去泛素化酶OTUD3基因缺失诱导食管鳞癌淋巴结转移的分子机制

Lymph node metastasis is the most common manner of metastasis in esophageal cancer, and is closely correlated with poor prognosis. Thus idenifying key molecular factor in lymphatic metastasis process would be of great value for early diagnosis and therapy. Recently, using the microarray screening, we found that the deubiquitinase OTUD3 is roubustly downregulated in the esophageal cancer tissues with lymph node metastasis. Furthermore, the immunoprecipitation assay revealed that OTUD3 interacts with multiple proteins including IκB-α. We then found that OTUD3 deubiquitinated the K48-linked ubiqutin chain of IκB-α, leading to its stability. As expected, high expression of OTUD3 inhibits NF-κB signaling. Moreover, we demonstrate that overexpression of OTUD3 supresses the capabilities of esophageal cancer cells to induce matrix degradation, lymphangiogenesis and transendothelial migration. Importantly, OTUD3 inhibits lymph node metastasis and distant metastasis in vivo. Finally, we identify that OTUD3 locus is lost in a large portion of esophageal cancer patients. Thus, our recent results suggest an important role of OTUD3 in lymph node metastasis. In the current project, we will combine in vivo and in vitro experimental systems with data from clinical samples, in order to investigate the multi-dimensional mechanisms of OTUD3 loss-induced esophageal cancer lymphatic metastasis. Our work may help identify novel diagnostic markers and therapeutic targets for esophageal cancer.

淋巴结转移是食管鳞癌最主要的转移方式，直接影响治疗的效果及患者的生存预后。寻找淋巴结转移的关键调控分子对食管癌的早期诊断和治疗具有重要的意义。近期，通过基因表达谱芯片筛选，我们发现去泛素化酶OTUD3在淋巴结转移性样品中表达明显下调。免疫共沉淀及质谱分析表明OTUD3与多个重要蛋白相互作用。其中，OTUD3可抑制IκB-α蛋白泛素化降解。OTUD3表达下调激活NF-κB信号通路。进一步预实验结果显示：上调 OTUD3表达可通过抑制①细胞外基质降解、②淋巴管新生、③肿瘤跨内皮细胞迁移等功能，进而多层面调控淋巴结转移的发生。并且，我们发现OTUD3基因存在广泛缺失，与其表达下调和肿瘤淋巴结转移显著相关。这些结果提示OTUD3基因缺失在食管鳞癌淋巴结转移中发挥重要作用。本研究将承前启后，深层次解析OTUD3基因缺失诱导淋巴结的分子机制，并与临床相结合，为食管鳞癌早期诊断及治疗提供新的理论依据。

转移是肿瘤致死的最主要原因。项目以OTUD3调控食管癌淋巴结转移的机制为出发点，深入探讨肿瘤转移的调控机制，鉴定了多个重要的转移调控因子，并阐明其作用机制以及临床意义，可能为肿瘤的临床诊治提供潜在标志物或者靶点。取得的主要成果包括：1.我们发现烟草中的尼古丁可通过PI3K/AKT/FOXO1信号通路显著抑制OTUD3的转录。OTUD3表达与吸烟患者的不良预后密切相关。OTUD3与IκBα相互作用并通过泛素化的β-TrCP介导的K48多聚泛素化来稳定IκBα。从而，尼古丁介导的OTUD3的下调维持了NF-κB信号传导并促进了细胞侵袭和癌症相关的淋巴管生成，从而导致体内食管癌的淋巴转移增加；另外，尼古丁可激活STAT3信号通路而上调RNA结合蛋白ILF2。ILF2通过结合THOC4促进多潜能基因mRNA出核而维持食管癌肿瘤干细胞特性。2. 我们发现SYNPO2抑制LATS2的泛素化降解，从而抑制YAP/TAZ信号激活以及肿瘤转移（J Pathol 2018）；相反，TRIB3抑制TAZ蛋白的泛素化降解，从而激活TAZ信号并诱导食管癌产生放疗抵抗（Oncogene 2020）。3.另外，我们的研究还发现多个OTUD3相关蛋白在肿瘤转移中的功能。其中，HOMER3可作为支架蛋白促进生长因子介导的β-Catenin酪氨酸磷酸化，促进肿瘤侵袭（J Hematol Oncol 2021）；ROPN1结合并激活RHOA促进肿瘤细胞迁移（FASEB J 2020）；CGI-99通过诱导IL-6因子自分泌增强肿瘤定植能力（Oncogene 2017）；SALL2基因甲基化可诱导乳腺癌产生激素治疗耐受（EMBO Mol Med 2019）。项目已发表第一/通讯作者论文5篇，其中2篇IF > 10。这些研究结果从多个不同层面揭示了肿瘤转移的调控机制，可能为肿瘤的临床诊治提供潜在标志物或者靶点。