背景:癌症仍然是一个全球性的健康问题,是延长预期寿命的重要障碍。恶性细胞迅速发展耐药性,导致许多临床治疗失败。药用植物作为经典药物发现的替代来对抗癌症的重要性是众所周知的。布鲁氏菌是一种传统上用于治疗癌症的非洲药用植物,痢疾,疟疾,腹泻,胃痛,蠕虫感染,发烧,和哮喘。本工作旨在鉴定布鲁氏菌抗痢疾在广泛的癌细胞系上的细胞毒性成分,并证明最活跃样品的凋亡诱导模式。
方法:通过柱色谱法从布鲁氏菌抗痢疾的叶(BAL)和茎(BAS)提取物中分离出7种植物化学物质,并使用光谱技术进行结构阐明。粗提物和化合物对9种人癌细胞系的抗增殖作用通过刀天青减少试验(RRA)来评价。通过Caspase-Glo测定评估细胞系中的活性。细胞周期分布,通过碘化丙啶(PI)染色凋亡,线粒体膜电位(MMP)通过5,5',6,6'-四氯-1,1',3,3'-四乙基苯并咪唑基碳花青碘化物(JC-1)染色,和活性氧(ROS)通过2',7'-二氯二氢荧光二乙酸酯(H2DCFH-DA)染色,通过流式细胞术进行了研究。
结果:对植物药(BAL和BAS)的植物化学研究导致了7种化合物的分离。BAL及其成分3,(3-(3-甲基-1-氧代-2-丁烯基))1H吲哚(1)和hydnocarpin(2),以及参考化合物,阿霉素,对9种癌细胞系具有抗增殖活性。BAL的IC50值从17.42µg/mL(针对CCRF-CEM白血病细胞)到38.70µg/mL(针对HCT116p53-/-结肠腺癌细胞)不等,化合物1从19.11μM(针对CCRF-CEM细胞)到47.50μM(针对MDA-MB-231-BCRP腺癌细胞),化合物2从4.07μM(针对MDA-MB-231-pcDNA细胞)到11.44μM(针对HCT116p53+/+细胞)。有趣的是,还观察到抗性癌细胞对化合物2的超敏反应。半胱天冬酶激活介导BAL和hydnocarpin诱导CCRF-CEM细胞凋亡,MMP的改变,并增加ROS水平。
结论:BAL及其成分,主要是化合物2,是布鲁氏菌抗痢疾的潜在抗增殖产品。从发现新的抗增殖剂以对抗对抗癌药物的抗性的角度来看,其他研究将是必要的。
BACKGROUND: Cancer remains a global health concern and constitutes an important barrier to increasing life expectancy. Malignant cells rapidly develop drug resistance leading to many clinical therapeutic failures. The importance of medicinal plants as an alternative to classical drug discovery to fight cancer is well known. Brucea antidysenterica is an African medicinal plant traditionally used to treat cancer, dysentery, malaria, diarrhea, stomach aches, helminthic infections, fever, and asthma. The present work was designed to identify the cytotoxic constituents of Brucea antidysenterica on a broad range of cancer cell lines and to demonstrate the mode of induction of apoptosis of the most active samples.
METHODS: Seven phytochemicals were isolated from the leaves (BAL) and stem (BAS) extract of Brucea antidysenterica by column chromatography and structurally elucidated using spectroscopic techniques. The antiproliferative effects of the crude extracts and compounds against 9 human cancer cell lines were evaluated by the resazurin reduction assay (RRA). The activity in cell lines was assessed by the Caspase-Glo assay. The cell cycle distribution, apoptosis via propidium iodide (PI) staining, mitochondrial membrane potential (MMP) through 5,5\',6,6\'-tetrachloro-1,1\',3,3\'-tetraethylbenzimidazolylcarbocyanine iodide (JC-1) staining, and the reactive oxygen species (ROS) via 2´,7´-dichlorodihydrofluoresceine diacetate (H2DCFH-DA) staining, were investigated by flow cytometry.
RESULTS: Phytochemical studies of the botanicals (BAL and BAS) led to the isolation of seven compounds. BAL and its constituents 3, (3-(3-Methyl-1-oxo-2-butenyl))1H indole (1) and hydnocarpin (2), as well as the reference compound, doxorubicin, had antiproliferative activity against 9 cancer cell lines. The IC50 values varied from 17.42 µg/mL (against CCRF-CEM leukemia cells) to 38.70 µg/mL (against HCT116 p53-/- colon adenocarcinoma cells) for BAL, from 19.11 µM (against CCRF-CEM cells) to 47.50 µM (against MDA-MB-231-BCRP adenocarcinoma cells) for compound 1, and from 4.07 µM (against MDA-MB-231-pcDNA cells) to 11.44 µM (against HCT116 p53+/+ cells) for compound 2. Interestingly, hypersensitivity of resistant cancer cells to compound 2 was also observed. BAL and hydnocarpin induced apoptosis in CCRF-CEM cells mediated by caspase activation, the alteration of MMP, and increased ROS levels.
CONCLUSIONS: BAL and its constituents, mostly compound 2, are potential antiproliferative products from Brucea antidysenterica. Other studies will be necessary in the perspective of the discovery of new antiproliferative agents to fight against resistance to anticancer drugs.