Eldecalcitol (ED-71) is a new type of vitamin D analog, and vitamin D has been reported to have therapeutic effects in infectious disease, autoimmune disease, and cancer. However, the anti-cancer effect of ED-71 remains unclear. The objective of this study was to explore the anti-cancer effect of ED-71 in human osteosarcoma cells and to identify the related mechanism. The CCK8 assay results showed that ED-71 inhibited MG-63 cell viability in dose and time dependent manners. Cloning and Transwell invasion assays showed that ED-71 inhibited clonal and invasion ability of MG-63 cells. Flow cytometry results showed ED-71 the G2/M cycle arrest rate, apoptosis, and intracellular ROS. Western blot was used to detect cleaved-caspase-3, Bax, Bcl-2, LC3-II/LC3-I, and P62 levels and the mTOR pathway. The increase of LC3-II and P62 indicated that ED-71 induced the formation of autophagosomes and inhibited autophagy flux. Furthermore, ED-71-induced apoptosis was weakened after adding 3-methyladenine and ED-71-induced early autophagy was weakened by caspase-3 inhibitor (Z-VAD-FMK), which indicated the two processes active each other in the presence of ED-71. Furthermore, N-acetylcysteine (NAC) pretreatment reversed the ED-71-treatment outcomes, including increased apoptosis and autophagy and inhibition of the PI3K/Akt/mTOR pathway. In conclusion, our results reveal that ED-71 induced G2/M arrest, apoptosis and autophagy in MG-63 cells by accumulating ROS to suppress the PI3K/Akt/mTOR signaling pathway

Eldecalcitol (ED-71) 是一种新型的维生素 D 类似物，据报道维生素 D 对感染性疾病、自身免疫性疾病和癌症具有治疗作用。然而，ED-71 的抗癌作用仍不清楚。本研究的目的是探讨ED-71对人骨肉瘤细胞的抗癌作用并确定相关机制。CCK8测定结果显示ED-71以剂量和时间依赖性方式抑制MG-63细胞活力。克隆和Transwell侵袭实验表明ED-71抑制MG-63细胞的克隆和侵袭能力。流式细胞术结果显示 ED-71 具有 G2/M 周期阻滞率、细胞凋亡和细胞内 ROS。蛋白质印迹用于检测 cleaved-caspase-3、Bax、Bcl-2、LC3-II/LC3-I 和 P62 水平以及 mTOR 通路。LC3-II和P62的增加表明ED-71诱导了自噬体的形成并抑制了自噬通量。此外，加入 3-甲基腺嘌呤后 ED-71 诱导的细胞凋亡减弱，而 caspase-3 抑制剂减弱了 ED-71 诱导的早期自噬。Z -VAD-FMK），这表明两个进程在 ED-71 存在下相互激活。此外，N-乙酰半胱氨酸（NAC）预处理逆转了 ED-71 治疗的结果，包括增加细胞凋亡和自噬以及抑制 PI3K/Akt/mTOR 通路。总之，我们的结果表明，ED-71 通过积累 ROS 抑制 PI3K/Akt/mTOR 信号通路诱导 MG-63 细胞的 G2/M 期阻滞、凋亡和自噬