Bone Marrow Stroma Confers Resistance to Apo2 Ligand/TRAIL in Multiple Myeloma in Part by Regulating c-FLIP

Abstract

Apo2 ligand (Apo2L)/TRAIL induces apoptosis of cancer cells that express the specific receptors while sparing normal cells. Because the tumor microenvironment protects myeloma from chemotherapy, we investigated whether hemopoietic stroma induces resistance to Apo2L/TRAIL apoptosis in this disease. Apo2L/TRAIL-induced death was diminished in myeloma cell lines (RPMI 8226, U266, and MM1s) directly adhered to a human immortalized HS5 stroma cell line but not adhered to fibronectin. In a Transwell assay, with myeloma in the upper well and HS5 cells in the lower well, Apo2L/TRAIL apoptosis was reduced when compared with cells exposed to medium in the lower well. Using HS5 and myeloma patients’ stroma-conditioned medium, we determined that soluble factor(s) produced by stroma–myeloma interactions are responsible for a reversible Apo2/TRAIL apoptosis resistance. Soluble factor(s) attenuated procaspase-8, procaspase-3, and poly(ADP-ribose) polymerase cleavage and diminished mitochondrial membrane potential changes without affecting Bcl-2 family proteins and/or Apo2L/TRAIL receptors. Soluble factor(s) increased the baseline levels of the anti-apoptotic protein c-FLIP in all cell lines tested. Inhibition of c-FLIP by means of RNA interference increased Apo2/TRAIL sensitivity in RPMI 8226 cells. Unlike direct adhesion to fibronectin, soluble factor(s) have no impact on c-FLIP redistribution within cellular compartments. Cyclohexamide restored Apo2L/TRAIL sensitivity in association with down-regulation of c-FLIP, suggesting that c-FLIP synthesis, not intracellular traffic, is essential for soluble factor(s) to regulate c-FLIP. Additionally, IL-6 conferred resistance to Apo2L/TRAIL-mediated apoptosis in association with increased c-FLIP levels. In conclusion, the immune cytotoxic effect of Apo2L/TRAIL can be restored at least in part by c-FLIP pathway inhibitors.

Aurora and IKK kinases cooperatively interact to protect multiple myeloma cells from Apo2L/TRAIL

“…Furthermore, exogenous IL-6 conferred resistance to TRAIL-mediated apoptosis in RPMI 8226 and OPM-2, but pretreatment with pan-AKIs was able to restore TRAIL sensitivity in both HMCLs ( Figure 1B). Furthermore, consistent with previous studies, we found that adherence of MM cells to BMSCs significantly increased their resistance to TRAIL-induced cell death compared with exogenous IL-6 16,17 ( Figure 1B). However, pan-AKIs were able to completely or almost completely reverse the BMSC-mediated TRAIL resistance in RPMI 8226/R5 and OPM-2 ( Figure 1B), whereas in RPMI 8226 and U266 HMCLs that was previously reported to strongly increase the caspase-8 inhibitor FLIP when cocultured with BMSCs, 16 panAKIs partially (but significantly) overcame stroma-mediated TRAIL resistance when TRAIL was used at fourfold higher concentrations (9.6 or 19.2 ng/mL, respectively) than those used in the absence of stroma ( Figure 1B).…”

γ-Tocotrienol Promotes TRAIL-Induced Apoptosis through Reactive Oxygen Species/Extracellular Signal-Regulated Kinase/p53–Mediated Upregulation of Death Receptors

“…Among the antiapoptotic genes, cellular FLICE-like inhibitory protein (c-FLIP; ref. 5), survivin, Bcl-2, bcl-xL, inhibitor of apoptosis (IAP)-1, IAP-2, and X-linked IAP (XIAP; ref. 6) play major roles in regulating TRAIL-induced apoptosis.…”

Inflammatory Cytokines Determine the Susceptibility of Human CD8 T Cells to Fas-mediated Activation-induced Cell Death through Modulation of FasL and c-FLIPS Expression

“…Although very little is known about the mechanisms by which IL-6 induces c-FLIP expression, Perez et al (36) have shown that IL-6 can induce c-FLIP L and c-FLIP S expression in multiple myeloma. Further, STAT3 induces c-FLIP expression in cardiomyocytes (37).…”