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研究生(外文):Tzu-Chao Chuang
論文名稱(外文):Study on HER2/neu-targeting Cancer Gene Therapy
指導教授(外文):Ming-Ching Kao
中文關鍵詞:神經膠原致癌基因猿病毒腫瘤抗原EB 病毒核抗原-1微脂粒基因治療合併基因治療
外文關鍵詞:HER2/neuSV40LTEBNA1liposomegene therapycombined gene therapy
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文獻報告神經膠原致癌基因的基因調控失常會導致許多惡性腫瘤的產生,若能抑制此致癌基因的表現,即能抑制細胞的轉形、癌化與惡性轉移。因此,神經膠原致癌基因是一個發展抗癌藥物的良好標的。本實驗室即以此為目標,目前已有下列五項成果:第一、發現猿病毒腫瘤抗原氮端部位(LT425)除了能抑制神經膠原致癌基因過度表現的卵巢癌細胞之轉形與癌化作用,同時也是神經膠原致癌基因過度表現的乳癌細胞之轉形與轉移抑制劑;第二、建立卵巢癌腫瘤之裸鼠動物模式,發現利用自製的微脂粒攜帶 LT425 進行神經膠原致癌基因標的基因治療,可達50﹪效果,且安全性高;第三、發現 EB 病毒核抗原-1(EBNA1)能有效抑制神膠原致癌基因之轉錄表現及癌化表徵,其作用機轉與延長 G2/M 細胞週期以及降低抗藥性有關;第四、發現EB 病毒核抗原-1之氮端部位(EBNA1-NT)即能作為神經膠原致癌基因之轉形抑制劑,大幅減低神經膠原致癌基因過度表現的卵巢癌細胞之抗藥性;第五、評估合併 LT425 與 E1A 進行基因治療之可行性。綜合本論文之結果可提供為將來臨床上治療因神經膠原致癌基因過度表現所引發之人類癌症的有益參考資料。
It is known that the deregulation of the HER2/neu oncogene will cause many types of human cancers. Repression of this oncogene expression will suppress the transformation, tumorigenicity and malignant metastasis of the cancer cells. Thus, the HER2/neu oncogene is a good target for developing anti-cancer agents. Our lab has achieved the following five discoveries by making an attemp at approaching this goal. First, the N-terminal domain of the SV40 large T antigen (LT425) may act as HER2/neu-mediated transforming and metastasis suppressor both in ovarian and breast cancers. Second, using the lab-made SP/DOPE-LT425 lipoplex-mediated HER2/neu-targeting approach, 50% of the mice would survive for more than one year in an ovarian tumor-bearing nude mouse model. No significant toxic effects to the mice were observed in the entire lipoplex treated process. Third, the wild-type Epstein-Barr virus nuclear antigen-1 (EBNA1) could inhibit the HER2/neu oncogene expression at the transcriptional level and subsequently repress the HER2/neu-mediated malignant phenotypes. The repression mechanism was proposed to be related to the prolongation of G2/M phase and sensitize the cancer cells to the topoisomerase-IIa-targeting drugs. Fourth, the N-terminal domain only of EBNA1 (EBNA1-NT) could also act as a transforming suppressor of the HER2/neu oncogene. It could sensitize the HER2/neu-overexpressed ovarian cancer cells to anti-cancer drugs. Finally, the possibility of a combined gene therapy approach was evaluated by using LT425 and E1A genes. Taken together, the results of this thesis may provide beneficial evidences of using liposome-mediated HER2/neu-targeting approaches to the human cancer gene therapy.
第一節、研究緣起與目的 ………………………………………………….1
第二節、神經膠原致癌基因與癌症 ………… ……………………………4
第三節、癌症基因治療 ……………………………………………………22
2.1.1. 實驗材料與方法 ……………………………………………………32
2.1.2. 結果與討論 …………………………………………………………36
2.1.3. 結果圖表 ……………………………………………………………41
2.2.1. 實驗材料與方法 ……………………………………………………48
2.2.2. 結果與討論 …………………………………………………………51
2.2.3. 結果圖表 ……………………………………………………………55
的轉形能力並增加抗癌藥物的活性 ………………………………………67
3.1.1. 實驗材料與方法 ……………………………………………………72
3.1.2. 結果與討論 …………………………………………………………78
3.1.3. 結果圖表 ……………………………………………………………82
細胞的G2/M細胞週期並增加對拓普脢 II a 標的藥及Taxol等藥物敏感性
3.2.1. 實驗材料與方法 ……………………………………………………92
3.2.2. 結果與討論 …………………………………………………………96
3.2.3. 結果圖表 …………………………………………………………102
現之卵巢癌細胞的轉形能力並減低抗藥性 ……………………………110
3.3.1. 實驗材料與方法 …………………………………………………110
3.3.2. 結果與討論 ………………………………………………………112
3.3.3. 結果圖表 …………………………………………………………117
第四章:合併性基因治療之評估 ………………………………………126
4.1. 實驗材料與方法 ……………………………………………………127
4.2. 結果與討論 …………………………………………………………130
4.3. 結果圖表 ……………………………………………………………132
第五章:結論 ………………………………………………………………134
參考文獻 ……………………………………………………………………135
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