臺灣博碩士論文加值系統

一些從柑橘類水果純化出來的多甲氧基黃酮類化合物，例如sinensetin (5,6,7,3',4'-pentamethoxyflavone)，被研究具有抗癌活性，且其毒性不高。本篇研究以3,4,5-trimethoxyphenol為起始物合成了一個sinensetin的結構類似物4'-hydroxy-5,6,7,3'-tetramethoxyflavone，並以洋菜膠細胞群落形成試驗測試此化合物與其前驅物等之抗腫瘤生成活性，結果顯示除了3,4,5-trimethoxyphenol 與1-(6-Hydroxy-2,3,4-trimethoxy-phenyl)-ethanone對胰臟癌細胞PANC-1之腫瘤生成無顯著抑制效果外，其餘數種化合物皆可抑制胰臟癌細胞之腫瘤形成。如果同時以胰臟癌纖維母細胞PC-015Ct共同培養進行胰臟癌細胞PANC-1之洋菜膠細胞群落形成試驗，則會顯著地減低抗癌藥物gemcitabine的抑制腫瘤生成效果，顯示纖維母細胞可增進癌細胞之抗藥性。以此檢視本研究使用或合成出來的化合物之抗腫瘤生成活性，發現這些化合物的抑癌活性並不受纖維母細胞PC-015Ct共同培養的影響。本篇研究顯示數種新合成之多甲氧基黃酮類化合物具有抑制胰臟癌腫瘤生成的活性，然其作用機制尚待進一步研究探討。

Some polymethoxylated flavones, such as sinensetin (5,6,7,3',4'-pentamethoxyflavone), isolated from citrus, have been shown to possess anti-cancer activity and lower toxicity. We synthesized 4'-hydroxy-5,6,7,3'-tetramethoxyflavone, a sinensetin analog, from 3,4,5-trimethoxyphenol. The anti-tumorigenesis activity of compounds used and produced in the process of synthesis reactions was tested by soft agar colony formation assay. The results showed that all the compounds, but not 3,4,5-trimethoxyphenol and 1-(6-Hydroxy-2,3,4-trimethoxy-phenyl)-ethanone, had inhibitory effects on colony formation of pancreatic cancer PANC-1 cells. The anti-tumorigenesis activity of gemcitabine was significantly decreased by fibroblasts, which was revealed by colony formation assay of pancreatic cancer PANC-1 cells with co-culture of pancreatic cancer-associated fibroblast PC-015Ct. The results showed the drug resistance occurred when pancreatic cancer cells grew with surrounding fibroblasts. However, the anti-tumorigenesis activity of tested compounds was not influenced with co-culture of PC-015Ct fibroblasts. In this study, we showed some novel polymethoxyflavone derivatives possess anti-tumorigenesis activity towards pancreatic cancer. The inhibitory mechanism of these compounds will be still elucidated.

中文摘要 II
英文摘要 III
目錄 V
表目錄 VII
縮寫表 X
第一章 緒論 1
第一節 類黃酮 1
第二節 黃酮 3
第三節 川陳皮素 (Nobiletin) 4
第四節 甜橙黃酮 (Sinensetin) 5
第五節 癌症特性 5
第六節 胰臟癌特性 7
第七節 Gemcitabine 10
第八節 微環境 11
第九節 纖維母細胞 13
第十節 研究動機 14
第二章 材料與方法 16
第一節 藥品 16
第二節 置與儀器 18
第三節 合成方法 20
第四節 細胞株 27
第五節 細胞培養液配置 27
第六節 洋菜膠腫瘤群落形成試驗 (Soft agar colony formation assay, SACF): 27
第七節 統計方法 29
第三章 結果 30
第四章 討論 37
第五章 結論 40
第六章 參考文獻 41
表 47
圖 51

1 Yamamoto Y, Gaynor RB. Therapeutic potential of inhibition of the NF-κB pathway in the treatment of inflammation and cancer. The Journal of clinical investigation. 2001;107(2):135-42.
2 Cazarolli LH, Zanatta L, Alberton EH, et al. Flavonoids: prospective drug candidates. Mini reviews in medicinal chemistry. 2008;8(13):1429-40.
3 Ruela de Sousa RR, Queiroz KCS, Souza ACS, et al. Phosphoprotein levels, MAPK activities and NFκB expression are affected by fisetin. Journal of enzyme inhibition and medicinal chemistry. 2007;22(4):439-44.
4 Ravishankar D, Rajora AK, Greco F, Osborn HM. Flavonoids as prospective compounds for anti-cancer therapy. The international journal of biochemistry & cell biology. 2013;45(12):2821-31.
5 Cushnie TT, Lamb AJ. Recent advances in understanding the antibacterial properties of flavonoids. International journal of antimicrobial agents. 2011;38(2):99-107.
6 Manner S, Skogman M, Goeres D, Vuorela P, Fallarero A. Systematic exploration of natural and synthetic flavonoids for the inhibition of Staphylococcus aureus biofilms. International journal of molecular sciences. 2013;14(10):19434-51.
7 Cushnie TT, Lamb AJ. Antimicrobial activity of flavonoids. International journal of antimicrobial agents. 2005;26(5):343-56.
8 Friedman M. Overview of antibacterial, antitoxin, antiviral, and antifungal activities of tea flavonoids and teas. Molecular nutrition & food research. 2007;51(1):116-34.
9 Schuier M, Sies H, Illek B, Fischer H. Cocoa-related flavonoids inhibit CFTR-mediated chloride transport across T84 human colon epithelia. The Journal of nutrition. 2005;135(10):2320-5.
10 Tangney CC, Rasmussen HE. Polyphenols, inflammation, and cardiovascular disease. Current atherosclerosis reports. 2013;15(5):324.
11 van Dam RM, Naidoo N, Landberg R. Dietary flavonoids and the development of type 2 diabetes and cardiovascular diseases: review of recent findings. Current opinion in lipidology. 2013;24(1):25-33.
12 Siasos G, Tousoulis D, Tsigkou V, et al. Flavonoids in atherosclerosis: an overview of their mechanisms of action. Current medicinal chemistry. 2013;20(21):2641-60.
13 柯萬盛. 生物類黃酮（Bioflavonoids）. 2008 [cited; Available from: http://www.ktgh.com.tw/HygieneArticle_look.asp?NewsID=233
14 國民健康/健康檢查/健康知識_有健康網. 生物類黃酮. 2013 [cited; Available from: http://www.uuuwell.com/mytag.php?id=71470
15 Benavente-Garcia O, Castillo J. Update on uses and properties of citrus flavonoids: new findings in anticancer, cardiovascular, and anti-inflammatory activity. Journal of agricultural and food chemistry. 2008;56(15):6185-205.
16 Havsteen BH. The biochemistry and medical significance of the flavonoids. Pharmacology & therapeutics. 2002;96(2):67-202.
17 Galati E, Monforte M, Kirjavainen S, et al. Biological effects of hesperidin, a citrus flavonoid.(Note I): antiinflammatory and analgesic activity. Farmaco (Società chimica italiana: 1989). 1994;40(11):709.
18 Jang S, Kelley KW, Johnson RW. Luteolin reduces IL-6 production in microglia by inhibiting JNK phosphorylation and activation of AP-1. Proceedings of the National Academy of Sciences of the United States of America. May 27.2008;105(21):7534-9.
19 Salakka AK, Jokela TH, Wahala K. Multiple hydride reduction pathways in isoflavonoids. Beilstein journal of organic chemistry. 2006;2:16.
20 Lei W, Xue-su S, Chen-mei F, Bi-ning J. Research Progress on Bio-logical Activities and Applications of Polymethoxyflavonoids from Citrus. Food Science. 2009;30(7):285-90.
21 Li S, Pan M-H, Lo C-Y, et al. Chemistry and health effects of polymethoxyflavones and hydroxylated polymethoxyflavones. Journal of Functional Foods. 2009;1(1):2-12.
22 行政院國家科學委員會. 多甲氧基類黃酮維持基因體穩定性的機制探討.
23 《科學發展》2012年7月，475期，72~ 73頁.
24 Manthey JA, Guthrie N, Grohmann K. Biological properties of citrus flavonoids pertaining to cancer and inflammation. Current medicinal chemistry. 2001;8(2):135-53.
25 Kawaii S, Tomono Y, Katase E, Ogawa K, Masamichi Y. Antiproliferative activity of flavonoids on several cancer cell lines. Bioscience, biotechnology, and biochemistry. 1999;63(5):896-9.
26 Cheng HL, Hsieh MJ, Yang JS, et al. Nobiletin inhibits human osteosarcoma cells metastasis by blocking ERK and JNK-mediated MMPs expression. Oncotarget. Jun 07.2016;7(23):35208-23.
27 Zhang Y, Dou H, Li H, He Z, Wu H. The citrus flavonoid nobiletin inhibits proliferation and induces apoptosis in human pancreatic cancer cells in vitro. Food Science and Biotechnology. 2014;23(1):225-9.
28 Choi S-Y, Hwang J-H, Ko H-C, Park J-G, Kim S-J. Nobiletin from citrus fruit peel inhibits the DNA-binding activity of NF-κB and ROS production in LPS-activated RAW 264.7 cells. Journal of ethnopharmacology. 2007;113(1):149-55.
29 Lin N, Sato T, Takayama Y, et al. Novel anti-inflammatory actions of nobiletin, a citrus polymethoxy flavonoid, on human synovial fibroblasts and mouse macrophages. Biochemical pharmacology. 2003;65(12):2065-71.
30 Yoshigai E, Machida T, Okuyama T, et al. Citrus nobiletin suppresses inducible nitric oxide synthase gene expression in interleukin-1β-treated hepatocytes. Biochemical and biophysical research communications. 2013;439(1):54-9.
31 Wu X, Song M, Zheng J, Guo S, Xiao H. In vitro and in vivo anti-inflammatory effect of 4′-hydroxylnobiletin, a major colonic metabolite of nobiletin. The FASEB Journal. 2013;27(1 Supplement):862.26-.26.
32 Laavola M, Nieminen R, Yam MF, et al. Flavonoids eupatorin and sinensetin present in Orthosiphon stamineus leaves inhibit inflammatory gene expression and STAT1 activation. Planta medica. May.2012;78(8):779-86.
33 Lam IK, Alex D, Wang YH, et al. In vitro and in vivo structure and activity relationship analysis of polymethoxylated flavonoids: identifying sinensetin as a novel antiangiogenesis agent. Molecular nutrition & food research. Jun.2012;56(6):945-56.
34 Akowuah G, Zhari I, Norhayati I, Sadikun A, Khamsah S. Sinensetin, eupatorin, 3′-hydroxy-5, 6, 7, 4′-tetramethoxyflavone and rosmarinic acid contents and antioxidative effect of Orthosiphon stamineus from Malaysia. Food Chemistry. 2004;87(4):559-66.
35 Shin HS, Kang SI, Yoon SA, Ko HC, Kim SJ. Sinensetin attenuates LPS-induced inflammation by regulating the protein level of IkappaB-alpha. Bioscience, biotechnology, and biochemistry. 2012;76(4):847-9.
36 Dong Y, Ji G, Cao A, et al. Effects of sinensetin on proliferation and apoptosis of human gastric cancer AGS cells. Zhongguo Zhong yao za zhi= Zhongguo zhongyao zazhi= China journal of Chinese materia medica. 2011;36(6):790-4.
37 Androutsopoulos VP, Ruparelia K, Arroo RR, Tsatsakis AM, Spandidos DA. CYP1-mediated antiproliferative activity of dietary flavonoids in MDA-MB-468 breast cancer cells. Toxicology. 2009;264(3):162-70.
38 衛生福利部. 104年國人死因統計結果. Aug 08.2016 [cited; Available from: http://www.mohw.gov.tw/news/572256044
39 Institute. NC. Cancer. February 9, 2015 [cited; Available from: https://www.cancer.gov/about-cancer/understanding/what-is-cancer
40 Organization. WH. Cancer February 2015 [cited; Available from: http://www.who.int/mediacentre/factsheets/fs297/en/
41 維基百科. 癌症. [cited; Available from: https://zh.wikipedia.org/wiki/%E7%99%8C%E7%97%87
42 Berk A, Zipursky S, Lodish H. Molecular Cell Biology 4th edition. National Center for Biotechnology InformationÕs Bookshelf; 2000.
43 Perera FP. Environment and cancer: who are susceptible? Science. 1997;278(5340):1068-73.
44 Mitchell R, Popham F. Effect of exposure to natural environment on health inequalities: an observational population study. The Lancet. 2008;372(9650):1655-60.
45 Society. AC. Family Cancer Syndromes. June 25, 2014 [cited; Available from: http://www.cancer.org/cancer/cancer-causes/genetics/family-cancer-syndromes.html
46 JOURNAL. TM. 腫瘤微環境簡介. 2015, Vol.58, No.1.
47 黃燦龍. 胰臟癌. [cited; Available from: http://www.canceraway.org.tw/cancerpageshow.asp?IDno=537
48 台灣癌症防治網. 認識胰臟癌. [cited; Available from: http://cisc.twbbs.org/lib/project/books/book02.php
49 Hwang RF, Moore T, Arumugam T, et al. Cancer-associated stromal fibroblasts promote pancreatic tumor progression. Cancer research. 2008;68(3):918-26.
50 Rucki AA, Zheng L. Pancreatic cancer stroma: understanding biology leads to new therapeutic strategies. World journal of gastroenterology. Mar 7.2014;20(9):2237-46.
51 Wang Z, Li J, Chen X, et al. Disrupting the balance between tumor epithelia and stroma is a possible therapeutic approach for pancreatic cancer. Medical science monitor : international medical journal of experimental and clinical research. 2014;20:2002-6.
52 Lunardi S, Muschel RJ, Brunner TB. The stromal compartments in pancreatic cancer: are there any therapeutic targets? Cancer letters. 2014;343(2):147-55.
53 Bachem MG, Zhou S, Buck K, Schneiderhan W, Siech M. Pancreatic stellate cells--role in pancreas cancer. Langenbeck's archives of surgery / Deutsche Gesellschaft fur Chirurgie. Nov.2008;393(6):891-900.
54 Masamune A, Shimosegawa T. Pancreatic stellate cells: A dynamic player of the intercellular communication in pancreatic cancer. Clinics and research in hepatology and gastroenterology. Sep.2015;39 Suppl 1:S98-103.
55 林口長庚紀念醫院血液腫瘤科陳仁熙. 胰臟癌的化學治療簡介.
56 維基百科. gemcitabine. [cited; Available from: https://en.wikipedia.org/wiki/Gemcitabine
57 光田綜合醫院. Gemcitabine. [cited; Available from: http://www.ktgh.com.tw/Medicament_tbDrug_Look.asp?CatID=12&ModuleType=&NewsID=599&Ordid=30247
58 Plunkett W, Huang P, Xu Y-Z, et al. Gemcitabine: metabolism, mechanisms of action, and self-potentiation. Seminars in oncology; 1995; 1995. p. 3-10.
59 Plunkett W, Huang P, Searcy CE, Gandhi V. Gemcitabine: preclinical pharmacology and mechanisms of action. Seminars in oncology; 1996; 1996. p. 3-15.
60 Hu M, Polyak K. Microenvironmental regulation of cancer development. Current opinion in genetics & development. 2008;18(1):27-34.
61 Hu M, Yao J, Cai L, et al. Distinct epigenetic changes in the stromal cells of breast cancers. Nature genetics. 2005;37(8):899-905.
62 Li H, Fan X, Houghton J. Tumor microenvironment: the role of the tumor stroma in cancer. Journal of cellular biochemistry. 2007;101(4):805-15.
63 Fiegl H, Millinger S, Goebel G, et al. Breast cancer DNA methylation profiles in cancer cells and tumor stroma: association with HER-2/neu status in primary breast cancer. Cancer research. 2006;66(1):29-33.
64 Trimboli AJ, Cantemir-Stone CZ, Li F, et al. Pten in stromal fibroblasts suppresses mammary epithelial tumours. Nature. 2009;461(7267):1084-91.
65 Diaz-Montero CM, Salem ML, Nishimura MI, et al. Increased circulating myeloid-derived suppressor cells correlate with clinical cancer stage, metastatic tumor burden, and doxorubicin–cyclophosphamide chemotherapy. Cancer Immunology, Immunotherapy. 2009;58(1):49-59.
66 Matsumoto K, Nakamura T. Hepatocyte growth factor and the Met system as a mediator of tumor–stromal interactions. International journal of cancer. 2006;119(3):477-83.
67 Nakamura T, Matsumoto K, Kiritoshi A, Tano Y, Nakamura T. Induction of hepatocyte growth factor in fibroblasts by tumor-derived factors affects invasive growth of tumor cells: in vitro analysis of tumor-stromal interactions. Cancer research. 1997;57(15):3305-13.
68 Johnson M, Koukoulis G, Matsumoto K, Nakamura T, Iyer A. Hepatocyte growth factor induces proliferation and morphogenesis in nonparenchymal epithelial liver cells. Hepatology. 1993;17(6):1052-61.
69 Bottaro DP, Rubin JS. Identification of the Hepatocyte Growth Factor Receptor as the c-met Porto-Oncogene Product. Science. 1991;251(4995):802.
70 Importance of the B Ring and Its Substitution on the -Glucosidase
Inhibitory Activity of Baicalein, 5,6,7-Trihydroxyflavon.
71 Tyan S-W, Kuo W-H, Huang C-K, et al. Breast cancer cells induce cancer-associated fibroblasts to secrete hepatocyte growth factor to enhance breast tumorigenesis. PloS one. 2011;6(1):e15313.