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標題: | 賀癌平包覆之氧化鐵奈米粒子之製備與生醫上的應用 On the preparation of Herceptin-coated magnetite nanoparticles and their biomedical application |
作者: | Kuan-Yu Chen 陳冠羽 |
指導教授: | 何國川(Kuo-Chuan Ho) |
關鍵字: | 生物共軛物,賀癌平,四氧化三鐵,標的治療, bioconjugate,Herceptin,magnetite,targeting therapy, |
出版年 : | 2008 |
學位: | 碩士 |
摘要: | 在高頻交流磁場下,氧化鐵磁性奈米粒子因磁滯損耗所產生之熱能可能用於臨床上對於腫瘤組織的高溫熱治療。然而卻因磁性奈米粒子對於腫瘤細胞的特異性不佳,限制了此種治療方式的發展。在本研究當中,引入了對人類乳癌細胞上的人類表皮生長因子第二接受體 (HER2/neu)有特異性的人工合成單株抗體,賀癌平 (Trastuzumab,Herceptin®)。本研究將賀癌平分子連接到氧化鐵奈米粒子的表面,增進氧化鐵粒子對於腫瘤細胞的標定效果。
作為核心的氧化鐵奈米粒子乃是利用共沈澱法,使部份的三價鐵離子還原而形成的四氧化三鐵奈米粒子。隨後加入聚丙烯酸進行奈米粒子的表面修飾,聚丙烯酸分子所帶有的羧基不但能夠減少奈米粒子的聚集情況,亦使奈米粒子能夠進行更進一步的合成。而聚丙烯酸包覆之氧化鐵奈米粒子與賀癌平分子之間,則是利用NHS/EDC方法所形成的胜肽鍵加以鍵結。接著討論經賀癌平分子包覆後的奈米粒子其各項性質,包含了不同pH值下的粒徑分佈、飽和磁矩及在交流磁場下的加熱效果等。為了測試賀癌平包覆之氧化鐵奈米粒子其標定效果以及細胞毒性,本研究將不同濃度經賀癌平修飾之奈米粒子與人類乳癌細胞 (SK-BR3)一同培養,並使用ICP檢驗經賀癌平分子辨識後吸附於腫瘤細胞的氧化鐵吸附量。此外,奈米粒子亦與正常的人類纖維母細胞一同培養,用來測試所合成奈米粒子的細胞毒性。 由FTIR光譜當中碳氧雙鍵特徵峰的偏移,可以確認賀癌平分子與氧化鐵之間的鍵結。此外,由TGA儀器所偵測到的熱損失亦能夠提供更進一步的證據。賀癌平包覆之氧化鐵奈米粒子其平均粒徑及飽和磁矩分別為 26 nm及 30 emu/g。由ICP所得結果可知賀癌平包覆之氧化鐵奈米粒子具有對HER2的特異性,且其標定效率約為2%。在細胞毒性實驗結果顯示人類纖維母細胞對於氧化鐵奈米粒子可容忍的最大濃度為0.05 mg/ml。本研究顯示賀癌平包覆之氧化鐵奈米粒子可望達到對於乳癌局部熱治療的功效。 AC magnetic field induces magnetite nanoparticles (MNPs) to generate heat which could be used as heat source for hyperthermia, but poor targeting ability of MNPs to the tumor cells needs to be improved. To solve this problem, Trastuzumab, Herceptin®, a monoclonal antibody directed against the HER2/neu receptor overexpressed on breast tumor cell, was applied to this study. In this research, Trastuzumab was used as a targeting moiety to enhance the targeting ability of iron oxide nanoparticles. The MNPs were synthesized by the coprecipitation method, and surface modification of MNPs was done by the poly(acrylic acid) (PAA) which not only prevents nanoparticles from aggregation but provides carboxyl groups for further synthesis. After the synthesis of PAA-coated MNP, the attachment of Herceptin was reached by the peptide bonding synthesis, N-hydroxysuccinimide (NHS) and 1-ethyl-3-[3-dimethylaminopropyl] -carbodiimide hydrochloride (EDC) methods. Furthermore, the basic properties of Herceptin-coated magnetite nanoparticles (HMNPs), such as particle size under different pH values, saturated magnetization and hyperthermia ability, were discussed. To further examine the targeting ability of HMNPs, the SK-BR3, human breast cancer cells were cultured with HMNPs, and inductive coupled plasma (ICP) was used to estimate the iron uptake amount. Additionally, the cytoxicity tests of HMNPs were achieved by in vitro test, in which human fibroblast served as normal cells. From the FTIR spectrum, the antibody, Herceptin, was successfully attached onto the MNP surface due to the shift of C=O. Furthermore, the mass loss in TGA analysis gave identical conclusion. The particle size and saturated magnetization of HMNPs were 26 nm and 30 emu/g, respectively. ICP gave proof that HMNPs targeted human breast cancer cells with targeting efficiency about 2%. In vitro cytoxicity test showed the maximum tolerance concentration of HMNPs to human fibroblast was 0.05 mg/ml. The HMNPs are expected to achieve local hyperthermia for breast cancer therapy in future applications. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/26671 |
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顯示於系所單位: | 化學工程學系 |
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