FBXO6介导ZEB2泛素化抑制结直肠癌侵袭转移的分子机制研究

Colorectal cancer is one of the malignant tumors that seriously endanger human health. Epithelial-mesenchymal transition (EMT) is an important link in its invasion and metastasis, which ultimately leads to the death of patients. More and more evidences show that F-box proteins are critical in tumor invasion and metastasis, but its specific molecular mechanism remains unclear. We found that FBXO6, as a classical E3 ligase in the process of ubiquitination, decreased the expression level in colorectal cancer, relieved the ubiquitination degradation of ZEB2, promoted the expression of important EMT proteins E-cadherin, ZO-1, N-cadherin and Vimentin, and ultimately led to poor prognosis and distant metastasis of colorectal cancer. Therefore, we speculate that FBXO6 regulates the expression of important proteins on the EMT/ZEB2 signal axis through ubiquitination regulation, thereby inhibiting the invasion and metastasis of colorectal cancer. Based on previous studies, this project applies a variety of biological research methods to explain FBXO6-mediated ZEB2 ubiquitination, regulation of EMT, and inhibition of colorectal cancer invasion and metastasis at the cellular, molecular, animal and clinical levels. This study will provide novel therapeutic strategies and targets for the treatment of distant metastasis of colorectal cancer.

结直肠癌是严重危害人类健康的恶性肿瘤之一，上皮间质转化（EMT）是其发生侵袭转移重要的环节，最终导致患者死亡。越来越多的证据表明，F-box蛋白在肿瘤侵袭转移过程中至关重要，但其具体的分子机制仍不明确。我们研究发现：FBXO6作为泛素化过程中经典的E3连接酶，在结直肠癌中表达水平降低，减弱了ZEB2的泛素化降解，促进EMT重要蛋白E-cadherin、ZO-1、N-cadherin、Vimentin的表达，最终导致结直肠癌的不良预后和远处转移。故我们推测，FBXO6通过泛素化调控来降低EMT/ZEB2信号轴上重要蛋白的表达，从而抑制结直肠癌的侵袭转移。本课题拟在前期研究基础上，应用多种生物学研究方法，在细胞水平、分子机制水平、动物水平和临床水平阐述FBXO6介导ZEB2泛素化，调控EMT，抑制结直肠癌侵袭转移的这一机制。本研究有望为结直肠癌远处转移的治疗提供新思路和新靶点。

背景：氧化应激、细胞焦亡和炎症被认为是溃疡性结肠炎（UC）发展的重要致病因素，最近有报道传统的抗酒精中毒药物双硫仑（DSF）对上述所有细胞功能发挥调节作用，这使得DSF成为UC治疗的理想治疗剂，但这个问题尚未得到充分研究。.方法：分别建立了C57BL/6J小鼠中葡聚糖硫酸钠（DSS）诱导的动物模型和结肠细胞系（HT-29和Caco-2）中脂多糖（LPS）诱导的UC细胞模型。细胞因子分泌由ELISA测定。通过MTT测定和EdU测定评估细胞活力和增殖。采用实时qPCR、蛋白质印迹、免疫荧光染色测定和免疫组织化学（IHC）评估基因表达。采用皮尔逊相关分析临床组织中基因的相关性。.结果：DSF在体外和体内抑制UC模型中的氧化应激，焦亡细胞死亡和细胞炎症，并通过N-乙酰基-L-半胱氨酸（NAC）消除活性氧（ROS）拯救LPS处理的结肠细胞（HT-29和Caco-2）中的细胞活力。进一步的实验表明，糖原合酶激酶-3β（GSK-3β）/Nrf2/NLRP3信号级联反应在这一过程中起了关键作用。从机制上讲，DSF下调了GSK-3β和NLRP3，而在LPS处理的结肠细胞中上调了Nrf2。此外，DSF对Nrf2和NLRP3的调节作用通过上调GSK-3β而被消除。此外，GSK-3β的上调消除了DSF对LPS处理的结肠细胞的保护作用。.结论：综上所述，本研究的数据表明，DSF通过调节GSK-3β/Nrf2/NLRP3通路抑制氧化损伤相关的焦亡细胞死亡和炎症，从而抑制LPS诱导的UC发展。

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