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http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/7342完整後設資料紀錄
| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 梁博煌 | |
| dc.contributor.author | Ming-Xian Yang | en |
| dc.contributor.author | 楊明憲 | zh_TW |
| dc.date.accessioned | 2021-05-19T17:41:48Z | - |
| dc.date.available | 2024-07-10 | |
| dc.date.available | 2021-05-19T17:41:48Z | - |
| dc.date.copyright | 2019-07-10 | |
| dc.date.issued | 2019 | |
| dc.date.submitted | 2019-06-26 | |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/7342 | - |
| dc.description.abstract | 癌症轉移在腫瘤發展的過程中是個難以治療與預測的現象，現有療法仍無法有效防止癌症轉移。目前對於抑制癌症轉移的標靶性療法仍有所不足。我們研究團隊先前發現一個擁有iodoacetamide基團的共價抑制劑I-Trp能夠對β-tubulin 354號位置的半胱胺酸進行烷基化，因此破壞β-tubulin和CCT-β間的交互作用，更進一步誘導CCT-β過度表現的癌細胞進行細胞凋亡，其中包含一株三陰性乳癌細胞株MDA-MB-231，而此株乳癌細胞株於本研究中用於測試I-Trp對其轉移能力之影響。本研究發現，低於半抑制濃度的I-Trp便能抑制MDA-MB-231細胞遷移和侵襲能力約7至8成。為了找出I-Trp於癌細胞中影響轉移能力的分子標的，本研究中利用帶有螢光基團的I-Trp探針，標記細胞中I-Trp的標靶蛋白質，再表現與癌細胞轉移有關的蛋白質進行烷基化位點研究。實驗發現被標記的其中一個蛋白為Rab5c，而Rab5c於先前報導顯示有調控癌細胞移動的能力。接著，將Rab5c的半胱胺酸位點進行突變，並表現於MDA-MB-231乳癌細胞中，測試I-Trp對其抑制遷徙和侵襲能力。實驗發現，Rab5c 20號以及64號位置的半胱胺酸會被I-Trp所烷基化，此外，若將這兩個位置的半胱胺酸突變為絲胺酸時，I-Trp便會失去抑制MDA-MB-231細胞遷徙和侵襲能力的藥效。總結而論，Rab5c以及先前研究發現的β-tubulin是I-Trp於細胞中的分子標的，而前者遭I-Trp所標的會影響癌症轉移能力，後者遭I-Trp標的則會誘導癌細胞凋亡。 | zh_TW |
| dc.description.abstract | Metastasis is a formidable process during tumor progression, often leading existing therapies to failure. As a result, there has been an unmet need for targeted therapies to effectively inhibit cancer metastasis. An iodoacetamide-based covalent inhibitor, I-Trp, was previously demonstrated to be capable of alkylating Cys354 of β-tubulin to disrupt the protein-protein interaction between β-tubulin and CCT-β, thus inducing apoptosis of CCT-β overexpressed cancer cells, including MDA-MB-231, a TNBC, which was used to test I-Trp on inhibiting its metastasis. In this study, a sub-IC50 concentration of I-Trp was found to decrease the migration and invasion of MDA-MB-231 by 70-80%. To identify the potential target for I-Trp inhibiting cell migration and invasion, total cellular proteins were treated with an I-Trp-derived fluorescent probe. The proteins labeled by I-Trp were further expressed in MDA-MB-231 to study their alkylating sites of I-Trp. One of the proteins, Rab5c, which has been reported to involve in metastasis, was mutated at the alkylating sites and transfected into MDA-MB-231 to test its role in I-Trp inhibiting cell migration and invasion. Cys20 and Cys64 of Rab5c were found to be alkylated by I-Trp and the alkylation resulted in suppression of MDA-MB-231 migration and invasion. In conclusion, this study combined with our previous findings show that I-Trp targets β-tubulin and Rab5c, thereby inducing cancer cell apoptosis and inhibiting cancer cell migration and invasion. | en |
| dc.description.provenance | Made available in DSpace on 2021-05-19T17:41:48Z (GMT). No. of bitstreams: 1 ntu-108-R06b46015-1.pdf: 2174032 bytes, checksum: aaa355fa0200b606ed2e029dfabffb03 (MD5) Previous issue date: 2019 | en |
| dc.description.tableofcontents | 中文摘要 1
Abstract 2 1. Introduction 3 1.1 Cancer metastasis: reasons and facts 3 1.2 Chemotherapy and targeted therapies: pros and cons 3 1.3 Target identification 4 1.4 Rab5c: a player in focal adhesion disassembly and the link with tumor cell migration and invasion 5 1.5 Previous and the present studies 6 2. Materials and methods 8 2.1 Cell culture 8 2.2 Chemicals and antibodies 8 2.3 Immunoblotting 9 2.4 Wound healing assay 9 2.5 Transwell invasion assay 10 2.6 Mass spectrometric analysis 10 2.7 Cloning of rab5c, tubb and their derivatives 11 2.8 Expression and purification of Rab5c 11 2.9 Rab5c-GTP pulldown assay 12 2.10 Integrin internalization assay 13 2.11 Capture ELISA assay 13 2.12 Production of cDNA-expressing viruses 14 2.13 Statistical analysis 15 3. Results 16 3.1 I-Trp inhibited MDA-MB-231 migration and invasion 16 3.2 Identification of the potential target of I-Trp for inhibiting MDA-MB-231 migration and invasion 16 3.3 I-Trp suppressed Rab5c activity and Rab5c-mediated integrin β1 internalization 18 3.4 Determination of alkylating sites of I-Trp in Rab5c 19 3.5 Validation of alkylating sites of I-Trp in Rab5c in MDA-MB-231 19 4. Discussion 21 Tables 24 Figures 29 Reference 41 | |
| dc.language.iso | en | |
| dc.subject | 小分子抑制劑 | zh_TW |
| dc.subject | 細胞遷移 | zh_TW |
| dc.subject | 癌症轉移 | zh_TW |
| dc.subject | Rab5c | zh_TW |
| dc.subject | 分子標的鑑定 | zh_TW |
| dc.subject | 細胞侵襲 | zh_TW |
| dc.title | 鑑定抑制MDA-MB-231遷移與侵襲能力之分子標的 | zh_TW |
| dc.title | Identification of cellular targets for inhibiting MDA-MB-231 migration and invasion | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 107-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 吳世雄,林源峰 | |
| dc.subject.keyword | 癌症轉移,小分子抑制劑,分子標的鑑定,Rab5c,細胞遷移,細胞侵襲, | zh_TW |
| dc.subject.keyword | cancer metastasis,small molecule inhibitor,target identification,Rab5c,cell migration,cell invasion, | en |
| dc.relation.page | 45 | |
| dc.identifier.doi | 10.6342/NTU201901077 | |
| dc.rights.note | 同意授權(全球公開) | |
| dc.date.accepted | 2019-06-27 | |
| dc.contributor.author-college | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 生化科學研究所 | zh_TW |
| dc.date.embargo-lift | 2024-07-10 | - |
| 顯示於系所單位： | 生化科學研究所 | |
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