背景:现代药理研究已经证实,葛花(PF)的植物衍生化合物具有针对肝脏损伤的显着生物活性,肿瘤和炎症。Kakkatin是从PF花中分离出的异黄酮多酚化合物。然而,kakkatin及其衍生物的抗肿瘤作用尚未得到很好的研究。
目的:设计并合成kakkatin衍生物[6-(hept-6-yn-1-基氧基)-3-(4-羟苯基)-7-甲氧基-4H-色烯-4-酮(HK)],探讨其抗肿瘤生物学活性。
方法:在kakkatin苯酚的羟基位置引入Hept-6-yn-1-烷基乙磺酸盐来代替氢,并制备了kakkatin的衍生物;3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四唑溴化物用于检测细胞活力,采用克隆形成试验检测细胞增殖,凋亡,坏死,通过膜联蛋白V/碘化丙啶染色和流式细胞术分析细胞周期。通过细胞划痕试验和transwell试验评价细胞迁移和侵袭能力。通过网络药理学和分子对接探讨HK对肝癌SMMC-7721细胞的潜在作用机制,最后采用实时荧光定量PCR方法对其潜在靶标进行验证,并对HK的生物活性进行评价。
结果:与kakkatin相比,修饰HK对胃癌MGC803细胞的抑制活性没有显著提高,但肝癌SMMC-7721细胞的抑制活性增加了约30倍,IC50值为2.5μM,抑瘤效果优于顺铂,这可以显著抑制克隆,肝癌SMMC-7721细胞的侵袭和转移,并诱导细胞凋亡和G2/M周期阻滞。其作用机制主要与cAMP通路中PDE3B和NFKB1靶蛋白的上调有关。
结论:HK对肝癌SMMC-7721细胞有明显的抑制作用,它们的作用靶点可能是cAMP通路中的PDE3B和NFKB1蛋白,使其成为治疗HCC的良好先导药物。
BACKGROUND: Modern pharmacological studies have confirmed that plant-derived compounds from Puerariae flos (PF) has significant biological activities against liver damage, tumors and inflammation. Kakkatin is an isoflavone polyphenolic compound isolated from PF flower. However, the effect of kakkatin and its derivatives on anti-tumor has not been well explored.
OBJECTIVE: To design and synthesize a kakkatin
derivative [6-(hept-6-yn-1-yloxy)-3-(4-hydroxyphenyl)-7-methoxy-4H-chromen-4-one (HK)] to explore its anti-tumor biological activity.
METHODS: Hept-6-yn-1-yl ethanesulfonate was introduced to replace hydrogen at the hydroxyl position of kakkatin phenol, and the
derivative of kakkatin was prepared; the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide was used to detect cell viability, a clone formation assay was adopted to detect cell proliferation, apoptosis, necrosis, and cell cycles were analyzed by Annexin V/propidium iodide staining and flow cytometry. Cell migration and invasion ability were evaluated by cell scratch assay and transwell assay. The potential mechanism of HK on hepatocellular carcinoma (HCC) SMMC-7721 cells was explored through network pharmacology and molecular docking, and finally real-time PCR assays was used to verify the potential targets and evaluate the biological activity of HK.
RESULTS: Compared with kakkatin, the modified HK did not significantly increase the inhibitory activity of gastric cancer MGC803 cells, but the inhibitory activity of HCC SMMC-7721 cells was increased by about 30 times, with an IC50 value of 2.5 μM, and the tumor inhibition effect was better than cisplatin, which could significantly inhibit the cloning, invasion and metastasis of HCC SMMC-7721 cells, and induce apoptosis and G2/M cycle arrest. Its mechanism of action is mainly related to the upregulation of PDE3B and NFKB1 target proteins in the cAMP pathway.
CONCLUSIONS: HK have a significant inhibitory effect on HCC SMMC-7721 cells, and the targets of their action may be PDE3B and NFKB1 proteins in the cAMP pathway, making it a good lead drug for the treatment of HCC.