Background and Purpose

Chemotherapy resistance is a major obstacle for the effective treatment of cancers. Although several studies have described the anticancer properties of rosemary extract and its components, the detailed mechanisms of action are poorly understood.

Methods

Activity-guided fractionation and repeated chromatographic separation of the n-hexane fraction of the aqueous methanol extract over silica gel, RP C18, and Sephadex LH-20 led to the isolation of three compounds. The structures of the compounds were determined using ¹H, ¹³C, and two-dimensional nuclear magnetic resonance spectroscopy, mass spectroscopy, and infrared spectroscopy. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay was used to evaluate the cytotoxicity of these compounds. Cell cycle, apoptotic cell populations, and mitochondrial membrane potential were analyzed by flow cytometry. Western blot analysis was conducted to detect apoptosis-related proteins.

Results

An abietane diterpenoid, sageone (1), an icetexane diterpenoid, (–)-barbatusol (2), and a monoterpene, (+)-verbenone (3), were identified. Of these compounds, sageone (1) showed cytotoxicity against SNU-1 cells with an IC₅₀ of 9.45 ± 1.33 µM. Sageone reduced the expression of Akt dramatically, as opposed to cisplatin, which increased phosphorylated Akt. Sageone combined with a subtoxic dose of cisplatin had synergistic effects on apoptosis induction in SNU-1 cells, as confirmed by calculating the combination index. Co-treatment was significantly more effective than monotherapy at reducing cell viability and inducing apoptosis, as determined by analyzing DNA fragmentation. The combined treatment of sageone and cisplatin markedly reduced Akt expression and phosphorylation, accompanied by increases in cleaved caspase-3, -9 and PARP.

Conclusion

This is the first time compounds 1 and 2 have been isolated from R. officinalis. Sageone induced apoptosis in SNU-1 human gastric cancer cells and notably enhanced the cytotoxicity of cisplatin in SNU-1 cells, which are known to be resistant to cisplatin. These findings suggest that sageone represents a promising anticancer agent against gastric cancer that warrants further study.

中文翻译：

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迷迭香中的二萜Sageone与顺铂在SNU-1人胃癌细胞中的细胞毒性协同作用

背景和目的

耐药性是有效治疗癌症的主要障碍。尽管有几项研究描述了迷迭香提取物及其成分的抗癌特性，但对其详细的作用机理了解甚少。

方法

活动引导分级和重复色谱分离Ñ含水甲醇萃取物，用硅胶RP C18的己烷馏分，和Sephadex LH-20导致了三种化合物的分离。使用¹ H，¹³ C和二维核磁共振波谱，质谱和红外光谱确定化合物的结构。使用3-（4,5-二甲基噻唑-2-基）-2,5-二苯基溴化四唑测定这些化合物的细胞毒性。通过流式细胞仪分析细胞周期，凋亡细胞群和线粒体膜电位。进行蛋白质印迹分析以检测凋亡相关蛋白。

结果

确定了松果油二萜，鼠尾草酮（1），冰tex烷二萜，（-）-巴贝妥（2）和单萜（+）-马鞭酮（3）。在这些化合物中，sageone（1）对ICU ₅₀对SNU-1细胞具有细胞毒性为9.45±1.33 µM。与顺铂相反，Sageone显着降低了Akt的表达，而顺铂则增加了磷酸化的Akt。通过计算组合指数证实，骨酮与亚毒性剂量的顺铂联合对SNU-1细胞的凋亡诱导具有协同作用。通过分析DNA片段测定，共治疗在降低细胞活力和诱导细胞凋亡方面比单药治疗更为有效。骨酮和顺铂的联合治疗显着降低了Akt表达和磷酸化，并伴有裂解的caspase-3，-9和PARP的增加。

结论

这是首次从鼠李糖中分离出化合物1和2。姜黄素诱导SNU-1人胃癌细胞凋亡，并显着增强顺铂在SNU-1细胞中的细胞毒性，已知SNU-1细胞对顺铂具有抗性。这些发现表明，age骨酮是一种有前途的抗胃癌抗癌药，值得进一步研究。