Scutebarbatine A (SBT-A), a diterpenoid alkaloid, has exerted cytotoxicity on hepatocellular carcinoma cells in our previous works. Here, the antitumor activity of SBT-A in breast cancer cells and the underlying mechanism were explored. The anti-proliferative effect of SBT-A was measured by trypan blue staining, 5-ethynyl-2′-deoxyuridine (EdU) incorporation and colony formation assay. DNA double-strand breaks (DSBs) were evaluated by observing the nuclear focus formation of γ-H2AX. Cell cycle distribution was assessed by flow cytometry. Apoptosis was determined by a TUNEL assay. Intracellular reactive oxygen species (ROS) generation and superoxide production were measured with 2′, 7′-dichlorofluorescein diacetate (DCFH-DA) and dihydroethidium (DHE) staining, respectively. The results indicated that SBT-A showed a dose-dependent cytotoxic effect against breast cancer cells while revealing less toxicity toward MCF-10A breast epithelial cells. Moreover, SBT-A remarkably induced DNA damage, cell cycle arrest and apoptosis in both MDA-MB-231 and MCF-7 cells. SBT-A treatment increased the levels of ROS and cytosolic superoxide production. Pretreatment with N-acetyl cysteine (NAC), a ROS scavenger, was sufficient to block viability reduction, DNA damage, apoptosis and endoplasmic reticulum (ER) stress caused by SBT-A. By exposure to SBT-A, the phosphorylation of c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (p38MAPK) was upregulated, while the phosphorylation of extracellular signal-regulated kinase (ERK) was downregulated. In addition, SBT-A inhibited the EGFR signaling pathway by decreasing EGFR expression and phosphorylation of Akt and p70S6K. As mentioned above, SBT-A has a potent inhibitory effect on breast cancer cells through induction of DNA damage, apoptosis and ER stress via ROS generation and modulation of MAPK and EGFR/Akt signaling pathway.

Scutebarbatine A (SBT-A) 是一种二萜类生物碱，在我们之前的研究中对肝细胞癌细胞具有细胞毒性。在此，探讨了 SBT-A 在乳腺癌细胞中的抗肿瘤活性及其潜在机制。通过台盼蓝染色、5-乙炔基-2'-脱氧尿苷 (EdU) 掺入和集落形成测定来测量 SBT-A 的抗增殖作用。通过观察 γ-H2AX 的核焦点形成来评估 DNA 双链断裂 (DSB)。通过流式细胞术评估细胞周期分布。通过 TUNEL 测定确定细胞凋亡。用 2', 7'-二氯荧光素二乙酸酯 (DCFH-DA) 和二氢乙锭测量细胞内活性氧(ROS) 的产生和超氧化物的产生(DHE) 染色，分别。结果表明，SBT-A 对乳腺癌细胞显示出剂量依赖性细胞毒作用，同时显示出对 MCF-10A 乳腺上皮细胞的毒性较小。此外，SBT-A 在 MDA-MB-231 和 MCF-7 细胞中显着诱导 DNA 损伤、细胞周期停滞和细胞凋亡。SBT-A 处理增加了 ROS 和细胞溶质超氧化物产生的水平。用 ROS 清除剂 N-乙酰半胱氨酸 (NAC) 预处理足以阻止活力降低、DNA 损伤、细胞凋亡和由 SBT-A 引起的内质网 (ER) 应激。通过暴露于 SBT-A，c-Jun N 末端激酶 (JNK) 和 p38 丝裂原活化蛋白激酶 (p38MAPK) 的磷酸化被上调，而细胞外信号调节激酶 (ERK) 的磷酸化被下调。此外，SBT-A 通过降低 EGFR 表达和 Akt 和 p70S6K 的磷酸化来抑制 EGFR 信号通路。如上所述，SBT-A 通过 ROS 的产生和 MAPK 和 EGFR/Akt 信号通路的调节诱导 DNA 损伤、细胞凋亡和 ER 应激，从而对乳腺癌细胞具有有效的抑制作用。