量子点在胃癌的早期诊断、治疗和预防中的应用研究

发布时间：2018-06-08 22:44

本文选题：CdSe/ZnS + CuInS2/ZnS　；参考：《上海交通大学》2015年博士论文

【摘要】：胃癌是世界上常见的癌症之一,是继肝癌之后世界上致死率第二的恶性疾病。在中国,平均每年因胃癌死亡的人数约22.9万人,而我国胃癌的早期诊断率不到10%。现代医学要求针对胃癌建立高灵敏度和高特异性的分析与诊断方法,在所有传统及现代的分析方法中,光学成像具有独特的高分辨率优势。过去用于标记组织和细胞的有机荧光染料普遍存在量子产率低、发射光谱宽、易被光漂白等缺点。与传统的有机荧光染料相比,量子点(Quantum dots,QDs)具有许多优异的光谱性能,在生物化学、细胞生物学、分子生物学、分析化学等研究领域显示出极其广阔的应用前景。近年来,随着量子点修饰方法的发展及特异性靶点的发现,量子点已被广泛应用于细胞标记、单分子示踪、活体成像以及疾病靶向诊断和可视化治疗等研究领域。本文以功能化量子点为研究对象,研究了其光谱可控性合成、表面修饰、生物相容性、稳定性及其在胃癌的早期诊断和治疗中的应用,主要包括:胃癌细胞的高灵敏度、高特异性标记,体内肿瘤成像,胃癌疫苗在体内主动靶向免疫治疗及示踪等,具体如下:(1)在非极性溶剂中合成具有核壳结构的CdSe/ZnS量子点,设计制备了两亲性聚合物,该两亲性聚合物具有齿状的烷基配位键,配位键的另一端是具有生物功能的羧基,羧基同时为量子点提供了良好的水溶性基团。使用该聚合物进行CdSe/ZnS量子点的表面化学修饰,成功将量子点由非极性溶剂相转移至水相。评价了经相转移以后的量子点的表面电荷、羧基含量、水合粒径、离子和pH稳定性、体外细胞毒性等参数,为该材料在分子标记、细胞成像及体内示踪应用等,奠定了基础。(2)使用前述相转移后的CdSe/ZnS核壳结构量子点为标准材料,选择BRC AA1和Her2的单克隆抗体作为原型抗体,与量子点进行化学偶联后,制备了抗BRCAA1和抗Her2的两种量子点荧光探针。实验证实,这些探针可以对胃癌细胞MGC-803进行原位特异性靶向标记。此外,使用发射峰657nm量子点标记的MGC-803细胞可以在裸鼠体内通过活体荧光成像系统有效地进行跟踪。结果表明:本研究制备的量子点探针可实现胃癌细胞的体外特异性靶向分子标记,在胃癌模型动物体内可进行高灵敏度、高分辨率成像,在早期胃癌的诊断中具有一定的应用潜力。(3)从健康人类供体的外周血中分离出单个核细胞,在体外经细胞因子诱导,培养出异源树突状细胞。以这种异源性树突状细胞和胃癌细胞为基础,在体外制备了胃癌融合瘤疫苗。本研究合成了具有近红外发射光谱的CuInS2/ZnS核壳结构量子点,按前述方法使用该量子点对融合瘤疫苗进行了标记。将标记后的融合瘤疫苗免疫接种C57BL/6小鼠,用活体成像系统对融合瘤疫苗细胞进行了实时示踪,以评价融合瘤疫苗在体内的免疫治疗和免疫预防效果。结果显示,融合瘤疫苗细胞在体外对多种胃癌细胞系均具有较强的CTL杀伤效应,且可以主动靶向至肿瘤部位,从而实现活体肿瘤成像;另外,当使用适当的肿瘤抗原载体及相应的效应细胞(FC+CIK)免疫接种后,可以在实验动物体内引发至少三个月时间的免疫记忆功能。本研究结果表明:以树突状细胞为基础的胃癌融合瘤疫苗在胃癌的发生、发展、转移及预后等方面,具有良好的应用潜能。(4)制备了荧光磁性纳米粒子(FMNPs),采用FMNPs标记的诱导多能干细胞(iPS),选择胃癌作为体内靶向标志,系统地研究了FMNPs标记的iPS细胞在体内外与胃癌的相互作用,经体内荧光、磁共振成像及体外交变磁场作用下的产热效应等方法,确认了FMNPs标记的iPS细胞可以作为一种有良好治疗作用的生物制剂,可应用于胃癌的体内成像和靶向性热疗,有望推广至其他肿瘤的临床研究或治疗。
[Abstract]:Gastric cancer is one of the common cancers in the world. It is a malignant disease of second in the world after liver cancer. In China, the average number of deaths from gastric cancer is about 229 thousand in China, and the early diagnosis rate of gastric cancer in China is not 10%. modern medicine requires high sensitivity and specificity of analysis and diagnosis for gastric cancer. In the traditional and modern analytical methods, optical imaging has a unique high resolution advantage. In the past, organic fluorescent dyes used to mark tissue and cells have the disadvantages of low quantum yield, wide emission spectrum and easy to be photobleached. Compared with traditional organic fluorescent dyes, Quantum dots (QDs) has many excellent spectroscopic properties. In recent years, with the development of quantum dots modification methods and the discovery of specific targets, quantum dots have been widely used in cell marking, mono tracing, living imaging, and target diagnosis and visualization of disease. The purpose of this study is to study the spectral controllability, surface modification, biocompatibility, stability, and its application in the early diagnosis and treatment of gastric cancer, including high sensitivity, high specificity marker, tumor imaging in vivo, and active target of gastric cancer vaccine in the body. To immunotherapy and tracer, the following are as follows: (1) the synthesis of CdSe/ZnS quantum dots with nuclear shell structure in non polar solvents is designed to prepare two amphiphilic polymers. The two amphiphilic polymers have dentate alkyl coordination bonds, the other side of the coordination bond is a carboxyl group with biological function, and the carboxyl provides good water solubility for the quantum dots at the same time. The surface chemical modification of CdSe/ZnS quantum dots was performed using the polymer. The quantum dots were successfully transferred from nonpolar solvent phase to aqueous phase. The surface charge, carboxyl content, hydrous particle size, ion and pH stability and cytotoxicity in vitro were evaluated after the phase transfer of the quantum dots. In vivo tracing application and so on, laid the foundation. (2) using the CdSe/ZnS nuclear shell structure quantum dots as the standard material after the phase transfer, the monoclonal antibodies of BRC AA1 and Her2 were selected as the prototype antibody, and the quantum dots were chemically coupled with the quantum dots. The anti BRCAA1 and anti Her2 quantum dots fluorescent probes were prepared. In addition, the MGC-803 cells labeled with the emission peak 657nm quantum dots (MGC-803) can be traced effectively in vivo by the living fluorescence imaging system in nude mice. The results show that the specific target molecular marker of gastric cancer cells in vitro can be realized by the quantum dot probe prepared in this study. The model animals can have high sensitivity and high-resolution imaging in the diagnosis of early gastric cancer. (3) isolated mononuclear cells are isolated from the peripheral blood of the healthy human donor, and the heterologous dendritic cells are cultured in vitro by cytokine induction, based on this heterogenous dendritic cell and gastric cancer cells. The fusion tumor vaccine of gastric cancer was prepared in vitro. The CuInS2/ZnS nuclear shell structure quantum dots with near infrared emission spectrum were synthesized. The fusion tumor vaccine was labeled using the quantum dots. The labeled fusion tumor vaccine was inoculated to C57BL/6 mice, and the fusion tumor vaccine cells were carried out with the living body imaging system. The results show that the fusion tumor vaccine cells have a strong CTL killing effect on a variety of gastric cancer cell lines in vitro, and can actively target the tumor site to realize the living tumor imaging; in addition, when the appropriate tumor antigen carrier is used, The immunization of the corresponding effect cells (FC+CIK) can cause at least three months of immune memory function in the experimental animals. The results of this study show that the vaccine of gastric cancer fusion based on dendritic cells has good application potential in the occurrence, development, metastasis and prognosis of gastric cancer. (4) the fluorescence magnetism is prepared. Nanoparticles (FMNPs), using FMNPs labeled induced pluripotent stem cells (iPS), selected gastric cancer as a target marker in vivo, systematically studied the interaction between FMNPs labeled iPS cells and gastric cancer in vivo and in vivo, and confirmed the iPS finer by the methods of fluorescence, magnetic resonance imaging and external alternating magnetic field. Cell can be used as a biological agent with good therapeutic effect. It can be applied to the imaging of gastric cancer in vivo and targeted thermotherapy. It is expected to be popularized to the clinical research or treatment of other tumors.
【学位授予单位】：上海交通大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R735.2

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