EBV膜蛋白LMP1调控H3K4me3/H3K27me3表观重编程导致鼻咽癌耐药的机制研究

Chemoresistance of tumor cells is the main reason for the failure of advanced nasopharyngeal carcinoma treatment. The disorder of histone methylation modification in tumor cells is the key mechanism leading to chemoresistance. Our previous study found that LMP1 was able to regulate H3K4me3/H3K27me3 on the gene promoter and observed that high expression of LMP1 could induce resistance to cisplatin and 5-fluorouracil of nasopharyngeal carcinoma cells.Further analysis of the RNA-seq data, we found that tumor resistance related genes were significantly enriched in LMP1-positive cells, suggesting that histone methylation deregulation may be crucial in LMP1-induced chemoresistance. Therefore, this project aims to establish the pattern of LMP1-regulated H3K4me3/H3K27me3 by ChIP-Seq sequencing, and screen the key target genes regulated by LMP1-induced H3K4me3/H3K27me3 and combined analysis the RNA-seq data. Next, we would explore the molecular mechanism of LMP1 conferred drug resistance and to verify the effect of target genes in animal model and clinical sample. Our study will provide new potential therapeutic targets to bypass the chemoresistance of nasopharyngeal carcinoma.

肿瘤细胞耐药是晚期鼻咽癌治疗失败的主要原因，而组蛋白甲基化修饰紊乱是导致肿瘤细胞耐药的关键机制。我们前期研究发现LMP1能够调控基因启动子上H3K4me3/H3K27me3甲基化修饰，并观察到高表达LMP1可导致鼻咽癌细胞对顺铂和5-氟尿嘧啶耐药，RNA-seq进一步分析发现肿瘤耐药基因在LMP1阳性细胞中显著富集，提示组蛋白甲基化修饰紊乱可能是LMP1导致鼻咽癌耐药的关键因素。因此，本项目拟通过ChIP-Seq测序建立LMP1调控H3K4me3/H3K27me3的甲基化修饰谱，结合前期RNA-seq表达谱分析筛选出LMP1调控H3K4me3/H3K27me3导致鼻咽癌耐药的关键靶基因，进一步探索该基因导致鼻咽癌耐药的分子机制，并在动物模型和临床样本中验证该基因对鼻咽癌耐药的影响。本研究的顺利完成将为逆转鼻咽癌耐药提供新的理论依据和潜在治疗靶点。

鼻咽癌恶性程度高且与EB病毒感染显著相关。目前病毒感染诱发鼻咽癌进展的机制尚未明确，本研究构建了可诱导表达EB病毒致癌蛋白LMP1蛋白的鼻咽癌模型。体外功能实验和体内成瘤实验证明LMP1在鼻咽癌内表达能够显著促进肿瘤的生长和转移。进一步对LMP1表达的鼻咽癌进行全转录组测序分析显示LMP1能够广泛调控鼻咽癌基因表达，其中分化相关的重要的转录因子CEBPA被显著沉默。在鼻咽癌细胞中恢复CEBPA表达后，鼻咽癌细胞增殖显著减慢，肿瘤形成明显放缓，且恢复的CEBPA能够抑制LMP1在鼻咽癌中的促癌作用。CEBPA在癌细胞中通常通过启动子甲基化而被沉默，而鼻咽癌中LMP1沉默CEBPA转录的机制尚不清楚。在LMP1表达的鼻咽癌模型中使用小分子抑制剂抑制甲基转移酶并不能阻止CEBPA被LMP1沉默，并且LMP1表达后并未改变CEBPA启动子甲基化状态，提示LMP1沉默CEBPA通过非启动子甲基化途径。进一步使用去乙酰化酶抑制剂处理LMP1表达鼻咽癌证明LMP1通过诱导组蛋白去乙酰化沉默CEBPA。本研究揭示了EB病毒诱导鼻咽癌的新机制并为鼻咽癌治疗提供潜在的新策略。

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