Natural Products Institute, University of the West Indies, Mona, Kingston 7, Jamaica.
Biotechnolgy Centre, University of the West Indies, Mona, Kingston 7, Jamaica.
BMC Complement Med Ther. 2020 Dec 10;20(1):375. doi: 10.1186/s12906-020-03130-z.
Annona muricata L. was identified as a popular medicinal plant in treatment regimens among cancer patients in Jamaica by a previously conducted structured questionnaire. Ethnomedically used plant parts, were examined in this study against human prostate cancer cells for the first time and mechanisms of action elucidated for the most potent of them, along with the active phytochemical, annonacin.
Nine extracts of varying polarity from the leaves and bark of A. muricata were assessed initially for cytotoxicity using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay on PC-3 prostate cancer cells and the ethyl acetate bark (EAB) extract was identified as the most potent. EAB extract was then standardized for annonacin content using High-performance Liquid Chromatography - Mass Spectrometry (HPLC-MS) and shown to be effective against a second prostate cancer cell line (DU-145) also. The mode of cell death in DU-145 cells were assessed via several apoptotic assays including induction of increased reactive oxygen species (ROS) production, reduction of mitochondrial membrane potential, activation of caspases and annexin V externalization combined with morphological observations using confocal microscopy. In addition, the potential to prevent metastasis was examined via inhibition of cell migration, vascular endothelial growth factor (VEGF) and angiogenesis using the chorioallantoic membrane assay (CAM).
Annonacin and EAB extract displayed selective and potent cytotoxicity against the DU-145 prostate carcinoma cells with IC values of 0.1 ± 0.07 μM and 55.501 ± 0.55 μg/mL respectively, without impacting RWPE-1 normal prostate cells, in stark contrast to chemotherapeutic docetaxel which lacked such selectivity. Docetaxel's impact on the cancerous DU-145 was improved by 50% when used in combination with EAB extract. Insignificant levels of intracellular ROS content, depolarization of mitochondrial membrane, Caspase 3/7 activation, annexin V content, along with stained morphological evaluations, pointed to a non-apoptotic mode of cell death. The extract at 50 μg/mL deterred cell migration in the wound-healing assay, while inhibition of angiogenesis was displayed in the CAM and VEGF inhibition assays for both EAB (100 μg /mL) and annonacin (0.5 μM).
Taken together, the standardized EAB extract and annonacin appear to induce selective and potent cell death via a necrotic pathway in DU-145 cells, while also preventing cell migration and angiogenesis, which warrant further examinations for mechanistic insights and validity in-vivo.
安农拉库拉 L. 被鉴定为牙买加癌症患者治疗方案中的一种流行药用植物，此前进行的一项结构式问卷调查发现了这一点。本研究首次检测了该植物不同极性的叶和树皮提取物对人前列腺癌细胞的作用，同时还对最有效的提取物及其活性植物化学物质安诺那辛进行了作用机制的研究。
使用 MTT（3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四唑溴化物）测定法对安农拉库拉的 9 种不同极性提取物对 PC-3 前列腺癌细胞的细胞毒性进行了初步评估，结果表明，乙酸乙酯树皮提取物（EAB）最有效。然后，使用高效液相色谱-质谱联用（HPLC-MS）对 EAB 提取物进行了安诺那辛含量的标准化，并发现其对另一种前列腺癌细胞系（DU-145）也有效。通过几种凋亡测定法（包括诱导活性氧（ROS）产生增加、线粒体膜电位降低、半胱天冬酶激活和膜联蛋白 V 外化，并结合共聚焦显微镜下的形态学观察）评估 DU-145 细胞中的细胞死亡模式。此外，通过抑制细胞迁移、血管内皮生长因子（VEGF）和血管生成，使用鸡胚尿囊膜（CAM）测定法来检测预防转移的潜力。
安诺那辛和 EAB 提取物对 DU-145 前列腺癌细胞具有选择性和强效的细胞毒性，IC 值分别为 0.1±0.07μM 和 55.501±0.55μg/mL，而对 RWPE-1 正常前列腺细胞则没有影响，与缺乏这种选择性的化疗药物多西他赛形成鲜明对比。当与 EAB 提取物联合使用时，多西他赛对癌症 DU-145 的影响提高了 50%。细胞内 ROS 含量、线粒体膜去极化、半胱天冬酶 3/7 激活、膜联蛋白 V 含量以及染色形态学评估均表明存在非凋亡细胞死亡模式。提取物在 50μg/mL 时抑制了划痕愈合试验中的细胞迁移，而在 CAM 和 VEGF 抑制试验中，EAB（100μg/mL）和安诺那辛（0.5μM）均显示出抑制血管生成的作用。
综上所述，标准化的 EAB 提取物和安诺那辛似乎通过 DU-145 细胞的坏死途径诱导选择性和强效的细胞死亡，同时还能防止细胞迁移和血管生成，这需要进一步的机制研究和体内验证。
