多功能纳米粒子在肝癌诊治一体化中的应用研究

发布时间：2018-06-23 06:40

本文选题：肝癌 + 诊治一体化　；参考：《吉林大学》2015年博士论文

【摘要】：肝癌是严重危害我国人民健康的恶性肿瘤之一，尽管以手术切除为基础、并辅助化疗、热疗等综合治疗部分改善了肝癌的治疗效果，但是肝癌患者的长期存活率仍然较低。由于大多数原发性肝癌患者的病情隐匿、潜伏期长、肿瘤生长迅速导致早期诊断困难，而且治疗后的耐药、复发和转移成为了肝癌患者死亡的主要原因。因此，早期精确诊断、提高药物的靶向性、降低治疗的毒副作用成为目前肝癌诊治的挑战。近年来，纳米技术的兴起为攻克临床上肝癌诊治的难题带来了新的机遇，而作为纳米技术和生物医学的结晶，纳米医学已成为当前最具有转化潜力的交叉学科之一。纳米药物载体是为纳米医学领域研究的热点，其中无机纳米粒子作为纳米材料领域的后起之秀，以其独特的纳米结构和性能以及良好的稳定性、高产量、低成本等优势备受纳米医学工作者的关注。目前整合了诊断和治疗特点的多功能纳米平台引起了研究者们的广泛关注，此类多功能无机纳米材料须具备以下三个特点：（1）通过多种分子影像学手段实现对肿瘤的早期精确诊断。（2）联合多种治疗手段协同治疗肿瘤，在提高治疗效果的同时降低副作用。（3）对药物的肿瘤靶向运输、释放和治疗效果进行实时示踪，指导并调整给药剂量，实现肿瘤的个体化治疗。基于无机纳米材料的上述优点，本博士论文紧绕改善肝癌诊治效果这一中心目标，，针对临床肝癌诊治中早期诊断难、传统化疗毒性大、效果差以及基因治疗靶向性差三大挑战，分别以量子点和磁性介孔二氧化硅两种无机纳米粒子为基础，发展智能分子设计、纳米特性控制、靶向分子修饰等纳米药物技术，构筑高效、安全的多功能纳米平台，开发具有靶向肝癌的基因治疗和降低肝癌化疗毒副作用的新型纳米药物，揭示其选择性杀伤肝癌细胞的分子机制；利用分子影像学技术对药物在体内的运输和释放过程进行实时示踪，并对肝癌治疗效果与生物安全性进行系统性评价；终而实现肝癌的诊治一体化。本论文主要创新性研究成果概括如下：（1）为了实时示踪HSV-TK/GCV自杀基因系统，我们成功的将量子点偶联TK基因，证实生物偶联既不影响量子点的发光特性，也不影响TK基因的生物学活性，实时示踪发现TK基因于转染24h进入胞核并表达TK蛋白，确定24h为GCV的最佳给予时间，并在体内外成功对HSV-TK/GCV自杀基因系统的抗肝癌效果进行监测，提示量子点可以作为示踪HSV-TK/GCV自杀基因系统的理想载体。（2）为了实现自杀基因的靶向肝癌治疗，我们成功构筑叶酸脂质体担载量子点自杀基因复合体（FL/QD-TK），证实其可在体外靶向和选择性杀伤叶酸受体高表达肝癌细胞，并可在体内外可视化示踪TK基因运输和治疗效果，同时具备了较好的生物安全性，提示FL/QD-TK作为一种潜在的高效低毒的纳米基因药物有望应用于肝癌的诊治一体化。（3）为了驾驭镉系量子点的生物毒性用于肝癌治疗，我们成功的制备出量子点脂质复合体（QD-LC），证实其可通过巨胞饮途径大量内吞进入肝癌细胞产生大量的ROS，通过线粒体依赖的Caspase凋亡途径选择性杀伤肝癌细胞。QD-LC还可抑制肝癌微小瘤灶的形成，并在实现肝癌荷瘤有效治疗的同时具备了较好的生物安全性，提示可利用镉系量子点内源性毒性实现肝癌的选择性治疗。（4）为了提高化疗药物治疗肝癌的有效性并降低毒副作用，我们通过优化反应条件制备出粒径合适、形貌均一的Janus型磁性介孔二氧化硅纳米粒子（M-MSNs），担载化疗药物DOX得到的Janus型M-MSNs-DOX具备了外加磁场介导的作用，证实Janus型M-MSNs-DOX可在体内外实现选择性肝癌治疗的效果，并具备了较好生物安全性。我们还证实外加磁场可增强Janus型M-MSNs-DOX的肝癌细胞内吞作用和在肿瘤部位的富集，且不影响其生物安全性。最后我们在体内外成功实现了肝癌诊治一体化的目标，提示Janus型M-MSNs有望解决肝癌化疗靶向性差、毒副作用大的弊端，实现肝癌的诊治一体化。综上所述，本论文通过构筑多功能纳米诊治平台，对肝癌靶向自杀基因治疗的监测、利用纳米毒性选择性治疗肝癌以及实现高效低毒的肝癌化疗策略进行了系统而又深入的研究，不仅成功实现了肝癌的诊治一体化，也为早日实现肝癌的早期诊断和个体化治疗提供新方法和新思路。
[Abstract]:Liver cancer is one of the malignant tumors that seriously harm the health of the people of our country. The long-term survival rate of the patients with liver cancer is still low, although the combined treatment of surgical resection and adjuvant chemotherapy and thermotherapy has improved the therapeutic effect of liver cancer. Early diagnosis is difficult, and drug resistance, recurrence and metastasis after treatment are the main causes of death in the patients with liver cancer. Therefore, early accurate diagnosis, improving the targeting of drugs and reducing the toxic and side effects of the treatment have become the challenge for the diagnosis and treatment of liver cancer. As a new opportunity, nanomedicine, as the crystallization of nanotechnology and biomedicine, has become one of the most promising interdisciplinary subjects. Nano drug carriers are the hot spots in the field of nanomedicine. Inorganic nanoparticles are the following show in the field of nanomaterials, with their unique nanostructures and properties as well as good properties. The advantages of stability, high output, low cost and other advantages have attracted the attention of nanomedicine workers. At present, the multi-functional nano platform which integrates the characteristics of diagnosis and treatment has attracted the attention of researchers. This kind of multifunctional inorganic nanomaterial must have the following three characteristics: (1) through a variety of molecular imaging methods to realize the early stage of the tumor Accurate diagnosis. (2) combined with multiple treatments to treat tumors together to improve the effect of the treatment and reduce the side effects. (3) real-time tracing of the target transport, release and treatment effect of the drug, guiding and adjusting the dosage and realizing the individualized treatment of the tumor. Based on the above advantages of inorganic nanomaterials, this doctoral thesis is tightly wound around In order to improve the central target of the diagnosis and treatment of liver cancer, it is difficult to diagnose the early diagnosis in the diagnosis and treatment of the clinical liver cancer, the traditional chemotherapy is big, the effect is poor and the target of the gene therapy is poor three major challenges. Based on the quantum dots and the magnetic mesoporous silica two inorganic nanoparticles, the intelligent molecular design, the nano characteristic control and the target molecular repair are developed respectively. To build an efficient and safe multi-functional nanoplatform, to develop a novel and multi-functional nanoplatform for targeting liver cancer and to reduce the side effects of hepatoma chemotherapy, and to reveal the molecular mechanism of the selective killing of liver cancer cells, and the use of molecular imaging techniques to carry out real-time transport and release of drugs in the body. A systematic evaluation of the therapeutic effect and biological safety of liver cancer, and the integration of the diagnosis and treatment of liver cancer. The main innovative research results in this paper are summarized as follows:
(1) in order to trace the HSV-TK/GCV suicide gene system in real time, we successfully coupled the quantum dots with the TK gene to verify that the biological coupling does not affect the luminescence characteristics of the quantum dots and does not affect the biological activity of the TK gene. The real-time tracer found that the TK gene was transfected into the nucleus and reached the TK protein by transfecting 24h to determine the optimum giving time of the 24h as GCV, and The anti hepatoma effect of the HSV-TK/GCV suicide gene system was monitored in vitro and in vivo, suggesting that the quantum dots could be used as an ideal carrier for tracing the HSV-TK/GCV suicide gene system.
(2) in order to achieve the target liver cancer therapy of suicide gene, we successfully constructed the folic acid liposome loaded quantum dot suicide gene complex (FL/QD-TK), and confirmed that it can target and selectively kill the liver cancer cells in vitro and selectively kill the folic acid receptor, and can visualize the transport and treatment effect of TK gene in vivo and in vivo. Physical safety suggests that FL/QD-TK as a potential high effective and low toxic nanomaterials is expected to be applied in the diagnosis and treatment of liver cancer.
(3) in order to control the biotoxicity of the cadmium system quantum dots for the treatment of liver cancer, we have successfully prepared the quantum dot lipid complex (QD-LC), and confirmed that it can produce a large number of ROS by massive endocytosis into the hepatoma cells through the mega drink pathway, and selectively kill the liver cancer cell.QD-LC through the mitochondrial dependent Caspase apoptosis pathway to inhibit the liver cancer micro The formation of small tumors and the effective treatment of hepatoma bearing tumor have good biological safety. It is suggested that the endogenous toxicity of cadmium system quantum dots can be used to achieve selective treatment of liver cancer.
(4) in order to improve the efficacy and lower toxic side effects of chemotherapeutic drugs in the treatment of liver cancer, we have prepared Janus type magnetic mesoporous silica nanoparticles (M-MSNs) with suitable particle size and homogeneous morphology by optimizing the reaction conditions. The Janus M-MSNs-DOX obtained by carrying chemotherapeutic drug DOX has the effect of applied magnetic field mediated, and the Janus type M-MSNs- is confirmed. DOX can be used in the treatment of selective liver cancer in vivo and in vivo, and has good biological safety. We also confirm that the applied magnetic field can enhance the endocytosis and enrichment of Janus type M-MSNs-DOX in the tumor site, and do not affect its biological safety. Finally, we have successfully realized the goal of the integration of liver cancer in the body and the body, It is suggested that Janus M-MSNs is expected to solve the problems of poor targeting and side effects of chemotherapy for hepatocellular carcinoma and achieve the integration of diagnosis and treatment of HCC.
To sum up, this paper makes a systematic and in-depth study on the monitoring of targeted suicide gene therapy for liver cancer by constructing a multi-functional nano diagnosis and treatment platform, using nano toxic selective treatment of liver cancer and realizing high efficiency and low toxicity of liver cancer. It not only successfully realized the integration of the diagnosis and treatment of liver cancer, but also realized the liver cancer at an early date. Early diagnosis and individualized treatment provide new methods and new ideas.
【学位授予单位】：吉林大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R735.7

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