ERα and Wnt/β-catenin pathways are critical for the progression of most endometrial cancers. We aimed to investigate the cytotoxic and apoptotic effects of tamoxifen and quinazoline derivative drugs of doxazosin and erlotinib, and their roles in ERα and Wnt/β-catenin signaling pathways in human endometrial cancer RL 95-2 cell. 3-(4,5-Dimethylthiazol-2yl)−2,5-diphenyltetrazolium bromide assay and xCELLigence systems were performed to evaluate cytotoxicity. Furthermore, apoptotic induction was tested by Annexin V analysis. Caspase-3 and -9 activity and changes in the mitochondrial membrane potential were evaluated. The level of reactive oxygen species was measured by incubating with dichlorofluorescein diacetate. Protein ratios of p-ERα/ERα, GSK3β/p-GSK3β, and p-β-catenin/β-catenin and expression levels of ESR1, EGFR, c-Myc genes were evaluated to elucidate mechanisms in signaling pathways. We found that the tested drugs showed cytotoxic and apoptotic effects in the cells. Doxazosin significantly reduced ESR1 expression, slightly reduced the p-β-catenin/β-catenin ratio and c-Myc expression. Erlotinib significantly increased c-Myc expression while significantly decreasing the p-β-catenin/β-catenin and p-ERα/ERα ratio, and ESR1 expression. However, we observed that the cells develop resistance to erlotinib over a certain concentration, suggesting that ERα, ESR1, EGFR, and c-Myc may be a new target for overcoming drug resistance in the treatment of endometrial cancer. We also observed that erlotinib and doxazosin play an important role in the ERα signaling pathway and can act as potent inhibitors of PKA and/or tyrosine kinase in the Wnt/β-catenin signaling pathway in RL 95-2 cell. In conclusion, doxazosin and erlotinib may have a possible therapeutic potential in human endometrial cancer.

中文翻译：

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多沙唑嗪和厄洛替尼在子宫内膜癌细胞中具有抗癌作用，并在 ERα 和 Wnt/β-catenin 信号通路中发挥重要作用

ERα 和 Wnt/β-catenin 通路对大多数子宫内膜癌的进展至关重要。我们旨在研究他莫昔芬和多沙唑嗪和厄洛替尼的喹唑啉衍生物药物的细胞毒性和凋亡作用，以及它们在人子宫内膜癌RL 95-2细胞中ERα和Wnt/β-catenin信号通路中的作用。进行了 3-(4,5-Dimethylthiazol-2yl)−2,5-diphenyltetrazolium bromide 测定和 xCELLigence 系统以评估细胞毒性。此外，通过膜联蛋白 V 分析测试了凋亡诱导。评估了 Caspase-3 和 -9 的活性以及线粒体膜电位的变化。通过与二氯荧光素二乙酸盐一起孵育来测量活性氧的水平。p- ERα /ERα 、 GSK3β / p-GSK3的蛋白质比例评估了β和 p- β -catenin/ β -catenin 以及ESR1、EGFR、c-Myc基因的表达水平，以阐明信号通路中的机制。我们发现受试药物在细胞中表现出细胞毒性和凋亡作用。多沙唑嗪显着降低ESR1表达，略微降低 p - β - catenin/ β -catenin 比率和c-Myc表达。厄洛替尼显着增加c-Myc表达，同时显着降低 p- β -catenin/ β -catenin 和 p-ER α /ER α比率，以及ESR1表达。然而，我们观察到细胞在一定浓度下对厄洛替尼产生耐药性，提示ERα 、ESR1、EGFR和c-Myc可能是克服子宫内膜癌治疗耐药性的新靶点。我们还观察到厄洛替尼和多沙唑嗪在 ER α信号通路中发挥重要作用，并且可以作为RL 95-2 细胞Wnt/ β-连环蛋白信号通路中 PKA 和/或酪氨酸激酶的有效抑制剂。总之，多沙唑嗪和厄洛替尼可能具有治疗人类子宫内膜癌的潜力。