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http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/78743完整後設資料紀錄
| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 王逸平 | |
| dc.contributor.author | Kai-Chi Chen | en |
| dc.contributor.author | 陳凱琪 | zh_TW |
| dc.date.accessioned | 2021-07-11T15:16:12Z | - |
| dc.date.available | 2029-07-31 | |
| dc.date.copyright | 2019-08-28 | |
| dc.date.issued | 2019 | |
| dc.date.submitted | 2019-07-25 | |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/78743 | - |
| dc.description.abstract | 在全球癌症統計中，頭頸部鱗狀細胞癌(HNSCC)是第六個最常見的惡性腫瘤，平均5年存活率為百分之五十。根據全外顯體定序結果(whole exome sequencing)，發現基底型(basal type)HNSCC具有 NRG1及HER3高度表現之現象，但由於台灣本土頭頸癌之表現及其調控機制尚不明確。因此，為了探討此議題，我們收集了一百位口腔癌(OSCC)患者的檢體，進行核酸定量分析(Quanti Gene Plex assay)及免疫組織染色(IHC)，發現有7成以上的樣本，其HER3及NRG1蛋白高度表現。此外，我們發現HER3及NRG1的低度表達與淋巴轉移及晚期病程相關。此外，在癌組織中HER3及NRG1的表現在空間位置上具有互斥之現象。HER3主要表現於腫瘤巢(tumor nest)中間分化良好的細胞中；反之，NRG1多於tumor nest外圍的較差分化之類基底細胞(basaloid)群中。因此，我們想進一步探討HER3及NRG1在腫瘤細胞中表現互斥的機制。我們挑選高度表現NOTCH1的HNSCC細胞株FaDu及Cal 27，利用shRNA及γ-secretase抑制劑分別抑制NOTCH1的表現及功能後，HER3的蛋白表現量增加；另外，將NOTCH1過量表現後，HER3的蛋白表現量下降。然而，利用即時聚合酶連鎖反應(RT-PCR)及啟動子活性分析(promotor activity assay)，發現NOTCH1不影響HER3 mRNA的表現及啟動子活性。隨後，在NOTCH1 knockdown的細胞中，發現AKT 的活性上升會使HER3泛素化之修飾增加，且蛋白穩定性下降。最後，利用siRNA抑制USP8在NOTCH1 knockdown細胞中表現，發現AKT會透過調控USP8的活性進而影響HER3蛋白之穩定性。綜合上述結果，我們發現台灣的OSCC之淋巴轉移及癌症晚期患者，其與HER3及NRG1低表現有顯著相關。並且，我們證實了在HNSCC中，NOTCH1透過抑制AKT磷酸化進而維持HER3的蛋白穩定性。未來，我們將進一步探討NOTCH1調控AKT及NRG1的機制，期望能找到更專一的生物標誌並更有效的診斷及治療頭頸癌。 | zh_TW |
| dc.description.abstract | Head and neck cancer squamous cell carcinoma (HNSCC) is the sixth most common malignancy in the global cancer statistics, with an average 5-year survival rate of 50%. According to the whole exome sequencing, it is found that NRG1 and HER3 are highly expressed in the basal type HNSCC. In this study, we intend to explore the correlations between the expressions of NRG1 and HER3 and the mechanism of HNSCC in Taiwan. 100 specimens of patients with OSCC obtained from the National Taiwan University Hospital from 2006 to 2012 were collected for QuantiGene Plex assay and immunohistochemical staining (IHC). Proteins of HER3 and NRG1 are overexpressed with prevalence of 70-90%. In addition, low mRNA and protein expressions of HER3 and NRG1 are associated with lymphatic involvement and late stage progression. Moreover, the protein expression of HER3 and NRG1 in tumor are mutually exclusive in spatial distribution. HER3 is mainly expressed in the control of tumor nest which defines as well differentiated cells. In contrast, NRG1 is expressed in the basaloid cell which located in the peripheral cell of tumor nest, which also is defined as poor differentiated cells. Therefore, we would like to further explore the mechanism by which HER3 and NRG1 are mutually exclusive in tumor cells. The expression of HER3 protein is increased after shRNA and γ-secretase inhibitor inhibiting the expression and function of NOTCH1 respectively in FaDu and Cal 27. In contrast, the protein expression of HER3 is decreased upon NOTCH1-overexpressed. However, validation using the promoter activity assay revealed that HER3 promoter activity is not changed in either in NOTCH1 knockdown or overexpressed cells. Subsequently, increasing AKT activity in NOTCH1 knockdown cells it is found that leads to enhance ubiquitination modification and decrease protein stability of HER3. Furthermore, silencing of USP8 decreases protein expression of HER3 and increase poly-ubiquitination of HER3 in NOTCH1 knockdown cells. Based on the above results, we find that patients with lymphatic involvement and late stage in Taiwan OSCC have a significant correlation with low expressions of HER3 and NRG1. Furthermore, we confirm that in HNSCC, NOTCH1 inhibits AKT phosphorylation activity, and induces USP8-dependent deubiquitination of HER3, thereby maintaining the stability of HER3 protein. In the future, we will further explore the mechanism through which NOTCH1 regulates AKT activation and NRG1 in detail. We expect to find out more specific biomarkers, which can be developed into more effective diagnosis and treatment of HNSCC. Thus, our study serves as the impetus to conduct future studies to uncover the underlying molecular mechanisms of these biomarkers. | en |
| dc.description.provenance | Made available in DSpace on 2021-07-11T15:16:12Z (GMT). No. of bitstreams: 1 ntu-108-R06450006-1.pdf: 7064888 bytes, checksum: 26dd69af1154831384b35034414b4e42 (MD5) Previous issue date: 2019 | en |
| dc.description.tableofcontents | 論文口試委員審定書 i
致謝 ii 中文摘要 iii Abstract iv Content vi Content of figures ix Content of tables x List of abbreviations xi Introduction 1 1.1 Head and neck squamous cell carcinoma 1 1.2 Human epidermal growth factor receptor 3 3 1.2.1 Neuregulin 1 3 1.2.2 HER family and HER3 4 1.2.3 HER3 signaling pathway 5 1.2.4 HER3 expression in HNSCC 6 1.3 NOTCH1 10 1.3.1 The structure and signaling of NOTCH 10 1.3.2 NOTCH1 expression in HNSCC 12 1.4 Specific aims 15 Materials and methods 17 2.1 Clinical specimen acquisition and clinicopathological staging 17 2.2 Quanti Gene Plex Assay 17 2.3 Immunohistochemical staining 18 2.4 Cell line and culture condition 19 2.5 Plasmid construction 19 2.6 Lentivirus infection 20 2.7 Luciferase reporter assay 21 2.8 Immunoprecipitation assay 21 2.9 Western blot analysis 22 2.10 RNA extraction and real-time polymerase chain reaction 22 2.11 Statistical analysis 23 Results 24 3.1 The expression pattern of HER3 and NRG1 in clinical specimens of OSCC and their correlation to clinicopathological parameter 24 3.2 Spatial discrepancy between HER3 and NRG1 in HNSCC. 26 3.3 Suppression of NOTCH1 downregulates HER3 production in HNSCC cell lines 26 3.4 NOTCH1 contributes to protein stability and post-translational modification of HER3 29 3.5 NOTCH1 regulates HER3 protein stability via phosphorylation of AKT 29 3.6 USP8 is involved in the regulation of HER3 by NOTCH1 31 Discussion 33 Figures 45 Tables 58 Appendix 65 Reference 67 | |
| dc.language.iso | en | |
| dc.title | 研究頭頸部鱗狀細胞癌HER3及NRG1之表達及調控機制 | zh_TW |
| dc.title | Analysis of expression status and regulators of HER3 and NRG1 in head and neck squamous cell carcinoma | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 107-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.coadvisor | 吳漢忠 | |
| dc.contributor.oralexamcommittee | 謝明書,蕭培文 | |
| dc.subject.keyword | 頭頸部鱗狀細胞癌,淋巴轉移,人類表皮生長因子受體3,神經調節蛋白1,NOTCH1,空間互斥分佈,泛素化, | zh_TW |
| dc.subject.keyword | head and neck squamous cell carcinoma,lymphatic metastasis,NOTCH1,HER3,NRG1,spatial mutually exclusive distribution,ubiquitination, | en |
| dc.relation.page | 81 | |
| dc.identifier.doi | 10.6342/NTU201901769 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2019-07-25 | |
| dc.contributor.author-college | 醫學院 | zh_TW |
| dc.contributor.author-dept | 口腔生物科學研究所 | zh_TW |
| dc.date.embargo-lift | 2029-07-31 | - |
| 顯示於系所單位： | 口腔生物科學研究所 | |
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