The IL-6/sIL-6R treatment of a malignant melanoma cell line enhances susceptibility to TNF-α-induced apoptosis

Melanoma is an intractable tumor that has shown very impressive and promising response to local administration of high dose recombinant TNF-α in combination with IFN-γ in clinical studies. In this study, we investigated the effect of IL-6/sIL-6R on TNF-α-resistant B16/F10.9 melanoma cells. A low dose of TNF-α or IL-6/sIL-6R had minimal affect on the cell growth. However, the highly active fusion protein of sIL-6R and IL-6 (IL6RIL6), covalently linked by a flexible peptide, sensitized TNF-α-resistant F10.9 melanoma cells to TNF-α-induced apoptosis. Stimulation of the cells with IL6RIL6 plus TNF-α resulted in both the activation of caspase-3 and the reduction of bcl-2 expression. Flow cytometry analysis showed that IL6RIL6-upregulated TNF-R55 and TNF-R75 expression, suggesting an increase in TNF-α responsiveness by IL6RIL6 resulting from the induction of TNF receptors. Moreover, exposure of F10.9 cells to neutralizing antibody to TNF-R55 significantly inhibited IL6RIL6/TNF-α-induced cytotoxicity. These results suggest that the IL6/sIL6R/gp130 system, which sensitizes TNF-α-resistant melanoma cells to TNF-α-induced apoptosis, may provide a new target for immunotherapy.