Zerumbone suppresses IKKα, Akt, and FOXO1 activation, resulting in apoptosis of GBM 8401 cells

Background: Zerumbone, a sesquiterpene compound isolated from subtropical ginger, Zingiber zerumbet Smith, has been documented to exert antitumoral and anti- inflammatory activities. In this study, we demonstrate that zerumbone induces apoptosis in human glioblastoma multiforme (GBM8401) cells and investigate the apoptotic mechanism. Methods: We added a caspase inhibitor and transfected wild-type (WT) IKK and Akt into GBM 8401 cells, and measured cell viability and apoptosis by MTT assay and flow cytometry. By western blotting, we evaluated activation of caspase-3, dephosphorylation of IKK, Akt, FOXO1 with time, and change of IKK, Akt, and FOXO1 phosphorylation after transfection of WT IKK and Akt. Results: Zerumbone (10 similar to 50 mu M) induced death of GBM8401 cells in a dose-dependent manner. Flow cytometry studies showed that zerumbone increased the percentage of apoptotic GBM cells. Zerumbone also caused caspase-3 activation and poly (ADP-ribose) polymerase (PARP) production. N-benzyloxycarbonyl -Val-Ala-Asp- fluoromethylketone (zVAD-fmk), a broad-spectrum caspase inhibitor, hindered zerumbone-induced cell death. Transfection of GBM 8401 cells with WT IKK alpha inhibited zerumbone-induced apoptosis, and zerumbone significantly decreased IKK alpha phosphorylation levels in a time-dependent manner. Similarly, transfection of GBM8401 cells with Akt suppressed zerumbone-induced apoptosis, and zerumbone also diminished Akt phosphorylation levels remarkably and time-dependently. Moreover, transfection of GBM8401 cells with WT IKK alpha reduced the zerumbone-induced decrease in Akt and FOXO1 phosphorylation. However, transfection with WT Akt decreased FOXO1, but not IKK alpha, phosphorylation. Conclusion: The results suggest that inactivation of IKK alpha, followed by Akt and FOXO1 phosphorylation and caspase-3 activation, contributes to zerumbone-induced GBM cell apoptosis.