Department of Respiratory and Critical Care Medicine, The Affiliated Hospital of Hangzhou Normal University, 310000 Hangzhou, Zhejiang, China.
Discov Med. 2024 Nov;36(190):2300-2308. doi: 10.24976/Discov.Med.202436190.211.
Epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) is considered highly effective treatment for advanced non-small cell lung cancer (NSCLC), who often develop drug resistance after 10 months of treatment. Herein, the aim was to unravel the mechanism behind the resistance to icotinib in NSCLC.
Establishment of icotinib-resistant PC-9 cells (PC-9R) was achieved through repeated exposure to increasing concentrations of icotinib for more than 12 months. PC-9R cells were transfected with programmed cell death ligand 1 () knockdown plasmid (-KD)/overexpression plasmid (-OE), and treated with Wnt pathway agonist CHIR99021 or β-catenin antagonist ICG-001. 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H tetrazolium assay was employed for detecting cell sensitivity to icotinib. The invasion and migration abilities of the cells were evaluated using Transwell and scratch assays. Quantification of , matrix metalloproteinase (MMP)-2, MMP-9 and Wnt/β-catenin pathway-related proteins was conducted by means of quantitative real-time polymerase chain reaction or Western blotting.
Half-maximal inhibitory concentrations (IC) of PC-9 and PC-9R cells to icotinib were 1.73 μM and 25.18 μM, respectively. The expression of , Wnt family member 1 (Wnt1) and β-catenin was higher in PC-9R cells than in PC-9 cells ( < 0.05). The transfection of -OE resulted in elevated IC, migration, invasion, and MMP-2 and MMP-9 expression in PC-9R cells ( < 0.05), while transfection with -KD had the opposite effect ( < 0.05). The expression of , β-catenin, MMP-2 and MMP-9, and IC, migration and invasion was increased following PC-9R cells treatment with CHIR99021 ( < 0.05). These impacts were observed to be in direct contrast in the case of ICG-001 treatment ( < 0.05).
Activation of the Wnt/β-catenin pathway mediates the high expression of to promote the resistance of NSCLC cells to icotinib. Thus, targeted inhibition of expression is of benefit for the treatment of NSCLC.
表皮生长因子受体酪氨酸激酶抑制剂（EGFR-TKI）被认为是治疗晚期非小细胞肺癌（NSCLC）的高度有效药物，患者在治疗 10 个月后常出现耐药性。本研究旨在揭示 NSCLC 对伊可替尼耐药的机制。
通过反复暴露于伊可替尼，建立伊可替尼耐药 PC-9 细胞（PC-9R），时间超过 12 个月。将程序性死亡配体 1 () 敲低质粒 (-KD)/过表达质粒 (-OE) 转染至 PC-9R 细胞，并用 Wnt 通路激动剂 CHIR99021 或 β-连环蛋白拮抗剂 ICG-001 处理。采用 3-(4,5-二甲基噻唑-2-基)-5-(3-羧甲氧基苯基)-2-(4-磺苯基)-2H 四唑比色法检测细胞对伊可替尼的敏感性。用 Transwell 和划痕试验评估细胞的侵袭和迁移能力。通过实时定量聚合酶链反应或 Western 印迹法定量检测 、基质金属蛋白酶（MMP）-2、MMP-9 和 Wnt/β-连环蛋白通路相关蛋白的表达。
PC-9 细胞和 PC-9R 细胞对伊可替尼的半抑制浓度（IC）分别为 1.73 μM 和 25.18 μM。PC-9R 细胞中 的表达、Wnt 家族成员 1（Wnt1）和 β-连环蛋白的表达均高于 PC-9 细胞（ < 0.05）。-OE 转染可使 PC-9R 细胞的 IC、迁移、侵袭以及 MMP-2 和 MMP-9 的表达升高（ < 0.05），而 -KD 转染则产生相反的效果（ < 0.05）。PC-9R 细胞用 CHIR99021 处理后， 的表达、β-连环蛋白、MMP-2 和 MMP-9 以及 IC、迁移和侵袭均增加（ < 0.05）。而用 ICG-001 处理时，情况则完全相反（ < 0.05）。
Wnt/β-连环蛋白通路的激活介导 高表达，促进 NSCLC 细胞对伊可替尼的耐药。因此，靶向抑制 的表达有助于治疗 NSCLC。
