. The in vitro drug release experiments showed that in the acidic environment simulation cell, hydrazone The bond cleavage of TPE micelle swelling and rapid release of the adriamycin. The cell experiment showed that, the polymer micelles have AIE good imaging effect and can effectively inhibit the proliferation of HepG2 cells. At the same time, DLS results showed that phosphate containing FBS in micellar Bile Alkaline DMEM culture medium can maintain long-term stability, and plasma protein BSA and FBG were co incubated with 24h, the content of micelle adsorption protein remained at a low level. Further animal experiments of phosphocholine micelles can transfer.4. to adriamycin target tumor tissue to enrichment and to further explore the feasibility through simple polymerization step and a single near infrared dye load choline phosphate preparation treatment the integration of the homopolymer micelles. By alkyl chain length growth of phosphorylcholine monomer MPC obtained two amphiphilic monomer MDPC by one-step polymerization of RAFT can get two amphiphilic homopolymer PMDPC. the two amphiphilic homopolymer PMDPC can also self assemble to form stable micelle, and the effective load of near infrared dye IR-780. The homopolymer micelle load, light stability of near infrared dye IR-780 was significantly enhanced, and have high photothermal conversion effect, can effectively kill BxPC-3 pancreatic cancer cells. Animal experiment further, IR-780 loaded the phosphorylcholine homopolymer of micelles could effectively accumulate in the tumor region, with near infrared imaging of high resolution and long-term.

【学位授予单位】：浙江大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R73-3;O631.3

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