臺灣博碩士論文加值系統

2011年台灣衛生福利部公告國人死因第一名為惡性腫瘤，男性第一名癌症死因為肝癌；女性則為乳腺癌，目前治療方式多以手術、放射性、化療、標靶治療等方式，雖然治癒率與五年存活率逐漸升高，但治療期間為病人帶來的副作用極其不舒服，近年來有學者提出：使用天然物併用臨床用藥可減輕藥物的副作用， 黃酮類化合物為植物次生代謝產物具有非常良好的抗發炎、抗氧化和干擾癌細胞週期生長等功能。為了開發天然藥物於癌症治療上的應用，我們藉由併用兩種天然物黃酮類化合物以低濃度達到比單一用藥更好的效果，我們以廣泛存在於食物當中的芹菜素 (Apigenin)和白楊素 (Chrysin)應用在肝癌與乳癌中，其中三陰性乳腺癌目前尚無有適合的標靶藥物。最近臨床研究表示在許多癌症中SKP2過度表達或Wnt/β-catenin不正常的被活化，然而目前並無針對此兩個蛋白的標靶藥物，因此開發SKP2及Wnt/β-catenin蛋白的抑制劑是很重要的。我們以Apigenin 併用Chrysin處理肝癌細胞HepG2與三陰性乳癌細胞MDA-MB-231在細胞存活率試驗中發現能有效減少癌細胞存活率；由西方墨點法結果得知Apigenin 併用Chrysin能明顯減少Wnt/β-catenin 訊號途徑中LRP6、phosph-LRP6、β-catenin的蛋白質表現量，並抑制調控細胞週期S期SKP2表現量，且近期研究指出Wnt/β-catenin及SKP2與細胞凋亡相關，在細胞實驗中發現Apigenin 併用Chrysin可藉由Caspase Pathways達到促進HepG2和MDA-MB-231凋亡的效果，由此可證SKP2不只可以調控細胞週期亦能促進細胞凋亡。為了進一步佐證藥物的效果，我們將MDA-MB-231細胞異體移植至免疫缺陷鼠體內產生腫瘤進行動物實驗，結果顯示併用兩種天然物可明顯使腫瘤縮小。由本實驗證明Apigenin併用Chrysin能有效抑制HepG2與MDA-MB-231的增生並抑制腫瘤生長，因此我們認為Apigenin併用Chrysin具有成為治療藥物的潛力，在未來癌症研究上，值得進一步研究其作用機制。

According to the Ministry of Health and Welfare, cancer is first leading cause death in Taiwan in 2011. Hepatocellular carcinoma (HCC, also called malignant hepatoma) is one of the most common tumors worldwide, and males usually are more affected than females. Breast cancer is the most universal cancer in women. Some patients may be given chemotherapy or radiation therapy after surgery to kill any cancer cells. Although multivariate analysis demonstrated that the overall five-year cancer survival rate for HCC and breast cancer has been increasing, but the drug treatments often cause uncomfortable side effects in patients. Recent clinical studies have shown that overexpression of S-phase kinase protein 2 (SKP2) and Wnt/β-catenin have been observed in many cancers. Unfortunately, specific drugs that target SKP2 and Wnt/β-catenin are unavailable at present.
Therefore, it is important to develop SKP2 and Wnt/β-catenin inhibitors as chemopreventive agents. Some scholars have proposed that the use of phytochemicals with clinical trials reduce the side effects of the drug treatments. Many studies have shown that the combined use of several drugs with different mechanisms of action can exhibit synergistic effects. In order to develop medicine in cancer treatment, we use two phytochemicals at low concentrations to achieve better results than a single drug.
Apigenin and chrysin (5,7 - dihydroxyflavone) are flavonoids, which have been reported in previous studies of anti-inflammatory, antioxidant, and can promote cell death or slow growth in many human cancers cells. However, one death signal transduction mechanisms apigenin inhibit the growth of liver cancer cells and breast cancer cells are still poorly understood. Although the anti-cancer activities of Apigenin and chrysin have attracted scientific attention, the detailed mechanisms of Apigenin and chrysin in HCC and breast cancer prevention have not been fully deciphered.
In this study, we want to investigate the combined effects of co-treatment with Apigenin and chrysin on HCC and breast cancer. MTT assay indicated that apigenin combined with chrysin can reduced HepG2 and MDA-MB-231 cell viability. The effect of chrysin combined apigenin on cell cycle phase distribution in breast cancer cells and liver cancer were analyzed by flow cytometry. After treatment of each cell line with these compounds, we found that Apigenin combined with chrysin synergisticallyinduced apoptosis in HepG2 and HMDA-BM-231 cells through downregulation of SKP2 and low-density lipoprotein receptor-related protein 6 (LRP6) expression. Furthermore, the combination of Apigenin and chrysin also suppressed tumor growth in nude mice with xenografted MDA-MB-231 cells through the down-regulation of SKP2 protein. To further evidence the effect of the drug, we MDA-MB-231 cell xenograft in immune-deficient nude mice to produce tumors 28 days after injection, tumors were observed changes. The experimental results show that apigenin combined with chrysin can reduce the HepG2 and MDA-MB-231 proliferation and induce apoptosis. It also offers opportunities for exploring new drug targets, and further investigations are underway in this regard.

目次
中文摘要1
英文摘要2
縮寫對照表4

第一章 緒論6
肝癌與乳癌7
標靶蛋白12
黃酮類化合物14

第二章 研究目的16
第三章 實驗架構18
第四章 實驗藥品與器材20
第五章 實驗方法27
第六章 實驗結果36
第七章 討論45
第八章 實驗結果圖表49
第九章 參考附圖77
第十章 參考文獻81

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