细胞外基质蛋白FGA调控肺腺癌培美曲塞耐药的作用和机制研究

Drug resistance of tumor is the main cause of treatment failure and disease progression. Pemetrexed is the first choice of chemotherapy drug for lung adenocarcinoma, thus it will be great theoretical and clinical value of pemetrexed to clarify the mechanism of its drug resistance and establish measures to reverse drug resistance. Secreted protein is highly related to acquired drug resistance of chemotherapy drugs and other pathological conditions. Our previous studies have found that the expression of extracellular matrix protein-Fibrinogen Alpha Chain (FGA) was significantly increased in lung adenocarcinoma patients with primary or secondary pemetrexed resistant, thus FGA may be an important regulatory molecule for pemetrexed resistant of lung adenocarcinoma. Meanwhile, there is an interaction between FGA and transmembrane receptor integrin β1, which suggests that FGA can transmit extracellular matrix signals through transmembrane receptor integrin. Therefore, it is proposed that extracellular matrix protein FGA may cause pemetrexed-resistance in the chemotherapy of lung adenocarcinoma through regulating transmembrane receptor integrin mediated-transmission of extracellular matrix signals. In this project, we aim to elucidate the molecular mechanism of FGA/transmembrane receptor integrin β1 signaling pathway on drug resistance in patients with lung adenocarcinoma from evidences in cell culture, animal model and clinical patients. It will provide the theoretical basis to reveal the molecular mechanism of drug resistance in lung adenocarcinoma patients with chemotherapy, and is expected to develop new treatment strategies to predict and improve chemotherapy curative effect in these patients.

肿瘤耐药是导致治疗失败和疾病进展的主要原因，培美曲塞是肺腺癌的首选化疗药物，明确耐药机制建立逆转耐药措施有重要的理论和临床应用价值。分泌蛋白与化疗药物的获得性耐药等病理状态密切相关。前期研究发现，细胞外基质蛋白-纤维蛋白原α(FGA)在原发或继发性培美曲塞耐药的肺腺癌患者中均表达升高，提示FGA可能是肺腺癌化疗中培美曲塞耐药的重要调控分子。由于FGA与跨膜受体整合素β1相互作用，提示FGA可能通过跨膜受体整合素传递胞外基质信号。由此我们提出：细胞外基质蛋白FGA可能通过调控跨膜受体整合素传递的胞外基质信号，从而导致肺腺癌化疗中培美曲塞耐药。本课题将从调控分子-细胞系-动物模型-患者样本等四个层次阐明FGA/整合素β1信号级联调节肺腺癌化疗中培美曲塞耐药的分子机制。本研究将为揭示肺腺癌患者化疗药物耐药的分子机制提供理论依据，并有望为预测和改善肺腺癌患者的化疗疗效提供新的治疗策略。

肺癌是我国最常见和死亡率最高的恶性肿瘤之一，其中肺腺癌占肺癌的30%-40%。肿瘤的复发、转移和耐药是导致治疗失败和疾病进展的主要原因。本研究建立了肺腺癌培美曲塞稳定耐药细胞株A549/PEM和H1299/PEM。并通过CCK-8法检测并对比筛选组与对照组对培美曲塞的敏感性，明确了筛选组细胞已经转化为较稳定的A549培美曲塞耐药细胞株。通过RNA-seq对两组细胞进行差异显著表达基因分析后，发现对比对照组，筛选组细胞有838个基因发生了表达上调，而1113个基因则发生了表达下调。结果显示，对比对照组细胞，A549耐培美曲塞细胞株中，包括氨基酸合成，氮代谢，丙氨酸、天冬氨酸等多种氨基酸代谢，轴突导向等相关基因发生显著上调；而造血细胞相关基因，细胞因子-细胞因子受体互作，IL-17 ( Interleukin-17 ) 相关信号通路以及TNF ( tumor necrosis factor ) 相关信号通路则发生了明显下调。细胞外基质蛋白与肿瘤血管新生、分化、侵袭转移及化疗与靶向药物的获得性耐药等病理状态密切相关。通过后续实验进一步分析表明细胞外基质蛋白FGA及其相关信号通路参与了培美曲塞耐药的发生。细胞实验发现 FGA在LUAD细胞中能够抑制肿瘤的生长和转移；在肺腺癌中FGA与整合素β1结合并减少AKT的磷酸化,导致mTOR信号受到抑制，从而调控肿瘤的生长和转移；肺癌患者的FGA表达下调与p-AKT的上调可能是肺癌预后不良的因素。本研究将为揭示肺腺癌患者发生发展及耐药的分子机制提供理论依据，为其进一步在临床的转化研究奠定基础。