Department of Oncology, Zhujiang Hospital, Southern Medical University, Guangzhou, Guangdong 510282, P.R. China.
Int J Oncol. 2020 Sep;57(3):858-870. doi: 10.3892/ijo.2020.5086. Epub 2020 Jun 22.
Erlotinib, an epidermal growth factor receptor tyrosine kinase inhibitor (EGFR‑TKI), is widely applied as a first‑line treatment for non‑small cell lung cancer (NSCLC) and greatly improves the clinical outcomes of patients. However, acquired resistance to EGFR‑TKIs remains a major clinical challenge. Here, we identified guanylate‑binding protein‑1 (GBP1) as a novel protein related to erlotinib resistance, and explored the specific mechanism by which GBP1 is involved in erlotinib resistance. First, the human NSCLC cells PC9ER and HCC827ER were generated by exposing cells to increasing concentrations of erlotinib over 6 months. We screened different genes between erlotinib‑sensitive and erlotinib‑resistant cells with data from the Gene Expression Omnibus database and detected the expression of these genes in erlotinib‑resistant and erlotinib‑sensitive cells by quantitative real‑time polymerase chain reaction (qPCR). Moreover, we constructed GBP1‑knockdown and GBP1‑overexpressing cells to determine the IC50 value of erlotinib, to perform an apoptosis assay and to examine cell cycle distribution. A subcutaneous tumorigenesis test was used to analyze how GBP1 affects erlotinib resistance. Then, mass spectrometry analysis and coimmunoprecipitation were performed to verify the interaction between GBP1 and phosphoglycerate kinase 1 (PGK1). Changes in epithelial‑mesenchymal transition (EMT)‑related markers were observed following the upregulation and downregulation of PGK1 expression. Finally, a rescue experiment was used to determine whether GBP1 regulates EMT through PGK1. In the present study, GBP1 was significantly upregulated in erlotinib‑resistant cells, compared with erlotinib‑sensitive cells. In vitro and in vivo experiments showed that upregulated GBP1 expression contributed to erlotinib resistance, while decreased GBP1 expression had the opposite effect. As shown by performing survival analysis, high GBP1 expression predicted poor prognosis in patients with lung adenocarcinoma. Furthermore, the interaction between GBP1 and PGK1 was confirmed, and a rescue experiment revealed that GBP1 regulates EMT via PGK1. Finally, functional experiments showed that EMT is involved in erlotinib resistance. Our study suggests that GBP1 regulates erlotinib resistance via PGK1‑mediated EMT signaling, suggesting GBP1 as a potential therapeutic target in erlotinib‑resistant NSCLC.
厄洛替尼是一种表皮生长因子受体酪氨酸激酶抑制剂（EGFR-TKI），广泛应用于非小细胞肺癌（NSCLC）的一线治疗，极大地改善了患者的临床结局。然而，获得性 EGFR-TKI 耐药仍然是一个主要的临床挑战。在这里，我们鉴定了鸟嘌呤核苷酸结合蛋白 1（GBP1）作为一种与厄洛替尼耐药相关的新型蛋白，并探讨了 GBP1 参与厄洛替尼耐药的具体机制。首先，通过将细胞暴露于逐渐增加的厄洛替尼浓度 6 个月，生成人 NSCLC 细胞 PC9ER 和 HCC827ER。我们使用来自基因表达综合数据库的数据筛选出厄洛替尼敏感和耐药细胞之间的不同基因，并通过实时定量聚合酶链反应（qPCR）检测这些基因在厄洛替尼耐药和敏感细胞中的表达。此外，我们构建了 GBP1 敲低和过表达细胞，以确定厄洛替尼的 IC50 值，进行凋亡测定并检查细胞周期分布。进行皮下肿瘤发生测试以分析 GBP1 如何影响厄洛替尼耐药性。然后，进行质谱分析和免疫共沉淀以验证 GBP1 和磷酸甘油酸激酶 1（PGK1）之间的相互作用。观察到 PGK1 表达上调和下调后上皮-间充质转化（EMT）相关标志物的变化。最后，进行挽救实验以确定 GBP1 是否通过 PGK1 调节 EMT。在本研究中，与厄洛替尼敏感细胞相比，厄洛替尼耐药细胞中 GBP1 的表达显著上调。体内外实验表明，上调 GBP1 表达有助于厄洛替尼耐药，而下调 GBP1 表达则产生相反的效果。通过进行生存分析，高 GBP1 表达预示着肺腺癌患者的预后不良。此外，证实了 GBP1 和 PGK1 之间的相互作用，并通过挽救实验揭示了 GBP1 通过 PGK1 调节 EMT。最后，功能实验表明 EMT 参与了厄洛替尼耐药。我们的研究表明，GBP1 通过 PGK1 介导的 EMT 信号通路调节厄洛替尼耐药，提示 GBP1 可能成为厄洛替尼耐药 NSCLC 的潜在治疗靶点。
