LncTGFB1调控AXL表达促进肺癌脑转移的机制研究

Our previous study indicated that lung cancer brain metastasis patients with high AXL expression have a worse prognosis. The LncRNA and mRNA transcriptomes were sequenced in every 3 cases of lung cancer brain metastasis patients in the high and low AXL expression groups. Multiple LncRNAs were highly expressed in the high AXL expression group. LncTGFB1that may regulate the AXL expression was found through multiple databases and bioinformatics analysis. Further high and low expression experiments showed that LncTGFB1can regulate the expression of AXL, and it can promote more expression of AXL if we high express the LncTGFB1and SP1 at the same time. Then, RNA pull down, ChIP, and ChIRP were used to verify the LncTGFB1binding protein to confirm further how LncTGFB1-4 regulates the AXL expression. Finally, animal models and lung cancer clinical specimens were used to verify further the LncTGFB1/SP1/AXL axis that can induce invasion, migration, and metastasis of lung cancer cells. To find out the new mechanism of brain metastasis of lung cancer, and it can provide some preliminary data for the development of new drugs and clinical treatment.

我们前期研究提示高表达AXL的肺癌脑转移患者预后更差。选择高低AXL表达组的肺癌脑转移患者各3例进行LncRNA及mRNA转录组测序，发现多个LncRNA在AXL高表达组中高表达，通过多个数据库及生物信息学分析找到可能通过调节AXL表达的LncTGFB1。进一步高/低表达实验发现LncTGFB1和SP1能够调节AXL表达，且同时高表达LncTGFB1及SP1后更能促进AXL表达。然后采用双荧光素酶报告基因，RNA pull down、ChIP、ChIRP等验证与LncTGFB1结合的蛋白从而进一步证实LncTGFB1是如何调节AXL表达的密切关系。最后采用动物模型和肺癌临床标本来进一步验LncTGFB1/SP1/AXL轴诱导肺癌细胞的侵袭，迁移和转移。以期发现肺癌脑转移的新机制，为下一步研发新药物及临床治疗提供一定的前期数据。

【背景】EGFR阳性晚期肺癌患者靶向治疗有效，但因为耐药问题导致生存期短，另外肺癌脑转移患者预后极差。找到导致肺癌EGFR-TKI耐药机制是目前临床的重点及难点。我们在前期研究中发现AXL高表达是EGFR突变肺癌患者耐药原因之一，进一步研究发现LncRNA可以调节AXL的表达从而导致耐药的发生；另外，我们也发现AXL与肺癌脑转移有关，且高表达AXL/GAS6患者更容易出现脑转移。【结果】1.研究发现不同LncRNA可以通过不同方式调节AXL表达，LncTGFB1-4定位于细胞核，通过调节AXL转录从而影响AXL表达；而LncRNA12.4/LncRNAB9.1定位于胞浆，能够通过抑制miRNA的水平，从而调控AXL的表达。2.另外，我们还发现，癌肉瘤肺癌患者，癌成分中的AXL表达量高于肉瘤成分AXL表达，提示可能AXL可能是癌肉瘤的一个潜在生物标记物或治疗靶标。【意义】本研究确定不同LncRNA在不同层面调节AXL表达，从而影响EGFR靶向治疗耐药及脑转移，为后续针对开展相应治疗提供一定前期理论基础。另外癌肉瘤里AXL表达差异，可能成为一个新的生物标记物。