Identification of inhibitors of TRAIL-induced death (ITIDs) in the TRAIL-sensitive colon carcinoma cell line SW480 using a genetic approach

The tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a po tent inducer of apoptosis in tumor cell lines, whereas normal cells appear to be protected from its cytotoxic effects. Therefore TRAIL holds promise a s a potential therapeutic agent against cancer. To elucidate some of the cr itical factors that contribute to TRAIL resistance, we performed a genetic screen in the human colon carcinoma cell line SW480 by infecting this TRAIL -sensitive cell line with a human placental cDNA retroviral library and iso lating TRAIL-resistant clones. Characterization of the resulting clones for inhibitors of TRAIL-induced death (ITIDs) led to the isolation of c-FLIPS, Bax inhibitor 1, and Bcl-X-L as candidate suppressors of TRAIL signaling. We have demonstrated that c-FLIPS and Bcl-X-L are sufficient when overexpre ssed to convey resistance to TRAIL treatment in previously sensitive cell l ines. Furthermore both c-FLIPS and Bcl-X-L protected against overexpression of the TRAIL receptors DR4 and KILLER/DR5. When c-FLIPS and Bcl-X-L were o verexpressed together in SW480 and HCT 116, an additive inhibitory effect w as observed after TRAIL treatment suggesting that these two molecules funct ion in the same pathway in the cell lines tested. Furthermore, we have demo nstrated for the first time that a proapoptotic member of the Bcl-2 family, Bax, is required for TRAIL-mediated apoptosis in HCT 116 cells. Surprising ly, we have found that the serine/threonine protein kinase Akt, which is an upstream regulator of both c-FLIPS and Bcl-X-L, is not sufficient when ove rexpressed to protect against TRAIL in the cell lines tested. These results suggest a key role for c-FLIPS, Bcl-X-L, and Bax in determining tumor cell sensitivity to TRAIL.