MTA1蛋白在食管鳞癌血管生成过程中的作用机制研究

Squamous cell cancer has been the most common histological type among primary esophageal cancer in China, despite major advances have been made in the diagnosis and therapy, the 5-year survival rate of patients are still dismal. Invasion and metastasis are the leading cause that results in treatment failure and tumor-related death. Angiogenesis plays vitally important roles in the process of invasion and metastasis of esophageal squamous cell cancer, therefore, investigation the molecular mechanisms of angiogenesis is expected to bring a breakthrough for the treatment of esophageal squamous cell cancer. On the basis of the available data, it is very likely that MTA1 protein plays important and critical roles in the angiogenesis of a wide variety of human malignant tumors, MTA1 protein could regulate tumor angiogenesis by means of regulating its downstream molecules, such as VEGF-A. Indicating that MTA1 protein might be a potentially key molecule involving in the angiogenesis of malignant tumors, MTA1 protein has been anchored as a new therapeutic target for anti-tumor angiogenesis. Our initial research confirmed that MTA1 protein high expression was common in esophageal squamous cell cancer tissues for the first time, and MTA1 protein high expression was closely associated with increased tumor angiogenesis. However, the specific signal control mechanisms of MTA1 protein in angiogenesis of esophageal squamous cell cancer have not yet well been addressed. In the present study, we will thoroughly and systematically investigate the specific molecular mechanisms of MTA1 protein regulating the angiogenesis of esophageal squamous cell cancer through in vivo and in vitro experiments, so as to provide new ideas and theoretical basis for the clinically targeted therapy of esophageal squamous cell cancer.

鳞癌是我国原发性食管癌中最常见的组织学类型，患者的5年生存率仍处于较低的水平。肿瘤血管生成在食管鳞癌的侵袭转移过程中发挥了极其重要的促进作用，因此，研究血管生成的分子机制有望为食管鳞癌的治疗带来突破。近年来的研究表明肿瘤转移相关蛋白1（MTA1）与人类多种恶性肿瘤的血管生成密切相关，并能通过调控VEGF-A等下游分子来调节肿瘤血管生成，提示MTA1蛋白可能是一个潜在的促进恶性肿瘤血管生成的关键分子，已被锚定为抗肿瘤血管生成治疗的新靶点。我们前期的研究首次证实MTA1蛋白在食管鳞癌组织中存在高表达，并与血管生成密切相关，但其具体的信号调控机制尚不明确，为进一步的分子机制研究及以MTA1蛋白为靶点的治疗研究打下了良好的前期基础。本研究将通过体内、外实验深入系统的探讨MTA1蛋白调控食管鳞癌血管生成的具体分子机制，以期为临床靶向治疗食管鳞癌提供新的思路和理论依据。

肿瘤血管生成是一个极其复杂的动态过程，涉及体内多种细胞并受多因子不同层次的调控。本研究的结果表明MTA1蛋白调控食管鳞癌裸鼠移植瘤血管生成的作用较弱，提示MTA1蛋白可能为一种下游的血管生成调节因子，而不是调控食管鳞癌血管生成相关因子的关键分子之一。由于实验结果显示MTA1蛋白表达与裸鼠移植瘤的生长增殖能力显著相关，遂对MTA1蛋白调控食管鳞癌裸鼠移植瘤的增殖机制进行相关的研究。主要研究内容：利用Affymetrix公司的高通量人基因表达谱芯片对KYSE410细胞裸鼠移植瘤进行差异表达基因的筛选，分析的结果显示共有779个基因的表达水平具有显著性差异，其中352个基因的表达水平显著上调，427个基因的表达水平显著下调；对筛选出的779个差异表达基因进一步进行Pathway富集分析及Gene Ontology（biological process, cellular component, molecular function）分析，结合Pubmed检索结果，从中筛选出30个尚未在食管鳞癌中报道的差异基因；针对每个基因设计3个RNAi靶点，并将携带不同靶点的3个质粒等比例混合进行慢病毒包装，从而确保目的基因的敲减效率；使用高内涵筛选的方法，通过比较基因敲减对KYSE410细胞增殖速度的影响，初步筛选出增殖抑制表型明显的基因SPRED1和KDM1B；对筛选出的SPRED1基因进行RNAi后，KYSE410细胞的增殖速率受到显著抑制、处于G1期的细胞显著减少、处于S期的细胞显著增多、细胞的凋亡数显著增多、克隆形成能力显著降低、增殖能力显著减缓，提示SPRED1基因与KYSE410细胞的增殖能力显著相关。综上所述，MTA1蛋白参与调控了KYSE410细胞裸鼠移植瘤内多种基因的表达，通过调控其下游基因SPRED1发挥对食管鳞癌KYSE410细胞的增殖调控作用。进一步研究MTA1蛋白调控SPRED1基因表达的分子机制及以MTA1蛋白为靶点，针对其信号传导通路的干预治疗可能有利于控制食管鳞癌细胞的增殖能力，从而为食管鳞癌的临床治疗提供新的思路。