YAP激活Notch通路诱导肿瘤干细胞形成进而参与食管癌放射抵抗

Radiation resistance is one of the major causes for esophageal cancer local relapse or metastasis after radiotherapy. According to the literature, cancer stem cells play a critical role in radiation resistance. Hippo-YAP signaling pathway promotes esophageal cancer progression and metastasis; however, whether YAP contributes to cancer stem cell self-renew and further participates in radiation resistance of esophageal cancer is relatively unknown. Our preliminary data showed that up-regulation of YAP in esophageal cancer cells after radiation induced the self-renew of cancer stem cells, and Notch signaling pathway was dramatically activated. Based on these data and relevant publications, we speculate that the major downstream effector of Hippo signaling pathway—YAP may induce the self-renew of tumor stem cells through Notch signaling pathway and further contributes to esophageal cancer radiation resistance; inhibition of YAP may increase the radiation sensitivity of esophageal cancer. In this project, we for the first time systemically study how YAP induces radiation resistance of esophageal cancer and the underlying signaling pathways involved in this process, especially the Notch signaling. In addition, we will try to explore the potential possibility and benefits of combining radiotherapy and YAP inhibitors for esophageal cancer treatment. Results of this project will help to further reveal the role of signaling pathways in esophageal cancer radiation resistance, enrich the theoretical basis for the participation of cancer stem cells in radiation resistance and provide novel targets for improving the radiation sensitivity of esophageal cancer.

放射抵抗是导致食管癌放射治疗后局部复发和远处转移的重要因素。肿瘤干细胞在食管癌放射抵抗中发挥关键作用。Hippo-YAP信号通路可促进食管癌的发生与进展，然而该通路是否参与肿瘤干细胞形成及食管癌放射抵抗尚不清楚。我们预实验发现放射后YAP增高导致食管癌中肿瘤干细胞标志物表达升高，且Notch信号通路显著激活。基于预实验及文献报道提出假说：YAP通过调控Notch通路诱导肿瘤干细胞形成，从而参与食管癌放射抵抗；降低YAP水平或活性可提高食管癌放射敏感性。本课题拟从细胞及分子生物学、组织病理学、临床资料等多层次系统研究YAP如何诱导肿瘤干细胞形成进而参与食管癌放射抵抗，阐明其上下游分子机制，进而探讨联合放射治疗及YAP抑制剂治疗食管癌的可能性。本课题将拓宽信号通路参与放射抵抗的视野、丰富肿瘤干细胞促进放射抵抗的理论，有助于为食管癌放射增敏提供新的靶点。

食管癌是我国最常见的恶性肿瘤之一。最新的流行病学资料显示，我国食管癌年发病人数为 47.79 万人，居各类恶性肿瘤第 3 位，年死亡人数为 37.5 万人，居第 4 位。我国食管癌患者 95%的病理类型为鳞癌，由于早期症状不典型和缺乏早期诊断的特异性肿瘤标志物，多数患者诊断为晚期，不能积极治疗，预后差，5年生存率仅为15%-25%。放射治疗是食管鳞癌的主要治疗方式之一，但50%-60%患者由于局部复发或未控而导致放疗失败，放射抵抗是肿瘤局部复发或未控的重要因素，是目前食管癌综合治疗亟待解决的问题。. 食管鳞癌患者手术标本的检测发现，YAP与患者无病生存期和总生存期呈现明显负相关，提示YAP表达或是患者不良预后的一个重要因素；放疗上调YAP并促进肿瘤干细胞的富集/自我更新以及Notch 通路的激活；机制研究中，敲减食管鳞癌细胞中的YAP能够通过抑制EMT水平来降低肿瘤细胞的迁移和侵袭能力。. Hippo信号通路下游，我们研究了Hippo信号通路的最终效应因子TAZ，在食管鳞癌的病理标本中，TAZ的过表达预示预后不良和不良的病理特征；细胞和动物模型中均证实，TAZ促进细胞增殖、迁移和放射抵抗。机制研究发现，TAZ通过促进非同源末端连接(NHEJ)相关基因的表达来诱导放射抵抗；食管癌TAZ基因的CpG岛在胸腺嘧啶DNA糖基化酶(TDG)的调控下发生去甲基化，从而促进食管癌生长、侵袭、放射抵抗等表型；由此证实了TDG/TAZ/NHEJ轴在食管癌进展和放射抵抗中的重要作用，为放疗增敏提供了潜在靶点。. Hippo信号通路上游，我们发现了一条YAP调控的新机制，发现昼夜节律相关蛋白RAI1可以作为转录因子上调Hippo通路上游分子NF2、LLGL1、FRMD6来激活Hippo通路，进而促进YAP和TAZ分子的磷酸化，导致YAP和TAZ入核减少，从而抑制肿瘤；m6A相关去甲基化酶ALKBH5可以上调RAI1表达，并通过体内外实验充分验证了ALKBH5/RAI1/Hippo/YAP调控轴线；YAP特异性抑制剂维替泊芬（VP）的体内外实验结果显示，ALKBH5低表达的食管癌细胞系和小鼠皮下肿瘤模型对于VP更敏感，说明ALKBH5通过YAP发挥肿瘤调控作用，提示YAP或是ALKBH5低表达食管癌患者的潜在新型治疗靶点。