阿莫替尼是一种重要的第三代表皮生长因子受体酪氨酸激酶抑制剂（EGFR-TKI），对 EGFR 致敏和 T790M 耐药突变表现出高选择性。阿莫替尼耐药是临床使用的主要障碍。黄芩素具有抗肿瘤特性，但其对阿莫替尼耐药的非小细胞肺癌（NSCLC）的抗肿瘤作用机制仍不清楚。采用CCK-8法检测H1975/AR和HCC827/AR细胞治疗后的存活率。使用不同浓度的黄芩素、阿莫替尼或其组合 24 小时。使用集落形成测定和流式细胞术分析处理细胞的集落形成能力、细胞凋亡和细胞内活性氧（ROS）水平的变化。通过Western blotting检测细胞内蛋白表达的变化。在黄芩素和/或阿莫替尼处理的细胞中观察到 NAC 预处理对增殖、凋亡和 PI3K/Akt 信号通路的影响。用黄芩素 (20 mg/kg) 或阿莫替尼 (15 mg/kg) 处理皮下 HCC827/AR 细胞异种移植物的裸鼠模型，并测量肿瘤体积和体重变化。黄芩素和阿莫替尼均抑制 HCC827 的活力/AR 和 H1975/AR 细胞呈浓度依赖性。与单独使用黄芩素或阿莫替尼相比，两种药物联合应用可显着减弱细胞增殖；引发细胞凋亡；导致 Caspase-3、PARP 和 Caspase-9 裂解；下调 p-PI3K 和 p-Akt 的蛋白表达；并显着抑制裸鼠肿瘤生长。此外，黄芩素与阿莫替尼联合导致活性氧（ROS）大量积累，与N-乙酰基-L-半胱氨酸（ROS去除剂）预孵育可防止增殖并抑制细胞凋亡诱导，部分恢复p-PI3K的下降黄芩素和阿莫替尼联合使用，可以通过ROS介导的PI3K/Akt通路提高对阿莫替尼耐药的NSCLC的抗肿瘤活性。版权所有©2024 Chen，Zhang，Cong，Wang，Li，Wang，Zhao和Sun。

Almonertinib is an important third-generation epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) exhibiting high selectivity to EGFR-sensitizing and T790M-resistant mutations. Almonertinib resistance is a major obstacle in clinical use. Baicalein possesses antitumor properties, but its mechanism of antitumor action against almonertinib-resistant non-small cell lung cancer (NSCLC) remains unelucidated.CCK-8 assay was used to examine the survival rate of H1975/AR and HCC827/AR cells following treatment for 24 h with different concentrations of baicalein, almonertinib or their combination. The changes in colony formation ability, apoptosis, and intracellular reactive oxygen species (ROS) levels of the treated cells were analyzed using colony formation assay and flow cytometry. Western blotting was performed to detect the changes in protein expressions in the cells. The effects of pre-treatment with NAC on proliferation, apoptosis, and PI3K/Akt signaling pathway were observed in baicalein- and/or almonertinib-treated cells. A nude mouse model bearing subcutaneous HCC827/AR cell xenograft were treated with baicalein (20 mg/kg) or almonertinib (15 mg/kg), and the tumor volume and body mass changes was measured.Both baicalein and almonertinib represses the viability of HCC827/AR and H1975/AR cells in a concentration-dependent manner. Compared with baicalein or almonertinib alone, the combined application of the two drugs dramatically attenuates cell proliferation; triggers apoptosis; causes cleavage of Caspase-3, PARP, and Caspase-9; downregulates the protein expressions of p-PI3K and p-Akt; and significantly inhibits tumor growth in nude mice. Furthermore, baicalein combined with almonertinib results in massive accumulation of reactive oxygen species (ROS) and preincubation with N-acetyl-L-cysteine (ROS remover) prevents proliferation as well as inhibits apoptosis induction, with partial recovery of the decline of p-PI3K and p-Akt.The combination of baicalein and almonertinib can improve the antitumor activity in almonertinib-resistant NSCLC through the ROS-mediated PI3K/Akt pathway.Copyright © 2024 Chen, Zhang, Cong, Wang, Li, Wang, Zhao and Sun.