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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 林淑萍 | |
| dc.contributor.author | Chih-Hsuan Kuo | en |
| dc.contributor.author | 郭芷瑄 | zh_TW |
| dc.date.accessioned | 2021-06-16T13:46:22Z | - |
| dc.date.available | 2018-09-24 | |
| dc.date.copyright | 2013-09-24 | |
| dc.date.issued | 2013 | |
| dc.date.submitted | 2013-07-08 | |
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Weng CJ, Fang PS, Chen DH, Chen KD, Yen GC (2010) Anti-invasive effect of a rare mushroom, Ganoderma colossum , on human hepatoma cells. J Agric Food Chem 58: 7657-7663. 58. Koizumi F, Shimoyama T, Taguchi F, Saijo N, Nishio K (2005) Establishment of a human non-small cell lung cancer cell line resistant to gefitinib. Int J Cancer 116: 36-44. 59. Comoglio PM, Giordano S, Trusolino L (2008) Drug development of MET inhibitors: targeting oncogene addiction and expedience. Nat Rev Drug Discov 7: 504-516. 60. Yamauchi M, Yoshino I, Yamaguchi R, Shimamura T, Nagasaki M, et al. (2011) N-cadherin expression is a potential survival mechanism of gefitinib-resistant lung cancer cells. Am J Cancer Res 1: 823-833. 61. Cobaleda C, Perez-Caro M, Vicente-Duenas C, Sanchez-Garcia I (2007) Function of the zinc-finger transcription factor SNAI2 in cancer and development. Annu Rev Genet 41: 41-61. 62. 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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/62403 | - |
| dc.description.abstract | 非小細胞肺癌 (Non-small-cell lung cancer, NSCLC)在肺癌當中佔了80-85%，常帶有epidermal growth factor receptor (EGFR)突變。Gefitinib是一個EGFR tyrosine kinase的抑制劑，用於標靶性治療EGFR突變的非小細胞肺癌特別有效，但臨床上使用後產生抗藥性是有待解決的問題。因此，發展提升Gefitinib在具抗藥性的細胞之效能的治療策略是很重要的。靈芝是一種在亞洲國家被廣泛使用的傳統藥材，含多樣性生物活性的小分子。在此實驗當中，我們從黃芝萃取三萜類GC538，並研究其合併Gefitinib施用的抗癌效用。PC9/gef細胞株比PC9細胞株對Gefitinib具有抗性，其抑制50%細胞活性的濃度 (IC50)相差近兩千倍 (分別為22 μM以及10 nM)。處理Gefitinib在兩個細胞中皆可以抑制EGFR、AKT和ERK的磷酸化，此結果指出PC9/gef對Gefitinib的抗性並不是透過維持EGFR路徑的活化。在PC9/gef細胞中，合併使用Gefitinib和GC538顯示出協同的細胞毒殺效能(combination index＜1)，以濃度50 μM GC538合併濃度10 μM Gefitinib可以明顯增加細胞凋亡：透過增加粒線體膜電位的喪失、DNA斷裂，以及caspase-9和caspase-3活性增加。研究報導指出上皮細胞間質轉換 (epithelial to mesenchymal transition， EMT)和細胞對Gefitinib產生抗藥性有關，但目前機制尚不清楚。實驗進一步研究，合併CG538與Gefitinib是否透過調控EMT的機制在具抗藥性的PC9/gef細胞增加細胞凋亡。細胞在GC538處理後，觀察到細胞由紡錘狀轉變為立方狀的形態，另外不管在mRNA及蛋白質層面，藥物處理後皆觀察到E-cadherin增加與slug下降。結果顯示GC538可以抑制PC9/gef之EMT並增強Gefitinib誘發之細胞凋亡。天然小分子GC538具有潛力成為一佐劑，在對Gefitinib具抗性的細胞株中提升Gefitinib細胞毒殺的效能。 | zh_TW |
| dc.description.abstract | Non-small-cell lung cancer (NSCLC) accounting for 80-85% of lung cancer incidence often harbors epidermal growth factor receptor (EGFR) mutations. Gefitinib is an EGFR tyrosine kinase inhibitor especially effective for NSCLC patients with EGFR mutation, but the development of drug resistance is a challenge in clinical practice. Therefore, therapeutic strategies enhancing efficacy of Gefitinib in the drug resistant cells would be valuable. Ganoderma is a well-known traditional Chinese medicinal herb been used for centuries in Asia. In this study, a triterpenoid, GC538, was isolated from Ganoderma colossum and its combination efficacy with Gefitinib in Gefitinib-resistant cells, PC9/gef, was studied. PC9/gef cells were less sensitive to Gefitinib than PC9 cells with the half maximal inhibitory concentration (IC50) at 22 μM and 10 nM, respectively. Gefitinib treatment inhibited the phosphorylation of EGFR, AKT and ERK in both cell lines, indicating that Gefitinib can still inhibit EGFR pathway in Gefitinib-resistance cells. In PC9/gef cells, the combined treatment of Gefitinib and GC538 displayed synergistic cytotoxicity (combination index＜1), and enhanced cell apoptosis as shown by the increases in the cell populations with lowered mitochondrial membrane potential, with fragmented DNA, with caspase-9 and caspase-3 activation. It has been reported that epithelial to mesenchymal transition (EMT) of cancer cells may cause Gefitinib-resistance, even though the mechanism is still unknown. Further, whether EMT is involved in Gefitinib-induced apoptosis was investigated. GC538 treatment altered cells from spindle-like shape to cubical morphology, increased epithelial maker E-cadherin expression and repressed EMT regulator slug in both mRNA and protein levels. These results showed that GC538 can inhibit EMT and enhance Gefitinib-induced apoptosis in Gefitinib-resistant cells. The compound GC538 isolated from G. colossum has the potential to be an adjuvant to enhance efficacy of Gefitinib. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-16T13:46:22Z (GMT). No. of bitstreams: 1 ntu-102-R00424022-1.pdf: 2527970 bytes, checksum: b4fcc2f89efaaee49e4b92a8723c83db (MD5) Previous issue date: 2013 | en |
| dc.description.tableofcontents | 中文摘要........................................................................................................................ I
英文摘要....................................................................................................................... II 英文名詞縮寫對照表.................................................................................................. IV 圖目錄....................................................................................................................... VIII 一、研究背景................................................................................................................ 1 1.1 肺癌.................................................................................................................... 1 1.2 艾瑞莎/吉非替尼 (Iressa/Gefitinib) ................................................................. 2 1.3 上皮細胞間質轉化 (epithelial to mesenchymal transition，EMT) ................ 4 1.4 靈芝 (Ganoderma) ............................................................................................ 5 1.5 細胞週期 (cell cycle) ........................................................................................ 6 1.6 細胞凋亡 (Apoptosis) ....................................................................................... 6 二、研究目標................................................................................................................ 8 三、材料與方法............................................................................................................ 9 3.1 GC538 的純化 ................................................................................................... 9 3.2 細胞培養............................................................................................................ 9 3.3 細胞活性測試 (ACP assay) ........................................................................... 10 3.4 細胞週期分析.................................................................................................. 10 3.5 細胞內Caspase 活性測定 .............................................................................. 11 3.6 粒線體膜電位測定.......................................................................................... 11 3.7 Annexin V 和PI 染色 ..................................................................................... 11 3.8 蛋白質萃取及西方墨點法.............................................................................. 12 3.9 基因表現分析.................................................................................................. 13 3.10 細胞傷口癒合試驗 (Wound healing assay) ................................................... 13 3.11 細胞骨架F-actin 染色 .................................................................................... 14 3.12 統計分析.......................................................................................................... 14 四、結果...................................................................................................................... 15 4.1 三萜類GC538 的純化及其對癌細胞之毒性 ................................................ 15 4.2 PC9/gef 的EGFR 活性被Gefitinib 抑制但對Gefitinib 具抗藥性 .............. 16 4.3 合併處理Gefitinib 與GC538 對於PC9/gef 的藥物協同作用 (synergistic) 16 4.4 GC538 增強Gefitinib 誘導細胞凋亡............................................................. 17 4.5 合併使用GC538 無法增強Gefitinib 造成G0/G1 arrest .............................. 18 4.6 GC538 改變PC9/gef 形態與抑制細胞爬行 .................................................. 18 4.7 GC538 可抑制EMT 相關分子slug 且促進E-cadherin 上升 ...................... 19 4.8 探討GC538 透過何種路徑調控slug ............................................................ 20 五、討論...................................................................................................................... 21 六、圖.......................................................................................................................... 24 七、參考文獻.............................................................................................................. 42 八、附錄...................................................................................................................... 49 8.1 附錄一. Primer 序列 ........................................................................................ 49 8.2 附錄二. slug 的轉譯調節及後轉錄修飾 ........................................................ 50 九、藥品材料清單...................................................................................................... 51 9.1 藥品與試劑...................................................................................................... 51 9.2 抗體.................................................................................................................. 52 | |
| dc.language.iso | zh-TW | |
| dc.subject | 非小細胞肺癌 | zh_TW |
| dc.subject | 吉非替尼抗藥性 | zh_TW |
| dc.subject | 黃芝 | zh_TW |
| dc.subject | 細胞凋亡 | zh_TW |
| dc.subject | 上皮細胞間質轉化 | zh_TW |
| dc.subject | non-small-cell lung cancer | en |
| dc.subject | Gefitinib resistant cells | en |
| dc.subject | Ganoderma colossum | en |
| dc.subject | apoptosis | en |
| dc.subject | epithelial mesenchymal transition | en |
| dc.title | 靈芝三萜GC538 促進吉非替尼於抗藥性肺癌細胞株之
抗癌活性 | zh_TW |
| dc.title | Ganoderma Triterpenoid GC538 Enhances Anticancer
Activity of Gefitinib in Drug-resistant Non-small-cell Lung Cancer Cell Line | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 101-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 楊雅倩,張雅雯,胡忠怡 | |
| dc.subject.keyword | 非小細胞肺癌,吉非替尼抗藥性,黃芝,細胞凋亡,上皮細胞間質轉化, | zh_TW |
| dc.subject.keyword | non-small-cell lung cancer,Gefitinib resistant cells,Ganoderma colossum,apoptosis,epithelial mesenchymal transition, | en |
| dc.relation.page | 53 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2013-07-08 | |
| dc.contributor.author-college | 醫學院 | zh_TW |
| dc.contributor.author-dept | 醫學檢驗暨生物技術學研究所 | zh_TW |
| 顯示於系所單位： | 醫學檢驗暨生物技術學系 | |
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