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研究生:魏于傑
研究生(外文):Yu-Chieh Wei
論文名稱:探討新合成1-benzyl indoles 衍生物 21-900 誘導人類血癌細胞凋亡之體外及體內的作用機轉
論文名稱(外文):21-900, a novel synthetic 1-benzyl indoles derivative, induces cell apoptosis in human leukemia in vitro and in vivo
指導教授:蕭哲志蕭哲志引用關係
指導教授(外文):George Hsiao
口試委員:皇甫維君劉景平黃聰龍
口試委員(外文):Wei-Chun HuangFuJing-Ping LiouTsong-Long Hwang
口試日期:2015-06-23
學位類別:碩士
校院名稱:臺北醫學大學
系所名稱:醫學科學研究所
學門:醫藥衛生學門
學類:醫學學類
論文種類:學術論文
論文出版年:2015
畢業學年度:103
語文別:中文
論文頁數:96
中文關鍵詞:去乙醯化酶微管細胞凋亡白血病癌症
外文關鍵詞:HDACmicrotubuleapoptosisleukemiacancer
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惡性腫瘤為造成國人死亡的主要原因之一。根據衛生福利部國民健康署統計癌症為十大死因之首有三十二年之久,當癌症無法以外科手術根治時,化學療法和標靶藥物治療即變成重要的治療方法。本研究的目的是探討新合成的雙重標靶pan-HDAC與Tubulin抑制劑21-900小分子藥物對於人類血癌細胞株之抗癌活性,並進一步探討in vitro以及in vivo 的藥理機轉。本實驗一開始先將這系列1-benzyl indoles藥物利用了SRB assay 及MTT assay 作用在人類多種癌細胞株中,結果可以看到 21-900 在多株癌細胞中抑制的效果最好,其中在血癌 (HL-60) 有更好的敏感度。接著利用MTT assay (3-[4,5-dimethythiazol-2-yl]-2,5-diphenyltetrazolium bromide) 的方式來評估21-900對於HL-60以及MOLT-4細胞毒殺的影響,發現在給予21-900 48小時後具有濃度相關性抑制細胞生長作用,其 IC50 分別為133 ± 4 nM及262 ± 14 nM。21-900亦具有抑制 HDACs 活性,其 IC50 為 610 nM。進一步利用 HDAC isoenzyme inhibition assay發現21-900抑制 HDAC6的活性 (IC50 = 64.5 nM) 最高,其對 HDAC1 (IC50 = 573.4 nM) , HDAC2 (IC50 = 1408.1 nM), 和HDAC8 (IC50 = 905.9 nM) 高出10-20倍的抑制能力。接著利用流式細胞儀分析發現,21-900使血癌細胞停滯在G2/M期,並且伴隨著濃度增加促使明顯的Sub G1增加。接著透過細胞外微管聚合實驗及細胞免疫螢光染色,發現21-900的作用類似於vincristine 也會抑制微管聚合,並且使M期指標蛋白MPM2磷酸化增加。另外21-900也會去影響Bcl-2 family 蛋白改變,例如:Bcl-2、Bid及Mcl-1減少,因此影響了粒線體膜通透度改變促使內生性凋亡路徑活化,誘發caspase-3、-7以及PARP的活化,促使呈現濃度相依性之細胞凋亡。接著使用了JNK的抑制劑SP600125發現可以反轉21-900所造成的細胞凋亡現象。最後在小鼠的人類異位移植腫瘤動物模式中證實,21-900可以抑制腫瘤生長且不影響小鼠的體重,腫瘤組織中證實,21-900會活化caspase-3與增加PARP cleavage、增加乙醯化的histone 3和??-tubulin的表現、也增加M期指標蛋白MPM2表現,這些作用的效果與我們在in vitro的結果相符合。結果顯示,21-900在人類白血病細胞株中,具有抑制其HDACs活性且影響了微管的動態平衡造成細胞週期停滯,且能夠使細胞走向凋亡。至今仍尚未有雙重抑制劑在臨床使用,期許21-900能成為治療血癌之候選藥物。
Malignant tumor has been the major cause of death in Taiwan. According to Health Promotion Administration, Ministry of Health and Welfare in Taiwan, cancer has been the first place of ten leading deaths for thirty-two years. When tumors are difficult to be removed by surgery simply, chemotherapies and targeted therapies become important treatments. In order to cure cancer, drug development is an important thing in cancer therapy. The purpose of this study is to evaluate the anti-tumor effects of a novel synthetic HDAC and tubulin dual inhibitor, 21-900, in human leukemia cells, and to further study the mechanism of action in vitro and in vivo. We first determined a series of 1-benzyl indoles as potent anti-cancer drugs which are more cytotoxic to human leukemia HL-60 cells. Among these compounds, 21-900 significant exhibits a cytotoxic effect in human leukemia cancer cell HL-60 and MOLT-4 with an IC50 of 133 ± 4 nM and 262 ± 14 nM, respectively, using the 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide (MTT) assay. 21-900 also exhibited as a potent HDAC inhibitor with IC50 value of 610 nM. In the HDAC isoenzyme inhibition assay, we found that 21-900 was a pan-HDAC inhibitor but 21-900 displayed selectivity toward HDAC6 (IC50 = 64.5 nM) over HDAC1 (IC50 = 573.4 nM), HDAC2 (IC50 = 1408.1 nM), and HDAC8 (IC50 = 905.9 nM) with 10-20-fold ratio. Cell cycle distribution was determined by flow cytometry, we demonstrated that 21-900 induced arrest of the cell cycle at G2/M phase in HL-60 and MOLT-4 cell line, with the increase of cell distribution at subG1 phase. Then tubulin polymerization assay and immunofluorescence stain suggested that 21-900 was similar to vincristine in causing depolymerization of microtubule. These results showed that 21-900 could affect microtubule dynamic and accompanied by the up-regulation of mitotic marker,MPM2.Further, 21-900 could influenced the Bcl-2 family proteins by decreasing Mcl-1, Bcl-2 and Bid and increasing p-Bcl-2. Moreover, 21-900 induced cell apoptosis in a concentration-dependent manner; it also increased the cleavage of caspase-3 and -7 and poly (ADP-ribose) polymerase (PARP) in HL-60 and MOLT-4 cells Treatment with SP600125 (JNK inhibitor) could reverse 21-900-induced apoptosis.Then 21-900 significantly inhibit HL-60 and MOLT-4 xenograft tumor growth without the loss of body weight in the SCID mice. 21-900 activated caspase-3 and increased the expression of cleavage- PARP, ac-tubulin, ac-H3 and MPM2 in vivo. These data correlated with in vitro results. All these findings suggest that 21-900 could be served as a new candidate in leukemia cancer treatment.
目錄
碩士學位考試委員審書................I
學位考試保密同意書暨簽到表.........II
電子暨紙本論文同意公開申請書.......IV
致謝................................V
縮寫表............................VII
中文摘要.........................VIII
英文摘要............................X
壹、研究動機與目的..................1
貳、文獻回顧........................3
參、材料與方法
第一節 實驗材料....................36
第二節 實驗方法....................38
肆、實驗結果.......................46
伍、討論...........................52
陸、結論與展望.....................58
柒、參考文獻.......................81
Annesley CE, Brown P (2015). Novel agents for the treatment of childhood acute leukemia. Therapeutic advances in hematology 6(2): 61-79.

Barneda-Zahonero B, Parra M (2012). Histone deacetylases and cancer. Molecular oncology 6(6): 579-589.

Barr AR, Gergely F (2007). Aurora-A: the maker and breaker of spindle poles. Journal of cell science 120(Pt 17): 2987-2996.

Bender T, Martinou JC (2013). Where killers meet--permeabilization of the outer mitochondrial membrane during apoptosis. Cold Spring Harbor perspectives in biology 5(1): a011106.

Bennett JM, Catovsky D, Daniel MT, Flandrin G, Galton DA, Gralnick HR, et al. (1976). Proposals for the classification of the acute leukaemias. French-American-British (FAB) co-operative group. British journal of haematology 33(4): 451-458.

Blander G, Guarente L (2004). The Sir2 family of protein deacetylases. Annual review of biochemistry 73: 417-435.

Bose P, Dai Y, Grant S (2014). Histone deacetylase inhibitor (HDACI) mechanisms of action: emerging insights. Pharmacology & therapeutics 143(3): 323-336.

Bradbury CA, Khanim FL, Hayden R, Bunce CM, White DA, Drayson MT, et al. (2005). Histone deacetylases in acute myeloid leukaemia show a distinctive pattern of expression that changes selectively in response to deacetylase inhibitors. Leukemia 19(10): 1751-1759.

Brunelle JK, Letai A (2009). Control of mitochondrial apoptosis by the Bcl-2 family. Journal of cell science 122(4): 437-441.

Chauhan D, Li G, Hideshima T, Podar K, Mitsiades C, Mitsiades N, et al. (2003). JNK-dependent release of mitochondrial protein, Smac, during apoptosis in multiple myeloma (MM) cells. The Journal of biological chemistry 278(20): 17593-17596.

Chen X, Wong P, Radany E, Wong JYC (2009). HDAC Inhibitor, Valproic Acid, Induces p53-Dependent Radiosensitization of Colon Cancer Cells. Cancer Biotherapy & Radiopharmaceuticals 24(6): 689-699.

Cornelissen JJ, Gratwohl A, Schlenk RF, Sierra J, Bornhauser M, Juliusson G, et al. (2012). The European LeukemiaNet AML Working Party consensus statement on allogeneic HSCT for patients with AML in remission: an integrated-risk adapted approach. Nature reviews. Clinical oncology 9(10): 579-590.

De Giorgi F, Lartigue L, Bauer MK, Schubert A, Grimm S, Hanson GT, et al. (2002). The permeability transition pore signals apoptosis by directing Bax translocation and multimerization. FASEB journal : official publication of the Federation of American Societies for Experimental Biology 16(6): 607-609.

Deng Y, Ren X, Yang L, Lin Y, Wu X (2003). A JNK-dependent pathway is required for TNFalpha-induced apoptosis. Cell 115(1): 61-70.

Dhanasekaran DN, Reddy EP (2008). JNK signaling in apoptosis. Oncogene 27(48): 6245-6251.

Dokmanovic M, Clarke C, Marks PA (2007). Histone deacetylase inhibitors: overview and perspectives. Molecular cancer research : MCR 5(10): 981-989.

Drummond DC, Noble CO, Kirpotin DB, Guo Z, Scott GK, Benz CC (2005). Clinical development of histone deacetylase inhibitors as anticancer agents. Annual review of pharmacology and toxicology 45: 495-528.

Duvic M, Vu J (2007). Update on the treatment of cutaneous T-cell lymphoma (CTCL): Focus on vorinostat. Biologics : targets & therapy 1(4): 377-392.

Elmore S (2007). Apoptosis: A Review of Programmed Cell Death. Toxicologic pathology 35(4): 495-516.

Franklin JL (2011). Redox regulation of the intrinsic pathway in neuronal apoptosis. Antioxidants & redox signaling 14(8): 1437-1448.

Fulda S, Debatin KM (2006). Extrinsic versus intrinsic apoptosis pathways in anticancer chemotherapy. Oncogene 25(34): 4798-4811.

Fuster JJ, Fernandez P, Gonzalez-Navarro H, Silvestre C, Nabah YN, Andres V (2010). Control of cell proliferation in atherosclerosis: insights from animal models and human studies. Cardiovascular research 86(2): 254-264.

Glozak MA, Seto E (2007). Histone deacetylases and cancer. Oncogene 26(37): 5420-5432.

Grisold W, Cavaletti G, Windebank AJ (2012). Peripheral neuropathies from chemotherapeutics and targeted agents: diagnosis, treatment, and prevention. Neuro-oncology 14 Suppl 4: iv45-54.

Groninger E, Meeuwsen-De Boer GJ, De Graaf SS, Kamps WA, De Bont ES (2002). Vincristine induced apoptosis in acute lymphoblastic leukaemia cells: a mitochondrial controlled pathway regulated by reactive oxygen species? International journal of oncology 21(6): 1339-1345.

Gross A, McDonnell JM, Korsmeyer SJ (1999). BCL-2 family members and the mitochondria in apoptosis. Genes & development 13(15): 1899-1911.

Harris MH, Thompson CB (2000). The role of the Bcl-2 family in the regulation of outer mitochondrial membrane permeability. Cell death and differentiation 7(12): 1182-1191.

Herman JG, Civin CI, Issa JP, Collector MI, Sharkis SJ, Baylin SB (1997). Distinct patterns of inactivation of p15INK4B and p16INK4A characterize the major types of hematological malignancies. Cancer research 57(5): 837-841.

Ishii S, Kurasawa Y, Wong J, Yu-Lee L-y (2008). Histone deacetylase 3 localizes to the mitotic spindle and is required for kinetochore–microtubule attachment. Proceedings of the National Academy of Sciences 105(11): 4179-4184.

Ishikawa Y, Kusaka E, Enokido Y, Ikeuchi T, Hatanaka H (2003). Regulation of Bax translocation through phosphorylation at Ser-70 of Bcl-2 by MAP kinase in NO-induced neuronal apoptosis. Molecular and cellular neurosciences 24(2): 451-459.

Izawa D, Pines J (2015). The mitotic checkpoint complex binds a second CDC20 to inhibit active APC/C. Nature 517(7536): 631-634.

Johnson NL, Gardner AM, Diener KM, Lange-Carter CA, Gleavy J, Jarpe MB, et al. (1996). Signal transduction pathways regulated by mitogen-activated/extracellular response kinase kinase kinase induce cell death. The Journal of biological chemistry 271(6): 3229-3237.

Johnstone RW (2002). Histone-deacetylase inhibitors: novel drugs for the treatment of cancer. Nature reviews. Drug discovery 1(4): 287-299.

Jones EV, Dickman MJ, Whitmarsh AJ (2007). Regulation of p73-mediated apoptosis by c-Jun N-terminal kinase. The Biochemical journal 405(3): 617-623.

Jordan MA, Wilson L (2004). Microtubules as a target for anticancer drugs. Nature reviews. Cancer 4(4): 253-265.

Kazantsev AG, Thompson LM (2008). Therapeutic application of histone deacetylase inhibitors for central nervous system disorders. Nature reviews. Drug discovery 7(10): 854-868.

Kim HJ, Bae SC (2011). Histone deacetylase inhibitors: molecular mechanisms of action and clinical trials as anti-cancer drugs. Am J Transl Res 3(2): 166-179.

Kramer OH, Mahboobi S, Sellmer A (2014). Drugging the HDAC6-HSP90 interplay in malignant cells. Trends in pharmacological sciences 35(10): 501-509.

Kramer OH, Muller S, Buchwald M, Reichardt S, Heinzel T (2008). Mechanism for ubiquitylation of the leukemia fusion proteins AML1-ETO and PML-RARalpha. FASEB journal : official publication of the Federation of American Societies for Experimental Biology 22(5): 1369-1379.

Kurosaka K, Takahashi M, Watanabe N, Kobayashi Y (2003). Silent cleanup of very early apoptotic cells by macrophages. Journal of immunology (Baltimore, Md. : 1950) 171(9): 4672-4679.

Lucas DM, Alinari L, West DA, Davis ME, Edwards RB, Johnson AJ, et al. (2010). The novel deacetylase inhibitor AR-42 demonstrates pre-clinical activity in B-cell malignancies in vitro and in vivo. PloS one 5(6): e10941.

Macurek L, Lindqvist A, Medema RH (2009). Aurora-A and hBora join the game of Polo. Cancer research 69(11): 4555-4558.

Marks P, Rifkind RA, Richon VM, Breslow R, Miller T, Kelly WK (2001). Histone deacetylases and cancer: causes and therapies. Nature reviews. Cancer 1(3): 194-202.

McDevitt MA (2012). Clinical applications of epigenetic markers and epigenetic profiling in myeloid malignancies. Seminars in oncology 39(1): 109-122.

McGrogan BT, Gilmartin B, Carney DN, McCann A (2008). Taxanes, microtubules and chemoresistant breast cancer. Biochimica et biophysica acta 1785(2): 96-132.

Minucci S, Pelicci PG (2006). Histone deacetylase inhibitors and the promise of epigenetic (and more) treatments for cancer. Nature reviews. Cancer 6(1): 38-51.

Mitsiades N, Mitsiades CS, Richardson PG, McMullan C, Poulaki V, Fanourakis G, et al. (2003). Molecular sequelae of histone deacetylase inhibition in human malignant B cells. Blood 101(10): 4055-4062.

Mollinedo F, Gajate C (2003). Microtubules, microtubule-interfering agents and apoptosis. Apoptosis : an international journal on programmed cell death 8(5): 413-450.

Montalban-Bravo G, Garcia-Manero G (2015). Novel drugs for older patients with acute myeloid leukemia. Leukemia 29(4): 760-769.

Musacchio A (2011). Spindle assembly checkpoint: the third decade. Philosophical transactions of the Royal Society of London. Series B, Biological sciences 366(1584): 3595-3604.

Oakley BR (2000). An abundance of tubulins. Trends in cell biology 10(12): 537-542.

Orr GA, Verdier-Pinard P, McDaid H, Horwitz SB (2003). Mechanisms of Taxol resistance related to microtubules. Oncogene 22(47): 7280-7295.

Pathak P, Hess R, Weiss MA (2014). Liposomal vincristine for relapsed or refractory Ph-negative acute lymphoblastic leukemia: a review of literature. Therapeutic advances in hematology 5(1): 18-24.

Petronczki M, Lénárt P, Peters J-M (2008). Polo on the Rise—from Mitotic Entry to Cytokinesis with Plk1. Developmental Cell 14(5): 646-659.

Piekarz RL, Frye AR, Wright JJ, Steinberg SM, Liewehr DJ, Rosing DR, et al. (2006). Cardiac studies in patients treated with depsipeptide, FK228, in a phase II trial for T-cell lymphoma. Clinical cancer research : an official journal of the American Association for Cancer Research 12(12): 3762-3773.

Pines J (2011). Cubism and the cell cycle: the many faces of the APC/C. Nature reviews. Molecular cell biology 12(7): 427-438.

Pradelli LA, Beneteau M, Ricci JE (2010). Mitochondrial control of caspase-dependent and -independent cell death. Cellular and molecular life sciences : CMLS 67(10): 1589-1597.

Prince HM, Dickinson M (2012). Romidepsin for cutaneous T-cell lymphoma. Clinical cancer research : an official journal of the American Association for Cancer Research 18(13): 3509-3515.

Quintas-Cardama A, Santos FP, Garcia-Manero G (2011). Histone deacetylase inhibitors for the treatment of myelodysplastic syndrome and acute myeloid leukemia. Leukemia 25(2): 226-235.

Sayers TJ (2011). Targeting the extrinsic apoptosis signaling pathway for cancer therapy. Cancer immunology, immunotherapy : CII 60(8): 1173-1180.

Seedhouse CH, Das-Gupta EP, Russell NH (2003). Methylation of the hMLH1 promoter and its association with microsatellite instability in acute myeloid leukemia. Leukemia 17(1): 83-88.

Seki A, Coppinger JA, Jang CY, Yates JR, Fang G (2008). Bora and the kinase Aurora a cooperatively activate the kinase Plk1 and control mitotic entry. Science (New York, N.Y.) 320(5883): 1655-1658.

Shamas-Din A, Brahmbhatt H, Leber B, Andrews DW (2011). BH3-only proteins: Orchestrators of apoptosis. Biochimica et biophysica acta 1813(4): 508-520.

Sharp DJ, Rogers GC, Scholey JM (2000). Microtubule motors in mitosis. Nature 407(6800): 41-47.

Siegel R, Ma J, Zou Z, Jemal A (2014). Cancer statistics, 2014. CA: a cancer journal for clinicians 64(1): 9-29.

Suto RK, Clarkson MJ, Tremethick DJ, Luger K (2000). Crystal structure of a nucleosome core particle containing the variant histone H2A.Z. Nature structural biology 7(12): 1121-1124.

Tapia C, Kutzner H, Mentzel T, Savic S, Baumhoer D, Glatz K (2006). Two mitosis-specific antibodies, MPM-2 and phospho-histone H3 (Ser28), allow rapid and precise determination of mitotic activity. The American journal of surgical pathology 30(1): 83-89.

Taylor RC, Cullen SP, Martin SJ (2008). Apoptosis: controlled demolition at the cellular level. Nature reviews. Molecular cell biology 9(3): 231-241.

Tenen DG (2003). Disruption of differentiation in human cancer: AML shows the way. Nature reviews. Cancer 3(2): 89-101.

Thompson CB (1995). Apoptosis in the pathogenesis and treatment of disease. Science (New York, N.Y.) 267(5203): 1456-1462.

Timofeev O, Cizmecioglu O, Settele F, Kempf T, Hoffmann I (2010). Cdc25 phosphatases are required for timely assembly of CDK1-cyclin B at the G2/M transition. The Journal of biological chemistry 285(22): 16978-16990.

Tournier C, Dong C, Turner TK, Jones SN, Flavell RA, Davis RJ (2001). MKK7 is an essential component of the JNK signal transduction pathway activated by proinflammatory cytokines. Genes & development 15(11): 1419-1426.

Vader G, Lens SM (2008). The Aurora kinase family in cell division and cancer. Biochimica et biophysica acta 1786(1): 60-72.

Viele CS (2003). Diagnosis, treatment, and nursing care of acute leukemia. Seminars in oncology nursing 19(2): 98-108.

Vucic D, Dixit VM, Wertz IE (2011). Ubiquitylation in apoptosis: a post-translational modification at the edge of life and death. Nature reviews. Molecular cell biology 12(7): 439-452.

Wang Y, Tjandra N (2013). Structural insights of tBid, the caspase-8-activated Bid, and its BH3 domain. The Journal of biological chemistry 288(50): 35840-35851.

Wei MC, Lindsten T, Mootha VK, Weiler S, Gross A, Ashiya M, et al. (2000). tBID, a membrane-targeted death ligand, oligomerizes BAK to release cytochrome c. Genes & development 14(16): 2060-2071.

West AC, Johnstone RW (2014). New and emerging HDAC inhibitors for cancer treatment. The Journal of clinical investigation 124(1): 30-39.

Xu WS, Parmigiani RB, Marks PA (2007). Histone deacetylase inhibitors: molecular mechanisms of action. Oncogene 26(37): 5541-5552.

Yamamoto S, Tanaka K, Sakimura R, Okada T, Nakamura T, Li Y, et al. (2008). Suberoylanilide hydroxamic acid (SAHA) induces apoptosis or autophagy-associated cell death in chondrosarcoma cell lines. Anticancer research 28(3a): 1585-1591.

Yang XJ (2004). The diverse superfamily of lysine acetyltransferases and their roles in leukemia and other diseases. Nucleic acids research 32(3): 959-976.

Yang XJ, Seto E (2007). HATs and HDACs: from structure, function and regulation to novel strategies for therapy and prevention. Oncogene 26(37): 5310-5318.

Youle RJ, Strasser A (2008). The BCL-2 protein family: opposing activities that mediate cell death. Nature reviews. Molecular cell biology 9(1): 47-59.

Zanke BW, Boudreau K, Rubie E, Winnett E, Tibbles LA, Zon L, et al. (1996). The stress-activated protein kinase pathway mediates cell death following injury induced by cis-platinum, UV irradiation or heat. Current biology : CB 6(5): 606-613.

Zitouni S, Nabais C, Jana SC, Guerrero A, Bettencourt-Dias M (2014). Polo-like kinases: structural variations lead to multiple functions. Nature reviews. Molecular cell biology 15(7): 433-452.
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