2010-08-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/647234摘要：癌轉移是造成癌症病患治療失敗以及死亡的主要因素。在癌細胞轉移過程中，癌細胞必須具備侵襲以及移動的能力，另外要能夠阻擋細胞凋亡的刺激，才能成功地發生癌轉移。Slug 是屬於轉錄抑制因子Snail family 中的一員，在許多癌症中slug 表現量的異常增加與其惡性癌變及癌轉移有關。Slug 可以直接抑制癌細胞E-cadherin 基因轉錄，因而使癌細胞喪失細胞與細胞間的連結而導致癌細胞改變表皮細胞型態，進而提高侵襲及轉移的能力。Slug 除了在細胞侵襲與移動的過程中扮演重要角色外也被認為可以影響細胞存活。實驗證據顯示在某些白血病中，Slug 會受到E2A-HLF oncoprotein 的調控而啟動它的表現，並使得細胞的存活率增加。此外，Slug 亦可透過抑制puma 基因的表現而對抗p53 所調控的細胞凋亡，使得hematopoietic progenitors 在hematopoiesis 過程中可以存活下來。根據這些發現使我們推論Slug 賦予癌細胞侵襲及轉移的能力，以及幫助細胞抵抗細胞凋亡刺激的攻擊。然而Slug 調控癌細胞轉移及存活的分子機制至今仍不清楚。為了釐清Slug 在促進癌轉移的分子機轉，我們利用yeast two-hybridscreening 發現homeodomain-interacting protein kinase-2 (HIPK2)可與Slug 蛋白相互作用。初步研究證實在酵母菌及哺乳動物細胞中 Slug 可以與HIPK2 相互結合。我們也發現HIPK2 可能導致Slug 蛋白的磷酸化，而HIPK2 可能促使Slug蛋白質降解。本計畫將進一步探討HIPK2 如何調控Slug 蛋白的磷酸化及HIPK2如何促使Slug 蛋白質降解，以及探討Slug 發生磷酸化改變後如何改變細胞的侵襲、移動以及對抗細胞凋亡的分子作用機轉。首先我們將會在in vitro 及in vivo中證實Slug 可以被HIPK2 磷酸化並鑑定出HIPK2 磷酸化的位置。其次我們將會深入探討HIPK2 與Slug 間的互相調控在癌細胞轉移及存活中所扮演的角色並解析其作用機轉。最後我們將比較HIPK2 與Slug 的調控路徑在原位癌及惡性轉移腫瘤間的差別，以釐清HIPK2 與Slug 的調控路徑在癌轉移的重要性。<br> Abstract: Cancer metastasis is the most common cause of treatment failure anddeath in cancer patients. The successful metastatic cancer cells must have thecharacteristics of high invasion and migration capabilities and can preventapoptotic stimuli during the process of metastasis. Slug is a member of theSnail family transcriptional repressors and aberrant upregulation of Slug isassociated with malignant transformation and metastatic progression in avariety of cancers. In human cancer, the ability of Slug to repress theE-cadherin transcription has been implicated in pathological alterations of theepithelial cell phenotype to acquire the invasive properties. Forced expressionof Slug can lead to changes in cell morphology, the loss of cell-cell contacts,and the acquisition of invasive and migratory abilities. In addition to its role incell invasion and migration, Slug has also been involved in cell survival. Slug isabnormally upregulated by the E2A-HLF oncoprotein in certain leukemias andincreased cell survival. Furthermore, Slug can antagonize p53-mediatedapoptosis of hematopoietic progenitors by repressing puma duringhematopoiesis. These findings raise the possibilities that Slug may conferinvasive potential as well as anti-apoptotic ability on tumor cells. The molecularmechanisms by which Slug leads to the invasive growth of tumor cells are stillnot clear. To in depth explore the molecular mechanisms of Slug in cancermetastasis, we performed a yeast two-hybrid screening to identify novelSlug-interacting factors. We found a recently identified serine/threonine kinase,homeodomain-interacting protein kinase-2 (HIPK2) is a Slug-interactingprotein. In the preliminary studies, we confirmed the interaction of Slug withHIPK2 in vivo. In addition, we found that HIPK2 may function as a kinase forSlug phosphorylation and co-expression of HIPK2 may promote theproteasomal degradation of Slug. To study the roles of HIPK2 in Slugregulation and cancer progression, this proposed project will dissect themechanisms how HIPK2 regulates Slug phosphorylation and degradation. Wewill examine whether the phosphorylation of Slug causes cancer cellphenotype changes in invasion and apoptosis during the process of metastasis.Firstly, we will prove the phosphorylation of Slug by HIPK2 in vitro and in vivo,and identify the specific HIPK2-phosphorylated sites. Secondly, we willelucidate the HIPK2-Slug regulatory pathway in tumor cell invasion andsurvival. Finally, we will investigate the clinical significance of the HIPK2-Slugregulatory pathway in primary and advanced human cancers.