Work

64: Interleukin-6 and hematopoietic soluble factors confers resistance to trail mediated apoptosis

Year: 2007

Type: article

Abstract: The bone marrow microenvironment provides tumor protection from chemotherapy and Fas-death receptor mediated apoptosis through a process known as environmental mediated death resistance (EM-DR). TRAIL, a member of the TNF superfamily, has anti-tumor activity sparing normal cells and mediates allogeneic graft-versus-tumor responses. We hypothesized that the tumor microenvironment influences TRAIL mediated killing conferring an immune resistance mechanism contributing to cancer progression in allogeneic transplantation. Experiments in a transwell (TW) assay with RPMI-8226-Myeloma cells in the upper well and HS5- hematopoietic stromal cells in the bottom well (TW+HS5) revealed that HS5 cells blocked recombinant human(rh-) TRAIL-induced apoptosis through soluble factors in a dose and time-dependent manner. RPMI-8226 treated in TW+HS5 exhibited attenuated pro-caspase-8, pro-caspase-3, PARP, and BID cleavage, with diminished mitochondrial membrane potential changes, without alterations on TRAIL receptors. Western blotting of RPMI-8226 cells showed that Fas-associated death domain like IL-1 converting enzyme-like inhibitory protein (FLIP), a regulatory factor that competes with caspase-8 inhibiting apoptosis, is increased in TW+HS5 treated cells. Subcellular fractionation of RPMI-8226 cells showed that FLIP is maintained in or associated with organelle membranes and is released to the cytosol when exposed to soluble factor (TW+HS5) suggesting that that soluble factor signaling may influence FLIP localization and availability. FLIP reduction by FLIP-RNA interference or with Bortezomib, NF-KB inhibitor that reduces FLIP levels, increases TRAIL apoptosis sensitivity of RPMI-8226 cells treated in TW+HS5. To explore whether IL-6, important myeloma survival factor, is involved in TRAIL-EMDR, RPMI-8226 treated with HS27a stroma that lack IL-6 secretion elicit a diminished apoptosis resistance compared to HS-5. In addition, RPMI-8226 treated in TW+HS5 with an anti-human IL-6 neutralizing antibody partially overcome the TRAIL apoptosis resistance. Furthermore, rh-IL-6 confers resistance to TRAIL mediated apoptosis in a dose-dependent manner associated with FLIP increase. Our results suggest that IL-6 and other soluble factors produced by marrow stromal cells promote myeloma cell survival by upregulating FLIP, thereby mitigating the influence of the microenvironment on TRAIL-induced apoptosis. The immune cytotoxic effect of TRAIL may be enhanced by FLIP inhibitors.

Source: Transplantation and Cellular Therapy

DOI: 10.1016/j.bbmt.2006.12.067