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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 吳嘉文 | zh_TW |
| dc.contributor.advisor | Kevin C.-W. Wu | en |
| dc.contributor.author | 鍾憶慈 | zh_TW |
| dc.contributor.author | Yi-Tzu Chung | en |
| dc.date.accessioned | 2025-07-30T16:10:01Z | - |
| dc.date.available | 2025-07-31 | - |
| dc.date.copyright | 2025-07-30 | - |
| dc.date.issued | 2025 | - |
| dc.date.submitted | 2025-07-23 | - |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/98162 | - |
| dc.description.abstract | 乳癌為全球女性最常見的癌症類型，在台灣亦長年位居女性癌症死因之首，其中三陰性乳癌（Triple-negative breast cancer, TNBC）由於缺乏雌激素受體、黃體素受體與HER2表現的侵略性乳癌亞型，治療困難且預後較差。目前臨床主要依賴化學治療，其中以紫杉醇（Paclitaxel）為臨床上最廣泛使用之藥物，然而，其神經毒性的副作用嚴重且容易產生抗藥性。
本研究選用三陰性乳癌細胞株MDA-MB-231進行紫杉醇處理，並透過微流體染色質免疫共沉澱定序技術（MOWChIP-seq）分析給藥前後細胞中與活性增強子相關的H3K27ac組蛋白修飾。MOWChIP技術具備僅需微量樣本及高靈敏度捕捉優勢，能有效減少細胞使用量並提升免疫共沉澱效率。藉由比較紫杉醇處理前後的H3K27ac表觀遺傳圖譜，預期可鑑別出與藥物反應相關之基因調控變化。 本研究旨在探索Paclitaxel於三陰性乳癌細胞中引發的基因調控之變化，揭示其可能的表觀遺傳機制。可作為未來發展低毒性、具標靶性的表觀遺傳治療藥物之參考依據，提升治療三陰性乳癌的成效與安全性。 | zh_TW |
| dc.description.abstract | Breast cancer is the most common type of cancer among women worldwide and has long been the leading cause of cancer-related deaths among women in Taiwan. Among its subtypes, triple-negative breast cancer (TNBC), characterized by the absence of estrogen receptor (ER), progesterone receptor (PR), and HER2 expression, is highly aggressive, difficult to treat, and associated with poor prognosis. Chemotherapy remains the primary clinical treatment, with Paclitaxel being one of the most widely used drugs. However, its application is often limited by severe neurotoxic side effects and the development of drug resistance.
In this study, the TNBC cell line MDA-MB-231 was treated with Paclitaxel, followed by epigenomic profiling using microfluidic oscillatory washing–based chromatin immunoprecipitation followed by sequencing (MOWChIP-seq) to examine changes in H3K27ac histone modifications associated with active enhancers before and after treatment. MOWChIP-seq enables highly sensitive ChIP-seq analysis with minimal cell input, improving enrichment efficiency while reducing sample consumption. By comparing genome-wide H3K27ac profiles before and after Paclitaxel treatment, we aim to identify transcriptional regulatory changes in response to the drug. This study seeks to explore the gene regulatory alterations induced by Paclitaxel in TNBC cells and to uncover the potential epigenetic mechanisms underlying its action. Our findings may provide a valuable reference for the development of novel epigenetic therapies with lower toxicity and greater specificity, thereby improving the safety and effectiveness of TNBC treatment. | en |
| dc.description.provenance | Submitted by admin ntu (admin@lib.ntu.edu.tw) on 2025-07-30T16:10:01Z No. of bitstreams: 0 | en |
| dc.description.provenance | Made available in DSpace on 2025-07-30T16:10:01Z (GMT). No. of bitstreams: 0 | en |
| dc.description.tableofcontents | 誌謝 i
摘要 ii Abstract iii Table of Contents v List of Figures ix List of Tables xii 1. Introduction 1 1.1. Breast Cancer and Triple-Negative Breast Cancer (TNBC) 1 1.2. Paclitaxel 4 1.3. Epigenetics 5 1.4. ChIP and MOWChIP 10 1.5. Sequencing 15 2. Objective 19 3. Experimental 21 3.1. Cell Line 21 3.2. Chemicals 22 3.2.1. Cell Culture 22 3.2.2. IC₅₀ Determination Using the CCK-8 Assay 23 3.2.3. Fabrication of The Microfluidic Device 23 3.2.4. Preparation of Buffers and Reagents 24 3.2.5. Coupling an Antibody to Superparamagnetic Beads 25 3.2.6. Chromatin Preparation by Lysis and MNase Digestion 25 3.2.7. Purification and Concentration of DNA 26 3.2.8. Enrichment Check Using qPCR 26 3.2.9. Library Preparation 26 3.3. Equipment 27 3.4. Methods 29 3.4.1. Cell Thawing 29 3.4.2. Cell Subculture 29 3.4.3. Cell Cryopreservation 31 3.4.4. Determination of the 72 h IC₅₀ Value of Paclitaxel in MDA-MB-231 Cells Using the CCK-8 Assay 32 3.4.5. Fabrication of The Microfluidic Device 33 3.4.6. Preparation of Buffers and Reagents 36 3.4.7. Coupling an Antibody to Superparamagnetic Beads 39 3.4.8. Chromatin Preparation by Lysis and MNase Digestion 40 3.4.9. MOWChIP 44 3.4.10. Oscillatory Washing 47 3.4.11. ChIP DNA isolation 50 3.4.12. Purification and Concentration of DNA 50 3.4.13. Primer Design 52 3.4.14. Enrichment check using qPCR 54 3.4.15. Library preparation 55 4. Results and Discussion 56 4.1. Determination of the 72 h IC₅₀ Value of Paclitaxel in MDA-MB-231 Cells Using the CCK-8 Assay 56 4.2. Primer Design 62 4.3. Enrichment Check Using qPCR 65 4.4. Data Quality Evaluation: Correlation Heatmap and PCA 72 4.5. Genomic Distribution of Differentially Enriched H3K27ac Peaks 76 4.6. Downregulated H3K27ac-Associated Genes Reveal Epigenetics Modulation by Paclitaxel 78 4.7. Upregulated H3K27ac-Associated Genes Reveal Epigenetic Activation by 7-Day Paclitaxel Treatment 86 5. Conclusion 93 6. Future Work 95 References 96 | - |
| dc.language.iso | en | - |
| dc.subject | 三陰性乳癌 | zh_TW |
| dc.subject | 紫杉醇 | zh_TW |
| dc.subject | 表觀遺傳學 | zh_TW |
| dc.subject | 組蛋白修飾 | zh_TW |
| dc.subject | H3K27ac | zh_TW |
| dc.subject | 染色質免疫共沉澱 | zh_TW |
| dc.subject | 微流體 | zh_TW |
| dc.subject | 次世代定序 | zh_TW |
| dc.subject | Next-generation sequencing (NGS) | en |
| dc.subject | Triple-negative breast cancer (TNBC) | en |
| dc.subject | Microfluidics | en |
| dc.subject | Paclitaxel | en |
| dc.subject | Epigenetics | en |
| dc.subject | H3K27ac | en |
| dc.subject | Chromatin immunoprecipitation (ChIP) | en |
| dc.subject | MOWChIP-seq | en |
| dc.title | 利用低樣本高通量 MOWChIP 系統剖析紫杉醇對三陰性乳癌治療中組蛋白修飾的表觀遺傳機制 | zh_TW |
| dc.title | Profiling the Epigenetic Mechanisms of Histone Modifications in Triple-Negative Breast Cancer Treatment with Paclitaxel Using a Low-Input, High-Throughput MOWChIP System | en |
| dc.type | Thesis | - |
| dc.date.schoolyear | 113-2 | - |
| dc.description.degree | 碩士 | - |
| dc.contributor.oralexamcommittee | 葉禮賢;謝元榜 | zh_TW |
| dc.contributor.oralexamcommittee | Li-Hsien Yeh;Yuan-Pang Hsieh | en |
| dc.subject.keyword | 三陰性乳癌,紫杉醇,表觀遺傳學,組蛋白修飾,H3K27ac,染色質免疫共沉澱,微流體,次世代定序, | zh_TW |
| dc.subject.keyword | Triple-negative breast cancer (TNBC),Paclitaxel,Epigenetics,H3K27ac,Chromatin immunoprecipitation (ChIP),MOWChIP-seq,Microfluidics,Next-generation sequencing (NGS), | en |
| dc.relation.page | 105 | - |
| dc.identifier.doi | 10.6342/NTU202502293 | - |
| dc.rights.note | 同意授權(全球公開) | - |
| dc.date.accepted | 2025-07-25 | - |
| dc.contributor.author-college | 工學院 | - |
| dc.contributor.author-dept | 化學工程學系 | - |
| dc.date.embargo-lift | 2025-07-31 | - |
| 顯示於系所單位： | 化學工程學系 | |
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