Alantolactone induces concurrent apoptosis and GSDME-dependent pyroptosis of anaplastic thyroid cancer through ROS mitochondria-dependent caspase pathway

• Published in: Phytomedicine (Stuttgart) 2023-01, Vol.108, p.154528-154528, Article 154528
• Main Authors: Hu, Yiqun, Wen, Qingliang, Cai, Yefeng, Liu, Yunye, Ma, Wenli, Li, Qinglin, Song, Fahuan, Guo, Yawen, Zhu, Lei, Ge, Jingyan, Zeng, Qian, Wang, Jiahui, Yin, Changtian, Zheng, Guowan, Ge, Minghua
• Format: Article
• Language: English
• Subjects: Alantolactone; Anaplastic thyroid cancer; animals; antineoplastic activity; Apoptosis; calreticulin; caspase-3; caspase-9; cell culture; cell cycle checkpoints; culture media; cyclins; enzyme activity; flow cytometry; fluorescent antibody technique; immunohistochemistry; immunotherapy; Inula helenium; lactate dehydrogenase; medicinal plants; membrane potential; mitochondrial membrane; Pyroptosis; ROS; terpenoids; thyroid neoplasms; Western blotting; xenotransplantation
• ISSN: 0944-7113; 1618-095X
• DOI: 10.1016/j.phymed.2022.154528

•Alantolactone exhibits considerable anti-tumor effects of anaplastic thyroid cancer both in vitro and in vivo without any adverse effects.•Alantolactone induces concurrent apoptosis and pyroptosis in anaplastic thyroid cancer cells through activated-caspase 3 mediated both cleavage of PARP and GSDME.•Alantolactone induces immunogenic cell death with cell membrane translocation of calreticulin and release of IL-1β. Anaplastic thyroid cancer (ATC) is one of the fatal cancers and has not effective treatments. Alantolactone (ATL), a terpenoid extracted from traditional Chinese medicinal herb Inula helenium L., confers significant anti-inflammatory, antibacterial and antitumor activity. However, the activity and mechanisms of ATL in ATC remain unclear. To investigate the potential anti-ATC effects in vitro and in vivo and the mechanisms involved. The anti-proliferative activity of Alantolactone (ATL) against ATC cells was analyzed through CCK-8 and colony formation assays. Flow cytometry assay was performed to assess the cell cycle, cell apoptosis, ROS, and mitochondrial membrane potential (ΔΨm), whereas the cellular localization of cytochrome c and calreticulin were determined using cellular immunofluorescence assays. The lactate dehydrogenase (LDH) enzyme activity in the cell culture medium was measured using a commercial LDH kit, whereas ELISA was conducted to assess the secretory function of IL-1β. Western blot assays were conducted to determine the expression or regulation of proteins associated with apoptosis and pyroptosis. Subcutaneous tumor model of nude mice was established to evaluate the anticancer activity of ATL in vivo. The expression of Ki67, cyclin B1, cleaved-PARP, cleaved-caspase 3, and IL-1β in the animal tumor tissues was profiled using immunohistochemistry analyses. Our data showed that ATL significantly inhibited the proliferation and colony formation activity of ATC cells. ATL induced ATC cell cycle arrest at G2/M phase, and downregulated the expression of cyclin B1 and CDC2. Furthermore, ATL induced concurrent apoptosis and pyroptosis in the ATC cells, and the cleavage of PARP and GSDME. It also significantly increased the release of LDH and IL-1β. Mechanically, ATL-mediated increase in ROS suppressed the Bcl-2/Bax ratio, downregulated the mitochondrial membrane potential and increased the release of cytochrome c, leading to caspase 9 and caspase 3 cleavage. We also found that ATL induced the translocation of an immunogenic cell death