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Intracellular regulation of TRAIL-induced apoptosis in human melanoma cells
T S Griffith1, W A Chin, G C Jackson
1Department of Immunobiology, Immunex Corp., Seattle, WA 98101, USA. tgriffith@immunex.com
Abstract:
The observation that TNF-related apoptosis-inducing ligand (TRAIL), a member of the TNF cytokine family, induces apoptosis in a number of different tumor cell types led us to compare the tumoricidal effects of TRAIL to those of other TNF family molecules on human melanoma cells. We found that a high proportion of the melanoma cell lines tested were killed by TRAIL, whereas all the melanoma lines were resistant to the other TNF family cytokines tested. TRAIL-induced death was characterized by caspase activation and cellular protein cleavage within minutes of TRAIL addition, and death could be completely inhibited by the caspase inhibitors Ile-Glu-Thr-Asp (IETD) and Val-Ala-Asp (VAD), indicating the presence of a TRAIL receptor signaling pathway similar to that identified for Fas and TNF receptors. Specific TRAIL receptor expression was determined by RT-PCR, and the presence of mRNA encoding the "protective" TRAIL receptors did not correspond to resistance or sensitivity to TRAIL-induced apoptosis. Addition of protein synthesis inhibitors to TRAIL-resistant melanomas rendered them sensitive to TRAIL, indicating that the presence or the absence of intracellular apoptosis inhibitors may mediate resistance or sensitivity to TRAIL-mediated apoptosis. Expression of one such inhibitor, FLICE-inhibitory protein (FLIP), was highest in the TRAIL-resistant melanomas, while being low or undetectable in the TRAIL-sensitive melanomas. Furthermore, addition of actinomycin D to TRAIL-resistant melanomas resulted in decreased intracellular concentrations of FLIP, which correlated with their acquisition of TRAIL sensitivity. Collectively, our results indicate that TRAIL-induced apoptosis occurs through a caspase signaling cascade and that resistance is controlled by intracellular regulators of apoptosis.
Insights
Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) effectively kills human melanoma cells, unlike other TNF family cytokines. Resistance to TRAIL-induced apoptosis in melanoma is linked to intracellular inhibitors like FLIP, not receptor expression.
Area of Science:
- Immunology
- Molecular Biology
- Cancer Research
Background:
- Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a TNF superfamily member known to induce apoptosis in various tumor cells.
- Human melanoma cells exhibit differential sensitivity to apoptosis-inducing agents.
Purpose of the Study:
- To compare the tumoricidal effects of TRAIL with other TNF family molecules on human melanoma.
- To elucidate the mechanisms of TRAIL-induced apoptosis and resistance in melanoma.
Main Methods:
- Human melanoma cell lines were treated with TRAIL and other TNF family cytokines.
- Caspase activation and protein cleavage were assessed.
- TRAIL receptor expression was analyzed using RT-PCR.
- The role of intracellular apoptosis inhibitors, specifically FLIP, was investigated.
Main Results:
- TRAIL demonstrated significant tumoricidal activity against a high proportion of melanoma cell lines, while other TNF cytokines were ineffective.
- TRAIL-induced apoptosis involved caspase activation and was inhibited by caspase inhibitors (IETD, VAD).
- Melanoma resistance to TRAIL was not correlated with specific TRAIL receptor mRNA expression but was associated with high levels of FLICE-inhibitory protein (FLIP).
- Inhibition of protein synthesis or reduction of FLIP levels sensitized resistant melanoma cells to TRAIL.
Conclusions:
- TRAIL induces apoptosis in human melanoma cells via a caspase-dependent signaling pathway.
- Resistance to TRAIL-mediated apoptosis in melanoma is primarily regulated by intracellular apoptosis inhibitors, notably FLIP.
- Targeting intracellular regulators like FLIP may represent a therapeutic strategy to enhance TRAIL sensitivity in resistant melanoma.