Ophiopogonin D' induces RIPK1-dependent necroptosis in androgen-dependent LNCaP prostate cancer cells

Ophiopogonin D' (OPD') is a natural compound extracted from Ophiopogon japonicus, which is a plant used in traditional Chinese medicine. Our previous study has indicated that OPD' exhibits antitumor activity against androgen-independent prostate cancer (PCa), but the effects and the underlying molecular mechanism of action of OPD' in androgen-dependent PCa were unclear. In the present study, OPD' induced significant necroptosis in androgen-dependent LNCaP cancer cells by activating receptor-interacting serine/threonine-protein kinase 1 (RIPK1). Exposure to OPD' also increased Fas ligand (FasL)-dependent RIPK1 protein expression. The OPD'-induced necroptosis was inhibited by a RIPK1 inhibitor necrostatin-1, further supporting a role for RIPK1 in the effects of OPD´. The antitumor effects of OPD' were also inhibited by a mixed lineage kinase domain-like protein (MLKL) inhibitor necrosulfonamide. Following treatment with inhibitors of RIPK1 and MLKL, the effects of OPD' on LNCaP cells were inhibited in an additive manner. In addition, co-immunoprecipitation assays demonstrated that OPD' induced RIPK3 upregulation, leading to the assembly of a RIPK3-MLKL complex, which was independent of RIPK1. Furthermore, OPD' increased the expression of Fas-associated death domain, which is required to induce necroptosis in LNCaP cells. OPD' also regulated the expression levels of FasL, androgen receptor and prostate-specific antigen in a RIPK1-dependent manner. These results suggested that OPD' may exhibit potential as an anti-PCa agent by inducing RIPK1- and MLKL-dependent necroptosis.