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系統識別號 U0026-1208201510510900
論文名稱(中文) HIP1在非小細胞肺癌轉移過程中所扮演之角色
論文名稱(英文) The Roles of HIP1 in the Metastasis of Non-Small Cell Lung Cancer (NSCLC)
校院名稱 成功大學
系所名稱(中) 基礎醫學研究所
系所名稱(英) Institute of Basic Medical Sciences
學年度 103
學期 2
出版年 104
研究生(中文) 許哲裕
研究生(英文) Che-Yu Hsu
學號 S58971154
學位類別 博士
語文別 英文
論文頁數 115頁
口試委員 指導教授-呂佩融
召集委員-劉校生
口試委員-蕭宏昇
口試委員-陳玉玲
口試委員-王憶卿
口試委員-洪義人
口試委員-黃奇英
中文關鍵字 亨廷頓結合蛋白1  非小細胞肺癌  預後生物指標  癌症轉移抑制因子  Akt參與的上皮-間質轉型過程 
英文關鍵字 HIP1  NSCLC  prognostic biomarker  metastatic suppressor  Akt-mediated EMT 
學科別分類
中文摘要 非小細胞肺癌常因癌症轉移而使病人存活率極低,且癌症轉移的分子機制到目前也仍不清楚。亨廷頓結合蛋白1 (HIP1)已發現在多種癌症中均有大量表現,但其病理角色在癌症進展的過程中仍是未知。我們用免疫組織染色分析臨床肺部檢體中HIP1的表現量,發現HIP1在正常肺組織中表現量極低,在癌生成的早期有極高的表現量,且隨著肺癌越往後期進展,其表現量也逐漸下降,並伴隨有癌症轉移的發生。我們也證明了HIP1是非小細胞肺癌早期的預後生物指標,隨著其表現量降低而有極差的預後。接著我們的研究顯示,HIP1在非小細胞肺癌中是個癌症轉移抑制因子,無論是在動物活體實驗中或體外實驗操作上,均可有效抑制肺癌細胞的活動。此外我們亦證實,HIP1可透過阻斷AKT-GSK3訊息傳遞路徑,進而抑制上皮-間質轉型過程,以達到緩止癌症轉移。最後我們發現,HIP1可藉由與Akt結合而降低其活性,且推論HIP1將Akt滯留在細胞質中以阻止其到細胞膜上被活化。總結來說,HIP1是個非小細胞肺癌早期的預後生物指標,也是肺癌轉移抑制因子,其表現量的降低可透過Akt參與的上皮-間質轉型訊息傳遞路徑,在非小細胞肺癌進展過程中,有助於發生遠端癌症轉移,和導致極差的預後。我們的發現,可在HIP1所參與的癌症轉移上提供一個嶄新的看法,也可在非小細胞肺癌的治療過程中,發展出一套新穎的預後評估應用。
英文摘要 Non-small cell lung cancer (NSCLC) carries a poor survival rate mainly due to metastasis, and the molecular mechanism of metastasis remains unclear. Huntingtin interacting protein-1 (HIP1) is overexpressed in many human tumors, but its pathological roles in the progression of lung cancer are unknown. We analyzed HIP1 expressions in clinical lung specimens using immunohistochemistry (IHC). The results revealed that HIP1 expressions were very low in the normal lung tissues, dramatically increased during the early tumorigenesis, and decreased with NSCLC progression accompanied with metastasis. We also proved that HIP1 was a NSCLC early-stage prognostic biomarker with downregulated expressions for a poor prognosis. Subsequently, our studies showed that HIP1 was a metastatic suppressor in NSCLC to repress significantly the mobility of lung cancer cells both in vitro and in vivo. Besides, we demonstrated that HIP1 repressed the epithelial-mesenchymal transition (EMT) to inhibit metastasis through blocking the AKT-GSK3β signaling axis. Finally, we found that HIP1 interacted with Akt to decrease its activity, and proposed that HIP1 restrained Akt in the cytoplasm from its activation on the plasma membrane. In conclusion, HIP1 serves as an early-stage prognostic biomarker of NSCLC and a metastatic suppressor with reduced expressions during NSCLC progression to develop late metastasis and cause a poor prognosis through regulating the Akt-mediated EMT signaling pathway. Our findings may put the new insight of HIP1-mediated metastasis and lead to a novel application in prognostic evaluation of NSCLC treatment.
論文目次 Abstract............................................I
中文摘要 (Chinese Abstract)............................................II
誌謝 (Acknowledgement)............................................III
Index............................................IV
Table List............................................VI
Figure List............................................VII
Appendix List............................................X
Abbreviation List............................................XI

1. Introduction............................................1

2. The Objective and Specific Aims............................................6

3. Materials and Methods............................................7
3-1. Cell culture, animal model and reagents
3-2. Plasmid construction, DNA transfection and lentivirus transduction
3-3. RT-PCR, quantitative real-time PCR (qPCR) and microarray analysis
3-4. Western blotting (WB), subcellular fractionation analysis and coimmunoprecipitation (co-IP)
3-5. Tissue-array, immunohistochemistry (IHC), hematoxylin/eosin stain (HE stain) and immunocytochemistry (ICC)/immunofluorescence (IF)
3-6. MTT assay and luciferase-reporter assay
3-7. Wound-healing, migration and invasion assays
3-8. Statistical analysis
3-9. Primers and antibodies

4. Results............................................23
4-1. Lung cancer exhibits high Htt and HIP1 expressions.
4-2. HIP1 expressions decrease during NSCLC progression and represent an early-stage prognostic biomarker.
4-3. HIP1 represses the mobility of NSCLC cells in vitro.
4-4. HIP1 acts as a metastatic suppresser of NSCLC in vivo.
4-5. HIP1 inhibits the EMT of NSCLC.
4-6. HIP1 blocks the Akt-mediated EMT signaling pathway.
4-7. HIP1 restrains Akt in the cytoplasm from its activation on the plasma membrane.

5. Discussion............................................36

6. Conclusion............................................43

References............................................44

Tables and Figures............................................54

Appendix............................................101
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