Notch和PI3K/Akt信号通路间交互对话对结直肠癌干细胞的调控及其机制研究

We have previously found that the colorectal patients with Notch1 highly expressed have a poor prognosis, which has been published in Clinical cancer research and other journals. However, the underlying mechanism for why Notch1 is highly expressed and how the increased Notch1 contributes to cancer progression is largely unknown. Further exploring these underlying mechanism is of essential importance for cancer biology and the prevention and therapy of colon cancer. Previous study revealed that activation of PI3K/Akt could increase the Notch signal, and our ongoing experiments reveal that Notch could transcriptionally repress the expression of PTEN and phosphorylate PTEN, both of which could lead to the activation of Akt. All of these suggest that Notch and Akt signal pathways could form a positive feedback through their complicated crosstalk, which thus play an essential role in colorectal cancer development. The proposed project here first aims to reveal how Notch regulates Akt signaling, by examine the status of Akt and related molecules upon RNAi or overexpression Notch related genes. Direct repression of PTEN by Notch1 and the detailed mechanism will further confirmed by ChIP, EMSA, IP assay. Next, we will test the role of the positive feedback based on the recently proposed "cancer stem cell" theroy. We will test the effects of Notch and(or) PI3K/Akt intervention on the self-renewal ability, migration and invasion ability, and secretion of VEGF of the cancer stem cell or bulk cancer cells.With these experiments we would expect to reveal the role of the positive feedback on the stemness of cancer cell. Finally, we would like to examine both the molecule and cancer stem cell relevance of our study at clinical settings, through enroll of about 200 tissue samples. Our study will reveal new knowledge on the crosstalk between Notch and Akt, and we also shed light on the pathology and therapy of colon cancer.

课题组前期利用大规模临床样本发现Notch1高表达的结直肠癌患者预后差（Clin Cancer Res等），但具体机制不详。深入探讨Notch1高表达的机制及其对肿瘤演变的具体影响，对结肠癌防治策略具有重要意义。既往研究表明PI3K/Akt信号可增强Notch表达，而我们正在进行的研究发现Notch可通过转录抑制和磷酸化修饰两种方式抑制PTEN、活化Akt，提示Notch和Akt之间可通过复杂的交互对话，形成正反馈，参与肿瘤演变。本课题拟首先通过基因干预和EMSA、ChIP等方法阐明Notch和Akt之间交互对话的具体机制；在此基础上，结合"肿瘤干细胞是决定肿瘤预后的种子细胞" 这一肿瘤演变新理论，探讨Notch和Akt交互对话对结肠癌干细胞自我更新、迁移、促血管因子分泌等生物学行为的影响，明确Notch和Akt交互对话是否通过调节肿瘤干细胞行为促进肿瘤演变；最后结合临床样本验证上述机制。

Notch信号在肿瘤发生发展中起重要作用。本课题组前期研究证明Notch信号结直肠癌在起促癌作用，与肿瘤的分化、侵袭、转移以及预后密切相关。 课题组前期对Notch信号途径的研究发现，Notch1可通过与PI3K/AKT通路形成正反馈环相互促进参与结直肠癌的发生发展，而Notch 1对PI3K/AKT的正调控很有可能通过转录抑制和磷酸化修饰PTEN两种方式进行。本课题在大样本临床标本中检测了Notch1、PTEN、p-PTEN、PI3K/Akt通路分子的表达及相关性，分析了各分子与肿瘤进展及预后的关系；利用腺病毒技术在细胞中改变了Notch1表达后检测PTEN、p-PTEN、PI3K/Akt通路分子的表达改变。初步证明了Notch1可通过与PI3K/AKT形成正反馈环路调控结直肠癌的发生和发展。