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研究生:吳永逸
研究生(外文):Yong-yi Wu
論文名稱:利用肺癌細胞建立荷瘤組織體外培養平台暨尾靜脈注射誘發肺癌動物模式來探討一系列新合成藥物之抗癌活性與分子標記
論文名稱(外文):Studies on anti-cancer activity and molecular marker of 5-substituted 2-cyanoimino-4-imidazodinone and 2-cyanoimino-4- pyrimidinone derivatives based on xenograft lung carcinoma histoculture and animal model via injecting cancer cells into tail vein
指導教授:褚俊傑褚俊傑引用關係
指導教授(外文):Jiunn-Jye Chuu
學位類別:碩士
校院名稱:南台科技大學
系所名稱:生物科技系
學門:生命科學學門
學類:生物科技學類
論文種類:學術論文
論文出版年:2006
畢業學年度:94
語文別:中文
論文頁數:84
中文關鍵詞:肺癌組織培養
外文關鍵詞:histoculturexenograftlung cancer
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根據世界衛生組織的統計,最近二十年來全世界的癌症死亡率正快速上升,其中以肺癌的死亡率增加最快,使得肺癌目前已躍升世界癌症第一殺手的地位,在台灣隨著抽煙人口增加,二手煙的危害以及女性吸煙人口逐年升高,肺癌人口急遽上升,同時罹患肺癌人口平均年齡也不斷下降,使得肺癌死亡人口更為主要癌症死亡原因。然而,進行手術或是化療的病患仍無法完全治癒甚至有復發的可能,化療藥物搭配預後治療成為趨勢,新抗癌藥物的研究更為重要,然而新的化合藥物被合成研究(5-substituted 2- cyanoimino-4-imidazodinone and 2-cyanoimino-4- pyrimidinone derivative of compounds),其有抗癌活性的效果。
我們將以此新化合藥物compound A﹐B﹐C(CP- A﹐B﹐C)對肺癌細胞(LL/2﹐A549)測試,進行細胞毒性分析(MTT)是否藥物有抗癌的效用、再以流式細胞儀技術分析藥物作用在細胞的週期變化及細胞凋亡分析藥物有否使癌細胞凋亡作用。另外,也以擬體內之體外組織培養方法進行組織活性分析,在組織和細胞等級差異的存在,利用(BrdU﹐TUNEL﹐MDR-1 labeling methods)免疫染色切片分析藥物作用在組織間的變化情形,最後再進行活體動物試驗來確定新化合藥物的作用。我們首先進行細胞活性分析在compound A對老鼠肺癌(LL/2) 72小時有最強的效果半致死濃度7.23μΜ(IC50),而compound C對人類肺癌(A549) 24小時有最強效果43.7μM(IC50),發現藥物對兩種細胞的敏感度並不相同,但都有抗癌活性的效果存在。在用流式細胞儀技術測試中,compound A﹐B﹐C使癌細胞週期停留在G1期卻沒有使細胞凋亡作用,在組織活性分析方面卻發現沒有如細胞活性的半致死濃度產生,僅在96小時最高濃度有明顯的差異出現,顯示組織在藥物作用效果上的確實不如細胞等級。
接著利用免疫切片分析發現藥物CP-C在組織間有抑制細胞生長的作用產生,隋著藥物濃度增加細胞增生染色(BrdU)數量顯示減少,在TUNEL免疫染色也發現藥物作用96小時有凋亡增加的情形,再利用膜外耐藥性蛋白(MDR-1)免疫染色分析發現隨著藥物濃度增加,MDR-1表現量減少,表示藥物傳送至細胞內達到抑制肺癌細胞生長的作用沒有受到MDR-1的影響。最後的活體動物實驗測試尾靜脈注射藥物CP-C能使誘導肺癌(LL/2)的ICR裸鼠存活天數增加,確定新化合藥物CP-C在肺癌作用上的活性效果。
Our present study aimed at evaluating putative possibility of using xenograft carcinoma as a model system to monitor effect of chemotherapy regiment of lung cancer. A series of novel synthesized, eg. 5-substituted 2-cyanoimino-4-imidazodinone and 2-cyanoimino-4-pyrimidinone derivative of compounds have been synthesized as anticancer agents.
The derivatives exhibited cytotoxicity against a variety of lung cancer cell lines (LL2 and A549) were evaluated in MTS assay and the cell cycle population using flow cytometry assay. In addition, xenograft nude mice model was determined to evaluate the ex vivo activity of anticancer agents with MTT while treated with these derivatives for 24-96 hr in a concentration range of 0.01-100μM. The MTT assay showed that the derivatives can effectively against tumor growth of human LL2 (IC50 = 2.39μM) while the derivatives cause G1 phase reset (100μM), but no apoptosis was detected via cell cycle assay. In histoculture model, the derivatives exhibited cytotoxicity and decreased the medium level of CEA in a concentration -dependent manner by means of on MTT assay. Our results suggest that the histoculture system may be further used for evaluation of anti-cancer agents for anti proliferation and cell death induction in tumor cells of the histocultured lung carcinoma.
中文摘要……………………………………………………………………………Ⅰ
Abstract…………………………………………………………………………Ⅲ
目次…………………………………………………………………………………Ⅳ
表目錄………………………………………………………………………………Ⅴ
圖目錄………………………………………………………………………………Ⅵ
縮寫對照表…………………………………………………………………………Ⅷ

壹﹑文獻整理………………………………………………1
貳﹑材料與方法……………………………………………21
參﹑實驗結果………………………………………………35
肆﹑討論……………………………………………………42
圖與表………………………………………………………44
參考文獻……………………………………………………81
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