To provide novel insight into approaches designed to combat
glioblastoma, the molecular details of the cytotoxicity of
gamabufotalin, were investigated in the human
glioblastoma cell line U-87. A dose-dependent cytotoxicity was observed in the cells, whereas no detectable toxicity was confirmed in mouse primary astrocytes. LDH leakage was only observed in the cells treated with a relatively high concentration (>80 ng/ml). Downregulation of the expression levels of Aurora B, cdc25A, cdc25C, cdc2,
Cyclin B1 and
survivin, and upregulation of the expression level of p21 were observed in treated cells and occurred in parallel with G2/M phase arrest. Treatment with
gamabufotalin also downregulated the expression level of uPA, CA9, and upregulated the expression level of TIMP3, all of which are closely associated with invasion/
metastasis. Autophagy induction was observed in the treated cells and the addition of
wortmannin, a potent autophagy inhibitor, significantly rescued U-87 cells. These results indicate that
gamabufotalin exhibits cytotoxicity against cancerous glial cells with high potency and selectivity through multiple cytotoxic signaling pathways. The activation of
p38 MAPK pathway along with the upregulation of
VEGF/VEGFR2 was observed in the treated cells, both of which are likely to be compensatory changes in response to
gamabufotalin treatment. Intriguingly, a specific inhibitor of
p38 MAPK enhanced the cytotoxicity of the drug, suggesting an important prosurvival role for
p38 MAPK. We thus suggest that developing a new combination regimen of
gamabufotalin plus a
p38 MAPK inhibitor and/or inhibitors for
VEGF/VEGFR could improve the efficacy of the drug, and may provide more therapeutic benefits to patients with
glioblastoma.