Cell-Based Drug and Health Product Development Research Unit, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Bangkok 10330, Thailand.
Department of Pharmacology and Physiology, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Bangkok 10330, Thailand.
Molecules. 2021 Jan 26;26(3):638. doi: 10.3390/molecules26030638.
Cancer metastasis is the major cause of about 90% of cancer deaths. As epithelial-to-mesenchymal transition (EMT) is known for potentiating metastasis, this study aimed to elucidate the effect of ovalitenone on the suppression of EMT and metastasis-related behaviors, including cell movement and growth under detached conditions, and cancer stem cells (CSCs), of lung cancer cells.
Cell viability and cell proliferation were determined by 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazo-liumbromide (MTT) and colony formation assays. Cell migration and invasion were analyzed using a wound-healing assay and Boyden chamber assay, respectively. Anchorage-independent cell growth was determined. Cell protrusions (filopodia) were detected by phalloidin-rhodamine staining. Cancer stem cell phenotypes were assessed by spheroid formation. The proteins involved in cell migration and EMT were evaluated by Western blot analysis and immunofluorescence staining.
Ovalitenone was used at concentrations of 0-200 μM. While it caused no cytotoxic effects on lung cancer H460 and A549 cells, ovalitenone significantly suppressed anchorage-independent growth, CSC-like phenotypes, colony formation, and the ability of the cancer to migrate and invade cells. The anti-migration activity was confirmed by the reduction of filopodia in the cells treated with ovalitenone. Interestingly, we found that ovalitenone could significantly decrease the levels of N-cadherin, snail, and slug, while it increased E-cadherin, indicating EMT suppression. Additionally, the regulatory signaling of focal adhesion kinase (FAK), ATP-dependent tyrosine kinase (AKT), the mammalian target of rapamycin (mTOR), and cell division cycle 42 (Cdc42) was suppressed by ovalitenone.
The results suggest that ovalitenone suppresses EMT via suppression of the AKT/mTOR signaling pathway. In addition, ovalitenone exhibited potential for the suppression of CSC phenotypes. These data reveal the anti-metastasis potential of the compound and support the development of ovalitenone treatment for lung cancer therapy.
癌症转移是导致约 90%癌症死亡的主要原因。由于上皮-间质转化（EMT）被认为可以增强转移，因此本研究旨在阐明卵形倍半萜酮对抑制肺癌细胞 EMT 和转移相关行为（包括在分离条件下的细胞运动和生长，以及癌症干细胞（CSC））的影响。
通过 3-[4,5-二甲基噻唑-2-基]-2,5 二苯基四唑溴盐（MTT）和集落形成测定来确定细胞活力和细胞增殖。通过划痕愈合测定和 Boyden 室测定分别分析细胞迁移和侵袭。测定无锚定依赖性细胞生长。通过鬼笔环肽罗丹明染色检测细胞突起（丝状伪足）。通过球体形成评估癌症干细胞表型。通过 Western blot 分析和免疫荧光染色评估参与细胞迁移和 EMT 的蛋白质。
使用 0-200μM 的卵形倍半萜酮。虽然它对肺癌 H460 和 A549 细胞没有细胞毒性作用，但卵形倍半萜酮显著抑制无锚定依赖性生长、CSC 样表型、集落形成以及癌细胞迁移和侵袭细胞的能力。通过用卵形倍半萜酮处理减少细胞中的丝状伪足证实了抗迁移活性。有趣的是，我们发现卵形倍半萜酮可以显著降低 N-钙粘蛋白、snail 和 slug 的水平，同时增加 E-钙粘蛋白，表明 EMT 抑制。此外，卵形倍半萜酮抑制粘着斑激酶（FAK）、ATP 依赖性酪氨酸激酶（AKT）、雷帕霉素靶蛋白（mTOR）和细胞分裂周期 42（Cdc42）的调节信号。
结果表明，卵形倍半萜酮通过抑制 AKT/mTOR 信号通路抑制 EMT。此外，卵形倍半萜酮表现出抑制 CSC 表型的潜力。这些数据揭示了该化合物的抗转移潜力，并支持开发卵形倍半萜酮治疗肺癌的治疗方法。
