CAFs源性Exosomal lncRNA-BX648637激活FoxM1/β-catenin通路参与胰腺癌干细胞干性维持的机制研究

Pancreatic cancer stem cell（PCSC）is the ‘seed cell’ for initiation, metastasis, and recurrence of pancreatic cancer. Though increasing evidence demonstrated a critical role of exosomes secreted by cancer associated fibroblasts (CAFs) in maintaining stemness of cancer stem cells, the underlying mechanism remains elusive. Our previous studies demonstrated that overexpression of FoxM1 promoted nuclear translocation of β-catenin and increased the stemness features of PCSC. lncRNA-BX648637 enriched in CAFs-derived exosomes is able to activate FoxM1/β-catenin signaling pathway, which is ceRNA mechanism dependent. Furthermore, a positive feedback loop was identified between FoxM1 and BX648637. Based on these results, we hypothesized that CAF-derived lncRNA-BX648637 can be delivered into PCSC through exosomes, leading to the ceRNA-dependent induction of FoxM1 and activation of a positive feedback loop, thus facilitating the nuclear translocation of β-catenin and ultimately, maintaining the stemness of PCSC. In this project, we will trace exosomes' role of signals transmission between CAFs and PCSCs through fluorescence labeling. Also we will try to determine the maintenance of PCSC’s stemness by exosomes and BX648637 through colony formation analysis and evaluation of markers for cancer stem cells. Furthermore, we will dissect the ceRNA mechanisms through which BX648637 upregulates FoxM1, as well as the transcriptional mechanisms through which FoxM1 upregulates BX648637. These two mechanisms form a positive feedback loop, which promotes the nuclear translocation of β-catenin, and hence, contributes to maintenance of PCSC stemness. Current project will propose a novel target for pancreatic cancer treatment in the future.

胰腺癌干细胞（PCSC）是胰腺癌成瘤、转移、复发的“种子细胞”。肿瘤相关成纤维细胞（CAF）源性外泌体参与PCSC干性调控，但具体机制未明。课题组前期研究发现：FoxM1/β-catenin通路参与PCSC干性调控；CAFs源性外泌体可通过BX648637激活FoxM1/β-catenin通路，该效应依赖于ceRNA机制；FoxM1与BX648637间可能存在正反馈调控环路。由此提出科学假说：CAFs源性外泌体携载BX648637进入PCSC，通过ceRNA机制上调FoxM1并与之形成正反馈调控环路，促进β-catenin入核，参与PCSC干性维持。本课题拟通过荧光标记示踪外泌体在CAFs与胰腺癌细胞间的信息传递过程；克隆形成、标志物检测等确定外泌体及BX648637对干性维持的诱导作用；信息学分析及验证确定BX648637与下游通路的具体作用机制。研究结果将为胰腺癌靶向治疗提供新思路。

胰腺导管腺癌（以下简称胰腺癌）恶性程度高、早期诊断困难、预后极差。胰腺癌干细胞（pancreatic cancer stem cell, PCSC）具有自我更新和多向分化潜能，靶向介导PCSC干性维持的相关通路具有重要的治疗价值。间质细胞（尤其是肿瘤相关成纤维细胞，CAF）在驯化胰腺癌细胞恶性表型，维持干细胞干性中发挥重要作用，但具体机制不明。外泌体（exosome）是细胞分泌的微囊泡，被新发现可介导细胞间的信息传递作用。通过与靶细胞相结合，exosome将核酸等物质传递给靶细胞，并调控其生物学功能。.本课题针对上述方向开展工作开展工作，发现：PCSC中过表达的FoxM1蛋白可通过促进β-catenin进核而维持其干细胞干性；CAFs源性exosome可通过激活FoxM1/β-catenin通路而提高PCSC的自我更新能力；lncRNA-BX648637在CAFs源性exosome中显著富集，并可以exosome为载体进入PCSC细胞，参与FoxM1/β-catenin通路的激活，上述过程依赖于ceRNA机制；FoxM1与BX648637间存在正反馈调控环路。由此，本研究形成如下科学结论：胰腺癌CAF源性的exosomes，可携载lncRNAs（BX648637）进入PCSC，通过miR-494介导的ceRNA机制上调FoxM1并形成正反馈调节回路，促进β-catenin入核，维持PCSC干性。课题组还进行了如下研究并形成结论：miR-10a-5p是CAF源性exosome中miRNome的主要组分，并介导维生素D对于CAF促癌表型的调控作用。KLF4介导的STK33过表达参与胃癌发生发展。PTTG3P/miR-132/212-3p/FoxM1构成正反馈环路，促进胰腺癌发生。上述研究围绕lncRNA、微环境等因素在肿瘤中的发病机制，为相关肿瘤的诊断及靶向治疗提供了新的靶标。