Shi Jingjing, Hao Shaoyu, Liu Xiantao, Li Yingying, Zheng Xin
College of First Clinical Medical, Shandong University of Traditional Chinese Medicine, Jinan, China.
Qingdao Hospital of Traditional Chinese Medicine (Qingdao Hiser Hospital), Qingdao, China.
Front Pharmacol. 2023 Jan 19;14:1093017. doi: 10.3389/fphar.2023.1093017. eCollection 2023.
Osimertinib is a potent epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) for the treatment of patients with EGFR-mutant non-small cell lung cancer (NSCLC). However, the emergence of acquired resistance due to the EGFR-Del19/T790M/C797S mutation limits the clinical application of osimertinib. Feiyiliu Mixture (FYLM), a clinical experience formula of Chinese medicine, was used to treat lung cancer with good clinical efficacy. In this study, we aimed to investigate the mechanism by which Feiyiliu Mixture delays osimertinib resistance in EGFR-mutant cell lines and EGFR-mutant cell tumor-bearing mice. The osimertinib-resistant cell models were established in mouse Lewis lung carcinoma (LLC) cells transfected with EGFR-Del19/T790M/C797S mutant lentivirus. In cell experiments, after 48 h of treatment with Feiyiliu Mixture-containing serum, MTT assay was used to detect the relative cell viability, and western blotting was used to detect EGFR protein phosphorylation expression. In animal experiments, C57BL/6J mice were subcutaneously injected with Lewis lung carcinoma cells stably expressing EGFR-Del19/T790M/C797S mutations to construct a xenograft model. After 2 weeks of Feiyiliu Mixture and/or osimertinib treatment, the expression of proliferation-related, apoptosis-related and PRC1/Wnt/EGFR pathway markers was detected by real-time qPCR, western blotting and immunohistochemistry. The results showed that when combined with osimertinib, Feiyiliu Mixture synergistically reduces proliferation and increases apoptosis to improve drug resistance. , Feiyiliu Mixture-containing serum reduced the EGFR phosphorylation. , Feiyiliu Mixture downregulated the expression of cyclin B1 and Bcl-2 while upregulating the level of cleaved Caspase-3 protein, indicating that Feiyiliu Mixture promotes apoptosis. Furthermore, Feiyiliu Mixture reduced the expression of p-EGFR, p-Akt, PRC1 and Wnt pathway-related proteins such as β-catenin, c-Myc and c-Jun. The present study identified that Feiyiliu Mixture inhibited PRC1/Wnt/EGFR pathway activation, reduced proliferation, and promoted apoptosis, thereby increasing the sensitivity of EGFR-mutant non-small cell lung cancer to osimertinib. Our study provided a new idea for Chinese medicine to play a role in enhancing efficacy and reducing toxicity in the treatment of non-small cell lung cancer.
奥希替尼是一种强效的表皮生长因子受体酪氨酸激酶抑制剂（EGFR-TKI），用于治疗EGFR突变的非小细胞肺癌（NSCLC）患者。然而，由EGFR-Del19/T790M/C797S突变导致的获得性耐药的出现限制了奥希替尼的临床应用。肺抑瘤合剂（FYLM）是一种中医临床经验方，用于治疗肺癌，临床疗效良好。在本研究中，我们旨在探究肺抑瘤合剂延缓EGFR突变细胞系和EGFR突变荷瘤小鼠对奥希替尼耐药的机制。通过用EGFR-Del19/T790M/C797S突变慢病毒转染小鼠Lewis肺癌（LLC）细胞建立奥希替尼耐药细胞模型。在细胞实验中，用含肺抑瘤合剂血清处理48小时后，采用MTT法检测相对细胞活力，并用蛋白质免疫印迹法检测EGFR蛋白磷酸化表达。在动物实验中，将稳定表达EGFR-Del19/T790M/C797S突变的Lewis肺癌细胞皮下注射到C57BL/6J小鼠体内，构建异种移植模型。在肺抑瘤合剂和/或奥希替尼治疗2周后，通过实时定量PCR、蛋白质免疫印迹法和免疫组织化学检测增殖相关、凋亡相关及PRC1/Wnt/EGFR通路标志物的表达。结果表明，肺抑瘤合剂与奥希替尼联合使用时，可协同降低细胞增殖并增加凋亡，从而改善耐药性。含肺抑瘤合剂血清降低了EGFR磷酸化水平。肺抑瘤合剂下调细胞周期蛋白B1和Bcl-2的表达，同时上调裂解的Caspase-3蛋白水平，表明肺抑瘤合剂促进凋亡。此外，肺抑瘤合剂降低了p-EGFR、p-Akt、PRC1以及Wnt通路相关蛋白如β-连环蛋白、c-Myc和c-Jun的表达。本研究发现肺抑瘤合剂抑制PRC1/Wnt/EGFR通路激活，降低细胞增殖并促进凋亡，从而提高EGFR突变的非小细胞肺癌对奥希替尼的敏感性。我们的研究为中药在非小细胞肺癌治疗中发挥增效减毒作用提供了新思路。
