lncRNA PVT1通过调控BTG3参与非小细胞肺癌顺铂耐药的机制研究

Cisplatin resistance is one of the main reasons for treatment failure of advanced non-small cell lung cancer (NSCLC). Long non-coding RNA (lncRNA) is closely related to tumor drug resistance, our preliminary findings showed that lncRNA PVT1 was expressed highly in cisplatin-resistant NSCLC cells and tissues. Interference of lncRNA PVT1 expression can reverse the sensitivity of cisplatin-resistant cells to cisplatin. High throughput sequencing results revealed that BTG3 transcription was regulated by lncRNA PVT1. Bioinformatics showed that BTG3 promoter region was rich in CpG island and lncRNA PVT1 could bind DNA methyltransferase (DNMT1). Interference of DNMT1 can increase BTG3 expression. Gene Set Enrichment Analysis (GSEA) showed that lncRNA PVT1 was associated with cell autophagy. Accordingly, the following hypothesis is proposed: lncRNA PVT1 binds DNMT1 which represses BTG3 transcription via DNA methylation and enhance the resistance of NSCLC by regulating cell autophagy. This study verified the hypothesis via clinical samples, and proved the above hypothesis through CHIP and RNA pull down assay, so as to provide theoretical basis and molecular targets for clinical reversal of drug resistance.

顺铂耐药是晚期非小细胞肺癌（NSCLC）患者治疗失败的主要原因之一。lncRNA与肿瘤耐药密切相关，我们发现lncRNA PVT1在NSCLC耐顺铂细胞及组织中高表达，干扰lncRNA PVT1可逆转耐药细胞对顺铂的敏感性，测序结果示lncRNA PVT1调控BTG3的转录。生物信息学发现BTG3基因启动子区富含CpG岛、lncRNA PVT1可绑定DNA甲基转移酶DNMT1。干扰DNMT1可上调BTG3表达，基因富集分析发现lncRNA PVT1与细胞自噬相关。因此我们提出以下科研假说：lncRNA PVT1绑定DNMT1介导BTG3启动子区DNA甲基化抑制其转录，通过调控细胞自噬进而增强NSCLC顺铂耐药细胞的耐药性。本课题组拟从组织标本验证，通过CHIP、RNA pull down等实验证实上述假说，为临床逆转耐药提供理论依据及分子靶标。

文献报道，LncRNA可通过表观调控下游靶基因参与非小细胞肺癌（NSCLC）顺铂耐药。本课题研究发现，lncRNA PVT1在NSCLC耐药组织和细胞中表达上调，过表达lncRNA PVT1可增加NSCLC细胞对顺铂的耐药性；BTG3在NSCLC耐药组织中表达下调，过表达BTG3，可抑制A549/DDP细胞生物学功能；机制实验研究发现，lncRNA PVT1位于A549/DDP细胞核并可绑定DNMT1蛋白，过表达lncRNA PVT1后，BTG3表达下降；过表达BTG3后，细胞自噬、凋亡等分子标记物表达发生变化。LncRNA PVT1负向调控BTG3的表达，部分通过细胞自噬途径参与NSCLC细胞对顺铂的耐药性。