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Celastrol inhibits growth and induces apoptotic cell death in melanoma cells via the activation ROS-dependent mitochondrial pathway and the suppression of PI3K/AKT signaling

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Abstract

Celastrol has been reported to possess anticancer effects in various cancers; however, the precise mechanism underlying ROS-mediated mitochondria-dependent apoptotic cell death triggered by celastrol treatment in melanoma cells remains unknown. We showed that celastrol effectively induced apoptotic cell death and inhibited tumor growth using tissue culture and in vivo models of B16 melanoma. In addition to apoptotic cell death in B16 cells, several apoptotic events such as PARP cleavage and activation of caspase were confirmed. Pretreatment with caspase inhibitor modestly attenuated the celastrol-induced increase in PARP cleavage and sub-G1 cell population, implying that caspases play a partial role in celastrol-induced apoptosis. Moreover, ROS generation was detected following celastrol treatment. Blocking of ROS accumulation with ROS scavengers resulted in inhibition of celastrol-induced Bcl-2 family-mediated apoptosis, indicating that celastrol-induced apoptosis involves ROS generation as well as an increase in the Bax/Bcl-2 ratio leading to release of cytochrome c and AIF. Importantly, silencing of AIF by transfection of siAIF into cells remarkably attenuated celastrol-induced apoptotic cell death. Moreover, celastrol inhibited the activation of PI3K/AKT/mTOR signaling cascade in B16 cells. Our data reveal that celastrol inhibits growth and induces apoptosis in melanoma cells via the activation of ROS-mediated caspase-dependent and -independent pathways and the suppression of PI3K/AKT signaling.

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References

  1. 1.

    Trott A, West JD, Klaić L, Westerheide SD, Silverman RB, Morimoto RI, Morano KA (2008) Activation of heat shock and antioxidant responses by the natural product celastrol: transcriptional signatures of a thiol-targeted molecule. Mol Biol Cell 19:1104–1112

  2. 2.

    Pinna GF, Fiorucci M, Reimund JM, Taquet N, Arondel Y, Muller CD (2004) Celastrol inhibits pro-inflammatory cytokine secretion in Crohn’s disease biopsies. Biochem Biophys Res Commun 322:778–786

  3. 3.

    Abbas S, Bhoumik A, Dahl R, Vasile S, Krajewski S, Cosford ND, Ronai ZA (2007) Preclinical studies of celastrol and acetyl isogambogic acid in melanoma. Clin Cancer Res 13:6769–6778

  4. 4.

    Zhang T, Hamza A, Cao X, Wang B, Yu S, Zhan CG, Sun D (2008) A novel Hsp90 inhibitor to disrupt Hsp90/Cdc37 complex against pancreatic cancer cells. Mol Cancer Ther 7:162–170

  5. 5.

    Yang H, Chen D, Cui QC, Yuan X, Dou QP (2006) Celastrol, a triterpene extracted from the Chinese “Thunder of God Vine,” is a potent proteasome inhibitor and suppresses human prostate cancer growth in nude mice. Cancer Res 66:4758–4765

  6. 6.

    Nagase M, Oto J, Sugiyama S, Yube K, Takaishi Y, Sakato N (2003) Apoptosis induction in HL-60 cells and inhibition of topoisomerase II by triterpene celastrol. Biosci Biotechnol Biochem 67:1883–1887

  7. 7.

    Sethi G, Ahn KS, Pandey MK, Aggarwal BB (2007) Celastrol, a novel triterpene, potentiates TNF-induced apoptosis and suppresses invasion of tumor cells by inhibiting NF-kappaB-regulated gene products and TAK1-mediated NF-kappaB activation. Blood 109:2727–2735

  8. 8.

    Ferreira CG, Epping M, Kruyt FA, Giaccone G (2002) Apoptosis: target of cancer therapy. Clin Cancer Res 8:2024–2034

  9. 9.

    Song Z, Steller H (1999) Death by design: mechanism and control of apoptosis. Trends Cell Biol 9:M49–M52

  10. 10.

    Kolenko VM, Uzzo RG, Bukowski R, Finke JH (2000) Caspase-dependent and independent death pathways in cancer therapy. Apoptosis 5:17–20

  11. 11.

    Slee EA, Adrain C, Martin SJ (1999) Serial killers: ordering caspase activation events in apoptosis. Cell Death Differ 6:1067–1074

  12. 12.

    Krantic S, Mechawar N, Reix S, Quirion R (2007) Apoptosis-inducing factor: a matter of neuron life and death. Prog Neurobiol 81:179–196

  13. 13.

    Corbiere C, Liagre B, Terro F, Beneytout JL (2004) Induction of antiproliferative effect by diosgenin through activation of p53, release of apoptosis-inducing factor (AIF) and modulation of caspase-3 activity in different human cancer cells. Cell Res 14:188–196

  14. 14.

    Susin SA, Lorenzo HK, Zamzami N, Marzo I, Snow BE, Brothers GM, Mangion J, Jacotot E, Costantini P, Loeffler M, Larochette N, Goodlett DR, Aebersold R, Siderovski DP, Penninger JM, Kroemer G (1999) Molecular characterization of mitochondrial apoptosis-inducing factor. Nature 397:441–446

  15. 15.

    Li LY, Luo X, Wang X (2001) Endonuclease G is an apoptotic DNase when released from mitochondria. Nature 412:95–99

  16. 16.

    Buttke TM, Sandstrom PA (1994) Oxidative stress as a mediator of apoptosis. Immunol Today 15:7–10

  17. 17.

    Seo WY, Goh AR, Ju SM, Song HY, Kwon DJ, Jun JG, Kim BC, Choi SY, Park J (2011) Celastrol induces expression of heme oxygenase-1 through ROS/Nrf2/ARE signaling in the HaCaT cells. Biochem Biophys Res Commun 407:535–540

  18. 18.

    Chen G, Zhang X, Zhao M, Wang Y, Cheng X, Wang D, Xu Y, Du Z, Yu X (2011) Celastrol targets mitochondrial respiratory chain complex I to induce reactive oxygen species-dependent cytotoxicity in tumor cells. BMC Cancer 11:170

  19. 19.

    Sung B, Park B, Yadav VR, Aggarwal BB (2010) Celastrol, a triterpene, enhances TRAIL-induced apoptosis through the down-regulation of cell survival proteins and up-regulation of death receptors. J Biol Chem 285:11498–11507

  20. 20.

    Yang HS, Kim JY, Lee JH, Lee BW, Park KH, Shim KH, Lee MK, Seo KI (2011) Celastrol isolated from Tripterygium regelii induces apoptosis through both caspase-dependent and -independent pathways in human breast cancer cells. Food Chem Toxicol 49:527–532

  21. 21.

    Madhunapantula SV, Mosca PJ, Robertson GP (2011) The Akt signaling pathway: an emerging therapeutic target in malignant melanoma. Cancer Biol Ther 12:1032–1049

  22. 22.

    Davies MA (2012) The role of the PI3K-AKT pathway in melanoma. Cancer J 18:142–147

  23. 23.

    Pang X, Yi Z, Zhang J, Lu B, Sung B, Qu W, Aggarwal BB, Liu M (2010) Celastrol suppresses angiogenesis-mediated tumor growth through inhibition of AKT/mammalian target of rapamycin pathway. Cancer Res 70:1951–1959

  24. 24.

    Kannaiyan R, Manu KA, Chen L, Li F, Rajendran P, Subramaniam A, Lam P, Kumar AP, Sethi G (2011) Celastrol inhibits tumor cell proliferation and promotes apoptosis through the activation of c-Jun N-terminal kinase and suppression of PI3K/Akt signaling pathways. Apoptosis 16:1028–1041

  25. 25.

    Ryu YB, Park SJ, Kim YM, Lee JY, Seo WD, Chang JS, Park KH, Rho MC, Lee WS (2010) SARS-CoV 3CLpro inhibitory effects of quinone-methide triterpenes from Tripterygium regelii. Bioorg Med Chem Lett 20:1873–1876

  26. 26.

    Skehan P, Storeng R, Scudiero D, Monks A, McMahon J, Vistica D, Warren JT, Bokesch H, Kenney S, Boyd MR (1990) New colorimetric cytotoxicity assay for anticancer-drug screening. J Natl Cancer Inst 82:1107–1112

  27. 27.

    Park SY, Cho SJ, Kwon HC, Lee KR, Rhee DK, Pyo S (2005) Caspase-independent cell death by allicin in human epithelial carcinoma cells: involvement of PKA. Cancer Lett 224:123–132

  28. 28.

    Ricote M, García-Tuñón I, Fraile B, Fernández C, Aller P, Paniagua R, Royuela M (2006) P38 MAPK protects against TNF-α-provoked apoptosis in LNCaP cancer cells. Apoptosis 11:1969–1975

  29. 29.

    Wan CK, Wang C, Cheung HY, Yang MS, Fong WF (2005) Triptolide induces Bcl-2 cleavage and mitochondria dependent apoptosis in p53-dependent HL-60 cells. Cancer Lett 241:1–11

  30. 30.

    Chen CY, Liu TZ, Liu YW, Tseng WC, Liu RH, Lu FJ, Lin YS, Kuo SH, Chen CH (2007) 6-Shogaol (alkanone from ginger) induces apoptotic cell death of human hepatoma p53 mutant Mahlavu subline via an oxidative stress-mediated caspase-dependent mechanism. J Agric Food Chem 55:948–954

  31. 31.

    Lee JH, Kishikawa M, Kumazoe M, Yamada K, Tachibana H (2010) Vitamin A enhances antitumor effect of a green tea polyphenol on melanoma by upregulating the polyphenol sensing molecule 67-kDa laminin receptor. PLoS ONE 5:e11051

  32. 32.

    Chang CP, Yang MC, Liu HS, Lin YS, Lei HY (2007) Concanavalin A induces autophagy in hepatoma cells and has a therapeutic effect in a murine in situ hepatoma model. Hepatology 45:286–296

  33. 33.

    Adam JM, Cory S (1998) The Bcl-2 protein family: arbiters of cell survival. Science 281:1322–1326

  34. 34.

    Tsujimoto Y (1998) Role of Bcl-2 family proteins in apoptosis: apoptosomes or mitochondria? Genes Cells 3:697–707

  35. 35.

    Budihardjo I, Oliver H, Lutter M, Luo X, Wang X (1999) Biochemical pathways of caspase activation during apoptosis. Annu Rev Cell Dev Biol 15:269–290

  36. 36.

    Gustafsson AB, Gottlieb RA (2007) Bcl-2 family members and apoptosis, taken to heart. Am J Physiol Cell Physiol 292:C45–C51

  37. 37.

    Kumar S (2007) Caspase function in programmed cell death. Cell Death Differ 14:32–43

  38. 38.

    Tan S, Sagara Y, Liu Y, Maher P, Schubert D (1998) The regulation of reactive oxygen species production during programmed cell death. J Cell Biol 141:1423–1432

  39. 39.

    Kluck RM, Bossy-Wetzel E, Green DR, Newmeyer DD (1997) The release of cytochrome c from mitochondria: a primary site for Bcl-2 regulation of apoptosis. Science 275:1132–1136

  40. 40.

    Lorenzo E, Ruiz-Ruiz C, Quesada AJ, Hernández G, Rodríguez A, López-Rivas A, Redondo JM (2002) Doxorubicin induces apoptosis and CD95 gene expression in human primary endothelial cells through a p53-dependent mechanism. J Biol Chem 277:10883–10892

  41. 41.

    Gottlieb E, Vander Heiden MG, Thompson CB (2000) Bcl-x(L) prevents the initial decrease in mitochondrial membrane potential and subsequent reactive oxygen species production during tumor necrosis factor alpha-induced apoptosis. Mol Cell Biol 20:5680–5689

  42. 42.

    Howard S, Bottino C, Brooke S, Cheng E, Giffard RG, Sapolsky R (2002) Neuroprotective effects of bcl-2 overexpression in hippocampal cultures: interactions with pathways of oxidative damage. J Neurochem 83:914–923

  43. 43.

    Chen KC, Chang LS (2009) Arachidonic acid-induced apoptosis of human neuroblastoma SK-N-SH cells is mediated through mitochondrial alteration elicited by ROS and Ca(2+)-evoked activation of p38alpha MAPK and JNK1. Toxicology 262:199–206

  44. 44.

    Choi WY, Choi BT, Lee WH, Choi YH (2008) Sulforaphane generates reactive oxygen species leading to mitochondrial perturbation for apoptosis in human leukemia U937 cells. Biomed Pharmacother 62:637–644

  45. 45.

    Li P, Nijhawan D, Budihardjo I, Srinivasula SM, Ahmad M, Alnemri ES, Wang X (1997) Cytochrome c and dATP-dependent formation of Apaf-1/caspase-9 complex initiates an apoptotic protease cascade. Cell 91:479–489

  46. 46.

    Muzio M, Stockwell BR, Stennicke HR, Salvesen GS, Dixit VM (1998) An induced proximity model for caspase-8 activation. J Biol Chem 273:2926–2930

  47. 47.

    Zhou LL, Lin ZX, Fung KP, Cheng CH, Che CT, Zhao M, Wu SH, Zuo Z (2011) Celastrol-induced apoptosis in human HaCaT keratinocytes involves the inhibition of NF-κB activity. Eur J Pharmacol 670:399–408

  48. 48.

    Mou H, Zheng Y, Zhao P, Bao H, Fang W, Xu N (2011) Celastrol induces apoptosis in non-small-cell lung cancer A549 cells through activation of mitochondria- and Fas/FasL-mediated pathways. Toxicol In Vitro 25:1027–1032

  49. 49.

    Simon HU, Haj-Yehia A, Levi-Schaffer F (2000) Role of reactive oxygen species (ROS) in apoptosis induction. Apoptosis 5:415–418

  50. 50.

    Green DR, Reed JC (1998) Mitochondria and apoptosis. Science 281:1309–1312

  51. 51.

    Kroemer G, Reed JC (2000) Mitochondrial control of cell death. Nat Med 6:513–519

  52. 52.

    Ma Q, Fang H, Shang W, Liu L, Xu Z, Ye T, Wang X, Zheng M, Chen Q, Cheng H (2011) Superoxide flashes: early mitochondrial signals for oxidative stress-induced apoptosis. J Biol Chem 286:27573–27581

  53. 53.

    Tait SW, Green DR (2008) Caspase-independent cell death: leaving the set without the final cut. Oncogene 27:6452–6461

  54. 54.

    Liu PL, Chen YL, Chen YH, Lin SJ, Kou YR (2005) Wood smoke extract induces oxidative stress-mediated caspase-independent apoptosis in human lung endothelial cells: role of AIF and EndoG. Am J Physiol Lung Cell Mol Physiol 289:L739–L749

  55. 55.

    Candé C, Cohen I, Daugas E, Ravagnan L, Larochette N, Zamzami N, Kroemer G (2002) Apoptosis-inducing factor (AIF): a novel caspase-independent death effector released from mitochondria. Biochimie 84:215–222

  56. 56.

    Candé C, Cecconi F, Dessen P, Kroemer G (2002) Apoptosis-inducing factor (AIF): key to the conserved caspase-independent pathways of cell death? J Cell Sci 115:4727–4734

  57. 57.

    Lorenzo HK, Susin SA (2004) Mitochondrial effectors in caspase-independent cell death. FEBS Lett 557:14–20

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Acknowledgments

This work was supported by National Research Foundation of Korean Grant funded by the Korean Government, Ministry of Education, Science and Technology (NRF-2010-355-F00049).

Conflict of interest

The authors declare that they no conflict of interest.

Author information

Correspondence to Kwon-Il Seo.

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Lee, J., Won, Y., Park, K. et al. Celastrol inhibits growth and induces apoptotic cell death in melanoma cells via the activation ROS-dependent mitochondrial pathway and the suppression of PI3K/AKT signaling. Apoptosis 17, 1275–1286 (2012). https://doi.org/10.1007/s10495-012-0767-5

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Keywords

  • Celastrol
  • Reactive oxygen species
  • Apoptosis
  • Apoptosis inducing factor
  • B16 melanoma