Inhibition of TACE Activity Enhances the Susceptibility of Myeloma Cells to TRAIL.

Abstract

TNF-related apoptosis-inducing ligand/Apo2 ligand (TRAIL/Apo2L) selectively induces apoptosis in various cancer cells including myeloma (MM) cells. Although TRAIL-mediated immunotherapy is as attractive maneuver against MM because of its tumor-specific cytotoxic activity, the susceptibility of MM cells to TRAIL has been demonstrated to vary among MM cells, which limits clinical applications of TRAIL-mediated immunotherapy. TRAIL is known to bind to 2 different proapoptotic receptors, death receptor 4 (DR4) and DR5. The membrane-bound TRAIL receptors have been reported to be subjected to proteolytic cleavage in melanoma cells in a manner inhibitable by tissue inhibitor of metalloproteinases-3 (TIMP-3), an endogenous inhibitor for TNF-alpha converting enzyme (TACE) (Ahonen M, Oncogene. 2003). Although TACE is regarded as a sheddase for TNF-like ligands/receptors, the role for TACE in TRAIL-mediated cytotoxicity in MM cells is largely unknown. In the present study, we therefore investigated the role for TACE in TRAIL and its receptor editing as well as the effect of TACE inhibition on TRAIL-triggered cytotoxicity in MM cells. MM cells constitutively expressed DR4 on their surface as determined in flow cytometry, while the expression of DR5 as well as TRAIL was negligible. TACE mRNA was constitutively overexpressed in all MM cell lines including RPMI8226, MM.1S, KMS12 and U266, whereas its endogenous inhibitor, TIMP-3, was not detected, suggesting enhanced TACE activity along with down-regulation of TIMP-3 in MM cells. TAPI-0, a TACE inhibitor, up-regulated the cell-surface expression of DR4 about 2-fold in RPMI8226 and U266 cells, while DR5 and TRAIL expression remained undetectable. Immunoblot analyses for DR4 revealed that conditioned media from RPMI8226 and U266 cells contained DR4 protein with a molecular size smaller than that observed in their cell lysates. Interestingly, TAPI-0 treatment down-regulated DR4 levels in culture supernatants of these MM cells, while increasing DR4 levels in their cell lysates as well as on their surface, suggesting ectodomain shedding of DR4 by TACE in MM cells. Importantly, TAPI-0 treatment potentiated TRAIL-triggered cytotoxicity against MM cells including RPMI8226 and MM.1S, which was almost completely antagonized by addition of osteoprotegerin, a soluble inhibitor for TRAIL, indicating a TRAIL-dependent mechanism. Taken together, the present observations demonstrate that cell-surface TRAIL receptor expression is post-transcriptionally down-regulated by endogenous TACE activity enhanced in MM cells, which may protect MM cells from TRAIL-mediated apoptosis by surrounding immune cells expressing TRAIL, and that up-regulation of TRAIL receptor expression by inhibiting TACE activity is able to restore the susceptibility of MM cells to TRAIL. Therefore, these preclinical data provide that TACE inhibitors in combination with TRAIL-based immunotherapy may be a novel therapeutic approach against MM which can not be cured by the present chemotherapeutic modalities.