Molecular Determinants of Response to TRAIL in Killing of Normal and Cancer Cells

The tumor necrosis factor-related apoptosis-inducing ligand (TRAIL or Apo2L) is a potent inducer of death of cancer but not normal cells, which suggests its potential use as a tumor-specific antineoplastic agent. TRAIL binds to the proapoptotic death receptors DR4 and the p53-regulated proapoptotic KILLER/DR5 as well as to the decoy receptors TRID and TRUNDD. In the present studies, we identified a subgroup of TRAIL-resistant cancer cell lines characterized by low or absent basal DR4 or high expression of the caspase activation inhibitor FLIP. Four of five TRAIL-sensitive cell lines expressed high levels of DR4 mRNA and protein, whereas six of six TRAIL-resistant cell lines expressed low or undetectable levels of DR4 (χ 2 ; P < 0.01). FLIP expression appeared elevated in five of six (83%) TRAIL-resistant cell lines and only one of five (20%) TRAIL-sensitive cells (χ 2 ; P < 0.05). Two TRAIL-resistant lines that expressed DR4 contained an A-to-G alteration in the death domain encoding arginine instead of lysine at codon 441. The K441R polymorphism is present in 20% of the normal population and can inhibit DR4-mediated cell killing in a dominant-negative fashion. The expression level of KILLER/DR5, TRID, TRUNDD or TRID, and TRUNDD did not correlate with TRAIL sensitivity ( P > 0.05). These results suggest that the major determinants for TRAIL sensitivity may be the expression level of DR4 and FLIP. TRAIL-resistant cells became susceptible to TRAIL-mediated apoptosis in the presence of doxorubicin. In TRAIL-sensitive cells, caspases 8, 9, and 3 were activated after TRAIL treatment, but in TRAIL-resistant cells, they were activated only by the combination of TRAIL and doxorubicin. Our results suggest: ( a ) evaluation of tumor DR4 and FLIP expression and host DR4 codon 441 status could be potentially useful predictors of TRAIL sensitivity, and ( b ) doxorubicin, in combination with TRAIL, may effectively promote caspase activation in TRAIL-resistant tumors.