Nlk经由Wnt通路调控EMT影响结直肠癌肝转移的分子机制研究

Epithelial-mesenchymal transitions (EMT) are implicated in the conversion of early stage tumors into invasive malignancies. The Wnt/β-catenin signaling pathway plays a pivotal role in EMT of human cancers. However, the regulatory mechanism of EMT in colorectal cancer (CRC) metastasis is still not yet clear. From previous results of 30 paired primary CRC and metastatic lesions, we found that the expression of nemo-like kinase (NLK) protein was significantly higher in the metastatic CRC lesions than in primary CRC tissues. Furthermore, we found that knockdown of NLK significantly hampered the growth and colony formatting ability of CRC cells, and inhibited tumor metastasis while overexpression of NLK changed the phenotype reversely. More importantly, down-regulation of NLK altered the cell morphology and expression of EMT markers. Theses function of NLK could be verified in migration model of nude mice. Based on our preliminary work, we will investigate the effect of NLK on the metastasis of CRC and the regulatory role of NLK-mediated Wnt signaling pathway in EMT process in CRC progression in the present study. Moreover, we will further explore the molecular mechanism by which NLK regulates the metastasis and EMT in CRC. Finally, we will perform a retrospective study on a cohort of patients with CRC to assess the correlation between NLK expression and the metastasis and recurrence of CRC and its clinical significance. Overall, this study could provide a new molecular target for the treatment of colorectal cancer liver metastases.

上皮间质转化（EMT）与肿瘤细胞侵袭转移关系密切，Wnt/β-catenin被认为是EMT效应中最重要的信号通路，但迄今对EMT在结直肠癌转移中的调控机制仍不够清楚。我们前期工作中在30例配对的大肠癌转移灶中发现Wnt/β-catenin信号通路中的关键分子NLK表达比原发灶显著升高，细胞实验证实NLK基因沉默后其生长显著减缓、克隆形成和转移能力降低，过表达该基因则表型相反。同时NLK能够介导细胞形态变化和EMT标志蛋白的表达改变。此外，我们在裸鼠转移模型中验证了下调NLK表达能够显著抑制结直肠癌肝转移。在此基础上，本项目拟进一步从分子、细胞、组织、动物水平多层次研究NLK与结直肠癌转移的功能和相关机制，深入探讨NLK基因调控EMT的相关信号通路和下游作用分子。并通过大样本组织芯片回顾性研究阐释NLK表达与结直肠癌临床病理与转移复发的相关性，明确其临床意义，为大肠癌肝转移提供新的治疗靶点。

摘要.既往研究证实nemo样激酶（NLK）在肿瘤的发生发展中起重要作用。有研究结果已经表明NLK过表达后肠癌细胞株中DLD-1生长活性下降，并可以诱导细胞凋亡，但是目前还没有报道系统的研究过在结直肠癌患者中NLK的表达情况。 NLK的表达是否是结直肠癌患者的局部复发或生存预后的因素还是未知的。为了探索NLK在CRC中预后的及在CRC的发生发展中的作用，我们通过免疫组化在一系列不同发展阶段的CRC （正常黏膜，腺瘤， I-IV期）患者中确定NLK表达。并将16例FAP患者和21例肠癌转移患者纳入研究。主要研究结论如下：.1、NLK的表达在正常黏膜、腺瘤、I~IV期肠癌的患者中逐渐升高。.2、NLK在FAP患者中正常黏膜、腺瘤、腺癌的表达逐渐升高。.3、NLK在肝转移的患者原发癌和转移癌中的表达高于正常黏膜，而原发癌与转移癌之间无差异。.4、NLK高表达与淋巴结转移、远处转移、高TNM分期、低分化及高复发率相关。 .5、NLK高表达的患者中OS 和DFS明显低于低表达的患者。.在我们的实验结论中， NLK蛋白的表达从正常黏膜腺瘤和Ⅳ期结直肠癌显着增加。我们还发现高表达NLK与预后差的临床变量包括淋巴结转移，高的TNM分期，分化程度差和较高的复发率显着相关。NLK可能是结直肠癌的一个癌基因,但是NLK参与肿瘤发生发展的机制还需要进一步深入和系统研究。