TRAIL signaling is mediated by DR4 in pancreatic tumor cells despite the expression of functional DR5
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Tumor necrosis factor related apoptosis-inducing ligand (TRAIL) and agonistic anti-DR4/TRAIL-R1 and anti-DR5/TRAIL-R2 antibodies are currently under clinical investigation for treatment of different malignancies. TRAIL activates DR4 and DR5 and thereby triggers apoptotic and non-apoptotic signaling pathways, but possible different roles of DR4 or DR5 in these responses has poorly been addressed so far. In the present work, we analyzed cell viability, DISC formation as well as IL-8 and NF–κB activation side by side in responses to TRAIL and agonistic antibodies against DR4 (mapatumumab) and against DR5 (lexatumumab) in pancreatic ductal adenocarcinoma cells. We found that all three reagents are able to activate cell death and pro-inflammatory signaling. Death-inducing signaling complex (DISC) analysis revealed that mapatumumab and lexatumumab induce formation of homocomplexes of either DR4 or DR5, whereas TRAIL additionally stimulated the formation of heterocomplexes of both receptors. Notably, blocking of receptors using DR4- and DR5-specific Fab fragments indicated that TRAIL exerted its function predominantly via DR4. Interestingly, inhibition of PKC by Goe6983 enabled DR5 to trigger apoptotic signaling in response to TRAIL and also strongly enhanced lexatumumab-mediated cell death. Our results suggest the existence of mechanisms that silence DR5 for TRAIL- but not for agonistic-antibody treatment.
KeywordsTRAIL DR4/DR5 Mapatumumab Lexatumumab Pancreatic adenocarcinoma
We thank Daniela Berg and Andreas Wicovsky for generation and characterization of TRAIL-receptor-specific antibodies and Fab fragments, Daniela Wesch for help with FACS analyses, and Robin Humphreys at Human Genome Sciences for supporting us with mapatumumab and lexatumumab antibodies. Some of the data are part of the doctoral thesis of Johannes Lemke. This work was supported by the DFG grant SFB 415/A3 given to Holger Kalthoff and DFG grants SCHU733/7-1 and SCHU 733/9-1 given to Stefan Schütze. In addition, the authors have no conflicts of interest to declare. All authors contributed to the preparation of the manuscript.
- 8.Koschny R, Holland H, Sykora J, Haas TL, Sprick MR, Ganten TM, Krupp W, Bauer M, Ahnert P, Meixensberger J et al (2007) Bortezomib sensitizes primary human astrocytoma cells of WHO grades I to IV for tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis. Clin Cancer Res 13:3403–3412PubMedCrossRefGoogle Scholar
- 15.Trauzold A, Wermann H, Arlt A, Schutze S, Schafer H, Oestern S, Roder C, Ungefroren H, Lampe E, Heinrich M et al (2001) CD95 and TRAIL receptor- mediated activation of protein kinase C and NF–kappaB contributes to apoptosis resistance in ductal pancreatic adenocarcinoma cells. Oncogene 20:4258–4269PubMedCrossRefGoogle Scholar
- 24.Schneider-Brachert W, Tchikov V, Neumeyer J, Jakob M, Winoto-Morbach S, Held-Feindt J, Heinrich M, Merkel O, Ehrenschwender M, Adam D et al (2004) Compartmentalization of TNF receptor 1 signaling: internalized TNF receptosomes as death signaling vesicles. Immunity 21:415–428PubMedCrossRefGoogle Scholar
- 28.Kelley RF, Totpal K, Lindstrom SH, Mathieu M, Billeci K, Deforge L, Pai R, Hymowitz SG, Ashkenazi A (2005) Receptor-selective mutants of apoptosis-inducing ligand 2/tumor necrosis factor-related apoptosis-inducing ligand reveal a greater contribution of death receptor (DR) 5 than DR4 to apoptosis signaling. J Biol Chem 280:2205–2212PubMedCrossRefGoogle Scholar
- 30.Wajant H, Moosmayer D, Wuest T, Bartke T, Gerlach E, Schonherr U, Peters N, Scheurich P, Pfizenmaier K (2001) Differential activation of TRAIL-R1 and -2 by soluble and membrane TRAIL allows selective surface antigen-directed activation of TRAIL-R2 by a soluble TRAIL derivative. Oncogene 20:4101–4106PubMedCrossRefGoogle Scholar
- 33.Shi RX, Ong CN, Shen HM (2005) Protein kinase C inhibition and x-linked inhibitor of apoptosis protein degradation contribute to the sensitization effect of luteolin on tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis in cancer cells. Cancer Res 65:7815–7823PubMedCrossRefGoogle Scholar