請用此 Handle URI 來引用此文件：
http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/74485完整後設資料紀錄
| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 梁博煌(Po-Huang Liang) | |
| dc.contributor.author | Chin-Jung Kuo | en |
| dc.contributor.author | 郭瑾融 | zh_TW |
| dc.date.accessioned | 2021-06-17T08:38:28Z | - |
| dc.date.available | 2022-08-16 | |
| dc.date.copyright | 2019-08-16 | |
| dc.date.issued | 2019 | |
| dc.date.submitted | 2019-08-08 | |
| dc.identifier.citation | 1. Brackley, K.I. and J. Grantham, Activities of the chaperonin containing TCP-1 (CCT): implications for cell cycle progression and cytoskeletal organisation. Cell stress & chaperones, 2009. 14(1): p. 23-31.
2. Llorca, O., et al., Eukaryotic chaperonin CCT stabilizes actin and tubulin folding intermediates in open quasi-native conformations. The EMBO journal, 2000. 19(22): p. 5971-5979. 3. Muñoz, I.G., et al., Crystal structure of the open conformation of the mammalian chaperonin CCT in complex with tubulin. Nature Structural &Amp; Molecular Biology, 2010. 18: p. 14. 4. Chen, S.-H., Small-molecule inhibitors of protein-protein interactions selectively induce chemoresistant/caspase-3-deficient cancer cell death, in Department of Biological Science and Technology. 2014, National Chiao Tung University. 5. Yokota, S., et al., Increased expression of cytosolic chaperonin CCT in human hepatocellular and colonic carcinoma. Cell stress & chaperones, 2001. 6(4): p. 345-350. 6. Lin, Y.-F., et al., Intracellular β-Tubulin/Chaperonin Containing TCP1-β Complex Serves as a Novel Chemotherapeutic Target against Drug-Resistant Tumors. Cancer Research, 2009. 69(17): p. 6879. 7. Liu, Y.-J., et al., Disrupting CCT-β : β-tubulin selectively kills CCT-β overexpressed cancer cells through MAPKs activation. Cell death & disease, 2017. 8(9): p. e3052-e3052. 8. Mimnaugh, E.G., et al., Endoplasmic Reticulum Vacuolization and Valosin- Containing Protein Relocalization Result from Simultaneous Hsp90 Inhibition by Geldanamycin and Proteasome Inhibition by Velcade. Molecular Cancer Research, 2006. 4(9): p. 667. 9. Wójcik, C., et al., Valosin-containing protein (p97) is a regulator of endoplasmic reticulum stress and of the degradation of N-end rule and ubiquitin-fusion degradation pathway substrates in mammalian cells. Molecular biology of the cell, 2006. 17(11): p. 4606-4618. 10. Lin, Y.F., Y.F. Lee, and P.H. Liang, Targeting β-tubulin:CCT-β complexes incurs Hsp90- and VCP-related protein degradation and induces ER stress-associated apoptosis by triggering capacitative Ca2+ entry, mitochondrial perturbation and caspase overactivation. Cell death & disease, 2012. 3(11): p. e434-e434. 11. Sano, R. and J.C. Reed, ER stress-induced cell death mechanisms. Biochimica et biophysica acta, 2013. 1833(12): p. 3460-3470. 12. Kuo, Y.-T., Disrupting β-tubulin/CCT-β complexes induces apoptosis and suppresses migration and invasion of CL1-5 cells through MMP-2 and AKT/GSK-3β inhibition, in Institute of Biochemical Sciences. College of Life Science. 2017, National Taiwan University. 13. Global Burden of Disease Cancer, C., et al., Global, Regional, and National Cancer Incidence, Mortality, Years of Life Lost, Years Lived With Disability, and Disability-Adjusted Life-Years for 29 Cancer Groups, 1990 to 2016: A Systematic Analysis for the Global Burden of Disease Study. JAMA oncology, 2018. 4(11): p. 1553-1568. 14. Li, G., K. Satyamoorthy, and M. Herlyn, N-Cadherin-mediated Intercellular Interactions Promote Survival and Migration of Melanoma Cells. Cancer Research, 2001. 61(9): p. 3819. 15. Felipe Lima, J., et al., EMT in Breast Carcinoma-A Review. Journal of clinical medicine, 2016. 5(7): p. 65. 16. Meighan, C.M. and J.E. Schwarzbauer, Temporal and spatial regulation of integrins during development. Current opinion in cell biology, 2008. 20(5): p. 520-524. 17. Jin, H. and J. Varner, Integrins: roles in cancer development and as treatment targets. British journal of cancer, 2004. 90(3): p. 561-565. 18. Alizadeh, A.M., S. Shiri, and S. Farsinejad, Metastasis review: from bench to bedside. Tumor Biology, 2014. 35(9): p. 8483-8523. 19. Parker, A.L., M. Kavallaris, and J.A. McCarroll, Microtubules and Their Role in Cellular Stress in Cancer. Frontiers in Oncology, 2014. 4(153). 20. Walter, P. and D. Ron, The Unfolded Protein Response: From Stress Pathway to Homeostatic Regulation. Science, 2011. 334(6059): p. 1081. 21. Liu, C.Y. and R.J. Kaufman, The unfolded protein response. Journal of Cell Science, 2003. 116(10): p. 1861. 22. Yoshida, H., et al., XBP1 mRNA Is Induced by ATF6 and Spliced by IRE1 in Response to ER Stress to Produce a Highly Active Transcription Factor. Cell, 2001. 107(7): p. 881-891. 23. Li, G., et al., Role of ERO1-alpha-mediated stimulation of inositol 1,4,5- triphosphate receptor activity in endoplasmic reticulum stress-induced apoptosis. The Journal of cell biology, 2009. 186(6): p. 783-792. 24. Li, J., B. Lee, and A.S. Lee, Endoplasmic Reticulum Stress-induced Apoptosis: MULTIPLE PATHWAYS AND ACTIVATION OF p53-UP-REGULATED MODULATOR OF APOPTOSIS (PUMA) AND NOXA BY p53. Journal of Biological Chemistry, 2006. 281(11): p. 7260-7270. 25. Timmins, J.M., et al., Calcium/calmodulin-dependent protein kinase II links ER stress with Fas and mitochondrial apoptosis pathways. The Journal of clinical investigation, 2009. 119(10): p. 2925-2941. 26. Han, J., et al., ER-stress-induced transcriptional regulation increases protein synthesis leading to cell death. Nature cell biology, 2013. 15(5): p. 481-490. 27. Su, J., et al., Bcl-2 family proteins are involved in the signal crosstalk between endoplasmic reticulum stress and mitochondrial dysfunction in tumor chemotherapy resistance. BioMed research international, 2014. 2014: p. 234370-234370. 28. McIlwain, D.R., T. Berger, and T.W. Mak, Caspase functions in cell death and disease. Cold Spring Harbor perspectives in biology. 7(4): p. a026716. 29. Boulares, A.H., et al., Role of Poly(ADP-ribose) Polymerase (PARP) Cleavage in Apoptosis: CASPASE 3-RESISTANT PARP MUTANT INCREASES RATES OF APOPTOSIS IN TRANSFECTED CELLS. Journal of Biological Chemistry, 1999. 274(33): p. 22932-22940. 30. Elmore, S., Apoptosis: a review of programmed cell death. Toxicologic pathology, 2007. 35(4): p. 495-516. 31. Garg, M., Epithelial-mesenchymal transition - activating transcription factors - multifunctional regulators in cancer. World journal of stem cells, 2013. 5(4): p. 188-195. 32. Jordà, M., et al., Upregulation of MMP-9 in MDCK epithelial cell line in response to expression of the Snail transcription factor. Journal of Cell Science, 2005. 118(15): p. 3371. 33. ten Berge, D., et al., Wnt signaling mediates self-organization and axis formation in embryoid bodies. Cell stem cell, 2008. 3(5): p. 508-518. 34. Howe, L.R., et al., Twist Is Up-Regulated in Response to Wnt1 and Inhibits Mouse Mammary Cell Differentiation. Cancer Research, 2003. 63(8): p. 1906. 35. Zhou, B.P., et al., Dual regulation of Snail by GSK-3β-mediated phosphorylation in control of epithelial–mesenchymal transition. Nature Cell Biology, 2004. 6(10): p. 931-940. 36. Rychahou, P.G., et al., Akt2 overexpression plays a critical role in the establishment of colorectal cancer metastasis. Proceedings of the National Academy of Sciences of the United States of America, 2008. 105(51): p. 20315- 20320. 37. Daugherty, R.L. and C.J. Gottardi, Phospho-regulation of Beta-catenin adhesion and signaling functions. Physiology (Bethesda, Md.), 2007. 22: p. 303-309. 38. G. Gritsenko, P., O. Ilina, and P. Friedl, Interstitial guidance of cancer invasion. The Journal of Pathology, 2012. 226(2): p. 185-199. 39. Wolf, K., et al., Collagen-based cell migration models in vitro and in vivo. Seminars in Cell & Developmental Biology, 2009. 20(8): p. 931-941. 40. Poincloux, R., F. Lizárraga, and P. Chavrier, Matrix invasion by tumour cells: a focus on MT1-MMP trafficking to invadopodia. Journal of Cell Science, 2009. 122(17): p. 3015. 41. Pietilä, M., J. Ivaska, and S.A. Mani, Whom to blame for metastasis, the epithelial–mesenchymal transition or the tumor microenvironment? Cancer Letters, 2016. 380(1): p. 359-368. 42. Guo, W. and F.G. Giancotti, Integrin signalling during tumour progression. Nature Reviews Molecular Cell Biology, 2004. 5(10): p. 816-826. 43. Krejci, P., et al., Receptor Tyrosine Kinases Activate Canonical WNT/β-Catenin Signaling via MAP Kinase/LRP6 Pathway and Direct β-Catenin Phosphorylation. PLOS ONE, 2012. 7(4): p. e35826. 44. Rangaswami, H., et al., Protein kinase G and focal adhesion kinase converge on Src/Akt/β-catenin signaling module in osteoblast mechanotransduction. The Journal of biological chemistry, 2012. 287(25): p. 21509-21519. 45. Zhang, F., et al., Distinct ligand binding sites in integrin alpha3beta1 regulate matrix adhesion and cell-cell contact. The Journal of cell biology, 2003. 163(1): p. 177-188. 46. Fujita, Y., et al., Hakai, a c-Cbl-like protein, ubiquitinates and induces endocytosis of the E-cadherin complex. Nature Cell Biology, 2002. 4(3): p. 222- 231. 47. Giubellino, A., T.R. Burke, Jr., and D.P. Bottaro, Grb2 signaling in cell motility and cancer. Expert opinion on therapeutic targets, 2008. 12(8): p. 1021-1033. 48. O'Connor, K. and M. Chen, Dynamic functions of RhoA in tumor cell migration and invasion. Small GTPases, 2013. 4(3): p. 141-147. 49. Fife, C.M., J.A. McCarroll, and M. Kavallaris, Movers and shakers: cell cytoskeleton in cancer metastasis. British journal of pharmacology, 2014. 171(24): p. 5507-5523. 50. Jayatilaka, H., et al., EB1 and cytoplasmic dynein mediate protrusion dynamics for efficient 3-dimensional cell migration. FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 2018. 32(3): p. 1207-1221. | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/74485 | - |
| dc.description.abstract | 在先前的研究中，我們發現破壞β-tubulin與CCT-β的蛋白質複合體可以做為新的抗癌方針。並且，經由虛擬篩選，我們從Sigma-Aldrich compound bank中找到一個非共價性的小分子藥物，3112210，可逆地結合於β-tubulin與CCT-β接觸面上的熱點。
本實驗中，我們將3112210測試於具有高度轉移性的非小細胞肺癌細胞株CL1-5。首先，在免疫共沈澱實驗結果顯示，經3112210處理後的細胞中，β- tubulin與CCT-β的蛋白質複合體會被破壞。另外，3112210藉由內質網壓力與細胞凋亡的機制使CL1-5肺小癌細胞死亡。除了探討3112210的癌細胞毒性，我們也以其EC20濃度進行細胞遷移與侵入實驗。實驗結果顯示，3112210可以抑制CL1-5細胞遷移與侵入的能力，並降低基質金屬蛋白酶MMP-2, -9的表現量與活性。在西方點墨法實驗與電泳酵素分析法中，我們推測此藥效是由整合素(Integrin)及其下游訊息傳遞帶動EMT調節因子的表現量改變所造成。 總結而論，3112210是一個新的非共價性地破壞CL1-5肺癌細胞內β-tubulin與CCT-β蛋白質複合體的小分子藥物，它對CL1-5細胞具有毒性並可以抑制其轉移能力。 | zh_TW |
| dc.description.abstract | Previously, we reported the protein-protein interaction (PPI) between β-tubulin and CCT-β complex as a potential anti-cancer chemotherapeutic target. Through virtual screening, a compound 3112210 from Sigma-Aldrich compound bank was identified to be a reversible inhibitor of the PPI by docking into hot spots on this PPI interface of β- tubulin.
In this study, 3112210 was tested on a highly metastatic non-small cell lung cancer (NSCLC) cell line, CL1-5. The co-IP experiments showed that, in 3112210-treated cancer cells, β-tubulin and CCT-β complex was disrupted. Furthermore, 3112210 caused CL1-5 cell death through ER stress and apoptosis. In addition to verifying its toxicity toward CL1-5, we performed migration and invasion assays using dosage at about IC20. The results indicated that 3112210 also inhibited cancer cell migration and invasion, and MMP-2, -9 were also inhibited. These anti-metastatic effects were endowed via integrin- related pathways and EMT transcriptional factors, as demonstrated by western blot experiments. To sum, 3112210 is a novel non-covalent inhibitor for β-tubulin:CCT-β complex in CL1-5 lung adenocarcinoma cells to induce cancer cell death and impeded cell metastasis. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-17T08:38:28Z (GMT). No. of bitstreams: 1 ntu-108-R06b46022-1.pdf: 2063572 bytes, checksum: 9884a9fa834e00590b496ea19184437c (MD5) Previous issue date: 2019 | en |
| dc.description.tableofcontents | TABLE OF CONTENTS .............................................................................................I
中文摘要....................................................................................................................IV ABSTRACT ............................................................................................................... V ABBREVIATIONS ...................................................................................................VI 1. INTRODUCTION .................................................................................................. 1 1.1 CHAPERONIN-CONTAINING TCP-1(CCT) .............................................................. 1 1.2 Β-TUBULIN:CCT-Β COMPLEX AS ANTI-CANCER CHEMOTHERAPEUTIC TARGET ......... 2 1.3 COMPOUND 3112210............................................................................................ 3 1.4 CANCER METASTASIS............................................................................................ 4 2. MATERIALS AND METHODS............................................................................. 6 2.1. CHEMICALS......................................................................................................... 6 2.2. CELL CULTURE .................................................................................................... 6 2.3. WESTERN BLOTTING ANALYSIS AND CO-IMMUNOPRECIPITATION (CO-IP) ASSAY ..... 7 2.4. MTT ASSAY ........................................................................................................ 8 2.5. FLOW CYTOMETRIC ANALYSIS .............................................................................. 9 2.6. PROTEASOME ACTIVITY ASSAY ........................................................................... 10 2.7. MEASUREMENT OF CYTOSOLIC CA2+ .................................................................. 10 2.8. WOUND-HEALING ASSAY ................................................................................... 11 2.9. MATRIGEL INVASION ASSAY ............................................................................... 12 2.10. GELATIN ZYMOGRAPHY ................................................................................... 12 3. RESULTS .............................................................................................................. 15 3.1. 3112210 INTERRUPTED THE PPI BETWEEN Β-TUBULIN AND CCT-Β IN CL1-5 CELLS. ............................................................................................................................... 15 3.2. 3112210 TREATMENT INHIBITED THE VIABILITY OF CL1-5 CELL. ......................... 15 3.3. 3112210 TRIGGERED UPR IN CL1-5 CELLS. ....................................................... 16 3.4. 3112210 INCREASED THE CONCENTRATION OF CYTOSOLIC CA2+ IN CL1-5 CELLS. 17 3.5. 3112210 INDUCED THE ACTIVATION OF CASPASES AND PRO-APOPTOTIC PROTEINS. 18 3.6. 3112210 INHIBITED MIGRATION OF CL1-5 CELLS. ............................................... 19 3.7. DECREASE IN INVASION POTENCY OF CL1-5 CELLS BY 3112210 .......................... 20 3.8. 3112210 TREATMENT SUPPRESSED EMT REGULATORS IN CL1-5 CELLS ............... 21 3.9. 3112210 DOWN-REGULATED THE EXPRESSION OF MMP-2, -9.............................. 21 3.10. 3112210 DOWN-REGULATES P-AKT/Β-CATENIN PATHWAY ................................ 22 4. DISCUSSION........................................................................................................ 25 FIGURE .................................................................................................................... 28 REFERENCE ........................................................................................................... 45 | |
| dc.language.iso | en | |
| dc.subject | 抗癌症小分子藥物 | zh_TW |
| dc.subject | β-tubulin:CCT-β蛋白質複合體 | zh_TW |
| dc.subject | 抗癌症轉移 | zh_TW |
| dc.subject | 細胞凋亡 | zh_TW |
| dc.subject | β-tubulin:CCT-β complex | en |
| dc.subject | anti-cancer small molecule | en |
| dc.subject | apoptosis | en |
| dc.subject | anti-metastasis | en |
| dc.title | 藉由非共價性破壞β-tubulin:CCT-β蛋白質複合體以引發CL1-5細胞凋亡並抑制其轉移能力的小分子藥物 | zh_TW |
| dc.title | A non-covalent small inhibitor blocking β-tubulin:CCT-β complex induces apoptosis and suppresses migration and invasion
in CL1-5 cells | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 107-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 李靜琪(Jinq-Chyi Lee),林源峰(Yuan-Feng Lin) | |
| dc.subject.keyword | 抗癌症小分子藥物,β-tubulin:CCT-β蛋白質複合體,細胞凋亡,抗癌症轉移, | zh_TW |
| dc.subject.keyword | anti-cancer small molecule,β-tubulin:CCT-β complex,apoptosis,anti-metastasis, | en |
| dc.relation.page | 52 | |
| dc.identifier.doi | 10.6342/NTU201902879 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2019-08-08 | |
| dc.contributor.author-college | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 生化科學研究所 | zh_TW |
| 顯示於系所單位： | 生化科學研究所 | |
文件中的檔案：
| 檔案 | 大小 | 格式 | |
|---|---|---|---|
| ntu-108-1.pdf 未授權公開取用 | 2.02 MB | Adobe PDF |
系統中的文件，除了特別指名其著作權條款之外，均受到著作權保護，並且保留所有的權利。