Dual inhibition of EGFR and MET by Echinatin retards cell growth and induces apoptosis of lung cancer cells sensitive or resistant to gefitinib.
- 作者列表："Oh HN","Lee MH","Kim E","Kwak AW","Seo JH","Yoon G","Cho SS","Choi JS","Lee SM","Seo KS","Chae JI","Shim JH
:Patients with non-small-cell lung cancer (NSCLC) containing epidermal growth factor receptor (EGFR) amplification or sensitive mutations initially respond to tyrosine kinase inhibitor gefitinib; however, the treatment is less effective over time. Gefitinib resistance mechanisms include MET gene amplification. A therapeutic strategy targeting MET as well as EGFR can overcome resistance to gefitinib. In the present study we identified Echinatin (Ecn), a characteristic chalcone in licorice, which inhibited both EGFR and MET and strongly altered NSCLC cell growth. The antitumor efficacy of Ecn against gefitinib-sensitive or -resistant NSCLC cells with EGFR mutations and MET amplification was confirmed by suppressing cell proliferation and anchorage-independent colony growth. During the targeting of EGFR and MET, Ecn significantly blocked the kinase activity, which was validated with competitive ATP binding. Inhibition of EGFR and MET by Ecn decreases the phosphorylation of downstream target proteins ERBB3, AKT and ERK compared with total protein expression or control. Ecn induced the G2/M cell cycle arrest, and apoptosis via the intrinsic pathway of caspase-dependent activation. Ecn induced ROS production and GRP78, CHOP, DR5 and DR4 expression as well as depolarized the mitochondria membrane potential. Therefore, our results suggest that Ecn is a promising therapeutic agent in NSCLC therapy.
: 含有表皮生长因子受体 (EGFR) 扩增或敏感突变的非小细胞肺癌 (NSCLC) 患者最初对酪氨酸激酶抑制剂吉非替尼有反应; 然而，随着时间的推移，治疗效果较差。吉非替尼耐药机制包括MET基因扩增。靶向MET以及EGFR的治疗策略可以克服对吉非替尼的耐药性。在本研究中，我们鉴定了甘草中的特征性查耳酮棘黄素 (Ecn)，其抑制EGFR和MET并强烈改变NSCLC细胞生长。Ecn对具有EGFR突变和MET扩增的吉非替尼敏感或耐药NSCLC细胞的抗肿瘤功效通过抑制细胞增殖和锚定非依赖性集落生长来证实。在靶向EGFR和MET期间，Ecn显著阻断激酶活性，这通过竞争性ATP结合来验证。与总蛋白表达或对照相比，Ecn对EGFR和MET的抑制降低了下游靶蛋白ERBB3 、AKT和ERK的磷酸化。Ecn通过半胱天冬酶依赖性激活的内在途径诱导G2/M细胞周期阻滞和凋亡。Ecn诱导ROS产生和GRP78 、CHOP、DR5 和DR4 表达以及线粒体膜电位去极化。因此，我们的研究结果表明，Ecn是一种有前途的NSCLC治疗药物。
METHODS::Pulmonary artery sling is a rare congenital anomaly of the origin and course of the left pulmonary artery. Patients with this condition typically present with respiratory failure in young infancy, and asymptomatic cases are uncommon. We describe the case of an adult patient with a lung adenocarcinoma of the right upper lobe, extending into the hilum and superior mediastinum, and with a previously unknown pulmonary artery sling anomaly. The local invasiveness of the tumor and the peculiar vascular anatomy contributed to a unique surgical scenario, wherein multiple reconstructive procedures were required.
METHODS::Patients with idiopathic pulmonary fibrosis (IPF) have higher risk of developing lung cancer, for example, squamous cell carcinoma (SCC), and show poor prognosis, while the molecular basis has not been fully investigated. Here we conducted DNA methylome analysis of lung SCC using 20 SCC samples with/without IPF, and noncancerous lung tissue samples from smokers/nonsmokers, using Infinium HumanMethylation 450K array. SCC was clustered into low- and high-methylation epigenotypes by hierarchical clustering analysis. Genes hypermethylated in SCC significantly included genes targeted by polycomb repressive complex in embryonic stem cells, and genes associated with Gene Ontology terms, for example, "transcription" and "cell adhesion," while genes hypermethylated specifically in high-methylation subgroup significantly included genes associated with "negative regulation of growth." Low-methylation subgroup significantly correlated with IPF (78%, vs. 17% in high-methylation subgroup, p = 0.04), and the correlation was validated by additional Infinium analysis of SCC samples (n = 44 in total), and data from The Cancer Genome Atlas (n = 390). The correlation between low-methylation subgroup and IPF was further validated by quantitative methylation analysis of marker genes commonly hypermethylated in SCC (HOXA2, HOXA9 and PCDHGB6), and markers specifically hypermethylated in high-methylation subgroup (DLEC1, CFTR, MT1M, CRIP3 and ALDH7A1) in 77 SCC cases using pyrosequencing (p = 0.003). Furthermore, low-methylation epigenotype significantly correlated with poorer prognosis among all SCC patients, or among patients without IPF. Multivariate analysis showed that low-methylation epigenotype is an independent predictor of poor prognosis. These may suggest that lung SCC could be stratified into molecular subtypes with distinct prognosis, and low-methylation lung SCC that significantly correlates with IPF shows unfavorable outcome.
METHODS::The role of Fyn-related kinase (FRK) in malignant tumors remains controversial. Our study investigated the function of FRK in lung cancer. Immunohistochemistry staining and generating a knockout of FRK by CRISPR/Cas9 in H1299 (FRK-KO-H1299) cells were strategies used to explore the role of FRK. Immunohistochemistry staining indicated that FRK expression was elevated in 223 lung cancer tissues compared to 26 distant normal lung tissues. FRK contributed to poor survival status in lung cancer patients and acted as a predictor for poor prognosis of lung cancer. Knockout of FRK by CRISPR/Cas9 markedly inhibited proliferation, invasion, colony formation and epithelial-mesenchymal transition (EMT) process in the lung cancer cell line H1299. Further exploration indicated that FRK-KO damaged the stemness phenotype of H1299 by inhibiting CD44 and CD133 expression. Seahorse detection and a U-13 C flux assay revealed that FRK-KO induced metabolism reprogramming by inhibiting the Warburg effect and changing the energy type in H1299 cells. Epidermal growth factor stimulation recovered the expression of FRK and biological functions, metabolic reprogramming and stemness phenotype of H1299 cells. FRK plays an oncogenic role in lung cancer cells via a novel regulation mechanism of enhancing the stemness of H1299 cells by inducing metabolism reprogramming, which finally promotes EMT and metastasis. Our study also indicates that FRK could be used as a potential therapeutic target for drug development.