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Smac mimetic sensitizes glioblastoma cells to Temozolomide-induced apoptosis in a RIP1- and NF-κB-dependent manner

Oncogene volume 32, pages 988–997 (2013)Cite this article

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Abstract

Inhibitor of apoptosis (IAP) proteins are expressed at high levels in many cancers and therefore represent attractive targets for therapeutic intervention. Here, we report for the first time that the second mitochondria-derived activator of caspases (Smac) mimetic BV6 sensitizes glioblastoma cells toward Temozolomide (TMZ), the first-line chemotherapeutic agent in the treatment of glioblastoma. BV6 and TMZ synergistically reduce cell viability and trigger apoptosis in glioblastoma cells (combination index <0.4–0.8), which is accompanied by increased loss of mitochondrial-membrane potential, cytochrome c release, caspase activation and caspase-dependent apoptosis. Analysis of the molecular mechanisms reveals that BV6 causes rapid degradation of cIAP1, leading to stabilization of NF-κB-inducing kinase and NF-κB activation. BV6-stimulated NF-κB activation is critically required for sensitization toward TMZ, as inhibition of NF-κB by overexpression of the mutant IκBα super-repressor profoundly reduces loss of mitochondrial membrane potential, cytochrome c release, caspase activation and apoptosis. Of note, BV6-mediated sensitization to TMZ is not associated with increased tumor necrosis factor alpha (TNFα) production. Also, TNFα, CD95 or TRAIL-blocking antibodies or knockdown of TNFR1 have no or little effect on combination treatment-induced apoptosis. Interestingly, BV6 and TMZ cooperate to trigger the formation of a RIP1 (receptor activating protein 1)/caspase-8/FADD complex. Knockdown of RIP1 by small interfering RNA significantly reduces BV6- and TMZ-induced caspase-8 activation and apoptosis, showing that RIP1 is necessary for apoptosis induction. By demonstrating that BV6 primes glioblastoma cells for TMZ in a NF-κB- and RIP1-dependent manner, these findings build the rationale for further (pre)clinical development of Smac mimetics in combination with TMZ.

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References

  1. DeAngelis LM . Brain tumors. N Engl J Med 2001; 344: 114–123.

    Article  CAS  PubMed  Google Scholar 

  2. Stupp R, Hegi ME, Gilbert MR, Chakravarti A . Chemoradiotherapy in malignant glioma: standard of care and future directions. J Clin Oncol 2007; 25: 4127–4136.

    Article  CAS  PubMed  Google Scholar 

  3. Hanahan D, Weinberg RA . Hallmarks of cancer: the next generation. Cell 2011; 144: 646–674.

    Article  CAS  PubMed  Google Scholar 

  4. Fulda S . Tumor resistance to apoptosis. Int J Cancer 2009; 124: 511–515.

    Article  CAS  PubMed  Google Scholar 

  5. Fulda S, Debatin KM . Extrinsic versus intrinsic apoptosis pathways in anticancer chemotherapy. Oncogene 2006; 25: 4798–4811.

    Article  CAS  PubMed  Google Scholar 

  6. Ashkenazi A . Targeting the extrinsic apoptosis pathway in cancer. Cytokine Growth Factor Rev 2008; 19: 325–331.

    Article  CAS  PubMed  Google Scholar 

  7. Fulda S, Galluzzi L, Kroemer G . Targeting mitochondria for cancer therapy. Nat Rev Drug Discov 2010; 9: 447–464.

    Article  CAS  PubMed  Google Scholar 

  8. LaCasse EC, Mahoney DJ, Cheung HH, Plenchette S, Baird S, Korneluk RG . IAP-targeted therapies for cancer. Oncogene 2008; 27: 6252–6275.

    Article  CAS  PubMed  Google Scholar 

  9. Varfolomeev E, Blankenship JW, Wayson SM, Fedorova AV, Kayagaki N, Garg P et al. IAP antagonists induce autoubiquitination of c-IAPs, NF-kappaB activation, and TNFalpha-dependent apoptosis. Cell 2007; 131: 669–681.

    Article  CAS  PubMed  Google Scholar 

  10. Vince JE, Wong WW, Khan N, Feltham R, Chau D, Ahmed AU et al. IAP antagonists target cIAP1 to induce TNFalpha-dependent apoptosis. Cell 2007; 131: 682–693.

    Article  CAS  PubMed  Google Scholar 

  11. Bertrand MJ, Milutinovic S, Dickson KM, Ho WC, Boudreault A, Durkin J et al. cIAP1 and cIAP2 facilitate cancer cell survival by functioning as E3 ligases that promote RIP1 ubiquitination. Mol Cell 2008; 30: 689–700.

    Article  CAS  PubMed  Google Scholar 

  12. Fulda S, Wick W, Weller M, Debatin KM . Smac agonists sensitize for Apo2L/TRAIL- or anticancer drug-induced apoptosis and induce regression of malignant glioma in vivo. Nat Med 2002; 8: 808–815.

    Article  CAS  PubMed  Google Scholar 

  13. Giagkousiklidis S, Vogler M, Westhoff MA, Kasperczyk H, Debatin KM, Fulda S . Sensitization for gamma-irradiation-induced apoptosis by second mitochondria-derived activator of caspase. Cancer Res 2005; 65: 10502–10513.

    Article  CAS  PubMed  Google Scholar 

  14. Vellanki SH, Grabrucker A, Liebau S, Proepper C, Eramo A, Braun V et al. Small-molecule XIAP inhibitors enhance gamma-irradiation-induced apoptosis in glioblastoma. Neoplasia 2009; 11: 743–752.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Karl S, Pritschow Y, Volcic M, Hacker S, Baumann B, Wiesmuller L et al. Identification of a novel pro-apoptotic function of NF-kappaB in the DNA damage response. J Cell Mol Med 2009; 13: 4239–4256.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Shirley S, Micheau O . Targeting c-FLIP in cancer. Cancer Lett 2010 (e-pub ahead of print 9 November 2010).

  17. Fulda S, Vucic D . Targeting IAP proteins for therapeutic intervention in cancer. Nat Rev Drug Disc 2012; 11: 109–124.

    Article  CAS  Google Scholar 

  18. Wang L, Du F, Wang X . TNF-alpha induces two distinct caspase-8 activation pathways. Cell 2008; 133: 693–703.

    Article  CAS  PubMed  Google Scholar 

  19. Festjens N, Vanden Berghe T, Cornelis S, Vandenabeele P . RIP1, a kinase on the crossroads of a cell's decision to live or die. Cell Death Differ 2007; 14: 400–410.

    Article  CAS  PubMed  Google Scholar 

  20. Probst BL, Liu L, Ramesh V, Li L, Sun H, Minna JD et al. Smac mimetics increase cancer cell response to chemotherapeutics in a TNF-alpha-dependent manner. Cell Death Differ 2010; 17: 1645–1654.

    Article  CAS  PubMed  Google Scholar 

  21. Straub CS . Targeting IAPs as an approach to anti-cancer therapy. Curr Top Med Chem 2011; 11: 291–316.

    Article  CAS  PubMed  Google Scholar 

  22. Fulda S, Friesen C, Los M, Scaffidi C, Mier W, Benedict M et al. Betulinic acid triggers CD95 (APO-1/Fas)- and p53-independent apoptosis via activation of caspases in neuroectodermal tumors. Cancer Res 1997; 57: 4956–4964.

    CAS  PubMed  Google Scholar 

  23. Fulda S, Strauss G, Meyer E, Debatin KM . Functional CD95 ligand and CD95 death-inducing signaling complex in activation-induced cell death and doxorubicin-induced apoptosis in leukemic T cells. Blood 2000; 95: 301–308.

    CAS  PubMed  Google Scholar 

  24. Mohr A, Zwacka RM, Debatin KM, Stahnke K . A novel method for the combined flow cytometric analysis of cell cycle and cytochrome c release. Cell Death Differ 2004; 11: 1153–1154.

    Article  CAS  PubMed  Google Scholar 

  25. Opel D, Westhoff MA, Bender A, Braun V, Debatin KM, Fulda S . Phosphatidylinositol 3-kinase inhibition broadly sensitizes glioblastoma cells to death receptor- and drug-induced apoptosis. Cancer Res 2008; 68: 6271–6280.

    Article  CAS  PubMed  Google Scholar 

  26. Gonzalez P, Mader I, Tchoghandjian A, Enzenmüller S, Cristofanon S, Basit F et al. Impairment of lysosomal integrity by B10, a glycosylated derivative of betulinic acid, leads to lysosomal cell death and converts autophagy into a detrimental process. Cell Death Differ 2012; 19: 1337–1346.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Hacker S, Dittrich A, Mohr A, Schweitzer T, Rutkowski S, Krauss J et al. Histone deacetylase inhibitors cooperate with IFN-gamma to restore caspase-8 expression and overcome TRAIL resistance in cancers with silencing of caspase-8. Oncogene 2009; 28: 3097–3110.

    Article  CAS  PubMed  Google Scholar 

  28. Berger R, Jennewein C, Marschall V, Karl S, Cristofanon S, Wagner L et al. NF-{kappa}B is required for Smac mimetic-mediated sensitization of glioblastoma cells for (gamma)}-irradiation-induced apoptosis. Mol Cancer Ther 2011; 10: 1867–1875.

    Article  CAS  PubMed  Google Scholar 

  29. Chou TC . The median-effect principle and the combination index for quantitation of synergism and antagonism. In: Chou TC (ed). Synergism and Antagonism in Chemotherapy. Academic Press: San Diego, CA, USA, 1991, pp 61–102.

    Google Scholar 

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Acknowledgements

We thank C Hugenberg for expert secretarial assistance. This work has been partially supported by grants from the Deutsche Forschungsgemeinschaft, European Community (ApopTrain, APO-SYS) and IAP6/18 (to SF) and by a scholarship from the International Graduate School in Molecular Medicine Ulm, Ulm University (to LW). Kerry Zobel, Kurt Deshayes, and Domagoj Vucic are employees of Genentech, Inc.

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Authors and Affiliations

  1. Department of Pediatrics and Adolescent Medicine,

    L Wagner, S Karl, K-M Debatin & S Fulda

  2. University Medical Center Ulm,, Ulm, Germany

    L Wagner, S Karl, K-M Debatin & S Fulda

  3. Institute for Experimental Cancer Research in Pediatrics, Goethe-University Frankfurt, Frankfurt, Germany

    V Marschall, S Cristofanon & S Fulda

  4. Genentech, Inc., 1 DNA Way, South San Francisco, CA, USA

    K Zobel, K Deshayes & D Vucic

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  1. L Wagner
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Correspondence to S Fulda.

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Wagner, L., Marschall, V., Karl, S. et al. Smac mimetic sensitizes glioblastoma cells to Temozolomide-induced apoptosis in a RIP1- and NF-κB-dependent manner. Oncogene 32, 988–997 (2013). https://doi.org/10.1038/onc.2012.108

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