XPC/Snail通路对肺癌干细胞亚群的影响及调控机制研究

Lung cancer is the leading cause of cancer-related deaths in the world. Lung cancer stem cells (LCSC) are the major reason for tumor relapse and metastasis which causes lung cancer patients’ mortality. Xeroderma pigmentosum group C (XPC) is a DNA repair factor mainly involving in the nucleotide excision repair pathway. The low XPC expression level in human lung adenocarcinomas is correlated with a poor outcome of patients, indicating that XPC may play a critical role in suppressing lung cancer progression. Our lab has demonstrated that XPC knockdown is able to increase the abundance of cancer stem cells (CSCs) defined as CD133+ in lung cancer cell lines, and to promote the generation of CD133+ cells from CD133- cell population. Furthermore, our preliminary study revealed that XPC is capable of repressing the expression of epithelial-to-mesenchymal transition (EMT) inducer Snail. Therefore, we hypothesize that XPC depletion in lung cancers promotes non-CSC-to-CSC conversion through enhancing the expression of Snail. The expanded CSC pool is responsible for the enhanced tumorigenic and metastatic potential of lung cancers characterized with low XPC expression. A series of in vitro and in vivo experiments are proposed to test our hypothesis. The elucidation of the significance of XPC in the regulation of the CSC population represents an opportunity to eradicate CSCs and improve the survival of lung cancer patients.

肺癌是全球致死率最高的恶性肿瘤之一。肺癌干细胞(LCSC)的存在是肺癌高复发转移的主要原因。人类着色性干皮病C组蛋白(XPC)的缺失或突变与肺癌的预后不良正相关。我们发现，下调XPC能促进CD133-肺癌细胞向CD133+转化，增强其致瘤能力；下调XPC还能增加上皮间皮转化关键蛋白Snail的表达。鉴于Snail在促进癌细胞干细胞化中的作用，推测下调XPC可通过增加Snail表达，促进非LCSC向LCSC转化，导致肺癌复发转移。本研究利用已建立的Tet-on和IPTG双调控XPC和Snail表达的稳定肺癌细胞株，体内外实验检测XPC和/或Snail对肺癌细胞干细胞化及LCSC数量、致瘤性和转移能力的影响；采用基因芯片、蛋白质组、染色质免疫共沉淀及色谱分析等方法，研究XPC调控Snail以及XPC/Snail信号调控LCSC亚群的分子机制，为靶向减少LCSC提高肺癌治疗水平提供理论依据。

本研究首先建立了肺癌组织芯片，检测XPC在肺癌组织和癌旁组织的差异性表达并检测干细胞标志物CD133、CD44、Oct4等在肺癌和癌旁组织中的表达及与XPC表达和患者预后的相关性。建立XPCsiRNA瞬时敲低和XPCshRNA稳定敲除的肺癌细胞株A549、H1299、H1650以及高表达pCDNA3.1XPC 的A549、H1299、H1650肺癌细胞株。在细胞水平上检测XPC表达对肺癌干细胞数量的影响，以及对STAT信号通路因子的影响。结果显示癌组织XPC低表达的患者预后较好，XPC表达与干细胞标志物CD133表达负相关。细胞学实验结果显示，下调XPC表达可以增加肺癌细胞中肿瘤干细胞标志物CD133，OCT4，SOX2的表达，并且XPC低表达可以增加STAT信号通路分子pSTAT1、pSTAT2和pSTAT3的表达。细胞划痕和细胞克隆实验证明，XPC表达降低可以促进肺癌细胞的迁移和增殖，进而导致肺癌的复发和转移。因此我们认为，XPC调控肺癌干性与STAT信号通路有关，这为临床早期筛查肺癌以及肺癌的治疗提理论依据。