Obatoclax enhances TRAIL-induced apoptosis in non-small-cell-lung-cancer cells by increasing TRAIL receptor expression, priming for mitochondrial apoptosis and reducing XIAP levels

Abstract

Aim: Antitumor activity of TNF-related apoptosis-inducing ligand (TRAIL) can be inhibited by anti-apoptotic Bcl-2 family members. Here, we examined the potential of the BH-3 (bcl-2 homolog domain 3) mimetic obatoclax to sensitize non-small cell lung cancer (NSCLC) cells for TRAIL-induced apoptosis.

Methods: The sensitivity to obatoclax treatment in combination with TRAIL was measured with the tetrazolium-based 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay. TRAIL-induced apoptosis was evaluated with flow cytometric analysis. Mitochondrial and cytosolic components were fractionated and western blotting was performed to determine the level of Cytochrome c, GAPDH, and Cox IV. Knockdowns were performed with small interfering RNA (siRNA). TRAIL receptor protein levels were assessed with fluorescence intensity index.

Results: Synergistic interactions were observed both in TRAIL-resistant (SW1573, A549) and TRAIL-sensitive (H460) cells, characterized by enhanced caspase-dependent apoptosis. Although concurrent treatment with obatoclax and TRAIL was sufficient for apoptosis activation in SW1573 cells, A549 cells required 48-h pre-incubation with obatoclax, during which TRAIL-R2 (TRAIL receptor 2) cell surface expression increased and X-linked inhibitor of apoptosis protein (XIAP) levels decreased. Interestingly, the knockdown of XIAP was sufficient to sensitize A549 cells for concurrent treatment.

Conclusion: Obatoclax is an effective sensitizer for TRAIL-induced apoptosis in NSCLC, although displaying cell type-specific effects.

Keywords

TRAIL / obatoclax / apoptosis / NSCLC / XIAP

References

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