Suppression of cFLIP is sufficient to sensitize human melanoma cells to TRAIL- and CD95L-mediated apoptosis.

Death ligands such as tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) and certain forms of CD95L are attractive therapeutic options for metastatic melanoma. Since knowledge about the regulation of death receptor sensitivity in melanoma is sparse, we have analysed these signaling pathways in detail. The loss of CD95 or TRAIL-R1, but not of TRAIL-R2, surface expression correlated with apoptosis sensitivity in a panel of melanoma cell lines. In contrast, the expression of proteins of the apical apoptosis signaling cascade (FADD, initiator caspases-8 and cFLIP) did not predict apoptosis sensitivity. Since both TRAIL-R1 and -R2 transmit apoptotic signals, we asked whether cFLIP, highly expressed in several of the cell lines tested, is sufficient to maintain resistance to TRAIL-R2-mediated apoptosis. Downregulation of cFLIP in TRAIL-R2-positive, TRAIL-resistant IGR cells dramatically increased TRAIL sensitivity. Conversely ectopic expression of cFLIP in TRAIL-sensitive, TRAIL-R2-expressing RPM-EP melanoma cells inhibited TRAIL- and CD95L-mediated cell death. Thus, modulation of cFLIP is sufficient to sensitize TRAIL-R2-expressing cells for TRAIL. Taken together, albeit expressing all proteins necessary for death receptor-mediated apoptosis, TRAIL-R1 negative melanoma cells cannot undergo TRAIL- or CD95L-induced apoptosis due to expression of cFLIP. Hence, cFLIP represents an attractive therapeutic target for melanoma treatment, especially in combination with TRAIL receptor agonists.

Basal cell adhesion molecule is inversely associated with apoptosis, but plays a limited role for protection against apoptotic stimuli.

Basal cell adhesion molecule (B-CAM) is strongly upregulated in epithelial skin cancer cells in vivo and in vitro. We have tested here whether B-CAM is (1) inversely associated with or (2) functionally involved in apoptosis. Towards this end, B-CAM expression was assessed in HaCaT transfectants overexpressing murine Bcl-2 and untransfected HaCaT cells exposed to various proapoptotic stimuli. In another series of experiments, we overexpressed B-CAM in HaCaT cells and different fibroblast lines, and stimulated various apoptotic pathways in the transfectants and control cells. In addition, apoptosis was assessed after an antibody-mediated B-CAM blockade. We could demonstrate that expression of B-CAM is inversely associated with the susceptibility of cells to apoptosis. However, overexpression or antibody- mediated inhibition of B-CAM had only limited functional effects on cellular apoptosis.