Death receptor 5 is activated by fucosylation in colon cancer cells

Abstract

The remarkable pro‐apoptotic properties of tumour necrosis factor (TNF)‐related apoptosis‐inducing ligand (TRAIL) have led to considerable interest in this protein as a potential anticancer therapeutic. However, TRAIL is largely ineffective in inducing apoptosis in certain cancer cells, and the mechanisms underlying this selectivity are unknown. In colon adenocarcinomas, posttranslational modifications including O‐ and N‐ glycosylation of death receptors were found to correlate with TRAIL‐induced apoptosis. Additionally, mRNA levels of fucosyltransferase 3 (FUT3) and 6 (FUT6) were found to be high in the TRAIL‐sensitive colon adenocarcinoma cell line COLO 205. In this study, we use agonistic receptor‐specific TRAIL variants to dissect the contribution of FUT3 and FUT6‐mediated fucosylation to TRAIL‐induced apoptosis via its two death receptors, DR4 and DR5. Triggering of apoptosis by TRAIL revealed that the low FUT3/6‐expressing cells DLD‐1 and HCT 116 are insensitive to DR5 but not to DR4‐mediated apoptosis. By contrast, efficient apoptosis is mediated via both receptors in high FUT3/6‐expressing COLO 205 cells. The reconstitution of FUT3/6 expression in DR5‐resistant cells completely restored TRAIL sensitivity via this receptor, while only marginally enhancing apoptosis via DR4 at lower TRAIL concentrations. Interestingly, we observed that induction of the salvage pathway by external administration of l‐fucose restores DR5‐mediated apoptosis in both DLD‐1 and HCT 116 cells. Finally, we show that fucosylation influences the ligand‐independent receptor association that leads to increased death inducing signalling complex (DISC) formation and caspase‐8 activation. Taken together, these results provide evidence for the differential impact of fucosylation on signalling via DR4 or DR5. These findings provide novel opportunities to enhance TRAIL sensitivity in colon adenocarcinoma cells that are highly resistant to DR5‐mediated apoptosis.

Artemisinin Derivatives Stimulate DR5-Specific TRAIL-Induced Apoptosis by Regulating Wildtype P53

“…Then, we analyzed the effect of ATS and DHA with the combination of DR5-specific TRAIL variant DHER and DR4-specific TRAIL variant 4C7 in the mentioned colon cancer cell lines (except for COLO 205, which is already highly sensitive to TRAIL). Both HCT116 and DLD-1 are sensitive to the cell death induced by 4C7 but resistant to DHER ( Figure 3 A,B, black bar), which corresponds to our previous report [ 28 ]. The cell viability observed in the single treatment with ATS (grey bar) or DHA (white bar) is ~70% for HCT116 and ~90% for DLD-1, respectively.…”

“…For clinical therapy, TRAIL and the death receptor agonistic antibodies are under phase II clinical trials in colorectal cancer treatment due to TRAIL’s harmfulness to cancer cells compared with healthy cells [ 28 , 30 ]. However, the recombinant human TRAIL dulanermin already showed reduced efficacy in phase I clinical trial as a result of low bioavailability and decoy receptor binding [ 31 , 32 ].…”

“…Cells were harvested and lysed in RIPA buffer with Complete Protease Inhibitor Cocktail, EDTA-Free (Roche, Basel, Switzerland) inside. Protein concentrations were determined using the Pierce BCA Protein Assay Kit (Thermo Fisher Scientific, Rockford, IL, USA) and Western blot analyses were performed as previously described [28]. PARP-1, caspase-3 (Cell Signaling Technology, Leiden, The Netherlands), DR4 (Novus Biologicals, Abingdon, UK), DR5, P21, P53 (abcam, Cambridge, UK) primary antibodies were used.…”

Natural Product Mediated Regulation of Death Receptors and Intracellular Machinery: Fresh from the Pipeline about TRAIL-Mediated Signaling and Natural TRAIL Sensitizers

“…Fucosylation is a post-translational modification of critical importance that plays a crucial role in improving TRAIL-mediated apoptosis [19]. Stably and transiently overexpressed FUT3 and FUT6 dramatically enhanced TRAIL-mediated apoptosis in HCT116 and DLD-1 cells.…”

“…Activation and cleavage mechanisms of caspase-8 and PARP-1 were noted to be more pronounced in cells overexpressing FUT6 and FUT3. More importantly, a significantly higher fraction of signalosomes was noticed, as evidenced by highly increased DISC-associated caspase-8 complexes in FUT3-overexpressing cells [19]. Harakiri (HRK), a BH3-only protein of the Bcl-2 family, has been shown to promote TRAIL-mediated apoptosis in glioblastoma cells [20].…”