多环芳香烃受体介导的肺癌EGFR-TKIs耐药新机制实验研究

EGFR-TKIs are outstanding breakthrough in lung cancer individualized treatment, while the subsequent drug resistance significantly faded their efficiency in the clinical settings. Apart from EGFR T790M mutation and Met gene amplification, the resistant mechanisms are not fully understood. Recent studies indicate cancer cells can catabolize tryptophan into kynurenine and kynurenine acts as a high affinity endogenous ligand for aryl hydrocarbon receptor (AhR). Our preliminary studies show tryptophan metabolic enzyme and AhR are overexpressed in the T790M mutated H1975 cells. Inhibition of AhR signaling by pharmacological and molecular approaches partially reverse resistance to EGFR-TKIs. We also notice overexpression of AhR in lung cancer cells without T790M mutation or Met gene amplification induces a resistant phenotype, suggesting AhR might be a new resistant mechanism to EGFR-TKIs. The present study will focus on underlying mechanism of AhR-induced resistance. By using the inducible transgenetic mice, AhR gene knock-out mice and resistant samples, we will speculate whether AhR act as a new target for reversing EGFR-TKIs resistance. It is reasonable to believe AhR is a promising candidate for overcoming EGFR-TKIs resistance according to the present study.

表皮生长因子受体酪氨酸激酶抑制剂（EGFR-TKIs）是肺癌个体化治疗的里程碑，但耐药难题极大的限制其临床疗效，除了EGFR T790M突变和Met基因扩增等已知机制，仍有一部分耐药病例机制不明。最新研究提示肿瘤细胞可以摄取色氨酸代谢为犬尿氨酸，后者是多环芳香烃受体（AhR）高亲和力内源性配体。我们的研究发现T790M突变的H1975细胞高表达色氨酸代谢酶和AhR，药物或shRNA抑制AhR信号可以逆转H1975细胞EGFR-TKIs耐药性。我们还发现无T790M突变或Met基因扩增的肺癌细胞过表达AhR后出现耐药表型，提示AhR可能是引起EGFR-TKIs耐药的独立因素。本课题将深入研究AhR导致EGFR-TKIs耐药的分子机制，利用可诱导转基因小鼠、AhR基因敲除小鼠和肺癌患者耐药标本全面评估AhR新靶点逆转耐药的作用，为最终克服EGFR-TKIs耐药提供理论和实验基础。

多环芳香烃受体（aryl hydrocarbon receptor, AhR）是一个受配体活化的胞内受体，在过去的近40年里人们一直以为AhR信号通路活化后转录激活下游代谢酶，其全部功能由转录因子活性介导；近些年研究发现AhR还有一些生物学功能不依赖其转录活性，且AhR调控细胞增殖、分化、凋亡、周期转化等新功能与代谢酶表达无关。AhR在多种实体瘤组织和细胞系中过度表达，回顾性研究提示AhR表达水平与肿瘤恶性程度正相关。课题组前期研究发现AhR蛋白在TKIs耐药的患者标本中表达亦增加，但其意义并不清楚。本文揭示了AhR蛋白负向调控NSCLC TKIs敏感性的作用，提出活化的AhR作为衔接蛋白（adaptor protein）招募Src和Jak2两种激酶，形成的Jak2-AhR-Src复合物可以绕过EGFR旁路活化下游PI3K/Akt和MEK/Erk信号通路导致TKIs耐药；在高表达内源性AhR蛋白的耐药细胞用药物或shRNA抑制AhR可以逆转TKIs耐药性，而在敏感细胞过表达AhR则导致TKIs耐药性；此外，AhR与Src和Jak2蛋白相互作用不依赖其转录因子活性，活化的AhR瞬时转位到近膜侧，携带胞浆内Src蛋白与Jak2结合，促使Jak2磷酸化Src，Jak2 TKIs和Src TKIs可以阻断AhR蛋白介导的激酶旁路。本研究发现AhR蛋白招募激酶的新功能，阐明了TKIs耐药患者AhR表达增加的意义及其导致耐药的分子机制，为克服EGFR TKIs耐药提供新的思路。