胰腺导管腺癌来源的外泌体通过下调SIRT6活化CAF中转移相关超级增强子的作用与机制研究

Pancreatic cancer，a serious disease greatly threaten human health, was characterized by high metastasis rate which become leading cause of death. In this process， tumor associated fibroblasts（CAF） play a vital role, yet the mechanisms of CAF recruitment，differentiation and molecular modulation remain largely unknown. In the previous research in our lab，the gene expression profiling reveals pancreatic fibroblasts treated with tumor-derived exosomes performs gene expression pattern similar to CAF. This phenomenon is highly correlated with downregulation of SIRT6 which might promote epigenetic remodeling via its histone deacetylation activity. Next, H3K27ac ChIP – Seq was performed and eight SIRT6-dependent super-enhancers (SEs) were identified to be abnormal activation in tumor-derived exosome treated pancreatic fibroblasts. Therefore, we propose an research plan on mechanism of “downregulation of SIRT6 which mediated by pancreatic ductal adenocarcinoma exosomes promotes metastasis related super-enhancers activation in CAF”. In brief, The writer hopes to clarify and biological function of SIRT6 and SEs regulated genes via Chromatin Conformation Capture. CRISPRi and RNAi would be preformed to intervene SEs activation or genes expression to uncover the mechanism of CAF formation in the development of pancreatic cancer. In sum, this application aims to find novel theoretical basis and new therapeutic target of pancreatic cancer metastasis.

胰腺癌高转移率是其主要致死原因。在此过程中肿瘤相关成纤维细胞（CAF）发挥重要作用，而其分化招募途径与分子调控机制尚不明确。前期探索中通过表达谱芯片，申请人发现胰腺成纤维细胞与肿瘤来源的外泌体共培养后表现出与CAF相似的基因表达模式，并提示下调的SIRT6通过组蛋白去乙酰化活性介导的表观重塑影响了细胞命运。进一步通过ChIP-Seq检测细胞中H3K27ac的变化，发现外泌体处理胰腺成纤维细胞后有8个SIRT6相关的超级增强子（SEs）出现异常活化。基于此我们提出研究胰腺癌来源的外泌体通过下调SIRT6活化CAF中超级增强子介导基质重塑这一课题，拟从SEs活化角度阐明SIRT6的生物学功能，通过染色质构象捕获技术发现SEs所调控的基因，并使用CRISPRi干扰SEs或RNAi干扰特定基因，阐明胰腺癌发展过程中肿瘤相关成纤维细胞的分化形成机制，旨在为胰腺癌侵袭和转移提出新的理论依据和干预靶点。

胰腺癌是恶性程度极高的肿瘤，转移是其致死的主要原因，肝脏作为胰腺癌最易转移的靶器官，探究胰腺癌肝转移发生的机制是亟待解决的科学问题。本课题聚焦于胰腺癌来源外泌体促进胰腺癌恶性生物学行为尤其是胰腺癌肝转移形成。在课题进展过程中我们发现胰腺癌来源的外泌体中富含生物活性物质，并被肝脏中Kuffer细胞摄取。通过构建胰腺癌肝转移小鼠模型，分离纯化其中免疫细胞进行单细胞测序，系统性分析Kupffer细胞的表型及功能变化。发现肝转移微环境中特异性的Mertk阳性Kupffer细胞亚群群。机制表明胰腺癌外泌体能够激活肝脏Kupffer细胞中MAPK信号通路，并诱导MertkhiKupffer细胞亚群的产生，上调多种趋化因子进一步募集MDSCs到达肝组织，促进了胰腺癌肝转移前微环境形成。本研究发现并鉴定了胰腺癌肝转移过程中Mertkhigh Kupffer细胞新亚群的，证实胰腺癌来源外泌体能够诱导该亚群的产生，并探讨了该亚群在肝脏转移前微环境形成过程中的功能和机制，为胰腺癌肝转移的发生发展及早期干预提供了新思路。