FOXC2招募LSD1表观遗传调控FBP1激活糖代谢通路促进结肠癌侵袭转移的实验研究

Epithelial-mesenchymal transition (EMT) makes colon cancer cells acquire invasive properties and is associated with poor prognosis. In fact, the EMT process is indeed highly inefficient. The matrix detachment of cancer cells rapidly leads to metabolic stress. Metabolic reprogramming to glycolysis mainly-dependent manner plays a vital role in cancer metastases. Our previous study revealed that EMT interacts with glycolysis to promote colon cancer cells progressing, and FOXC2 has been proved to be a key factor that facilitates both EMT and glycolysis. Mass Spectrometry and Proteomics analysis found that LSD1, as a chromatin modifier, is a potential interacting protein of FOXC2, and PCR array analysis of glycolysis related enzymes indicated that FBP1 is a potential target gene of FOXC2. The present study is oriented towards the molecular, cellular and animal levels, as well as utilizing colorectal tissue samples to further clarify the following issues on the basis of previous research: ①To validate the interaction between FOXC2 and LSD1; ②To investigate whether FOXC2 activates glucose metabolism and promotes metastasis by recruiting the LSD1 to epigenetically co-repress FBP1 transcription; ③To intervene the expression of these genes in vitro in order to provide a new theoretical and experimental basis for repression and intervention of the development, progression and metastasis of colorectal cancer in clinics.

上皮间质转化（EMT）是结肠癌获得侵袭潜能、导致预后不良的重要原因之一。实质上，EMT是一个低效的过程，脱离基底膜的肿瘤细胞很快进入能量不足状态。糖酵解为主的代谢模式重塑在侵袭转移中起着重要的作用。我们前期研究发现：结肠癌中EMT和糖酵解相互促进，从而促使了结肠癌的恶性转化；而FOXC2则是促进结肠癌EMT和获得糖酵解代谢优势的关键分子。高通量蛋白质组学筛选发现染色质重塑蛋白LSD1可与FOXC2相互作用，葡萄糖代谢基因PCR array筛选提示FBP1是FOXC2潜在的靶基因。本课题拟在前期工作基础上，从分子、细胞、动物及临床标本水平进行实验进一步明确：①证实FOXC2与LSD1的相互作用关系；② 探究FOXC2是否通过招募LSD1表观遗传调控FBP1激活糖代谢、促进结肠癌的恶性转化；③通过体外干预相关基因的表达，为临床阻遏和干预结肠癌发生发展以及转移提供新的理论与实验依据。

FOXC2是经典的转录因子，参与了结直肠癌的进展的调节。但FOXC2参与结直肠癌进展和有氧糖酵解的机制尚不清楚。本研究旨在阐述FOXC2调节结直肠癌有氧糖酵解和侵袭转移的机制。我们首先分析了FOXC2在TCGA数据库中的表达情况，发现FOXC2高表达和常见的9中恶性肿瘤不良预后密切相关。在复旦大学附属肿瘤医院的组织芯片检测中，FOXC2在癌组织中表达显著高于癌旁组织，并且FOXC2在被证实为是影响结直肠癌根治术后的独立预后因子。更有意思的是FOXC2表达水平和初诊结直肠癌患者SUVmax数值密切相关。SUVmax被认为是有氧糖酵解在体内活性的反应。体内和体外实验证实，结肠癌细胞中沉默FOXC2表达后可显著降解有氧糖酵解活性和侵袭能力。过表达FOXC2有相反的作用。RNA sequence发现FBP1是FOXC2的下游靶基因。Luciferase和ChIP实验证实FOXC2可以与FBP1启动子特异性结合抑制FBP1的转录活性。高通量蛋白质谱及免疫共沉淀、细胞免疫荧光实验等证实FOXC2通过招募LSD1至FBP1启动子区域影响H3K27me3在启动子区域的富集从而发挥转录调控作用。功能学实验也证实LSD1与FOXC2功能存在一致性。干预FBP1的表达水平可以影响FOXC2作用的发挥。总之，FOXC2是结直肠癌患者中一个新型预后标志物，通过招募LSD1至FBP1启动子区域发挥对FBP1的转录抑制作用促进结直肠癌的有氧糖酵解和侵袭转移的发生，这有望成为结直肠癌诊断和干预治疗的一个新型标志物。