臺灣博碩士論文加值系統

中文摘要
近年來隨著生活及飲食西方化，台灣大腸直腸癌發生率逐年增加，根據行政
院衛生署2002 公佈的統計資料，大腸直腸癌是癌症死因的第三位。曾有很多的
實驗結果及臨床上的報告指出，在多數癌症中，表皮生長因子接受器(EGFR)會活
化細胞內的訊息傳遞作用，控制細胞的增殖、凋亡、血管心生及轉移，但過度表
現會使癌症病患的預後較差,而且癌細胞容易轉移。5-Fluorouracil(5-FU)是治
療大腸直腸癌患者常用的化療藥物，主要作用機制是能夠有效阻斷DNA 合成及形
成結構不完全的RNA 而抑制癌細胞的分裂，化學治療失敗的主要因是癌細胞容易
產生抗藥性，而對於5-FU 的抗藥性目前仍不清楚，雖然有初步研究顯示EGFR
的活性抑制劑可以增強5-FU 的毒殺作用，但到目前為止EGFR 和5-FU 的抗藥性
關係並未被確立，在本研究中，我們將探討5-FU 與大腸直腸癌中EGFR 過度表現
的關係。我們利用了四株大腸直腸癌的細胞株，包含HCT8/Parental 和三株以
5-FU 養成的抗藥性細胞株，分別是HCT8/Yale、HCT8/7D 及HCT8/4h，我們以化
學免疫染色法和西方墨點法初步證實，表皮生長因子接受器的表現量增加和
5-FU 的抗藥性有正相關性，不論利用是Iressa 抑制EGFR 的激酶活性或是以
SiRNA Knockdown EGFR 的表現量，進行細胞群落試驗，均顯示會增加5-FU 對細
胞的毒殺作用，我們的結果證實了EGFR 過度表現會使細胞對於5-FU 產生抗藥
性，但其機制作用到目前仍不明瞭。

Abstract
In Taiwan, due to the westernized lifestyle and food consumption, the incidence
of colorectal cancer is increased yearly in recent years. According to the report from
the Department of Health of Executive Yuan 2002, colorectal cancer has become the
country’s third leading cause of cancer death. There were many studies and clinical
reports showed that over-expression Epidermal Growth Factor Receptor(EGFR)p;
could activate cancer cells signal transduction and lead to cancer cells proliferations,
apoptosis, angiogenesis and metastasis and therefore result in the poor prognosis in
cancer patients. The mechanisms of 5-fluorouracil (5-FU), which is a common choice
of chemotherapy for treating colon cancer patients, to inhibit growth of cancer cells
involve the ability to cease DNA synthesis and to interrupt RNA structures .Up to now,
there has been no established relationship between 5-FU and EGFR although 5-FU is
frequently included in the EGFR-based chemotherapy regimens for treating colon
cancer recently. In this study, we investigated the correlation between a novel
resistance to 5-FU and over-expression of EGFR in colon cancer. We used four colon
cancer cell lines including HCT8-parental and three derived 5-FU resistant cell lines
named HCT8-yale, HCT8-4h and HCT8-7d based on the different treatment with
5-FU in this study. Our results show that level of EGFR is increased corresponding
with the level of resistance to 5-FU by immunocytochemistry staining and Western
blotting. Pre-treatment with inhibitor (e.g. Iressa) that inhibits kinase activity of
EGFR indeed sensitizes the parental and 5-FU-resistant cells to 5-FU by clonogenic
assays. Moreover, knockdown of the expression level of EGFR by small interfering
RNA increases the cytotoxicity to 5-FU in the stable transfected parental and
5-FU-resistant cells. Thus our results collectively suggest that over-expression of
EGFR could result in the resistance of 5-FU and agents targeting EGFR enhance
5-FU mediated cytotoxity despite the molecular mechanism is not yet understood.

目次
第一章文獻回顧
第一節大腸直腸癌1
第二節表皮生長接受器（Epidermal Growth Factor Receptor EGFR） 10
第三節5-Fluorouracil (5-FU) 12
第二章材料與方法18
第三章結果34
第四章結論37
第五章討論38
第六章參考文獻41
附錄
圖一、5_FU 藥物作用機制圖47
圖二、細胞毒性試驗48
圖三、EGFR 機制圖49
圖四、細胞株EGFR 蛋白質的表現50
圖五、細胞群落分析試驗51
圖六、Iressa 機制圖52
圖七、EGFR Inhibitor 53
圖八、RNAi 的作用機制簡圖54
圖九、SiRNA 染色圖片55
圖十、Parental SiRNA cell lines IC50 57
圖十一、SiRNA cell lines EGFR level 58
圖十二、SiRNA cell lines 群落分析試驗61

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