新分子LYRM2促进大肠癌干细胞特性的分子机制及其关键作用靶蛋白鉴定研究

The incidence and mortality of colorectal cancer(CRC) still rise.Its initiation,relapse and metastasis are believed to be closely related with the existance of cancer stem cell (CSC).Clarification of its molecular mechanisms in tumorigenesis and identification of crucial molecular targets are the basis and prerequisite for removing CSC. Our group firstly found that LYRM2 expression was upregulated in CD133+ colorectal cancer stem cells（CCSC）. Studies in vitro and in vivo further confirmed that overexpression of LYRM2 was associated with characteristics of CSC stemness such as sphere-forming,cell migration and tumorigenesis. These results suggest that LYRM2 may be a novel molecule promoting the stemness of CCSC, and a new potential target for CCSC-targeted therapy. However, the concrete molecular mechanisms and biological functions of LYRM2 are still unknown. Based on our previous acheivements, we firstly screen and identify key target proteins interacting with LYRM2 by tandem affinity chromatography and yeast two-hybrid assays ; secondly analyze the molecular mechanisms regulating LYRM2 and its partners ,and then clarify their biological functions and evaluate their clinical significance, finally explore the possibility of LYRM2 and its partners as potential markers in the targeted therapy of CCSC .This study will enrich the CCS theory and lay a solid foundation for the discovery and development of CSC-targeted drugs.

大肠癌（CRC）发病率与死亡率均呈上升趋势，其发生、复发转移等与大肠癌干细胞(CCSC)存在密切相关。阐明CCSC致瘤性的分子机制并鉴定关键分子靶点是靶向清除CCSC的基础和前提。我们前期研究首次发现CD133+ CCSC高表达LYRM2，体内外实验证实LYRM2高表达与CCSC发生及其成瘤性、细胞迁移、克隆球形成等干细胞生物学特征高度相关，提示LYRM2可能是促进CCSC干性的重要分子之一,是潜在靶点。但LYRM2促进这些干性特征的分子机制及其功能迄今未见文献报告。本课题拟在前期研究的基础上，通过串联亲和层析及酵母双杂交等技术筛选和鉴定LYRM2关键作用蛋白并解析其生物学功能，明确调控LYRM2及其关键靶蛋白的分子机制和相关信息通路，并分析其临床生物学意义，由此探讨 LYRM2及其蛋白在靶向CCSC治疗中作为潜在靶点的可能性,为丰富肿瘤干细胞理论及开发CSC靶向药物提供依据。

大量研究表明肿瘤干细胞与肿瘤的发生、转移密切相关。先前大肠癌的研究中，课题组筛选出LYRM2为新的肿瘤干细胞相关分子，本项目在先前研究基础上，1、建立大肠癌的组织标本库及临床随访数据库，并基于GEO上的芯片数据，分析结果示LYRM2在大肠癌、胃癌、肝癌、肺癌、乳腺癌等多种肿瘤中低表达；2、MSP结果示LYRM2在肿瘤组织中的低表达与启动子区甲基化相关；3、体内外实验结果示LYRM2能明显促进大肠癌克隆球形成、迁移、致瘤等恶性生物学表型； 4、串联亲和层析分析LYRM2 的相互作用蛋白，从中筛选出重要的相关分子并进一步予以验证；5、探讨了LYRM2的胞内定位，seahorse检测发现LYRM2可能调控肿瘤细胞的有氧呼吸和无氧糖酵解；6、通过Talen技术以及morpholino建立LYRM2敲除的斑马鱼模型，发现LYRM2未对正常肠道早期胚胎发育阶段有明显影响。通过本研究可为丰富肿瘤干细胞理论及开发CSC 靶向药物提供依据。也为大肠癌的诊断、治疗提供新标记物、靶点，具有较大的科学意义与临床转化应用价值。