臺灣博碩士論文加值系統

研究分析2002-2010年期間三軍總醫院520位肺腺癌手術病患臨床資料及預後追蹤，94位病患於追蹤期間復發，復發危險因子包括腫瘤分化程度、腫瘤位置、淋巴血管侵犯與否、手術方式、淋巴腺侵犯與否、腫瘤分期、腫瘤大小、及正子攝影氟18標記葡萄糖攝取最大值，多變項回歸分析顯示腫瘤大小(大於2公分)、腫瘤分化程度(較差者)為復發危險因子，其風險分別為3.371及2.937倍。
在無復發病患組，病患5年存活率97.3%；復發組5年存活率為38.4%，腫瘤分化程度與正子攝影氟18標記葡萄糖攝取最大值(SUVmax)有高度相關，分化程度良好腫瘤其氟18標記葡萄糖攝取值較低(2.66 ± 2.64)；分化程度中等腫瘤其攝取值為4.75 ± 3.64；分化程度較差腫瘤氟18標記葡萄糖值較高(7.59 ± 5.25)，p < 0.001，進一步分析，腫瘤分化程度與存活率呈顯著相關，腫瘤分化良好病患有較佳5年存活率(86.8%)及五年無病存活率(95.1%)；腫瘤分化較差病患呈現較差5年存活率(58.6%)及五年無病存活率(50.4%)，達統計學顯著意義(p < 0.001)。
分析520病患檢體其第一型甲狀腺轉錄因子表現程度，依病理組織免疫化學染色區分為三個等級，第一型甲狀腺轉錄因子表現較強(score 3)腫瘤之病患有較佳五年存活率 (86.1%)；第一型甲狀腺轉錄因子表現較弱(score 1)腫瘤之病患五年存活率較差(60.0%)，達統計學顯著意義(p = 0.001)，腫瘤分化與第一型甲狀腺轉錄因子表現有相關，分化程度良好腫瘤其第一型甲狀腺轉錄因子在免疫化學染色表現較強，分化程度較差之腫瘤其第一型甲狀腺轉錄因子在免疫化學染色表現較弱，卡方檢定達統計學顯著意義(p < 0.001)。
利用肺癌細胞株研究第一型甲狀腺轉錄因子與腫瘤分化及侵襲能力關聯性，首先檢測常見肺癌細胞株(CL1-0、CL1-5、H1299、A549、HCC827、及HCC827GR)，分析細胞株第一型甲狀腺轉錄因子表現、侵襲能力，CL1-0細胞株第一型甲狀腺轉錄因子蛋白質、核糖核酸(RNA)表現均較CL1-5細胞株高，CL1-5為惡性度較高之細胞株，其上皮鈣粘蛋白(E-cadherin)表現量低，波形蛋白(Vimentin)、高移動蛋白(High Mobility group protein A2,HMGA2)、表皮生長因子(Epithelial Growth Factor Receptor, EGFR)表現量較高，近一步利用外加或去除第一型甲狀腺轉錄因子表現來驗證肺癌細胞株侵襲性表是否受影響，針對第一型甲狀腺轉錄因子表現高之細胞株(CL1-0、HCC827)，以小片段核糖核酸(short hairpin RNA, shRNA)處理後，CL1-0、HCC827細胞株第一型甲狀腺轉錄因子蛋白表現量下降，HMGA2、EGFR、間葉組織標誌蛋白(Vimentin)表現增加，上皮組織標誌蛋白表現量下降，細胞侵襲能力增加；針對第一型甲狀腺轉錄因子表現低之細胞株(CL1-5、HCC827GR)，外加第一型甲狀腺轉錄因子載體處理後，CL1-5、HCC827GR細胞株第一型甲狀腺轉錄因子蛋白表現量增加，HMGA2、EGFR、間葉組織標誌蛋白(Vimentin)表現下降；上皮組織標誌蛋白(E-cadherin)表現增加，細胞侵襲能力下降。
免疫沉澱分析發現第一型甲狀腺轉錄因子可能藉由與HMGA2啟動子(promotor)結合而進行調控，以shRNA處理CL1-0、HCC827細胞株，TTF-1與HMGA2免疫沉澱反應減弱，第一型甲狀腺轉錄因子表現高之細胞株(CL1-0、HCC827)以小片段核糖核酸處理後，miR33a蛋白表現減少；第一型甲狀腺轉錄因子表現高之細胞株(CL1-5、HCC827GR)以cDNA載體處理後，miR33a蛋白表現增加。
此臨床觀察、基礎細胞轉譯醫學研究顯示TTF-1/HMGA2調控在肺腺癌細胞分化、腫瘤復發扮演重要角色，影響肺腺癌病人預後

A total of 520 patients with clinical early stage lung adenocarcinoma who underwent surgical resection were reviewed retrospectively. Clinical data and outcomes were evaluated with an average follow-up of 117 months. The results were validated via lung cancer cell line studies. The clinical parameters did not differ between relapse and nonrelapse patients. Exceptions were tumor differentiation, lymphovascular space invasion, F18-fluorodeoxyglucose maximum standard uptake value, tumor size, and pathological stage (p < 0.001). Poor tumor differentiation was the independent prognostic factor (odds ratio: 2.937, p =0.026). The expression of TTF-1 was correlated with tumor differentiation in resected lung adenocarcinoma patients (p < 0.001). Five-year survival was 60.0% for score 1 TTF-1 expression patients, 80.1% for score 2 TTF-1 expression patients, and 86.1% for score 3 TTF-1 expression group patients. The lung cancer cell line study of knockdown and overexpression of TTF-1 revealed TTF-1 mediated High Mobility Group AT-Hook 2 (HMGA2) protein involved with epithelium-mesenchymal transformation. The chromatin immunoprecipitation revealed TTF-1 regulated HMGA2 via direct binding. TTF-1/HMGA2 axis was associated with tumor differentiation and mediated the aggressiveness of the tumor and prognosis.

壹、 中文摘要 p.1-4
貳、 緒論
第一節 研究動機與背景 p.5-11
第二節 問題及研究假說 p.12-13
參、 研究方法與材料 p.14-27
肆、 結果 p.28-32
伍、 討論 p.33-35
陸、 結論與未來研究發展 p.36
柒、 參考文獻 p.37-45
捌、 圖表 p.46-73
玖、 英文論文摘要 p.73-106

1.Yoshida, B.A., et al., Metastasis-suppressor genes: a review and perspective on an emerging field. JNatl Cancer Inst, 2000. 92(21): p. 1717-30.
2.Woodhouse, E.C., R.F. Chuaqui, and L.A. Liotta, General mechanisms of metastasis. Cancer, 1997.80(8 Suppl): p. 1529-37.
3.Kohn, E.C., Development and prevention of metastasis. Anticancer Res, 1993. 13(6B): p. 2553-9.
4.Duffy, M.J., Proteases as prognostic markers in cancer. Clin Cancer Res, 1996. 2(4): p. 613-8.
5.Ahmad, A., et al., Modulation of human stromelysin 3 promoter activity and gene expression byhuman breast cancer cells. Int J Cancer, 1997. 73(2): p. 290-6.
6.Folkman, J., How is blood vessel growth regulated in normal and neoplastic tissue? G.H.A. Clowes memorial Award lecture. Cancer Res, 1986. 46(2): p. 467-73.
7.Hanna, N., Role of natural killer cells in control of cancer metastasis. Cancer Metastasis Rev, 1982.1(1): p. 45-64.
8.Key, M.E., Macrophages in cancer metastases and their relevance to metastatic growth. Cancer Metastasis Rev, 1983. 2(1): p. 75-88.
9.Morris, V.L., et al., Sequential steps in hematogenous metastasis of cancer cells studied by in vivo videomicroscopy. Invasion Metastasis, 1997. 17(6): p. 281-96.
10.Bracke, M.E., F.M. Van Roy, and M.M. Mareel, The E-cadherin/catenin complex in invasion andmetastasis. Curr Top Microbiol Immunol, 1996. 213 (Pt 1): p. 123-61.
11.Okada, T., H. Okuno, and Y. Mitsui, A novel in vitro assay system for transendothelial tumor cell invasion: significance of E-selectin and alpha 3 integrin in the transendothelial invasion by HT1080 fibrosarcoma cells. Clin Exp Metastasis, 1994. 12(4): p. 305-14.
12.Morini, M., et al., The alpha 3 beta 1 integrin is associated with mammary carcinoma cell metastasis, invasion, and gelatinase B (MMP-9) activity. Int J Cancer, 2000. 87(3): p. 336-42.
13.Friedrichs, K., et al., High expression level of alpha 6 integrin in human breast carcinoma is correlated with reduced survival. Cancer Res, 1995. 55(4): p. 901-6.
14.Hendrix, M.J., et al., A simple quantitative assay for studying the invasive potential of high and low human metastatic variants. Cancer Lett, 1987. 38(1-2): p. 137-47.
15.Kramer, R.H., K.G. Bensch, and J. Wong, Invasion of reconstituted basement membrane matrix by metastatic human tumor cells. Cancer Res, 1986. 46(4 Pt 2): p. 1980-9.22.
16.Griese M. Pulmonary surfactant in health and human lung diseases: state of the art. Eur Respir J. 1999, 13 (6): 1455- 1476.
17.Gortner L, Hilgendorff A. Surfactant-associated proteins B and C: molecular biology and physiologic properties. Z Geburtshilfe Neonatol, 2004, 208(3): 91- 97.
18.Ferzli G, Yunis KA, Mroueh S. Surfactant protein B deficiency: a rare cause of respiratory failure in a Lebanese newborn [J]. Ann Saudi Med, 2006, 26(1): 69-70.
19.Ordonez, N. G. (2000) Thyroid transcription factor-1 is a marker of lung and thyroid carcinomas. Adv. Anat. Pathol. 7, 123–127
20.Zamecnik, J. and Kodet, R. (2002) Value of thyroid transcription factor-1 and surfactant apoprotein A in the differential diagnosis of pulmonary carcinomas: a study of 109 cases. Virchows Arch. 440, 353–361
21.Moldvay, J., Jackel, M., Bogos, K. et al. (2004) The role of TTF-1 in differentiating primary and metastatic lung adenocarcinomas. Pathol. Oncol. Res. 10, 85–88
22.Tan, D., Li, Q., Deeb, G., Ramnath, N. et al. (2003) Thyroid transcription factor-1 expression prevalence and its clinical implications in non-small cell lung cancer: a high-throughput tissue microarray and immunohistochemistry study. Hum. Pathol. 34, 597–604
23.Myong, N. H. (2003) Thyroid transcription factor-1 (TTF-1) expression in human lung carcinomas: its prognostic implication and relationship with expressions of p53 and Ki-67 proteins. J. Korean Med. Sci. 18, 494–500
24.Puglisi, F., Barbone, F., Damante, G. et al. (1999)　Prognostic value of thyroid transcription factor-1 in primary, resected, non-small cell lung carcinoma.Mod. Pathol. 12, 318–324
25.Haque, A. K., Syed, S., Lele, S. M., Freeman, D. H. and Adegboyega, P. A. (2002) Immunohistochemical study of thyroid transcription factor-1 and HER2/neu in non-small cell lung cancer: strong thyroid transcription factor-1 expression predicts better survival. Appl. Immunohistochem. Mol. Morphol. 10, 103–109
26.Saad, R. S., Liu, Y. L., Han, H., Landreneau, R. J. and Silverman, J. F. (2004) Prognostic significance of thyroid transcription factor-1 expression in both early-stage conventional adenocarcinoma and bronchioloalveolar carcinoma of the lung. Hum. Pathol. 35, 3–7
27.Weir BA, Woo MS, Getz G, et al. Characterizing the cancer genome in lung adenocarcinoma. Nature 2007; 450: 893–898.
28.Lockwood WW, Chari R, Coe BP, et al. DNA amplification is a ubiquitous mechanism of oncogene activation in lung and other cancers. Oncogene 2008; 27: 4615–4624.
29.Tanaka H, Yanagisawa K, Shinjo K, et al. Lineage-specific dependency of lung adenocarcinomas on the lung developmentregulator TTF-1. Cancer Res 2007; 67: 6007–6011
30.Yatabe Y, Kosaka T, Takahashi T, et al. EGFR mutation is specific for terminal respiratory unit type adenocarcinoma. Am J Surg Pathol 2005; 29: 633–639.
31.Morishita A, Zaidi MR, Mitoro A, Sankarasharma D, Szabolcs M, Okada Y, D'Armiento J, Chada K. HMGA2 is a driver of tumor metastasis. Cancer Res 2013; 73:4289-4299
32.Sarhadi VK, Wikman H, Salmenkivi K, Kuosma E, Sioris T, Salo J, Karjalainen A, Knuutila S, Anttila S. Increased expression of high mobility group A proteins in lung cancer. J Pathol. 2006 Jun;209(2):206-12.
33.Thuault S, Valcourt U, Petersen M, Manfioletti G, Heldin CH, Moustakas A. Transforming growth factor-beta employs HMGA2 to elicit epithelial-mesenchymal transition. J Cell Biol. 2006 Jul 17;174(2):175-83.
34.Winslow MM, Dayton TL, Verhaak RG, et al. Suppression of lung adenocarcinoma progression by　Nkx2-1. Nature. 473, 101–104 (2011).
35.Kondo T, Nakazawa T, Ma D, Niu D, Mochizuki K, Kawasaki T, Nakamura N, Yamane T, Kobayashi M, Katoh R. Epigenetic silencing of TTF-1/NKX2-1 through DNA hypermethylation and histone H3 modulation in thyroid carcinomas. Lab Invest. 2009 Jul;89(7):791-9.
36.Feng PH, Yu CT, Wu CY, Lee MJ, Lee WH, Wang LS, Lin SM, Fu JF, Lee KY, Yen TH. Tumor-associated macrophages in stage IIIA pN2 non-small cell lung cancer after neoadjuvant chemotherapy and surgery. Am J Transl Res 2014;6(5):593-603
37.Chao-Ying Liu, Juan-Ying Xu, Xiao-Yan Shi, Wei Huang, Ting-Yan Ruan, Ping Xie and Jun-Li Ding. M2-polarized tumor-associated macrophages promoted epithelial–mesenchymal transition in pancreatic cancer cells, partially through TLR4/IL-10 signaling pathway. Laboratory Investigation (2013) 93, 844–854
38.Fan QM, Jing YY, Yu GF, Kou XR, Ye F, Gao L, Li R, Zhao QD, Yang Y, Lu ZH, Wei LX. Tumor-associated macrophages promote cancer stem cell-like properties via transforming growth factor-beta1-induced epithelial-mesenchymal transition in hepatocellular carcinoma. Cancer Lett. 2014 Oct 1; 352(2):160-8.
39.Zeng TB, He HJ, Han ZB, Zhang FW, Huang ZJ, Liu Q, Cui W, Wu Q. DNA methylation dynamics of a maternally methylated DMR in the mouse Dlk1-Dio3 domain.FEBS Lett. 2014 Dec 20;588
40.Bonde AK1, Tischler V, Kumar S, Soltermann A, Schwendener RA. Intratumoral macrophages contribute to epithelial-mesenchymal transition in solid tumors. BMC Cancer. 2012 Jan 24;12:35
41.Yatabe Y, Hida T, Horio Y, Kosaka T, Takahashi T, Mitsudomi T. A rapid, sensitive assay to detect EGFR mutation in small biopsy specimens from lung cancer. J Mol Diagn. 2006 Jul;8(3):335-41.
42.Chen YY, Huang TW, Tsai WC, Lin LF, Cheng JB, Chang H, Lee SC. Risk factors of postoperative recurrence/metastasis for patients of clinical stage-I non-small-cell lung cancer. World Journal of Surgical Oncology 2014 Jan 10; 12:10
43.Chen YY, Huang TW, Tsai WC, Lin LF, Cheng JB, Lee SC, Chang H. Lymphovascular space invasion and tumor differentiation are predictors for postoperative recurrence in patients with pathological stage I nonsmall cell lung cancer. J Chin Med Assoc. 2014 Aug;77(8):416-21
44.American Joint Committee on Cancer. AJCC Cancer Staging Manual. 7th ed. New York: Springer. (2010).
45.Ikeda, S. et al. Immunostaining for thyroid transcription factor 1, Napsin A, p40, and cytokeratin 5 aids in differential diagnosis of non-small cell lung carcinoma. Oncol Lett. 2015; 9:2099–2104.
46.Yamaguchi T, Yanagisawa K, Sugiyama R, Hosono Y, Shimada Y, Arima C, Kato S, Tomida S, Suzuki M, Osada H, Takahashi T. NKX2-1/TITF1/TTF-1-Induced ROR1 is required to sustain EGFR survival signaling in lung adenocarcinoma. Cancer Cell. 2012 Mar 20;21(3):348-61