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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
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dc.contributor.advisor | 蔡丰喬 | |
dc.contributor.author | Li-Yun Chang | en |
dc.contributor.author | 張立昀 | zh_TW |
dc.date.accessioned | 2021-06-17T06:35:48Z | - |
dc.date.available | 2023-10-03 | |
dc.date.copyright | 2018-10-03 | |
dc.date.issued | 2018 | |
dc.date.submitted | 2018-08-16 | |
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Gyanchandani, R., et al., A Proangiogenic Signature Is Revealed in FGF-Mediated Bevacizumab-Resistant Head and Neck Squamous Cell Carcinoma. Molecular Cancer Research, 2013. 11(12): p. 1585-1596. 32. Gartside, M.G., et al., Loss-of-function fibroblast growth factor receptor-2 mutations in melanoma. Mol Cancer Res, 2009. 7(1): p. 41-54. 33. Ornitz, D.M. and N. Itoh, The Fibroblast Growth Factor signaling pathway. Wiley Interdiscip Rev Dev Biol, 2015. 4(3): p. 215-66. 34. Wu, Y.J., Z.J. Chen, and A. Ullrich, EGFR and FGFR signaling through FRS2 is subject to negative feedback control by ERK1/2. Biological Chemistry, 2003. 384(8): p. 1215-1226. 35. Yamada, H., et al., Dynasore, a dynamin inhibitor, suppresses lamellipodia formation and cancer cell invasion by destabilizing actin filaments. Biochem Biophys Res Commun, 2009. 390(4): p. 1142-8. | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/72330 | - |
dc.description.abstract | 頭頸部鱗狀上皮細胞癌在台灣有相當高的發病率及致死率,且十分容易復發及轉移,然而目前在頭頸癌的治療上並沒有很大的突破。因此,我們希望找到新的治療標的來阻止腫瘤的進展。纖維母細胞受器(Fibroblast growth factor receptor,FGFR)參與了胚胎發育、血管新生、傷口癒合等重要的生理功能,且在細胞增生、分化以及移動上都扮演了重要的角色。除此之外,近年來有研究也顯示了FGFR的異常和許多癌症的發生及其病程發展相關。然而,FGFR在頭頸癌病程上所扮演的角色為何,目前仍沒有詳盡的探討與瞭解。因此,我們實驗室先前利用FGFRs的抑制劑AZD4547來觀察對口腔鱗狀上皮癌細胞SAS移動的影響。有趣的是,我們發現到,在沒有FBS、並加入FGF1作刺激的環境下給予AZD4547,細胞移動速度會顯著的下降;然而,若是在有FBS的環境下給予AZD4547,卻發現無法達到抑制細胞遷移的效果。進一步去釐清FBS中的何種物質導致此現象的發生,發現到在沒有FBS、並加入EGF去刺激細胞的環境下,再給予AZD4547,能夠看到類似在FBS中的結果。藉此,我們推論EGFR和FGFR signaling之間可能存在交互作用。
為了進一步探討EGFR與FGFR signaling之間的交互作用,我們透過single cell tracing的技術去探討當其中一個signaling存在或被抑制時,對另一個signaling在細胞遷移上會造成何種影響。從實驗結果我們觀察到一個有趣的現象:EGF可以回復FGFRs抑制劑的抑制效果,卻無法回復FGFR2 knockdown的抑制效果。在加EGFR抑制劑及EGFR knockdown的實驗中也觀察到相似的情形。藉此推論EGFR和FGFR要在彼此的physical structure存在之下才會產生交互作用。我們因此透過FGFR kinase-dead construct來模擬抑制劑的效果,觀察到其結果與加抑制劑時相似,也更強化了我們的推論。 建立於此推論之上,我們進一步提出三個EGFR和FGFR signaling產生交互作用的可能機轉模型:receptor level model、transactivation model 以及 localization model,並透過不同的實驗去作驗證。我們目前的實驗結果指出:當FGFR的adaptor protein FRS2被knockdown時可以輕微減弱EGF在抑制劑上的回復效果,且從western blot結果則發現到FGFR2 knockdown時可以讓細胞中的Erk表現量減少,然而在加入FGFRs抑制劑時則看不到此現象的產生。以上,我們推論EGFR和FGFR signaling之間很可能是透過transactivation的過程來產生交互作用。而我們仍需再透過其他實驗來釐清並加強此部分的推論,期望能在有更充分的瞭解後,對頭頸癌的治療提供指引與方向。 | zh_TW |
dc.description.abstract | Head and neck squamous cell carcinoma (HNSCC) causes high rates of metastasis, recurrence and mortality in Taiwan, but effective treatments remain lacking. Thus searching for novel therapeutic targets against HNSCC may improve current therapeutics for HNSCC patients. We therefore focus on fibroblast growth factor receptor (FGFR), which is important in cancer cell proliferation, differentiation and migration. Since whether and how FGFR signaling affected HNSCC progression had not been well explored, we used FGFR inhibitor AZD4547 to treat SAS in cell migration assays. Interestingly, AZD4547 suppressed SAS migration speed stimulated by FGF1 but it failed to suppress SAS migration when FBS was present. Similarly, the presence of EGF also abrogated the suppressive effect of AZD4547 on SAS migration. Thus, we proposed EGFR and FGFR signaling interacted with each other during cell migration.
To further explore the crosstalk between EGFR and FGFR signaling, we used single cell tracing technique to analyze how FGFR/EGFR suppression changed the migration behaviors in the presence or absence of EGFR/FGFR signaling. We found it interesting that EGF rescued the suppressive effect of FGFR inhibitor on HNSCC cell migration, but EGF could not rescue FGFR2 knockdown. Similar though less dramatic results were also noted when we used FGF1 to rescue EGFR inhibitor or EGFR knockdown. Thus we speculate that interaction between EGFR and FGFR occurs through physical interactions between receptors. Furthermore, over-expressing the kinase-dead mutant of FGFR2 in SAS cells mimicked the suppressive effects of FGFR inhibitors, and the suppression could be rescued by the addition of EGF. Such results support that physical presence of receptor is important and required for signaling interaction between EGFR and FGFR. To explain how EGFR interacts with FGFR signaling, we proposed three hypothetical models : receptor level model、transactivation model and localization model. Because FRS2 knockdown showed modest inhibition on EGF stimulation and western blot showed that shFGFR2 decreased Erk expression but FGFR inhibitors did not, we favor the transactivation model. Further investigations are underway to verify our speculations, with the hope to benefit current HNSCC treatments. | en |
dc.description.provenance | Made available in DSpace on 2021-06-17T06:35:48Z (GMT). No. of bitstreams: 1 ntu-107-R05443008-1.pdf: 2778137 bytes, checksum: a33e9501cb0800c165150d69d523866c (MD5) Previous issue date: 2018 | en |
dc.description.tableofcontents | 誌謝 1
中文摘要 2 Abstract 4 目錄 6 圖目錄 8 表目錄 9 第一章、緒論 10 1. 頭頸部鱗狀上皮細胞癌(Head and neck squamous cell carcinoma, HNSCC)為全球及台灣應重視的疾病問題 10 2. 纖維母細胞生長因子受器(Fibroblast growth factor receptors,FGFRs)影響了EMT與癌症移轉,為頭頸癌治療有潛力的目標 12 3. 研究動機 15 第二章、材料與方法 17 1.細胞培養 17 2.細胞遷移實驗 (Cell migration assay) 17 3.細胞蛋白質萃取(Protein extraction)與西方墨點法(Western blotting) 18 4.核糖核酸萃取(RNA extraction)與反轉錄酶聚合酶鏈反應(Reverse transcription polymerase chain reaction, RT-PCR) 21 5 慢病毒基因靜默技術(Lentivirus shRNA knockdown) 23 6.質體DNA轉染 (Plasmid DNA transfection) 25 7.Constructs製作 25 8.免疫螢光染色 (Immunofluorescence assay) 27 第三章、結果 28 1. 透過抑制FGFR signaling探討其與EGFR signaling之間的交互作用 28 2. 透過抑制EGFR signaling探討其與FGFR signaling之間的交互作用 34 3. 驗證Hypothetical model 1 : receptor level model 37 4. 驗證Hypothetical model 2 : transactivation model 40 5. 驗證 Hypothetical model 3 : localization model 42 6. 透過overexpress pEYFP-P2A-FGFR2質體來驗證receptor physical structure對signaling pathway之間產生交互作用的重要性 45 7. 透過表現FGFR2 kinase-dead的質體來模擬FGFRs抑制劑的現象 48 第四章、討論與結論 51 1. 透過Western blot來確認加入生長因子以及抑制劑後的下游訊息傳遞表現 51 2. SAS以及HSC3在FGFR2 knockdown時,加入EGF刺激後對細胞移動的影響存在差異性 53 3. Overexpress FGFR2IIIb以及FGFR2IIIc無法增加對EGFR signaling在細胞移動上的效果 55 4. 在SAS細胞中過度表現 pEYFP-P2A-FGFR2IIIc-A648T後是否真的有發揮kinase-dead的效果 55 5. Knockdown FRS2後在SAS細胞中產生的抑制效果不顯著 56 6. Dynasore無法使EGF刺激回復FGFR2 knockdown的抑制效果 59 第五章、參考資料 61 第六章、附錄 64 | |
dc.language.iso | zh-TW | |
dc.title | 纖維母細胞生長因子受器與上皮細胞生長因子受器在頭頸部鱗狀細胞癌中的交互作用對細胞遷移的影響 | zh_TW |
dc.title | Signaling interaction between FGFR and EGFR in HNSCC: focusing on cell migration | en |
dc.type | Thesis | |
dc.date.schoolyear | 106-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 張玉芳,陳青周,涂熊林,鄭乃禎 | |
dc.subject.keyword | 頭頸部鱗狀細胞癌,EGFR,FGFR,細胞遷移,訊息傳遞, | zh_TW |
dc.subject.keyword | Head and neck squamous cell carcinoma (HNSCC), Epithelial growth factor receptor (EGFR),Fibroblast growth factor receptor (FGFR),Cell migration,Signal transduction, | en |
dc.relation.page | 80 | |
dc.identifier.doi | 10.6342/NTU201803551 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2018-08-16 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 藥理學研究所 | zh_TW |
顯示於系所單位: | 藥理學科所 |
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