致癌蛋白Eps8可以促進大腸癌細胞之增生與移行。為了探究Eps8在人類子宮頸癌中所扮演的角色，我們由45位屬於早期子宮頸癌的患者檢體切入，以免疫組織染色的方式檢視Eps8的表現。與周圍正常的子宮頸上皮組織相比，Eps8的表現在癌化組織中有增加的情形，此現象亦在in vitro細胞培養的條件下得到證實。進一步Kaplan-Meier分析法得到的結果顯示Eps8表現較高的病患檢體，患者的預後較差。這個結果顯示Eps8在子宮頸癌中有促進惡化的作用，因此我們便以siRNA的方式，進行一系列的研究。Eps8表現減少的HeLa與SiHa子宮頸癌細胞，無論是在in vitro細胞增生或in vivo腫瘤生長上，都比控制組細胞來得慢。細胞週期的分析結果顯示Eps8表現降低會造成G1-phase arrest，而Eps8減弱導致的p53蛋白囤積，p21Waf 1／Cip 1表現上升，cyclins D1，D3，E的表現下降及Rb hyperphosphorylation的減少也解釋了G1-phase arrest的形成原因。值得注意的是，Eps8減弱導致的p53蛋白囤積，至少部份是由於p53蛋白半衰期增長所致。同時，我們也證實了Eps8有促進子宮頸癌細胞化療抗藥性的作用，而p53蛋白堆積的增加及Src與AKT活性的下降可能是造成Eps8減弱所致之抗藥性降低的原因。而將Eps8送回Eps8表現減低之HeLa細胞則可使上述這些因Eps8減弱所產生的變化（包括對於細胞週期、腫瘤生長及化療敏感度的作用）有一定程度的回復，顯示eps8 siRNA的作用是具有專一性的。在細胞移行方面，Eps8減弱會降低HeLa及SiHa細胞的移行能力，FAK活性及表現，MMP-9表現及basal ERK活性，而在送回Eps8的HeLa細胞中則發現這些變化有回復的情形。同時，以PD98059抑制ERK活化可抑制HeLa及SiHa細胞中MMP-9的表現。進一步以PD98059處理，可以在程度上有差別的降低HeLa，Eps8減弱及回送Eps8之HeLa細胞的移行能力。以專一性的MMP-9抑制劑處理也可觀察到同樣的情形。綜合以上結果，我們在子宮頸癌細胞中確立了Eps8可經由活化ERK而促進MMP-9的表現；同時，此條訊息傳遞路徑參與在部份Eps8所調控的子宮頸癌細胞移行能力中。最後，本論文的這些新發現讓我們對Eps8在人類子宮頸癌上所扮演的病理角色有較深入的了解。

The oncoprotein Eps8 promotes colon cancer proliferation and migration. In an attempt to elucidate the roles of Eps8 in human cervical cancer, we began the study by examining immunohistochemically Eps8 expression in specimens of 45 early-stage cervical cancer patients. Eps8 expression was increased in cervical carcinoma compared to neighboring normal cervical epithelia. In vitro studies also indicated that Eps8 expression was upregulated in cervical carcinoma cells. Kaplan-Meier analysis of these 45 patients demonstrated an inverse relationship between the level of Eps8 protein expression and percentage of patient survival. By virtue of small interference RNA (siRNA) methodology, reduced in vitro proliferation and in vivo tumor formation were observed in Eps8-attenuated HeLa and SiHa cervical cancer cells. Cell cycle analysis demonstrated that Eps8 attenuation resulted in G1-phase arrest, which might be attributed to increased p53 protein accumulation and p21Waf1/Cip1 protein expression, reduced cyclins D1, D3, E expression and a concomitant decrease in Rb hyperphosphorylation. It was worth noting that decreased p53 protein turnover rate contributed to increased p53 protein accumulation resulted from Eps8 attenuation. Furthermore, we demonstrated for the first time that increased Eps8 expression conferred better chemoresistance on cervical cancer cells. Reduced chemoresistance in Eps8-attenuated HeLa and SiHa cells might be attributed to increased p53 protein accumulation and decreased Src and AKT activity. Ectopic Eps8 expression in Eps8-attenuated HeLa cells could reverse the aforementioned biological events, indicating that the effect of eps8 siRNA was specific. With respect to cellular mobility, Eps8 attenuation decreased motility, FAK activity and expression in cervical cancer cells. MMP-9 expression and basal ERK activity were also reduced in Eps8-attenuated cells. Restored Eps8 expression could recover MMP-9 expression, FAK activity and expression, and migratory ability in these cells. Suppression of ERK activity by PD98059 in cervical cancer cells repressed MMP-9 expression. Furthermore, PD98059 and MMP-9 specific inhibitor could decrease mobility differentially in HeLa and its derived cells. Taken together, we established a signaling cascade in which Eps8 could enhance MMP-9 expression by promoting ERK activation, and this cascade contributed to Eps8-mediated cervical cancer cell mobility. These new findings deepen our understanding towards the pathological roles of Eps8 in human cervical cancer.

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