携带Tspan8、CD44v6的肿瘤干细胞外泌体对胰腺癌侵袭性的机制研究

Cancer stem cells (CSC), a small subpopulation of cancer cells with capability of self-renewal, high differentiation potential and drug resistance, which are one of the major causes of metastasis and recurrence in pancreatic cancer (PaCa). Exosomes play an important role in the crosstalk between CSC and PaCa. Previously, we demonstrated that CD44v6 acts as a CSC marker in PaCa which playing an important role in premetastatic niche formation via exosomes. CD44v6-competent exosomes enhance the tumor cells migration and invasion, and promote distant metastasis in PaCa. In addition, the expression of Tspan8 also was downregulated in CD44v6-knockdown cell lines significantly. Therefore, we suppose that Tspan8 maybe feedback and impact CD44v6 expression, both of Tspan8 and CD44v6 jointly regulate PaCSC biological characteristics. We considered it important to control whether CD44v6/Tspan8 play as CSC biomarker in human PaCa, and promote tumor metastasis, finally, propose new therapeutic strategies for targeted therapy. So far, no evidences suggest CD44v6 or Tspan8 impacts the adhesion and cargoes of exosomes. In this program, based on the association between CD44v6/Tspan8 and PaCSC, to demonstrate the impact of CD44v6 or Tspan8 on the components of exosomes, and to predict the target miRNAs which play a role in the progression of PaCa.

肿瘤干细胞（CSC）是胰腺癌（PaCa）转移、复发的主要原因之一，同时外泌体（Exosomes）又在CSC与PaCa疾病进展中起着重要的“桥梁”作用。我们前期研究证实，CD44v6可作为PaCSC标志物，携带CD44v6的外泌体可增强肿瘤细胞的运动性及侵袭力，促进PaCa远处转移。此外，CD44v6基因缺失也会明显降低Tspan8的表达量，我们猜测也许Tspan8可以反向影响CD44v6表达，二者协同调节PaCSC生物特性，成为一个高效精准的CSC联动靶点，为PaCa精准靶向治疗找到重要突破口。既往尚未有关于CD44v6或Tspan8对外泌体黏附性及“运载物”调节、干扰的研究，而本课题将在确定CD44v6、Tspan8与PaCSC关系的基础上，进一步明确CD44v6、Tspan8对外泌体结构组分的影响，预测外泌体在调节PaCa进展中发挥作用的靶点miRNA。

背景：将肿瘤干细胞（CIC）外泌体（CIC-TEX）与非CIC（Non-CIC）共培养，可以对非CIC进行基因重编辑。但是信息传递和CIC标志物的参与方式仍存在争议。因此，在本研究中我们将详细阐述CD44v6和Tspan8 及其TEX对非 CIC 功能的影响。.方法：将非转移性CD44v6-和Tspan8-knockdown（kd）胰腺癌细胞用作Non-CIC。CIC-TEX与Non-CIC共培养后通过深度测序和功能分析，对Non-CIC的改变进行评估。 并通过裸鼠体内实验观察CIC-TEX处理期间的肿瘤进展变化。.结果：CIC-TEX共培养的CD44v6kd-Non-CIC通过深度测序发现信号传导、转运、转录和翻译中均存在明显的mRNA变化，改变的miRNA影响细胞代谢、信号传导和转录。CIC-TEX共培养诱导的Tspan8kd-Non-CIC变化主要依赖于CIC-TEX-Tspan8的特定靶向。CIC-TEX转递促进细胞凋亡抗性，并且在体外和体内显着促进非CIC的上皮间充质转化、迁移、侵袭和（淋巴）血管生成。更重要的是，CIC-TEX作为信息传递中枢，由CD44v6依赖性RTK，GPCR和整合素激活引发并涉及CD44v6辅助转录和RNA加工。因此，激酶抑制剂可阻碍CIC-TEX促进肿瘤的功能，主要为抗Tspan8抗体阻断CIC-TEX与肿瘤细胞的结合。.结论：关于CIC-TEX对CD44v6kd或Tspan8kd的非CIC的体外和体内影响的深入研究揭示了CIC-TEX的功能活性，强调了CIC 标志物CD44v6在信号级联激活、转录、翻译中的重要作用。那么，通过靶向抗CD44v6或Tspan8可阻断CIC-TEX结合/摄取，从而显著降低CIC-TEX对非CIC的影响作用。