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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
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dc.contributor.advisor | 俞松良(Song-Liang Yu) | |
dc.contributor.author | Jie-Ning Zhang | en |
dc.contributor.author | 章潔凝 | zh_TW |
dc.date.accessioned | 2021-07-10T22:01:11Z | - |
dc.date.available | 2021-07-10T22:01:11Z | - |
dc.date.copyright | 2019-03-11 | |
dc.date.issued | 2019 | |
dc.date.submitted | 2019-01-29 | |
dc.identifier.citation | 1.. Meyer T, Hart IR. Mechanisms of tumor metastasis. Eur. J. Cancer
1998, 34, 214–221. 2. Kent W Hunter, Nigel PS Crawford and Jude Alsarraj et al., Mechanisms of metastasis, Breast Cancer Research 2008, 10(Suppl 1):S2 3. Kevin J. Klos et al., Brain Metastases, The Neurologist 2004;10: 31–46. 4. Wang Chun Kwok, Terence Chi Chun Tam, Macy Mei Sze Lui, et al., Control of brain metastases with alectinib in anaplastic lymphoma kinase-rearranged lung cancer, Respirology Case Reports, 5 (3), 2017, e00224. 5 Alexander Chi 1, Ritsuko Komakii et al. , Treatment of Brain Metastasis from Lung Cancer, Cancers (Basel). 2010 Dec; 2(4): 2100–2137. 6. Shazia Ashraf, Heon Yung Gee,1Stephanie Woerner et al, ADCK4 mutations promote steroid-resistant nephrotic syndrome through CoQ10 biosynthesis disruption, J Clin Invest. 2013 Dec 2; 123(12): 5179–5189. 7. Tannishtha Reya, Sean J. Morrison, Michael F. Clarke & Irving L. Weissman et al.,Stem cells,cancer,and cancer stem cells [J].Nature,2001,414 (6859):105-111. 8 Shaheenah Dawood, Laura Austin, Massimo Cristo fanilli et al, Cancer stem cells: implications for cancer therapy[J].Oncology,2014,28(12):1101-1107,1110. 9. Marius Eugen Ciurea, Ada Maria Georgescu, Stefana Oana Purcaru,et al., Cancer stem cells: Biological functions and therapeutically targeting[J].Int J Mol Sci,2014,15(5):8169-8185. 10. Shadan Ali, Aamir Ahmad, Sanjeev Banerjee, et al. Gemcitabine sensitivity can be induced in pancreatic cancer cells through modulation of miR-200 and miR-21 expression by curcumin or its analogue CDF[J].Cancer Res,2010,70(9):3606-3617. 11. Ma Zhifang, Wei Liang, Zhang Wei, et al., The androgen receptor plays a suppressive role in epithelial- mesenchymal transition of human prostate cancer stem progenitor cells[J].BMC Biochem,2015,16(1):13-23. 12. Clarke MF1, Dick JE, Dirks PB et al, Cancer stem cells-perspectives on current status and future directions:AACR Workshop on Cancer Stem Cells[J].Cancer Res,2006,66(19):9339-9344 | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/77423 | - |
dc.description.abstract | 肺腺癌病人的存活期因為基因檢驗及新藥進步而延長,因此轉移癌的預防及治療更為重要。在肺腺癌好發的轉移中,顱內轉移對病患生活品質及壽命影響最嚴重,鑑定腦轉移有關的分子調控機制以協助評估病患腦轉移的風險、甚至選擇最佳的治療方式尤為重要。
在本研究中,我們利用慢性病毒(Lentivirus)將6000多組抑制人類Kinase及phosphatase的shRNA 基因組送入肺腺癌A549細胞中,使每一顆肺腺癌A549細胞隨機的帶有一種shRNA。將這群細胞由小鼠的內頸動脈注射,並於小鼠腦部長出腫瘤後取出腫瘤經培養種到下一批老鼠中,總計重複三次。接著將三次實驗的腫瘤以PCR與NGS定序的方式,得到各shRNA在腫瘤中所佔的比例。經Western及qPCR驗證A549中的ADCK4確實被抑制後,我們先進行了體外實驗,分別是Boyden chamber 癌細胞侵襲實驗 和 克隆形成實驗,均觀察到差異。 我們又在免疫缺陷鼠皮下注射該細胞,在皮下長成腫瘤後進行切除,模擬臨床上病人治療後的情況,觀察小鼠復發及轉移的情形。發現在shADCK4組別的腫瘤,除了生長速度較快,病理切片上可以看出分化較不完全,腫瘤細胞的核質比較高。在體外和體內的實驗中,都發現當A549細胞中的ADCK4被抑制後,與對照組相比有了更惡性的變化,這證明ADCK4靜默確實促進癌生成。另一方面,我們亦發現A549細胞被抑制掉ADCK4後,會擁有幹細胞特性,或許,ADCK4是透過影響肺癌細胞的幹性促進癌生成。 在之後的實驗中,我們選取不同的肺癌細胞,抑制掉ADCK4後進行相同的實驗,證明不僅僅是在A549細胞上才有此現象。最後,未來還需測量病人檢體上ADCK4的表現量高低,並與病人的轉移狀況與存活相比較,觀察ADCK4是否具有臨床應用價值。 | zh_TW |
dc.description.abstract | With the advances of gene testing and drug discovery, the survival of lung adenocarcinoma is significantly prolonged. The prevention and treatment of metastatic cancer become one of the most challenging subjects clinically. Brain metastasis occurs frequently in advanced lung adenocarcinoma, which affects patients’ life seriously. We would like to identify the underlying molecular mechanism of brain metastasis for new treatment development.
In this study, A549 cells were infected with a lentivirus library containing more than 6, 000 independent shRNA clones that target human kinases and phosphatases. After antibiotics selection, the resulting stable cell lines were injected into the brain of SCID mice through internal carotid artery. The tumor cells of metastatic loci were harvested from brain and cultured for next injection. The in vivo selections were repeated three times. Next, the shRNA clones of brain tumor lesions in the three times injections were amplified by PCR, and the proportion of individual shRNA in the tumor was calculated after NGS AmpliSeq. We found that ADCK4, a mitochondria protein, can inhibit cell invasion and tumorigenesis in vitro assessed by Boyden chamber invasion assay and colony formation assay. We confirmed the finding by an in vivo subcutaneous tumorigenesis assay. The tumor growth of A549 cells with shADCK4 was higher than the control, A549 cells with shLacZ. On the other hand, we found that knockdown of ADCK4 increases stemness in A549 cells. It is possible ADCK4 silencing increases tumorigenesis via stemness acquisition. In the following experiments, we selected different lung cancer cells and conducted the same experiment after inhibiting ADCK4, proving that this phenomenon was not only found on A549 cells. Finally, the further experiments should be performed to measure the level of ADCK4 expression on the patients’ lung cancer samples and to correlate with the patients’ metastasis status and survival, so as to observe whether it has the potential for clinical applications. | en |
dc.description.provenance | Made available in DSpace on 2021-07-10T22:01:11Z (GMT). No. of bitstreams: 1 ntu-108-R05424030-1.pdf: 1971281 bytes, checksum: 016c6c8af42c60fac4d94ce6b8336136 (MD5) Previous issue date: 2019 | en |
dc.description.tableofcontents | 致謝……………………………………………………………………………………II
中文摘要………………………………………………………………………………III 英文摘要………………………………………………………………………………IV 中文版…………………………………………………………………………………1 1. 緒論…………………………………………………………………………………1 1.1肺腺癌…………………………………………………………………………2 1.2 ADCK4…………………………………………………………………………2 1.3 輔酶Q10………………………………………………………………………3 2.實驗方法與材料……………………………………………………………………4 2.1 細胞培養………………………………………………………………………5 2.2腫瘤細胞侵犯力分析實驗……………………………………………………5 2.3細胞群落形成實驗……………………………………………………………5 2.4實驗動物………………………………………………………………………6 2.5 肺原位注射……………………………………………………………………6 2.6內頸動脈注射…………………………………………………………………7 2.7皮下注射………………………………………………………………………7 3.實驗結果……………………………………………………………………………9 3.1肺癌顱內轉移動物模型建立…………………………………………………10 3.2探討A549 shADCK4 細胞是否更容易轉移-皮下腫瘤形成試驗……………10 3.3探討A549 shADCK是否會增加細胞幹性…………………………………………11 3.4 A549 shADCK4更易在小鼠腦部形成腫瘤……………………………………12 3.5 A549 shADCK4的回復性實驗,並證明在其他細胞上也有相同的實驗結果…………………………………………………………………………………12 4.結論…………………………………………………………………………………13 5.討論…………………………………………………………………………………15 5.1肺癌顱內轉移動物模型的重要性………………………………………………16 5.2輔酶Q10的使用是否可以抑制腫瘤的生長及轉移?…………………………16 5.3 A549 shADCK4細胞,是否具有幹細胞特性?…………………………………16 6.圖片及表格…………………………………………………………………………18 圖片1肺癌顱內轉移動物模型建立…………………………………………………19 圖片2 A549中ADCK4 表現量的穩定性……………………………………………20 圖片3皮下腫瘤形成實驗結果分析…………………………………………………21 圖片4小鼠肺臟及腫瘤切片的HE 染色……………………………………………22 圖片5小鼠皮下腫瘤形成試驗………………………………………………………24 圖片6 A549 shADCK4 回復性試驗并在其他腫瘤細胞上進行ADCK4抑制………25 表格1皮下腫瘤形成實驗結果分析…………………………………………………26 表格2 小鼠皮下腫瘤形成試驗-細胞注射顆數……………………………………27 7.參考文獻……………………………………………………………………………28 英文版…………………………………………………………………………………31 1 Introduction……………………………………………………………………………31 1.1 Brain metastases of lung adenocarcinoma………………………………32 1.2 ADCK4……………………………………………………………………32 1.3 Coenzyme Q10…………………………………………………………33 2 Materials and Methods……………………………………………………………34 2.1 Cell culture…………………………………………………………………35 2.2 Cell invasion assay…………………………………………………………35 2.3 Soft agar(anchorage independent ) colony formation………………………35 2.4 Animal experiment……………………………………………………35 2.5 Lung orthotopic implantation……………………………………………36 2.6 Intracarotid artery injection………………………………………………36 2.7 Subcutaneousinjection……………………………………………………36 3 Results…………………………………………………………………………38 3.1 Candidate validation……………………………………………39 3.2 A549 shADCK4 are more easily to metastasis than A549 shLacZ in vivo…………………………………………………………………39 3.3 Stemness analysis in vivo…………………………………………………40 3.4 Intracarotid artery injection…………………………………………………………………………………41 3.5 ADCK4 restoration in A549 cells and knocwdown in other lung cancer cells…….……………………………………………………………………41 4 Conclusions………………………..……………………………………....………43 5 Discussion………………………………………………………………..……..…45 5.1 Importance of animal models of lung cancer brain metastasis ………………………………………………………………46 5.2 To investigate whether the use of coenzyme Q10 can inhibit tumor growth and metastasis……………………………………………………………………………………46 5.3 A549 shADCK4 cells possess stem cell properties?......................................46 6 Figures…………………………………………………………………………….48 6.1 Figure 1、Characterization of ADCK4 functions in vitro……….…….…49 6.2 Figure 2、The stability of ADCK4 in A549……………………….…..…51 6.3 Figure 3、Tumor of A549 shADCK4 growth faster than A549 shLacZ…………………………………………………………………52 6.4 Figure 4、Histological analysis of A549 shADCK4 cells by mouse subcutaneous implantation model………………………………………53 6.5 Figure 5、stemness of A549 shADCK4 cells assayed by the subcutaneous injection……………………………………………………………55 6.6 Fig6、Restoration and knocwdown of ADCK4 in other lung cancer cells………………………………………………………………………56 6.7 Table 1、The number of metastasis in A549 shADCK4 and A549 shLacZ……………………………………………………………………57 6.8 Table 2、Subcutaneous tumor formation experiment……………………58 7 References………………………………………………………………………59 | |
dc.language.iso | zh-TW | |
dc.title | ADCK4靜默透過增加幹性促進肺腺癌的生成 | zh_TW |
dc.title | Knockdown of ADCK4 enhances tumorigenesis via upregulating stemness in lung adenocarcinoma | en |
dc.type | Thesis | |
dc.date.schoolyear | 107-1 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 華國泰(Kuo-Tai Hua),蘇剛毅(Kang-Ii Su),李明學(Ming-Shyue Lee) | |
dc.subject.keyword | 非小細胞肺癌,腦轉移,ADCK4,腫瘤幹細胞, | zh_TW |
dc.subject.keyword | non-small cell lung cancer (NSCLC),brain metastasis,ADCK4,CSCs, | en |
dc.relation.page | 61 | |
dc.identifier.doi | 10.6342/NTU201900188 | |
dc.rights.note | 未授權 | |
dc.date.accepted | 2019-01-30 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 醫學檢驗暨生物技術學研究所 | zh_TW |
顯示於系所單位: | 醫學檢驗暨生物技術學系 |
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