Her2抗体介导的药物传递与靶向治疗
发布时间：2018-11-09 16:27
【摘要】：多年来,癌症一直对人类的健康构成巨大威胁。目前治疗癌症最常用的方法是化疗。但化疗存在很大的弊端,由于化疗药物在体内不能特异性识别肿瘤,在杀死肿瘤细胞的同时,也会杀伤正常组织,从而带来严重的毒副作用。光动力疗法(Photodynamic Therapy, PDT)是近年来兴起的一种治疗癌症的新方法。其中光敏剂是PDT的关键物质。然而绝大多数光敏剂都是不溶于水,或在生理条件下容易聚合。这大大限制PDT的发展。本实验以疏水性吲哚菁绿(ICG-ODA)为光敏剂,阿霉素(doxorubicin, DOX)为化疗药,以Her2抗体为靶头,采用硫酸铵梯度法制备了一种具有靶向特异性的光敏脂质体。本实验采用单因素法筛选处方,制备出了粒径为128.1±1.8nm的光敏脂质体(PSL),粒径大小均一,稳定性高。脂质体的结构清晰,形态良好。DOX包封率高达90%以上。对测定包封率的影响因素考察结果表明包封率的测定方法科学,结果准确、可靠。光敏脂质体的体外释放实验中,经近红外激光(NIR)照射后,DOX会快速且持续地从光敏脂质体中释放出来。结果显示在近红外激光照射条件下,6h内DOX的累积释放量比无近红外激光照射时提高了近20%,近红外激光触发DOX从脂质体释放的效果明显。在以上光敏脂质体的基础上,我们又以DSC为中间嫁接物,制备出了PEG化修饰的Her2抗体,再通过与光敏脂质体进行共孵育制备得到Her2靶向的光敏脂质体(Her2-PSL)。该靶向脂质体可以特异性地被MCF-7摄取。在MCF-7和A549细胞摄取差异实验中,MCF-7细胞中的荧光信号强度远高于A549细胞,表明Her2-PSL能够特异性地主动靶向至MCF-7细胞。荷MCF-7和荷A549肿瘤的动物模型体内分布研究显示,MCF-7肿瘤内荧光信号明显也强于A549肿瘤,表明携带Her2抗体的光敏脂质体能够更多地靶向至MCF-7实体瘤。在抗肿瘤药效学研究中,结果显示照射近红外激光后,治疗组肿瘤抑制率为93.6%,且体重曲线变化波动很小,展现出显著的抗肿瘤效果。实验结果表明,该光敏脂质体与Her2抗体结合后可以选择性靶向到MCF-7肿瘤部位,并在给予近红外激光照射后释放出大量DOX,实现化疗与PDT的协同治疗,疗效显著。二者协同治疗在抗肿瘤治疗领域有着光明的前景。
[Abstract]:Cancer has been a great threat to human health for many years. The most commonly used treatment for cancer is chemotherapy. However, chemotherapeutic drugs have great disadvantages. Because chemotherapy drugs can not specifically recognize tumor in vivo, it can kill tumor cells and kill normal tissues, which brings serious toxic side effects. Photodynamic therapy (Photodynamic Therapy, PDT) is a new method to treat cancer in recent years. Guang Min is the key substance of PDT. However, most Guang Min agents are insoluble in water or easily polymerized under physiological conditions. This greatly limits the development of PDT. Using hydrophobic indocyanine green (ICG-ODA) as Guang Min, doxorubicin (doxorubicin, DOX) as chemotherapeutic agent, and Her2 antibody as target, a kind of liposome with targeted specificity was prepared by ammonium sulfate gradient method. In this experiment, the single factor method was used to screen the prescription, and the liposome (PSL), with a diameter of 128.1 卤1.8nm was prepared with uniform particle size and high stability. The structure and morphology of liposomes were clear and the entrapment efficiency of DOX was over 90%. The results of investigation on the factors influencing the determination of encapsulation efficiency show that the determination method of encapsulation efficiency is scientific, and the results are accurate and reliable. In the in vitro release of Guang Min liposomes, DOX was released rapidly and continuously from Guang Min liposomes after near-infrared laser (NIR) irradiation. The results showed that under the condition of NIR laser irradiation, the cumulative release amount of DOX in 6 h was increased by nearly 20% than that without NIR laser irradiation, and the effect of NIR laser triggering DOX release from liposome was obvious. On the basis of the above Guang Min liposomes, we prepared the Her2 antibody modified by PEG with DSC as the intermediate grafted material. Then we co-incubated with Guang Min liposomes to obtain the Her2 targeted Guang Min liposomes (Her2-PSL). The targeted liposome can be specifically ingested by MCF-7. The fluorescence intensity of MCF-7 cells was much higher than that of A549 cells in the uptake difference assay between MCF-7 and A549 cells, which indicated that Her2-PSL could target MCF-7 cells specifically and actively. The distribution of MCF-7 and A549 tumor models in vivo showed that the fluorescence signal in MCF-7 tumor was stronger than that in A549 tumor, indicating that Guang Min liposome carrying Her2 antibody could target more MCF-7 solid tumor. In the study of antitumor pharmacodynamics, the results showed that the tumor inhibition rate of the treatment group was 93.6 after NIR laser irradiation, and the change of body weight curve was very small, showing significant antitumor effect. The results showed that the liposome combined with Her2 antibody could selectively target to the tumor site of MCF-7 and release a large amount of DOX, after irradiation with NIR laser to achieve the synergistic effect of chemotherapy and PDT. The synergistic therapy has a bright prospect in the field of anti-tumor therapy.
【学位授予单位】：哈尔滨商业大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R943
[Abstract]:Cancer has been a great threat to human health for many years. The most commonly used treatment for cancer is chemotherapy. However, chemotherapeutic drugs have great disadvantages. Because chemotherapy drugs can not specifically recognize tumor in vivo, it can kill tumor cells and kill normal tissues, which brings serious toxic side effects. Photodynamic therapy (Photodynamic Therapy, PDT) is a new method to treat cancer in recent years. Guang Min is the key substance of PDT. However, most Guang Min agents are insoluble in water or easily polymerized under physiological conditions. This greatly limits the development of PDT. Using hydrophobic indocyanine green (ICG-ODA) as Guang Min, doxorubicin (doxorubicin, DOX) as chemotherapeutic agent, and Her2 antibody as target, a kind of liposome with targeted specificity was prepared by ammonium sulfate gradient method. In this experiment, the single factor method was used to screen the prescription, and the liposome (PSL), with a diameter of 128.1 卤1.8nm was prepared with uniform particle size and high stability. The structure and morphology of liposomes were clear and the entrapment efficiency of DOX was over 90%. The results of investigation on the factors influencing the determination of encapsulation efficiency show that the determination method of encapsulation efficiency is scientific, and the results are accurate and reliable. In the in vitro release of Guang Min liposomes, DOX was released rapidly and continuously from Guang Min liposomes after near-infrared laser (NIR) irradiation. The results showed that under the condition of NIR laser irradiation, the cumulative release amount of DOX in 6 h was increased by nearly 20% than that without NIR laser irradiation, and the effect of NIR laser triggering DOX release from liposome was obvious. On the basis of the above Guang Min liposomes, we prepared the Her2 antibody modified by PEG with DSC as the intermediate grafted material. Then we co-incubated with Guang Min liposomes to obtain the Her2 targeted Guang Min liposomes (Her2-PSL). The targeted liposome can be specifically ingested by MCF-7. The fluorescence intensity of MCF-7 cells was much higher than that of A549 cells in the uptake difference assay between MCF-7 and A549 cells, which indicated that Her2-PSL could target MCF-7 cells specifically and actively. The distribution of MCF-7 and A549 tumor models in vivo showed that the fluorescence signal in MCF-7 tumor was stronger than that in A549 tumor, indicating that Guang Min liposome carrying Her2 antibody could target more MCF-7 solid tumor. In the study of antitumor pharmacodynamics, the results showed that the tumor inhibition rate of the treatment group was 93.6 after NIR laser irradiation, and the change of body weight curve was very small, showing significant antitumor effect. The results showed that the liposome combined with Her2 antibody could selectively target to the tumor site of MCF-7 and release a large amount of DOX, after irradiation with NIR laser to achieve the synergistic effect of chemotherapy and PDT. The synergistic therapy has a bright prospect in the field of anti-tumor therapy.
【学位授予单位】：哈尔滨商业大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R943
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