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研究生: 杜珮鈺
Tu, Pei-Yu
論文名稱: 發展具有診療合一多功能鐵鉑高分子複合奈米粒子於癌症治療上的應用
Development of Theranostic Multifunctional FePt-polymer Nanoparticles in Cancer Therapeutic Application
指導教授: 吳炳慶
Wu, Ping-Ching
學位類別: 碩士
Master
系所名稱: 工學院 - 生物醫學工程學系
Department of BioMedical Engineering
論文出版年: 2018
畢業學年度: 106
語文別: 英文
論文頁數: 62
中文關鍵詞: 奈米醫學奈米材料鐵鉑奈米粒子磁性奈米粒子聚乳酸-羥基乙酸共聚物紫杉醇化學治療放射治療協同作用
外文關鍵詞: Nanomedicine, nanomaterials, FePt nanoparticles, magnetic nanoparticles, PLGA, Paclitaxel, chemotherapy, radiation therapy, Synergistic inhibition
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  • 近年來奈米應用於醫學上出現革命性的變化,奈米技術對於癌症的早期預防、初期顯影偵測、精準定位、準確投藥,能夠形成一系列完整的療程。它被認為可以改善傳統診斷及治療的缺點,甚至降低醫療成本。奈米粒子透過其功能,可以被區分為顯影診斷用及藥物的傳遞輸送系統,如市場上常見的Feridex®顯影劑及輸送藥物的Lipo-Dox®。從磁性奈米粒子角度來看,粒子可以增強質子在特定組織的鬆弛,並適用於對分子成像中的顯影,如超音波、磁共振成像(MRI)、電腦斷層掃描(CT)、正電子發射斷層掃描(PET)、光學生物發光和螢光顯影劑等。
    由於它們獨特的特性和在細胞生物間相互作用的能力,這些磁性奈米粒子可以實時監測癌細胞變化及進行藥物治療。其中超順磁鐵鉑奈米粒子因為具有高穩定X-ray吸收能力結合,使得鐵鉑奈米粒子成為MRI及CT雙重顯影劑。在另一方面,奈米粒子的藥物遞送系統克服了傳統化療藥物於人體中的限制,透過高分子材料的包封,避免藥物在運送至腫瘤位置過程中,傷害正常組織,同時以延長藥物在腫瘤中的積累,這種現像被稱為增強滲透性和保留效應(EPR效應)。而本研究整體目標在於發展具有診療合一多功能鐵鉑高分子複合奈米粒子,以PLGA作為高分子材料包封一線化療藥物Paclitaxel,同時帶有具雙重顯影效果的鐵鉑奈米粒子,以作為輻射敏感劑增強放射療法之療效。
    根據我們的實驗結果發現:在沒有放射線的輔助治療下,鐵鉑奈米粒子可以增強化療藥物PLGA@Paclitaxel對於癌細胞的抑制效果。當1 mg/mL 鐵鉑奈米粒子鍵結於濃度為6.25 µg/mL的PLGA@Paclitaxel表面上時,對於癌細胞的抑制生長效果相當於濃度為25 µg/mL的PLGA@Paclitaxel;而0.5 mg/mL 鐵鉑奈米粒子鍵結於濃度為6.25 µg/mL的PLGA@Paclitaxel,癌細胞生長抑制率高達50%,因此在後續的測試,我們皆以0.5 mg/mL 鐵鉑奈米粒子與6.25 µg/mL 作為實驗組。而針對放射治療的測試,根據MTT毒性檢驗可以發現在2Gy時,實驗組對於4T1癌細胞具有相當顯著的差異。

    In recent years, nanotechnology has undergone revolutionary changes in medicine, especially in the case of cancer. Nanotechnology has formed a complete treatment plan from early prevention, initial development detection, precise positioning, and accurate drug administration. It is believed to improve the shortcomings of traditional diagnosis and treatment and may even reduce medical costs. Nanoparticles can be distinguished based on the functions of their diagnostic and delivery systems, such as Feridex® developer and the drug microsphere Dox®. From the perspective of magnetic nanoparticles, magnetic nanoparticles can enhance the relaxation of protons in specific tissues and are suitable for visualization in molecular imaging, such as ultrasound, magnetic resonance imaging (MRI), computed tomography (CT), positron emission tomography (PET), optical bioluminescence, and fluorescent developers.
    Because of their exceptional properties and ability to interact with cells, these magnetic nanoparticles can monitor changes in cancer cells and medications in real time. Among them, superparamagnetic platinum nanoparticles demonstrate highly stable seepage absorption, so iron-platinum nanoparticles can become a dual developer in MRI and CT. On the other hand, the drug delivery system of nanoparticles overcomes the limitations of traditional chemotherapeutic drugs in the human body. Through the encapsulation of polymer materials, the drug is prevented from being damaged during normal delivery to the tumor site, and at the same time, the drug is prolonged and accumulates in tumors. This phenomenon is known as enhanced permeability and retention effects (EPR effects).
    The overall goal of this study is to develop a multi-functional iron-platinum polymer nanoparticle with diagnosis and treatment capabilities to encapsulate the first-line chemotherapy drug paclitaxel with PLGA as a polymer material, and to develop iron-platinum nanoparticles with dual development effects. Radiation-sensitive agents enhance the efficacy of radiation therapy.
    According to our results, FePt nanoparticles can enhance the inhibitory effect of the chemotherapy drug, PLGA@Paclitaxel, on cancer cells without radiation. 1 mg/mL FePt-PLGA@Paclitaxel (6.25µg/mL PLGA@Paclitaxel) and the highest concentration of PLGA@Paclitaxel alone (25µg/mL) exhibited the same colony formation, which showed that FePt nanoparticles could weaken the proliferation of 4T1 cells and enhanced the inhibitory effect of the chemotherapy drug. At 0.5 mg/mL FePt-PLGA@Paclitaxel (6.25µg/mL PLGA@Paclitaxel), the growth inhibition rate of cancer cell is as high as 50%. Therefore, in the subsequent tests, we used 0.5 mg/mL FePt-PLGA@Paclitaxel (6.25µg/mL PLGA@Paclitaxel) as the experimental group. For the radiation test, the experimental group has a significant difference for 4T1 cancer cells at 2Gy by MTT assay.

    摘要 I Abstract III 誌謝 VI Contents VIII List of Tables XI List of Figures XII Abbreviation List XIII Chapter 1 Introduction 1 1.1 Nanotechnology 2 1.1.1 Cancer nanomedicine 2 1.1.2 The EPR effect for nanoparticles on tumor cells 4 1.2 Nanomaterials 5 1.2.1 Magnetic nanoparticles 7 1.2.1.1 Magnetic nanoparticles as contrast and therapy agents 9 1.2.2 Nanoparticles for targeted drug delivery systems 10 1.3 The cancer therapy 12 1.3.1 Surgery 12 1.3.2 Radiation therapy 12 1.3.3 Chemotherapy 13 1.4 Synergistic inhibition of tumor cells using radiation therapy and chemotherapy 14 1.5 Hypothesis and Specific Aims 15 Chapter 2 Materials and Methods 16 2.1 Materials 17 2.2 Synthesis of 12nm FePt nanoparticles 18 2.2.1 Ligand exchange of 12nm FePt nanoparticles 18 2.2.2 Characterization of FePt nanoparticles 19 2.3 Synthesis of Paclitaxel-loaded PLGA nanoparticles 19 2.3.1 Determination of paclitaxel content in PLGA NPs 20 2.3.2 Paclitaxel release from PLGA nanoparticles 21 2.4 FePt NPs conjugated with Paclitaxel-loaded PLGA NPs 21 2.5 Cell culture 22 2.5.1 Cell culture of 4T1 cells 22 2.5.2 Cell culture of Vero cells 22 2.5.3 Cell count 23 2.6 Cytotoxicity assay 24 2.7 Radiation test 25 2.7.1 Colony formation assay of 4T1 cells in vitro 25 2.7.2 Cellular uptake of Fe+/Pt+ 26 2.7.3 Apoptosis assay 26 2.7.4 ROS 27 Chapter 3 Results 28 3.1 Characterization of the FePt nanoparticles 29 3.2 Characteristics of the PLGA@Paclitaxel NPs 30 3.3 Characteristics of the FePt-PLGA@Paclitaxel NPs 30 3.4 The Cytotoxicity of FePt NPs 31 3.5 PLGA@Paclitaxel NPs influence the proliferation of 4T1 cells 31 3.6 FePt-PLGA@Paclitaxel NPs enhanced the chemotherapeutic sensitivity in 4T1 cells 31 3.7 FePt-PLGA@Paclitaxel NPs enhanced the radiation sensitivity in 4T1 cells 32 3.8 FePt-PLGA@Paclitaxel nanoparticles upregulated ROS 33 Chapter 4 Discussion and Conclusion 34 4.1 Discussion 35 4.2 Conclusion 38 Tables and Figures 39 Reference 59

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