乏氧和近红外光照双控的上转换纳米抗肿瘤诊疗试剂的构建及评价

发布时间：2018-03-15 11:35

本文选题：肿瘤　切入点：诊断治疗　出处：《河北大学》2017年硕士论文　论文类型：学位论文

【摘要】：诊断治疗学已成为药学、医学、化学等多学科交叉领域的前沿和热点。诊疗试剂能够为癌症的高效诊治提供一个集早期实时监测、定位诊断与个性化干预治疗于一体的多功能平台。抗肿瘤诊疗试剂主要通过内部刺激(如pH、乏氧等)或外部刺激(如光照、磁场等)来实现药物的可控释放。乏氧已被应用于肿瘤的诊断与治疗,但由于某些其他疾病也会引起组织乏氧,导致药物在非肿瘤部位释放从而引起毒副作用。光响应诊疗剂具有在特定的时间和空间实现药物可控释放的优点。上转换纳米发光材料(Upconversion Nanoparticles,UCNPs)能够将近红外光转换成可见光和紫外光。近红外光能够有效避免生物体自体荧光的干扰,对生物组织有较高的穿透能力,对生物样品的光损伤也比较小。然而,光响应诊疗剂所释放的药物不可避免会对所辐照区域的正常组织产生毒副作用。若将肿瘤的乏氧(内在特征)和光照(外部刺激)巧妙结合设计新型诊疗试剂,则可通过乏氧和光照双重控制药物的释放,达到降低抗肿瘤药物的毒副作用和精准治疗的目的。本文以氟脲苷(FDU)作抗肿瘤药物模型,Yb~(3+)、Tm~(3+)掺杂的NaYF_4上转换发光纳米颗粒(UCNP)作为光转换载体,取代E-邻羟基肉桂酸(CAE)作为光敏基团(荧光染料的前体),4-硝基苄醚作为乏氧响应基团,经亲核取代反应、Wittig反应、水解反应、酯化反应、缩合反应、Click反应等反应合成了乏氧和近红外光照双重控制的诊疗试剂UCNP-CAE-FDU/NO_2。采用TEM、NMR、MS及IR方法对诊疗剂及中间体的进行表征;采用HPLC及HRMS法研究了其释药机制;采用荧光光谱法探讨乏氧孵育及近红外光照时间对释药量的影响;采用MTT法探讨在不同乏氧加近红外光照条件下UCNP-CAE-FDU/NO_2对MCF-7细胞活力的影响;采用双光子激光共聚焦荧光成像法对乏氧细胞内药物释放情况进行检测。结果表明:纳米诊疗试剂UCNP-CAE-FDU/NO_2,分散性良好,尺寸单一,平均粒径为30 nm。经乏氧孵育和近红外光照射,能够释放出FDU,并同时生成香豆素衍生物;香豆素衍生物的荧光强度可以反映FD U的释放量,并且FDU释放量随乏氧孵育及光照时间的延长而增加;诊疗剂在乏氧和近红外光照条件下对MCF-7细胞产生毒性;释放出的香豆素衍生物可用于报告肿瘤细胞乏氧程度及细胞内药物释放情况。该诊疗试剂利用乏氧结合光照的策略可实现药物的精确释放,为实现乏氧肿瘤的诊断治疗一体化提供了一种新方法。
[Abstract]:Diagnostic therapeutics has become the frontier and hotspot in the interdisciplinary fields of pharmacy, medicine, chemistry and so on. The diagnostic and therapeutic reagents can provide an early and real-time monitoring for the effective diagnosis and treatment of cancer. A multifunctional platform for localizing diagnosis and individualized intervention therapy. Antitumor agents are mainly obtained through internal stimuli (e.g. pH, hypoxia, etc.) or external stimuli (e.g. light, light, etc.). Hypoxia has been used in the diagnosis and treatment of tumors, but some other diseases can also cause tissue hypoxia. Light response diagnostic agents have the advantage of controlled drug release at a specific time and space. Upconversion nanoluminescent material can be used to convert UCNPsto infrared light. The drug can be released in non-tumor sites and cause toxic side effects. Photo-responsive diagnostic agents have the advantage of controlled drug release at a specific time and space. The up-conversion nano-luminescence material can be used to convert UCNPsto near infrared light. Visible light and ultraviolet light. Near infrared light can effectively avoid the interference of organism autofluorescence. Higher penetration to biological tissue and less light damage to biological samples. However, Drugs released by photo-responsive diagnostic agents will inevitably produce toxic side effects on normal tissues in irradiated areas. If the tumor's hypoxia (intrinsic characteristics) and light (external stimuli) are skillfully combined to design new diagnostic and therapeutic reagents, The release of the drug can be controlled by both hypoxia and light. In order to reduce the toxicity and side effects of antitumor drugs and to treat them accurately, the NaYF_4 up-conversion luminescent nanoparticles (UCNP) doped with fluorouracil (FDU) as the antitumor drug model were used as optical conversion carrier. Substituted E- o-hydroxycinnamic acid (CAE) was used as Guang Min group (the precursor of fluorescent dyestuff, Con 4- nitrobenzyl ether) as anoxic responsive group, which was treated by nucleophilic substitution reaction, Wittig reaction, hydrolysis reaction and esterification reaction. UCNP-CAE-FDU / NOS2 was synthesized by condensation reaction and Click reaction. The drug release mechanism of UCNP-CAE-FDU / NOS2 was studied by HPLC and HRMS, and the diagnosis and treatment agents and intermediates were characterized by TEMP-NMRMS and IR methods, and the drug release mechanism of UCNP-CAE-FDU / NOS2 was studied by HPLC and HRMS. The effects of hypoxic incubation and near-infrared illumination time on drug release were investigated by fluorescence spectrometry, and the effects of UCNP-CAE-FDU/NO_2 on the activity of MCF-7 cells under different hypoxic and near-infrared illumination conditions were investigated by MTT method. Two-photon laser confocal fluorescence imaging was used to detect the drug release in hypoxic cells. The results showed that the nano-diagnostic reagent UCNP-CAE-FDU / no _ 2 had good dispersion, single size and an average particle size of 30 nm. The fluorescence intensity of coumarin derivative can reflect the amount of FDU release, and the amount of FDU release increases with the increase of hypoxia incubation and illumination time, and the fluorescence intensity of coumarin derivative can reflect the amount of FDU release, and the fluorescence intensity of coumarin derivative can reflect the amount of FDU release. The diagnostic and therapeutic agents were toxic to MCF-7 cells under hypoxia and near infrared illumination. The released coumarin derivatives can be used to report the degree of hypoxia and intracellular drug release in tumor cells. It provides a new method for the diagnosis and treatment of hypoxic tumor.
【学位授予单位】：河北大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R730.4;TB383.1

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