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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 周綠蘋(Lu-Ping Chow) | |
dc.contributor.author | Chao-Chi Yeh | en |
dc.contributor.author | 葉昭圻 | zh_TW |
dc.date.accessioned | 2021-06-15T16:31:07Z | - |
dc.date.available | 2020-09-25 | |
dc.date.copyright | 2015-09-25 | |
dc.date.issued | 2015 | |
dc.date.submitted | 2015-08-13 | |
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/52861 | - |
dc.description.abstract | 以質譜分析為基礎的定量蛋白質體學目前已廣泛地應用在生物醫學或臨床研究中,運用其系統化地解析不同樣品間的表現量差異,找尋生物標識物或探討致病機轉。本篇論文中我們利用此技術分別研究肝癌治療之生物標識物及microRNA (miRNA) 於胃癌致病過程中參與的角色。 蕾莎瓦為目前晚期肝細胞癌病患的標準治療方式。不幸地,大部份病患治療過程中往往會產生抗藥性。因此可預測蕾莎瓦效用的生物標誌物顯得十分重要。為了找尋衍生蕾莎瓦抗藥性中的相關蛋白,我們應用 SILAC 定量蛋白體學方法分析比較人類肝癌細胞株 HuH-7 及具蕾莎瓦抗藥性的 HuH-7R 中有表現差異的蛋白,另外配合 iTRAQ 定量蛋白體學方法鑑定由 HuH-7 和 HuH-7R 細胞植入小鼠皮下產生腫瘤之差異蛋白。藉由上述兩種定量蛋白體方法,並利用軟體運算分析差異蛋白得知有 10 個蛋白與癌症中的細胞貼附,移動與侵襲有關。其中,當我們減少HuH-7R細胞中的半乳糖凝集素-1 (galectin-1) 表現量,觀察到細胞增生與轉移受到抑制,同時也能恢復抗藥性細胞對蕾莎瓦的感受性。我們證實了galectin-1為可預測蕾莎瓦抗藥性的生物標誌物,且 galectin-1 是經由 AKT/mTOR/ HIF-1 訊息傳遞途徑而產生。此外,肝癌病患血液中的 galectin-1 高表現量與較差的腫瘤控制率及低反應率有關。我們也發現血液中galectin-1 含量高與不良的無惡化存活率與總體存活期存在著獨立因素關聯。顯示 galectin-1 可當作肝癌病人接受蕾莎瓦治療期間是否產生抗藥性反應的生物標誌物。藉此我們可協助區分肝癌治療效果及提供直接的個人化治療。 本篇論文的第二部分著重於胃癌。過去研究已證實,幽門螺旋桿菌 (Helicobacter pylori) 的感染確實與胃癌發生有關,然而其詳細的致癌機轉仍有待釐清,尤其與微核醣核酸 (miRNAs) 之關聯性鮮少被討論過。miRNAs 為一內生性且不被轉譯成蛋白的小分子 RNA,能藉由與 mRNA基因的 3 端不轉譯區 (3’UTR) 結合抑制轉譯或造成其裂解,進而調控相關的基因表現。本實驗室發現在幽門螺旋桿菌感染後 miR-21 大量表現,為了釐清 miR-21 與幽門螺旋桿菌之致癌機制,我們應用 SILAC 定量蛋白體學方法分析比較高度表現 miR-21 之胃線癌上皮細胞株 AGS 及對照組中有表現差異的蛋白。利用 miRNAs 目標預測軟體 (miRsystem) 與其中表現量降低的蛋白交集得 47 個 miR-21 的潛力目標,同時使用生物資訊學分析交集之蛋白得知與癌症中的細胞增生,移動與細胞凋亡有關。我們也藉由細胞實驗證實了 miR-21 高度表現下,細胞增生與轉移能力增強且能抑制細胞凋亡。其中選取了6個潛力 miR-21 目標: DAXX, MAP3K1, NFAT5, PDCD4, RASA1 及 TP53BP2,未來將利用報導基因分析 (luciferase report assay) 確定是否為 miR-21 之目標基因,同時藉由基因抑制 (knockdown) 觀察其參與之功能。藉此釐清在幽門螺旋桿菌感染時,大量表現的 miR-21 抑制其下游基因表現,使得原有的抑癌功能遭受破壞進而導致胃癌的產生。 | zh_TW |
dc.description.abstract | Quantitative proteomics have been utilized widely in biomedical or clinical research. According the different expressed proteins in diverse samples, we could discovery the biomarker from diseases or investigate the tumorigenic mechanisms in different cancers. In this thesis, we utilized quantitative proteomics respectively to investigate the biomarkers for hepatocellular carcinoma (HCC) and the role of miRNAs in Helicobacter pylori-induced gastric cancer (GC). Sorafenib has become the standard therapy for patients with advanced HCC. Unfortunately, most patients eventually develop acquired resistance. Therefore, it is important to identify potential biomarkers that could predict the efficacy of sorafenib. To identify target proteins associated with the development of sorafenib resistance, we applied SILAC-based quantitative proteomic approach to analyze differences in protein expression levels between parental HuH-7 and sorafenib-acquired resistance HuH-7 (HuH-7R) cells in vitro, combined with an iTRAQ quantitative analysis of HuH-7 and HuH-7R tumors in vivo. In silico analyses of these differentially expressed proteins predicted that 10 proteins were related to cancer with involvements in cell adhesion, migration, and invasion. Knockdown of one of these candidate proteins, galectin-1, decreased cell proliferation and metastasis in HuH-7R cells and restored sensitivity to sorafenib. We verified galectin-1 as a predictive marker of sorafenib resistance and a downstream target of the AKT/mTOR/HIF-1α signaling pathway. In addition, increased galectin-1 expression in HCC patients’ serum was associated with poor tumor control and low response rate. We also found that a high serum galectin-1 level was an independent factor associated with poor progression-free survival and overall survival. In conclusion, these results suggest that galectin-1 is a possible biomarker for predicting the response of HCC patients to treatment with sorafenib. As such, it may assist in the stratification of HCC and help direct personalized therapy. In the second part of the studies, although the etiology of gastric carcinogenesis is thought to be multifactorial, Helicobacter pylori (H. pylori)-related gastric mucosal inflammation seems to be the most important trigger. The role of H. pylori on gastroduodenal diseases has been proposed, but the detailed molecular pathway remains unclear. MicroRNAs (miRNAs) are a class of naturally occurring small non-coding RNAs that regulate gene expression by targeting the 3’- untranslated region (3’-UTR) of mRNAs for translational repression or cleavage. Recently accumulating evidence suggests that microRNAs (miRNAs) such as miR-21 are aberrantly over-expressed in AGS cells caused by H. pylori infection. Furthermore, we investigated the potential targets of miR-21 and the relevant pathway triggered by miR-21 using SILAC-based proteomics approach. A total of 47 down-regulated proteins (fold change < 0.77) were identified by both SILAC and miRsystem approaches. In addition, analyzing the quantified proteins by SILAC-labeled with IPA, led us to focus on proteins that could play a relevant role in proliferation, cell apoptosis and metastasis. We also found that miR-21 could also remarkably increase cell proliferation and inhibit apoptosis. In addition, we showed that overexpression of miR-21 could dramatically increase cell invasion and migration in AGS cells. After merging three datasets, we found six potential targets, such as DAXX, MAP3K1, NFAT5, PDCD4, RASA1 and TP53BP2. In the future, we will validate those potential targets by luciferase reporter and functional assays to elucidate the possible role of miR-21 in the Helicobacter pylori-induced gastric carcinogenesis. | en |
dc.description.provenance | Made available in DSpace on 2021-06-15T16:31:07Z (GMT). No. of bitstreams: 1 ntu-104-D97442008-1.pdf: 19152389 bytes, checksum: 375f5c9545264ad89e065b9339b7cc44 (MD5) Previous issue date: 2015 | en |
dc.description.tableofcontents | 中文摘要 iv Abstract vi Chapter I – Overview and Rationale 2 1.1 Quantitative proteomic mass spectrometry 3 1.2 The strategies of discovering protein biomarkers and elaborating tumorigenic mechanism by using mass spectrometry 5 1.3 Quantitative proteomics in biomarker discovery 6 1.4 Quantitative proteomics for the identification of miRNA targets 8 1.5 Specific aims 11 Chapter II –Dual quantitative proteomic analysis identified galectin-1 as potential biomarker for predicting sorafenib resistance in hepatocellular carcinoma 13 2.1 Introduction 14 2.2 Experimental Procedures 24 2.3 Results 34 2.4 Discussion 40 Chapter III – Using quantitative proteomic approach to investigate the potential targets of microRNA-21 in the Helicobacter pylori-induced gastric carcinogenesis 45 3.1 Introduction 46 3.2 Experimental Procedures 51 3.3 Results 58 3.4 Discussion 62 Chapter IV – Conclusion and Perspectives 65 List of Tables 69 List of Figures 99 References 142 Appendix I 155 Appendix II 163 | |
dc.language.iso | zh-TW | |
dc.title | 利用定量蛋白質體學平台找尋預測肝癌產生蕾莎瓦抗藥性之生物標記物及 幽門螺旋桿菌誘導胃癌之microRNA-21標的蛋白 | zh_TW |
dc.title | Using quantitative proteomic analysis identifies the potential biomarker for predicting sorafenib resistance in liver cancer and microRNA-21 targets in H. pylori-induced gastric cancer | en |
dc.type | Thesis | |
dc.date.schoolyear | 103-2 | |
dc.description.degree | 博士 | |
dc.contributor.oralexamcommittee | 林中梧(Chung-Wu Lin),徐志宏(Chih-Hung Hsu),蔡孟勳(Mong-Hsun Tsai),游佳融(Chia-Jung Yu) | |
dc.subject.keyword | 蕾莎瓦抗藥性,肝細胞癌,細胞培養穩定同位素標記,等重相對標記絕對定量,半乳糖凝集素-1,AKT/mTOR/HIF-1α 訊息傳遞途徑,生物標誌物,胃癌,幽門螺旋桿菌,miR-21, | zh_TW |
dc.subject.keyword | sorafenib resistance,hepatocellular carcinoma,SILAC,iTRAQ,galectin-1,AKT/mTOR/HIF-1α signaling pathway,biomarker,gastric cancer,Helicobacter pylori,miR-21, | en |
dc.relation.page | 164 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2015-08-13 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 生物化學暨分子生物學研究所 | zh_TW |
顯示於系所單位: | 生物化學暨分子生物學科研究所 |
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