Selective targeting of death receptor 5 circumvents resistance of MG-63 osteosarcoma cells to TRAIL-induced apoptosis

Tumor necrosis factor–related apoptosis-inducing ligand (TRAIL), a tumor necrosis factor superfamily member, targets death receptors and selectively kills malignant cells while leaving normal cells unaffected. However, unlike most cancers, many osteosarcomas are resistant to TRAIL. To investigate this resistance, we characterized the response of MG-63 osteosarcoma cells and hPOB-tert osteoblast-like cells to TRAIL and agonist antibodies to death receptor 4 (DR4) and death receptor 5 (DR5). We found that MG-63 osteosarcoma cells and hPOB-tert osteoblast-like cells show no or very little response to TRAIL or a DR4 agonist, but MG-63 cells undergo apoptosis in response to a DR5 agonist. Analysis of TRAIL receptor expression showed that normal osteoblastic and osteosarcoma cells express a variety of TRAIL receptors but this does not correlate to TRAIL responsiveness. Production of the soluble decoy receptor osteoprotegerin also could not explain TRAIL resistance. We show that TRAIL activates the canonical caspase-dependent pathway, whereas treatment with cycloheximide increases the sensitivity of MG-63 cells to TRAIL and anti-DR5 and can also sensitize hPOB-tert cells to both agents. Proapoptotic and antiapoptotic protein expression does not significantly differ between MG-63 and hPOB-tert cells or change following treatment with TRAIL or anti-DR5. However, sequencing the death domain of DR4 in several osteoblast-like cells showed that MG-63 osteosarcoma cells are heterozygous for a dominant-negative mutation, which can confer TRAIL resistance. These results suggest that although the dominant-negative form of the receptor may block TRAIL-induced death, an agonist antibody to the active death receptor can override cellular defenses and thus provide a tailored approach to treat resistant osteosarcomas. [Mol Cancer Ther 2007;6(12):3219–28]