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標題: | 鑑定抑制MDA-MB-231遷移與侵襲能力之分子標的 Identification of cellular targets for inhibiting MDA-MB-231 migration and invasion |
作者: | Ming-Xian Yang 楊明憲 |
指導教授: | 梁博煌 |
關鍵字: | 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, cancer metastasis,small molecule inhibitor,target identification,Rab5c,cell migration,cell invasion, |
出版年 : | 2019 |
學位: | 碩士 |
摘要: | 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。 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. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/7342 |
DOI: | 10.6342/NTU201901077 |
全文授權: | 同意授權(全球公開) |
電子全文公開日期: | 2024-07-10 |
顯示於系所單位: | 生化科學研究所 |
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ntu-108-1.pdf 此日期後於網路公開 2024-07-10 | 2.12 MB | Adobe PDF |
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