Molecular determinants of response to TRAIL in killing of normal and cancer cells

The tumor necrosis factor-related apoptosis-inducing ligand (TRAIL, or Apo2 L) is a potent inducer of death of cancer but not normal cells, which sugge sts 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 prese nt studies, we identified a subgroup of TRAIL-resistant cancer cell lines c haracterized by low or absent basal DR4 or high expression of the caspase a ctivation inhibitor FLIP. Four of five TRAIL-sensitive cell lines expressed high levels of DR4 mRNA and protein, whereas sis of six TRAIL-resistant ce ll lines expressed low or undetectable levels of DR4 (chi(2) P < 0.01). FLI P expression appeared elevated in five of six (83%) TRAIL-resistant cell li nes and only one of five (20%) TRAIL-sensitive cells (chi(2); P < 0.05), Tw o TRAIL-resistant lines that expressed DR4 contained an A-to-G alteration i n the death domain encoding arginine instead of lysine at codon 441. The K4 41R polymorphism is present in 20%, of the normal population and can inhibi t DR4-mediated cell killing in a dominant-negative fashion. The expression level of KILLER/DR5, TRID, TRUNDD or TRID, and TRUNDD did not correlate wit h TRAIL sensitivity (P > 0.05), These results suggest that the major determ inants for TRAIL sensitivity may be the expression level of DR4 and FLIP. T RAIL-resistant cells became susceptible to TRAIL-mediated apoptosis in the presence of doxorubicin. In TRAIL-sensitive cells, caspases 8, 9, and 3 wer e activated after TRAIL treatment, but in TRAIL-resistant cells, they were activated only by the combination of TRAIL and doxorubicin. Our results sug gest: (a) evaluation of tumor DR4 and FLIP expression and host DR4 codon 44 1 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.