EP1/EP4在ras致大腸直腸癌癌化之角色探討

腫瘤的發展過程中，原型致癌基因的活化與抑癌基因的不活化,扮演了相當重要的角色。K-ras的突變在大腸直腸癌中佔有相當高的比率。先前的研究指出腸上皮細胞的癌化是透過前列腺素E2 (prostaglandin E2；PGE2) 接受器EP1/EP4及活化其下游的訊息傳遞路徑。但目前為止，並沒有研究確切指出K-ras的突變與PGE2 /EP 訊息傳遞路徑之間是否具有相關性及分子機制為何。我們在這篇論文中主要的目的是找出K-ras與PGE2 接受器之間的關聯性。我們將pcDNA3.1載體及pcDNA-K-ras轉殖到HT29大腸癌細胞中，以生長曲線及西方墨點法進行分析。根據我們的研究結果推測K-ras會活化PI3K/Akt路徑，促使GSK3磷酸化與COX-2的產生，故我們推測GSK3不具活性下，無法降解β-catenin，而β-catenin會進入細胞核內與TCF4轉錄因子結合，誘導COX-2的產生，進而分泌PGE2，而PGE2活化接受器EP1/EP4，EP1/EP4又開始活化PI3K/Akt訊息傳遞路徑，所以這個正迴饋路徑調控著大腸癌細胞的增殖。但我們發現由K-ras引發的這條路徑無法誘導MMP2的過量分泌，推測促使大腸癌細胞遷徙有可能透過其他路徑。我們的發現提供了在治療由K-ras突變所引起的大腸直腸癌一個新思考點；透過EP1/EP4抑制劑的使用對於將來大腸直腸癌的預防也是一個可以考量的選擇。

Recent advances in molecular genetics have revealed that multiple genetic alterations including activation of oncogenes and inactivation of tumor suppressor genes are required for tumor development and progression. K-ras is frequently mutated in colorectal cancer. Previous studies have shown that prostaglandin E2 (PGE2) is involved in intestinal carcinogenesis through its binding to the PGE2 receptor subtypes EP1 and EP4 and activation of downstream pathways. But the molecular mechanisms that link K-ras and PGE2 receptor are currently undefined.The aim of the present work was to study the association between K-ras and PGE2 receptor .We transfer pcDNA3.1 and pcDNA-K-ras in to HT29 colon cancer cells. The transfected cells and the control cells (empty vector) were analyzed by growth assay and immunoblot. Our data showed overexpression Ras protein led to an increase in cell proliferation with activation of the phosphatidylinosotol-3 Kinase (PI3K)/Akt pathway, an effect likely to be due to inhibits GSK3β(Glycogen synthase kinase 3β)activity. Inhibition of GSK3 stabilizes β-catenin (decrease its degradation) and promotes β-catenin nuclear translocation and transcriptional activation of TCF-regulated gene further induction of COX-2 and activation EP1/EP4. We collected colorectal tumor tissues as well as to confirm tumor tissues were overexpression Ras, PI3K, pAkt and EP1/EP4 protein. In this study, we also confirm in the H29 cells, K-ras does not affect MMP2 secretion. Our result provided colorectal cancer causes K-ras mutation which one newly thinks in the treatment. In addition, EP1/EP4 inhibitor use regarding the colorectal cancer prevention also is the choice which may consider.