E2F1通过p53介导的自噬调控肺腺癌干细胞自我更新的机制研究

肿瘤干细胞参与了包括肺癌在内的人类癌症的各个方面，但其在癌症状态下自我更新的调控机制尚不明确。我们预实验发现，E2F1在肺腺癌干细胞中高表达，在表达野生型p53的肺腺癌干细胞中可调控自噬增强肺腺癌干细胞的干性。本项目以此为切入点，以分别表达p53野生型和p53缺失型的肺腺干细胞为研究对象；以E2F1调控自噬是否捕获p53泛素化降解，进而上调Nanog的表达促进肺腺癌干细胞干性为研究核心。通过建立E2F1，p53等基因shRNA或过表达质粒转染模型，采用RT-qPCR，western blot，自噬流检测和激光共聚焦显微镜等技术，探讨E2F1通过p53介导的自噬调控肺腺癌干细胞干性的分子机制，并在动物模型及临床样本上进行验证。以期回答E2F1可以通过调控自噬诱导p53的降解增强肺腺癌干细胞的干性，促进肺腺癌的恶性进展这一科学问题。为肺腺癌的临床诊断、治疗及预后提供新的理论构想和研究导向。

Accumulating evidence suggest cancer stem-like cells (CSCs), a group of cells with self-renewal capability, may play a role in initiation and progression of human cancers, including lung cancer. However, the cancer biology characteristics of CSCs have not been well elucidated due to the lack of stable cellular models of CSCs. Consequently, mechanisms that regulate CSC self-renewal are poorly defined. Our recent study showed the elevated expression of E2F1 in lung CSCs, which can enhance the self-renewal capability of lung CSCs with wild-type p53 through autophagy. In the present study, human lung CSC cell lines that has the wild-type p53 (A549) or in which p53 is deleted (H1229) as issue will be investigated to understand whether E2F1 augments the stemness of lung CSCs by degrading ubiquitinated p53 through autophagy, and Nanog is required for p53 regulation of CSC self-renewal. We would build up the cell models of shRNA/siRNA and overexpression about E2F1, p53 etc. genes, using RT-qPCR, western blot, autophagic flow detection and confocal system to explore the molecular mechanism of E2F1 regulating the self-renewal of lung adenocarcinoma stem cells by p53-mediated autophagy. We will also test in mouse model in vivo and clinical samples. We hope what we get from the study will answer this scientific question: E2F1 can enhance the self-renewal capability of lung CSCs by degrading p53 through autophagy, and then promote the malignant progression of lung adenocarcinoma. It provides a new theoretical conception and research direction for the clinical diagnosis, treatment and prognosis of lung adenocarcinoma.