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研究生: 陳玠瑋
Chen, Chieh-Wei
論文名稱: 金奈米棒於酸鹼及光熱應答型藥物載體設計及其於癌症標靶治療之應用
pH-Responsive Drug Release from Gold Nanorods by NIR irradiation for Targeted Drug Delivery
指導教授: 黃郁棻
Huang, Yu-Fen
口試委員: 黃志清
張建文
王福年
學位類別: 碩士
Master
系所名稱: 原子科學院 - 生醫工程與環境科學系
Department of Biomedical Engineering and Environmental Sciences
論文出版年: 2012
畢業學年度: 100
語文別: 中文
論文頁數: 99
中文關鍵詞: 金奈米棒藥物傳遞光熱轉換效應
外文關鍵詞: Gold nanorod, Drug delivery, Photothermal effect
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  • 激活性奈米材料泛指材料在受到外力刺激時,會產生物理或化學現象的改變,例如產生熱、光、化學反應、和體積及運動模式改變等,近年來在生醫領域備受矚目。本研究利用連續波型的808 nm雷射照射金奈米棒 (Au NRs),使Au NRs吸收雷射能量轉換而成的熱能,能加速其表面具有酸鹼應答特性之藥物–PDPH-Dox釋放行為,提高藥物治療果效。為減少藥物在傳遞過程因生理酸鹼值(pH 7.4)導致藥物滲漏產生之毒性,Dox以共價性鍵結修飾於Au NRs上。然而在酸性環境下(pH 4.5~5.5)藥物Dox和載體(Au NRs)之間的腙鍵結 (Hydrazone bond)會被酸解而從載體上釋放出藥物。為增進載體對於目標細胞(CCRF-CEM)的特異性標定,同時在Au NRs表面組裝眾多對於有強親和力的適體–Sgc8c。此外,因巨噬細胞可於腫瘤區域累積,為增進奈米藥物對於腫瘤的傳遞效率,將Sgc8c替換為玻尿酸的奈米藥物載體,可與巨噬細胞(RAW 264.7)表面的CD44藉由特異性標定累積於細胞內,形成一細胞型載體。研究結果顯示,本研究成功開發一兼具酸鹼值和近紅外光應答特性之新型奈米藥物載體,透過雷射光照射,可精準控制奈米載體在細胞內的釋藥行為,藉此雙重釋藥機制可結合藥物及光熱治療,提升毒殺癌細胞效率。同時利用細胞型載體以共培養方式,照射NIR雷射毒殺另一癌細胞Tramp-C1。


    In this study, a doxorubicin (Dox) conjugated gold nanorod (Dox-Au NR) which exhibits a pH-responsive drug release profile has been constructed successfully. The antitumor drug Dox was covalently linked to Au NRs to reduce the overall toxicity of the drug in physiological conditions. However, the acid-labile hydrazone linker, connecting the Dox-Au NR conjugates could be cleaved in an acidic environment. Furthermore, multiple aptamers, sgc8c have also been assembled on the NR surface, providing strong binding affinity with targeted cancer cells through simultaneous multivalent interactions with the cell membrane receptors. Owing to the photothermal effect that Au NR could convert the absorbed light energy to heat, CCRF-CEM cells incubated with Dox-Au NR conjugates were greatly damaged after near-infrared (NIR) light exposure. Our results demonstrated that these novel drug nanocarriers which combine pH- and NIR-responsiveness are highly promising for precise drug releasing in targeted drug delivery. Furthermore, an efficient approach for tumor-targeted drug delivery system was developed with macorphage (RAW 264.7) as the targeting vehicle and a hyaluronic acid conjugated gold nanorod (HA-Au NR) as the drug carrier. This novel strategy provides a great potential to actively deliver nano-therapeutics to tumors utilizing macrophages as tumor-tropic carriers. The synergistic NIR photothermally enhanced drug release of HA/Dox-Au NRs also provides a considerable contribution to enhanced tumor-cell(Tramp-C1) apoptosis and low systematic toxicity.

    總目錄 摘要 I Abstract II 致謝 III 總目錄 IV 圖目錄 IX 表目錄 XI 第一章 緒論 1 1.1 奈米科技與生醫應用 1 1.1.1 奈米科技簡介 1 1.1.2 奈米材料的特性 2 1.1.3 金屬奈米粒子的表面電漿共振 4 1.1.4 奈米藥物載體 7 1.1.5 觸發式的藥物釋放 15 1.2 癌症與治療 25 1.2.1 癌症 25 1.2.2 藥物治療 27 1.2.3 前驅藥物 29 1.2.4 溫熱治療 31 1.2.5 標靶治療 35 1.3 研究動機與目的 38 第二章 實驗材料與方法 44 2.1 實驗藥品與儀器 44 2.1.1 實驗藥品 44 2.1.2 緩衝溶液配置 47 2.1.3 細胞培養與使用 47 2.1.4 儀器 49 2.2 PDPH-Dox的合成與特性鑑定 50 2.2.1 PDPH-Dox的合成 50 2.2.2 透過RP-TLC及HPLC鑑定PDPH-Dox的合成 51 2.2.3 測試PDPH-Dox的酸解能力 51 2.2.4 測試PDPH-Dox的毒性 52 2.3 奈米藥物載體的合成 53 2.3.1 配置金奈米棒的晶種溶液 53 2.3.2 合成金奈米棒 53 2.3.3 合成金銀奈米棒 54 2.3.4 修飾Sgc8c於金銀奈米棒表面 54 2.3.5 將PDPH-Dox以及Sgc8c修飾於金銀奈米棒的表面 55 2.3.6 修飾HA於金銀奈米棒的表面 56 2.3.7 將PDPH-Dox以及HA修飾於金銀奈米棒的表面 56 2.4 增強載體的釋藥能力 57 2.4.1 NIR雷射增強載體上藥物釋放行為 57 2.4.2 流式細胞儀觀測於細胞內的藥物釋放行為 58 2.5 測試載體對於目標細胞CCRF-CEM的選擇性 58 2.5.1 利用螢光標記法測定載體對目標細胞的選擇性 58 2.5.2 利用原子光譜吸收儀測定目標細胞吞噬奈米載體的能力 59 2.6 測試藥物載體對於目標細胞CCRF-CEM的毒殺能力 60 2.6.1 載體照射NIR雷射前後的毒性比較 60 2.6.2 拉長NIR雷射的照射時間增強對目標細胞的毒殺性 61 2.7 利用巨噬細胞攜帶奈米藥物開發一新穎的細胞型載體 62 2.7.1 經由NIR雷射增強細胞型載體的毒殺能力 62 2.7.2 利用細胞型載體毒殺共培養(Co-culture)之癌細胞Tramp-C1 62 2.7.3 利用細胞螢光染色觀察細胞型載體毒殺共培養癌細胞之效果 63 第三章 實驗結果與討論 65 3.1 PDPH-Dox的特性 65 3.1.1 PDPH-Dox的合成鑑定 65 3.1.2 PDPH-Dox的酸鹼應答特性 66 3.2 奈米藥物載體的藥物釋放能力及其標靶治療效果 67 3.2.1 奈米藥物載體的建構 67 3.2.2 結合照射NIR雷射來增強藥物的釋放 68 3.2.3 奈米藥物載體的標靶性 70 3.2.4 奈米藥物載體對於癌細胞的毒性作用 71 3.2.5 比較光熱結合藥物治療的雙重治療優勢 73 3.3 利用細胞型載體傳遞奈米藥物並毒殺癌細胞 74 3.3.1 利用巨噬細胞RAW 264.7作為細胞型載體攜帶奈米藥物 74 3.3.2 透過細胞型載體毒殺共培養(Co-culture)之癌細胞Tramp-C1 75 3.3.3 利用細胞螢光染色觀察細胞型載體毒殺共培養癌細胞的效果 76 第四章 結論 77 參考文獻 89

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