ECSIT基因突变在NK/T细胞淋巴瘤中激活NF-κB信号通路的机制研究

Natural killer/T-cell lymphoma (NK/TCL) is a subtype of extranodal non-Hodgkin’s lymphoma. NK/TCL is primarily involved in nasopharynx, and sometimes in skin and intestinal tract. This kind of lymphoma is rare in western countries while it is more prevalent in China, Southeast Asia, Korea and Japan. Most physicians believe that NK/TCL is associated with aggressive behavior and poor prognosis. The 5-year OS of patients with early stage disease treated by chemo-radiotherapy is about 30-70%, while it is only 3-31% in patients with advanced stage disease. NK/TCL was previously named as lethal midline granuloma. Most patients presented with inflammatory symptoms such as fever and necrosis. Compared with other malignancies, the infiltration of inflammatory cells (macrophages, lymphocytes, plasma cells and granulocytes) is observed more significantly in the local focus. However, it is still unclear what exactly the role of inflammation is on the pathogenesis and development of NK/TCL.NF-kappaB signaling pathway provides a critical link between inflammation and cancer. A variety of recent studies in lymphoid cancers indicated that several genes in NF-κB pathway were frequently mutated, including MyD88, A20, CARD11, and CD79B. These discoveries offer us an opportunity to understanding the underlying mechanisms of the disease pathway and the design of potential therapeutic drugs.In this study, using next-generation sequencing technology and Sequenom platform, we have identified one mutation which caused ECSIT (Evolutionarily Conserved Signaling Intermediate in Toll pathway) gene V93A amino acid change occurred in about 30% NK/TCL samples. This mutation activated NF-κB signaling pathway when tested using luciferase reporter assay in 293T cell. Our following project will focus on how ECSIT V93A mutation activating NF-κB signaling pathway. Combine with animal model and clinical data analysis, this study will elucidate the molecular mechanism responsible for ECSIT V93A mutation activating NF-κB signaling pathway and inducing inflammation, thus providing a promising therapeutic target in the treatment of NK/TCL.

NK/T细胞淋巴瘤（NK/TCL）特异高发于中国，其强烈炎症反应引发的溃疡、穿孔及大面积组织坏死给临床治疗带来挑战。目前关于NK/TCL中炎症过度激活的遗传学背景及分子机制尚不明确。申请人具有良好的肿瘤遗传学和生物信息学研究基础（PLoS Genetics，2011），前期利用新一代测序技术和Sequenom质谱分析平台，发现NF-κB信号通路中ECSIT基因在约30%的NK/TCL患者中存在V93A位点的突变，同时证明该突变能过度激活NF-κB通路。综上研究结果，本项目将深入研究ECSIT突变通过激活NF-κB信号通路来诱导过度炎症反应的分子机制，利用动物模型验证该突变对NK/TCL细胞致瘤能力的调控作用，通过大规模临床样本分析评估该突变对生存及预后的影响，深入阐明ECSIT基因突变在NK/TCL发生发展中的重要调节机制，为NK/TCL的临床诊断和个性化治疗提供有效的干预靶点及治疗模式。

NK/T细胞淋巴瘤是在中国发病率较高（欧美国家极为罕见）的恶性非霍奇金淋巴瘤。近年来对于早期NK/T细胞淋巴瘤的治疗取得了明显进步，经放化疗后早期患者的5年总生存率可达70%。相对于早期患者，晚期患者预后差，5年生存率仅为7%-31%，且部分患者病情发展极为迅速，容易并发以过度炎症反应为特征的致死性综合症——“噬血综合症”。目前对于NK/T细胞淋巴瘤合并“噬血综合症”的发病机制尚不明确，也缺乏有效的治疗手段，“噬血综合症”发生后病人的平均中位生存时间仅为1-2个月。因此，深入研究这类疾病的发病机制并探讨其控制策略尤为重要。.本课题利用高通量测序技术联合质谱分析平台，在88例NK/T细胞淋巴瘤病人的肿瘤样品中发现17例样品的ECSIT（Evolutionarily Conserved Signaling Intermediate in Toll pathways）基因存在V140A（第140位缬氨酸（Val）突变为丙氨酸（Ala））的突变。ECSIT基因的V140A突变导致肿瘤细胞的NF-κB 信号通路持续激活，并分泌多种细胞因子。研究发现，V140A突变型ECSIT激活NF-κB 信号通路主要通过促进钙结合蛋白S100A8和S100A9形成稳定的二聚体并增强NADPH氧化酶的活性而完成。肿瘤细胞NF-κB 信号通路激活后分泌的炎症因子（主要为TNFα和IFNγ）会进一步促进肿瘤周围巨噬细胞的激活并分泌大量细胞因子，形成“细胞因子风暴”。随后，利用CRISPR/Cas9技术构建了ECSIT V140A点突变的小鼠模型和肿瘤移植模型验证了这一结果。进一步的研究发现，地塞米松和沙利多胺的联合用药能有效抑制ECSIT V140A突变引起的过度炎症反应。在两例NK/T细胞淋巴瘤合并“噬血综合症”的病人中，地塞米松和沙利多胺的联合用药显示出较好的疗效，初步验证这一治疗方案在控制“噬血综合症”中的可行性。.该研究成果是NK/T细胞淋巴瘤相关”噬血综合症”领域的重要进展，有助于了解NK/T细胞淋巴瘤中易发”噬血综合症”的遗传机制，并初步验证了控制这一致死性并发症的治疗策略，为下一步的大样本临床研究奠定了基础。