LINC01234通过RNA甲基化调控非小细胞肺癌吉非替尼耐药的机制研究

Secondary drug resistance of EGFR-TKI plagues clinical treatment of non-small cell lung cancer (NSCLC). lncRNA is closely related to gefitinib resistance. We found that LINC01234 is relatively highly expressed in NSCLC gefitinib-resistant cells and tissues. Inhibition of LINC01234 increased the sensitivity of drug-resistant cells to gefitinib. RNA pulldown and mass spectrometry analysis showed that LINC01234 can bind to RNA methyltransferase METTL3, transcriptome sequencing and RNA methylation sequencing found that RCC2 is the target gene of LINC01234, and RCC2 regulates autophagy. Therefore, we propose a scientific hypothesis: LINC01234 increases the mRNA stability and protein translation ability of RCC2 by binding to METTL3 protein-mediated RNA methylation modification, thereby inducing autophagy to promote the resistance of NSCLC to gefitinib. Our research team will verify the above hypothesis through clinical sample verification, using RIP, RNA pulldown, MeRIP and other experiments to provide a new basis and target for reversing targeted drug resistance.

EGFR-TKI继发性耐药困扰非小细胞肺癌（NSCLC）临床治疗，lncRNA与吉非替尼耐药密切相关。我们发现LINC01234在NSCLC吉非替尼耐药细胞和组织中相对高表达。干扰LINC01234增加了耐药细胞对吉非替尼的敏感性。RNA pulldown及质谱分析发现LINC01234可以与RNA甲基转移酶METTL3结合，转录组测序及RNA甲基化测序发现RCC2为LINC01234的靶基因，RCC2调控细胞自噬。据此提出假设：LINC01234通过结合METTL3蛋白介导RNA甲基化修饰提高RCC2的mRNA稳定性和蛋白翻译能力，从而诱发细胞自噬促进NSCLC吉非替尼的耐药性。本课题组将通过临床样本验证，运用RIP、RNA pulldown、MeRIP等实验证实以上假说，为逆转靶向耐药提供新依据及靶标。

EGFR-TKIs继发性耐药困扰非小细胞肺癌临床治疗，EGFR-TKIs的耐药是复杂的、多基因、多通路调控的结果，至今仍有30%的耐药机制尚未明确。我们研究发现LINC01234的异常表达可能与NSCLC细胞吉非替尼耐药有关，通过体内外实验证实LINC01234可影响NSCLC吉非替尼耐药细胞的增殖和对EGFR-TKIs的耐药性。通过RNA pulldown及质谱分析发现，LINC01234可与甲基转移酶METTL3结合，在PC9/GR细胞中进行转录组mRNA测序和全转录组RNA甲基化免疫共沉淀测序，发现RCC2是LINC01234的靶基因。在RCC2终止密码子附近的3’UTR区存在明显的m6A峰，提示RCC2与RNA甲基化修饰有关。敲低METTL3和LINC01234的表达，RCC2的m6A修饰水平、mRNA水平和稳定性显著降低。LINC0234可促进METTL3与RCC2的结合能力，使RCC2的m6A水平上调，稳定RCC2的表达。在PC9/GR细胞中抑制RCC2的表达可引起自噬相关蛋白差异表达，部分恢复耐药细胞对吉非替尼的敏感性。本课题深入探讨LINC01234调控NSCLC吉非替尼耐药的表型及分子机制，为临床逆转NSCLC吉非替尼耐药提供新的靶标和依据。