假基因DUXAP8调控RHOB促进NSCLC顺铂耐药机制研究

Drug resistance is one of the causes of cancer related death. There are studies revealed that pseudogenes contribute to cancer development and progression. However, the potential function and molecular mechanism of pseudogenes in non small cell lung cancer drug resistance is still not unclear. In this study, we analyzed the pseudogenes profiling in NSCLC cisplatin resistant cells compared with sensitive cell, and found that the pseudogene DUXAP8 is significantly upregulated in cisplatin resistant cells. We further confirmed the analysis result in NSCLC tissue samples. In addition, knockdown of UDXAP8 increase the sensitivity of A549/CDDP cells to cisplatin, and affect EMT markers protein levels. The results of RIP assays showed that DUXAP8 could directly bind with PRC2 and LSD1 in A549/CDDP cells. RNA sequencing showed that RHOB is the potential underlying target of DUXAP8, and knockdown of EZH2 or LSD1 also upreguated its expression level. Therefore, we propose our hypothesis that increased DUXAP8 promotes NSCLC cells EMT and induced cisplatin resistance by recruiting PRC2/LSD1 to RHOB promoter and repressing its transcription. Further mechanistic investigation will be performed to determine our hypothesis by using RNA pulldown, ChIP et al. Technique. Our study will further our understanding of the molecular mechanism of NSCLC cells resistance to cisplatin.

顺铂耐药是目前非小细胞肺癌（NSCLC）治疗中亟待解决的问题。假基因与肿瘤发生密切相关，但在NSCLC细胞耐药中的作用和机制尚不清楚。我们通过分析GEO数据库中顺铂敏感细胞和耐药细胞中差异表达的假基因，发现DUXAP8表达显著上调，在组织中得到验证。干扰DUXAP8后，A549/CDDP细胞对顺铂敏感性增加，并影响EMT相关分子表达。RIP实验证实DUXAP8与PRC2、LSD1蛋白绑定。RNA测序发现RHOB是DUXAP8的靶基因，且干扰EZH2，LSD1亦能上调其表达。据此，提出假设：NSCLC细胞中DUXAP8表达上调，发挥脚手架作用同时募集PRC2/LSD1抑制RHOB的表达，促进NSCLC细胞EMT，从而导致顺铂耐药。本课题组将通过临床样本验证，运用ChIP、RIP、RNA pulldown等技术证实上述假设，丰富NSCLC化疗耐药分子机制的认知，为解决临床化疗耐药提供新依据。

非小细胞肺癌（NSCLC）是全球范围内最常见的肿瘤之一。假基因与肿瘤发生密切相关，但在NSCLC顺铂耐药中的作用和机制尚不清楚。我们通过分析GEO数据库，发现假基因DUXAP8在NSCLC耐药细胞中表达显著上调，并在组织样本中进行了验证。DUAXP8高表达患者无病生存期更短，提示DUXAP8可作为NSCLC的预后标志物。敲除DUXAP8后，细胞增殖能力受损，凋亡增加。同时，DUXAP8抑制也会增加A549/CDDP和H460/CDDP细胞对顺铂敏感性。RIP实验证实在NSCLC耐药细胞中DUXAP8与EZH2和LSD1蛋白相结合。RNA测序发现RHOB是DUXAP8的靶基因。本课题组证实，DUXAP8通过招募组蛋白去甲基化酶LSD1和组蛋白甲基转移酶EZH2以降低抑癌蛋白RHOB的表达，从而促进NSCLC细胞顺铂耐药。该研究结果提示，针对DUXAP8进行治疗对改善NSCLC临床疗效具有重要意义。