Kruppel样因子5介导VRK2超级增强子形成促进鼻咽癌凋亡逃逸和转移的机制研究

Super enhancers (SEs) are enhancers cluster with extremely high activity, which could activate the transcription of oncogenes, thus promote the occurrence and evolution of malignancies. The proposer has picked up the nasopharyngeal carcinoma (NPC)-specific SEs related gene, VRK2, through integrating the sequencing data. The preliminary experiments demonstrated that VRK2 could increase the anti-apoptosis, invasion and migration abilities of NPC cells in vitro; and the expression of VRK2 in NPC tissues correlated with the prognosis of patients; moreover, JASPAR database suggested that the Kruppel like factor 5 (KLF5) might bind to VRK2 SEs region, and knock down the expression of KLF5 resulted in the inhibition of VRK2 expression. Therefore, we speculated that KLF5 mediated the formation of VRK2 SEs, which further led to the enhanced abilities of anti-apoptosis and metastasis of NPC cells. However, the necessity of KLF5 in VRK2 SEs and the detailed mechanisms of KLF5 in promoting the transcription of VRK2 are undetermined. The project will further investigate the role of KLF5 in VRK2 SEs, deeply explore the detailed mechanisms of KLF5 in driving the expression of VRK2, and determine the crucial signal transduction pathway of VRK2 in promoting the anti-apoptosis and metastasis of NPC, as well as demonstrate the preventive and therapeutic effect of KLF5 inhibitor in anti-apoptosis and metastasis abilities of NPC at the levels of molecule, cell, animal and clinical samples, aiming to discover new target for precise treatment in NPC patients with advanced disease.

超级增强子（super enhancers,SEs）是极度活跃的增强子簇，可激活癌基因表达促使恶性肿瘤发生和进展。申请人通过数据整合筛选出鼻咽癌特异性SEs相关基因VRK2，体外研究证实VRK2促进鼻咽癌细胞凋亡逃逸、侵袭及迁移，与患者的预后相关；JASPAR数据库提示KLF5可能结合于VRK2 SEs区段，且敲减KLF5可下调VRK2的转录。由此，我们推测KLF5介导VRK2 SEs形成，从而促进鼻咽癌凋亡逃逸及转移。然而，KLF5对于VRK2 SEs的必要性及其调控VRK2转录的详细机理仍不明确。本课题拟在分子、细胞、动物及临床组织层面深入阐释KLF5在VRK2 SEs中的地位，全面解析KLF5驱动VRK2转录的具体机制，明确VRK2促鼻咽癌凋亡逃逸及转移的关键信号通路，并探讨KLF5抑制剂对鼻咽癌凋亡逃逸和转移的防治作用，旨在发现精准治疗晚期鼻咽癌的新靶点。

在本项目中，我们通过整合鼻咽癌细胞系、异体移植组织、永生化鼻咽上皮细胞株与口腔上皮细胞株的H3K27ac CHIP-seq结果、欧洲核酸数据库及本课题组测序获得的鼻咽癌组织全转录组数据，筛选出鼻咽癌特异性的超级增强子相关基因VRK2。临床大样本研究发现鼻咽癌组织中VRK2表达水平与T分期弱相关，鼻咽癌局部晚期患者原发灶高表达VRK2的比例高于局部早期患者，且鼻咽癌原发灶VRK2高表达组患者的5年总生存率（OS）低于VRK2低表达组的患者（p = 0.028，n = 229）。有意思的是，在 NPC 细胞株SUNE1 和 S26 中过表达 VRK2抑制鼻咽癌细胞体外增殖、迁移和侵袭能力。相似地，裸鼠皮下成瘤模型及尾静脉-肺转移模型中，过表达 VRK2的 S26细胞在裸鼠中皮下成瘤及肺转移能力均较对照组降低。分子生物实验探索发现，VRK2可能通过调节foxm5及ki67的转录表达调节鼻咽癌细胞的增殖、迁移、侵袭和转移能力。综上，临床相关研究提示VRK2高表达可能与患者的不良预后相关，然而细胞功能实验与动物实验发现VRK2可能通过调节干性及增殖相关基因调节鼻咽癌的增殖、迁移、侵袭和转移，临床预后研究与细胞功能研究有矛盾的地方，具体机制仍待进一步阐明。该项目有可能为鼻咽癌的内科治疗提供潜在的靶点。