EMX2基因在结直肠癌中的临床意义及作用机理

发布时间：2018-05-17 23:24

  本文选题：EMX2基因 + 结直肠癌　； 参考：《北京协和医学院》2017年博士论文

【摘要】：·背景EMX2基因是同源框基因中的一员,在神经系统及泌尿生殖系统的胚胎发育中有广泛的作用。该基因在多癌种中均有表达调低,且表达抑制与预后不良或治疗耐药相关。该基因在肿瘤中突变较为少见,其表达降低主要是启动子区过甲基化的结果。EMX2有抑制肿瘤细胞增殖和侵袭的能力,可通过抑制经典Wnt通路发挥作用。本研究选择结直肠癌为研究对象,旨在探索EMX2基因在结直肠癌中的表观遗传学调控及其临床意义,并对该基因在结直肠癌中的功能及作用通路做进一步验证。·方法临床研究方面:研究对象为2012年于中国医学科学院肿瘤医院进行手术治疗的原发结直肠癌患者170例。应用甲基化特异性PCR(MSP)技术检测EMY2基因启动子区甲基化状态,并分析其与各项临床因素、预后及治疗的相关性。机制研究方面:检测该基因在结直肠癌细胞系中的表达情况,以及加入去甲基化试剂后表达水平的变化以明确其调控机制。在结直肠癌细胞系中过表达EMX2或用shRNA抑制其表达,检测细胞增殖和迁移能力的改变以及Wnt经典通路活性的改变,以明确该基因的功能和发挥作用的分子通路。·结果对1例正常结直肠组织和3例结直肠癌组织进行免疫组化染色的结果显示:EMX2基因在结直肠癌组织中有表达降低的趋势。对18例结直肠癌及相应正常组织的MSP结果显示:该基因在结直肠癌组织中有过甲基化的趋势(甲基化比例为83.3%vs 53.3%)。在170例结直肠癌标本中,61例(35.9%)EMX2基因甲基化阳性。甲基化阳性与年龄≥65岁(P=0.029)和KRAS基因突变(P=0.045)显著相关。预后方面,该基因甲基化阳性与总生存期的缩短有相关趋势,且甲基化阳性的患者倾向于出现远期复发和进展,对II期患者的单独分析中也显示类似的趋势。该基因甲基化阴性组靶向治疗可延长总生存期,阳性组治疗效果不明显。EMX2基因在结直肠癌细胞系中表达显著降低,且用去甲基化试剂处理后基因表达上调。过表达EMX2基因的HCT116细胞系增殖速度显著降低,而转染shRNA的低表达细胞系中细胞迁移能力有显著上调。此外,EMX2过表达细胞系中Wnt经典通路活性抑制,低表达细胞系中通路活性增强。·结论EMX2基因在结直肠癌中表达降低,主要是启动子区过甲基化的结果。该基因表达降低与总生存期缩短和病变的远期复发进展存在相关趋势,有作为II期肿瘤预后因子的可能性。该基因可抑制结直肠癌细胞的增殖和迁移,通过Wnt经典通路发挥作用,与EGFR等结直肠癌中常见致癌分子通路的关系有待进一步探索。
[Abstract]:Background EMX2 gene is a member of homobox gene and plays an important role in embryonic development of nervous system and genitourinary system. The expression of the gene was down-regulated in multiple carcinomas, and its inhibition was associated with poor prognosis or drug resistance. The decrease of the gene expression is mainly due to the hypermethylation of promoter region. EMX2 has the ability to inhibit the proliferation and invasion of tumor cells, which can play a role by inhibiting the classical Wnt pathway. The purpose of this study was to explore the epigenetic regulation of EMX2 gene in colorectal cancer and its clinical significance. Methods: 170 patients with primary colorectal cancer underwent surgical treatment in 2012 at the Cancer Hospital of the Chinese Academy of Medical Sciences. The methylation status of promoter region of EMY2 gene was detected by methylation specific PCR, and the correlation between methylation and clinical factors, prognosis and treatment was analyzed. Mechanism study: the expression of this gene in colorectal cancer cell line and the change of expression level after adding demethylation reagent were detected to clarify its regulatory mechanism. EMX2 was overexpressed or inhibited by shRNA in colorectal cancer cell lines. The changes of cell proliferation and migration ability and the activity of Wnt classic pathway were detected. Results Immunohistochemical staining of 1 normal colorectal tissue and 3 colorectal cancer tissues showed that the expression of the 1% EMX2 gene decreased in colorectal cancer tissues. The MSP results of 18 cases of colorectal cancer and the corresponding normal tissues showed that there was a tendency of hypermethylation of the gene in colorectal cancer tissues (83.3%vs 53.3%). EMX2 gene methylation was positive in 61 of 170 colorectal cancer specimens. There was a significant correlation between methylation positive and age 鈮，

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