Low-dose Diosbulbin-B (DB) Regulates Tumor-Intrinsic PD-L1/NLRP3 Signaling Pathway to Increase Cisplatin-Sensitivity in Gastric Cancer (GC)

Abstract

Background: Generation of cisplatin (DDP)-resistance by gastric cancer (GC) cells seriously limits the therapeutic efficacy of this drug in clinic, and recent data suggested that Diosbulbin-B (DB), the main anti-tumor compound of Dioscorea bulbifera L, was effective to improve cisplatin-sensitivity in GC cells. However, the underlying mechanisms are still largely unknown.Methods: Genes expressions were determined by Real-Time qPCR and Western Blot. Cell viability was evaluated by cell counting kit-8 and trypan blue staining assay. Annexin V-FITC/PI double staining assay was used to examine cell apoptosis. The Spheroid formation assay was used to evaluated cell stemness. Immunohistochemistry (IHC) was employed to examine the expressions and localization of Ki67 protein in mice tumor tissues.Results: Here we found that low-dose DB (12.5 μM) downregulated PD-L1 to activate NLRP3-medicated pyroptosis, and inhibited cancer stem cells (CSCs) properties, to sensitize cisplatin-resistant GC (CR-GC) cells to cisplatin. Mechanistically, the CR-GC cells were obtained, and either low-dose DB or cisplatin alone had little effects on cell viability in CR-GC cells, while low-dose DB significantly induced apoptotic cell death in cisplatin treated CR-GC cells. In addition, low-dose DB triggered cell pyroptosis in CR-GC cells stimulated with cisplatin, which were abrogated by silencing NLRP3. Next, CSCs tended to be enriched in CR-GC cells, instead of their parental cisplatin-sensitive GC (CS-GC) cells, and low-dose DB (12.5 μM) inhibited spheroid formation and stemness biomarkers (CD44, CD133, SOX2, OCT4 and Nanog) expressions to eliminate CSCs in CR-GC cells, which were reversed by upregulating programmed death ligand-1 (PD-L1). Furthermore, we proved that PD-L1 negatively regulated NLRP3 in CR-GC cells, and low-dose DB (12.5 μM) activated NLRP3-mediated pyroptotic cell death in cisplatin treated CR-GC cells by downregulating PD-L1. Also, low-dose DB aggravated the inhibiting effects of cisplatin on tumorigenesis of CR-GC cells in vivo. Conclusions: Collectively, low-dose DB (12.5 μM) regulated intrinsic PD-L1/NLRP3 pathway to improve cisplatin-sensitivity in CR-GC cells, and this study provided alternative therapy treatments for GC.