臺灣博碩士論文加值系統

動情素受體(estrogen receptor, ER)、黃體素受體(progesterone receptor, PR)及第二型人類上皮生長受體(human epidermal growth factor receptor 2, HER2)在大多乳癌組織中過量表現，然而在三陰性乳癌(triple-negative breast cancer, TNBC)佔乳癌病患數量的15%，這三個生物標記表現皆為陰性。先前有研究指出鋅化合物Zn-3有潛力用於乳癌治療，然而細節相關機制尚未清楚。本論文旨在評估Zn-3對三陰性乳癌細胞(HCC1937和MDA-MB-468）以及非三陰性乳癌細胞(MCF7和SKBR3)的抗增生作用，並探討其相關機制。使用MTS分析，相較於正常乳腺細胞(M10)，Zn-3明顯抑制乳癌細胞的增生，表現出了選擇性毒殺的能力。使用流式細胞儀，Zn-3增加活性氧化物和粒線體超氧化物產生，並且降低粒線體膜電位，顯示了Zn-3在乳癌細胞誘導氧化壓力的結果。與正常細胞相比，Zn-3還在乳癌細胞中增加了subG1比例、細胞凋亡和DNA損傷。然而，上述Zn-3促使的變化都可以被抗氧化劑NAC (N-acetylcysteine)部分抑制，表明了Zn-3抗增生能力是基於增加細胞氧化壓力。總之，鋅化合物藥Zn-3傾向在三陰性乳癌及非三陰性乳癌表現的抗增生能力，與氧化壓力、細胞凋亡和DNA損傷有關。

Estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) are generally overexpressed in most breast cancer tissues. Triple-negative breast cancer (TNBC) shows negative for these three markers and accounts for 15% of breast cancer patients. A previous report showed that the zinc complex-derived drug Zn-3 has the potential to treat breast cancer; however, the detailed mechanism remains unclear. This thesis aims to evaluate the antiproliferation effects and explore the mechanism of Zn-3 against TNBC (HCC1937 and MDA-MB-468) and non-TNBC (MCF7 and SKBR3) cells. Using MTS assay, Zn-3 inhibited more proliferation on tested breast cancer cells than on normal breast (M10) cells, showing selective killing effects. Using flow cytometry, the generation of reactive oxygen and mitochondrial superoxide and the depletion of mitochondrial membrane potential were induced by Zn-3, showing oxidative stress-inducible results in breast cancer cells. Zn-3 also increased more subG1 populations in breast cancer cells than normal cells. Zn-3 preferentially triggered the apoptosis and DNA damage effects in breast cancer cells but not normal cells in terms of annexin V and 8-hydroxy-2-deoxyguanosine analyses. Moreover, all the above Zn-3-induced changes were partly inhibited by the antioxidant N-acetylcysteine, indicating that Zn-3 exerts oxidative stress effects on antiproliferation mechanisms. In conclusion, the zinc complex drug Zn-3 exhibits a promising preferential antiproliferation effect on both TNBC and non-TNBC cells, associated with oxidative stress, apoptosis, and DNA damage.

目錄
中文摘要 5
英文摘要 6
第一章、背景介紹
1.1. 乳癌研究與發展 7
1.2. 三陰性乳癌研究發展與治療 8
1.3. 鋅-化學藥物 9
1.4. 鋅化合物Zn-3 10
1.5. 活性氧化物ROS與氧化壓力 11
1.6. 細胞凋亡Apoptosis 13
第二章、研究動機與目的 14
第三章、實驗材料與方法
3.1. 鋅化合物Zn-3合成製備 16
3.2. 細胞培養 17
3.3. 藥物處理 22
3.4. 細胞存活率測定(MTS assay) 24
3.5. 細胞週期測定(Cell cycle assay) 26
3.6. 細胞凋亡測定(Apoptosis assay) 29
3.7. 細胞凋亡蛋白酶測定(Pancaspase assay) 32
3.8. 活性氧化物測定(ROS assay) 34
3.9. 粒線體中超氧化物表現量測定(MitoSOX assay) 36
3.10. 粒線體膜電位測定(MMP assay) 38
3.11. 氧化性DNA傷害測定(8-OHdG assay) 40
第四章、結果
4.1. Zn-3選擇性毒殺乳癌細胞和正常細胞 42
4.2. Zn-3誘導乳癌細胞週期變化 43
4.3. Zn-3誘導乳癌細胞凋亡 43
4.4. Zn-3造成乳癌細胞凋亡蛋白酶活性增加 44
4.5. Zn-3誘導乳癌細胞中ROS含量上升 44
4.6. Zn-3造成乳癌細胞粒線體超氧化物含量上升 45
4.7. Zn-3造成乳癌細胞粒線體膜電位下降 46
4.8. Zn-3造成乳癌細胞DNA氧化性損傷 46
第五章、討論
5.1. Zn-3與生物成分的潛在化學作用 47
5.2. Zn-3傾向毒殺乳癌細胞 48
5.3. Zn-3傾向在乳癌細胞中產生ROS 48
5.4. Zn-3促使乳癌細胞細胞周期改變並細胞凋亡 49
5.5. Zn-3促使乳癌細胞發生DNA損傷 49
5.6. Zn-3在乳癌細胞中抗增生能力與氧化壓力有關 50
第六章、結論 51
第七章、結果圖
4.1. Zn-3抑制乳癌細胞存活率 52
4.2. Zn-3影響乳癌細胞subG1 % 53
4.3. Zn-3影響乳癌細胞凋亡比例 55
4.4. Zn-3處理乳癌細胞凋亡蛋白酶(Pancaspase)分析 58
4.5. Zn-3影響乳癌細胞ROS含量 61
4.6. Zn-3影響乳癌細胞粒線體中的超氧化物含量 64
4.7. Zn-3影響乳癌細胞粒線體膜電位 67
4.8. Zn-3處理乳癌細胞的氧化性DNA損傷分析 70
第八章、文獻 73

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