Chrysanthemulide A induces apoptosis through DR5 upregulation via JNK-mediated autophagosome accumulation in human osteosarcoma cells

Osteosarcoma is the most frequent malignant primary bone tumor, and it generally develops a multidrug resistance. Chrysanthemulide A (CA) is a sesquiterpenoid from the herb Chrysanthemum indicum that has demonstrated a great anti-osteosarcoma potential. In this study, CA-induced apoptotic cell death resulted in the activation of the caspase-8-mediated caspase cascade, as evidenced by the cleavage of the substrate protein Bid and the caspase-8 inhibitor Z-VAD-FMK. The CA treatment upregulated the expression of death receptor 5 (DR5) in both whole cells and the cell membrane. Blocking DR5 expression by the small interfering RNA (siRNA) treatment decreased the caspase-8-mediated caspase cascade and efficiently attenuated CA-induced apoptosis, suggesting the critical role of DR5 in CA-induced apoptotic cell death. CA-induced upregulation of the DR5 protein was accompanied by the accumulation of LC3B-II, indicating the formation of autophagosomes. Importantly, DR5 upregulation was mediated by transcriptionally controlled autophagosome accumulation, as blockade of autophagosomes by LC3B or ATG-5 siRNA substantially decreased DR5 upregulation. Furthermore, CA activated the c-Jun N-terminal kinase (JNK) signaling pathway, and treatment with JNK siRNAs or inhibitor SP600125 significantly attenuated CA-mediated autophagosome accumulation and DR5-mediated cell apoptosis. Finally, CA sensitized the osteosarcoma cells to the DR5 ligand tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptotic cell death. Above all, these results suggest that CA induces apoptosis through upregulating DR5 via JNK-mediated autophagosome accumulation and that combined treatment with CA and TRAIL might be a promising therapy for osteosarcoma.