Cyclovirobuxine D suppresses cancer stemness in osteosarcoma with implication of the noncanonical NF-kappaB pathway

Background Cyclovirobuxine D (CVB-D), an alkaloid from Buxus microphylla, exhibits anticancer effects in various tumors, but its role in osteosarcoma remains unexplored. This study investigates its efficacy and mechanism in osteosarcoma. Methods We screened a drug library and evaluated CVB-D’s effects on osteosarcoma cell lines using CCK-8 and colony formation assays. Saos2 and K7M2 cells were selected for further analysis. Flow cytometry assessed apoptosis and cell cycle. RNA-seq identified downstream pathways, including NF-kappaB, and stemness markers (CD24, ALDH1A1). Stemness was examined via serum-free suspension culture, and NF-kappaB pathway activator Diprovocim was used in rescue experiments. A xenograft mouse model validated the findings. Results CVB-D suppressed proliferation, stemness, and induced apoptosis in osteosarcoma. These effects may be partially mediated through the p-NF-kappaB2/NF-kappaB2 axis and were reversible upon NF-kappaB pathway activation. Conclusion CVB-D inhibits osteosarcoma possibly via the non-canonical NF-kappaB pathway, suggesting a potential therapeutic strategy.