Multiple cytotoxic effects of gamabufotalin against human glioblastoma cell line U-87

Chem Biol Interact. 2019 Dec 1;314:108849. doi: 10.1016/j.cbi.2019.108849. Epub 2019 Oct 11.

Abstract

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.

Keywords: Autophagy; G(2)/M arrest; Gamabufotalin; Glioblastoma; Metastasis; p38 MAPK.

MeSH terms

  • Animals
  • Apoptosis / drug effects*
  • Astrocytes / cytology
  • Astrocytes / drug effects
  • Astrocytes / metabolism
  • Autophagy / drug effects
  • Bufanolides / chemistry
  • Bufanolides / pharmacology*
  • Cell Line, Tumor
  • Cell Survival / drug effects
  • G2 Phase Cell Cycle Checkpoints / drug effects
  • Glioblastoma / metabolism
  • Glioblastoma / pathology
  • Humans
  • Mice
  • Protein Kinase Inhibitors / pharmacology
  • Proto-Oncogene Proteins c-akt / metabolism
  • Signal Transduction / drug effects
  • Up-Regulation / drug effects
  • Vascular Endothelial Growth Factor A / antagonists & inhibitors
  • Vascular Endothelial Growth Factor A / metabolism
  • Vascular Endothelial Growth Factor Receptor-2 / antagonists & inhibitors
  • Vascular Endothelial Growth Factor Receptor-2 / metabolism
  • Wortmannin / pharmacology
  • p38 Mitogen-Activated Protein Kinases / antagonists & inhibitors
  • p38 Mitogen-Activated Protein Kinases / metabolism

Substances

  • Bufanolides
  • Protein Kinase Inhibitors
  • Vascular Endothelial Growth Factor A
  • gamabufotalin
  • Vascular Endothelial Growth Factor Receptor-2
  • Proto-Oncogene Proteins c-akt
  • p38 Mitogen-Activated Protein Kinases
  • Wortmannin