Apoptosis

, Volume 18, Issue 5, pp 589–604 | Cite as

Targeting RET to induce medullary thyroid cancer cell apoptosis: an antagonistic interplay between PI3K/Akt and p38MAPK/caspase-8 pathways

  • Minakshi Mazumdar
  • Arghya Adhikary
  • Samik Chakraborty
  • Shravanti Mukherjee
  • Argha Manna
  • Shilpi Saha
  • Suchismita Mohanty
  • Amrita Dutta
  • Pushpak Bhattacharjee
  • Pallab Ray
  • Sreya Chattopadhyay
  • Shuvomoy Banerjee
  • Juni Chakraborty
  • Arun K. Ray
  • Gaurisankar Sa
  • Tanya Das
Original Paper

Abstract

Mutations in REarranged during Transfection (RET) receptor tyrosine, followed by the oncogenic activation of RET kinase is responsible for the development of medullary thyroid carcinoma (MTC) that responds poorly to conventional chemotherapy. Targeting RET, therefore, might be useful in tailoring surveillance of MTC patients. Here we showed that theaflavins, the bioactive components of black tea, successfully induced apoptosis in human MTC cell line, TT, by inversely modulating two molecular pathways: (i) stalling PI3K/Akt/Bad pathway that resulted in mitochondrial transmembrane potential (MTP) loss, cytochrome-c release and activation of the executioner caspases-9 and -3, and (ii) upholding p38MAPK/caspase-8/caspase-3 pathway via inhibition of Ras/Raf/ERK. Over-expression of either constitutively active myristoylated-Akt-cDNA (Myr-Akt-cDNA) or dominant-negative-caspase-8-cDNA (Dn-caspase-8-cDNA) partially blocked theaflavin-induced apoptosis, while co-transfection of Myr-Akt-cDNA and Dn-caspase-8-cDNA completely eradicated the effect of theaflavins thereby negating the possibility of existence of other pathways. A search for the upstream signaling revealed that theaflavin-induced disruption of lipid raft caused interference in anchorage of RET in lipid raft that in turn stalled phosphorylation of Ras and PI3Kinase. In such anti-survival cellular micro-environment, pro-apoptotic signals were triggered to culminate into programmed death of MTC cell. These findings not only unveil a hitherto unexplained mechanism underlying theaflavin-induced MTC death, but also validate RET as a promising and potential target for MTC therapy.

Keywords

Akt Caspase-8 MTC p38MAPK RET Theaflavins 

Abbreviations

cDNA

Complementary deoxyribonucleic acid

ERK

Extracellular signal-regulated protein kinases

IB

Immunoblotting

IP

Immunoprecipitation

MEN 2

Multiple endocrine neoplasia type 2

MTC

Medullary thyroid cancer

p38MAPK

p38 Mitogen-activated protein kinase

RET

REarranged during Transfection

siRNA

Short-interfering RNA

Notes

Acknowledgments

Authors are thankful to Ms. R. Sarkar for editing the manuscript. Thanks are also due to Mr. R. Dutta, Mr. S. Samanta, Mrs. S. Das for technical help. This work was supported by research grants from Department of Biotechnology, Department of Science and Technology, Council of Scientific and Industrial Research, Government of India.

Conflict of interest

None.

References

  1. 1.
    Stratakis CA, Ball DW (2000) A concise genetic and clinical guide to multiple endocrine neoplasias and related syndromes. J Pediatr Endocrinol Metab 13:457–465PubMedGoogle Scholar
  2. 2.
    Torino F, Paragliola RM, Barnabei A, Corsello SM (2010) Medullary thyroid cancer: a promising model for targeted therapy. Curr Mol Med 10:608–625PubMedCrossRefGoogle Scholar
  3. 3.
    Kebebew E, Clark OH (2000) Medullary thyroid cancer. Curr Treat Options Oncol 1:359–667PubMedCrossRefGoogle Scholar
  4. 4.
    Cakir M, Grossman AB (2009) Medullary thyroid cancer: molecular biology and novel molecular therapies. Neuroendocrinology 90:323–348PubMedCrossRefGoogle Scholar
  5. 5.
    Wells SA Jr, Santoro M (2009) Targeting the RET pathway in thyroid cancer. Clin Caner Res 15:7119–7123CrossRefGoogle Scholar
  6. 6.
    Kouvaraki MA, Shapiro SE, Perrier ND, Cote GJ, Gagel RF, Hoff AO et al (2005) RET proto-oncogene: a review and update of genotype-phenotype correlations in hereditary medullary thyroid cancer and associated endocrine tumors. Thyroid 15:531–544PubMedCrossRefGoogle Scholar
  7. 7.
    Takahashi M (2001) The GDNF/RET signaling pathway and human diseases. Cytokine Growth Factor Rev 12:361–373PubMedCrossRefGoogle Scholar
  8. 8.
    Cranston A, Carniti C, Martin S, Mondellini P, Hooks Y, Leyland J et al (2006) A novel activating mutation in the RET tyrosine kinase domain mediates neoplastic transformation. Mol Endocrinol 20:1633–1643PubMedCrossRefGoogle Scholar
  9. 9.
    Putzer BM, Drosten M (2004) The RET proto-oncogene: a potential target for molecular cancer therapy. Trends Mol Med 10:351–357PubMedCrossRefGoogle Scholar
  10. 10.
    Ichihara M, Murakumo Y, Takahashi M (2004) RET and neuroendocrine tumors. Cancer Lett 204:197–211PubMedCrossRefGoogle Scholar
  11. 11.
    Gallel P, Pallares J, Dolcet X, Llobet D, Eritja N, Santacana M et al (2008) Nuclear factor-kappaB activation is associated with somatic and germ line RET mutations in medullary thyroid carcinoma. Hum Pathol 39:994–1001PubMedCrossRefGoogle Scholar
  12. 12.
    Santoro M, Carlomagno F, Melillo RM, Fusco A (2004) Dysfunction of the RET receptor in human cancer. Cell Mol Life Sci 61:2954–2964PubMedCrossRefGoogle Scholar
  13. 13.
    de Groot JW, Links TP, Plukker JT, Lips CJ, Hofstra RM (2006) RET as a diagnostic and therapeutic target in sporadic and hereditary endocrine tumors. Endocinol Rev 27:535–560CrossRefGoogle Scholar
  14. 14.
    Santarpia L, Ye L, Gagel RF (2009) Beyond RET: potential therapeutic approaches for advanced and metastatic medullary thyroid carcinoma. J Intern Med 266:99–113PubMedCrossRefGoogle Scholar
  15. 15.
    Kuo CT, Hsu MJ, Chen BC, Chen CC, Teng CM, Pan SL et al (2008) Denbinobin induces apoptosis in human lung adenocarcinoma cells via Akt inactivation, Bad activation, and mitochondrial dysfunction. Toxicol Lett 177:48–58PubMedCrossRefGoogle Scholar
  16. 16.
    Yoon CH, Kim MJ, Park MT, Byun JY, Choi YH, Yoo HS et al (2009) Acivation of p38 mitogen-activated protein kinase is required for death receptor independent caspase-8 activation and cell death in response to sphingosine. Mol Cancer Res 7:361–370PubMedCrossRefGoogle Scholar
  17. 17.
    Xia Z, Dickens M, Raingeaud J, Davis RJ, Greenberg ME (1995) Opposing effects of ERK and JNK-p38 MAP kinases on apoptosis. Science 270:1326–1331PubMedCrossRefGoogle Scholar
  18. 18.
    Yana I, Nakamura T, Shin E, Karakawa K, Kurahashi H, Kurita Y et al (1992) Inactivation of the p53 gene is not required for tumorigenesis of medullary thyroid carcinoma or pheochromocytoma. Jpn J Cacer Res 83:1113–1116CrossRefGoogle Scholar
  19. 19.
    Pierchala BA, Milbrandt J, Johnson EM Jr (2006) Glial cell line-derived neurotrophic factor-dependent recruitment of Ret into lipid rafts enhances signaling by partitioning Ret from proteasome-dependent degradation. J Neurosci 26:2777–2787PubMedCrossRefGoogle Scholar
  20. 20.
    Kodama Y, Asai N, Kawai K, Jijiwa M, Murakumo Y, Ichihara M et al (2005) The RET proto-oncogene: a molecular therapeutic target in thyroid cancer. Cancer Sci 96:143–148PubMedCrossRefGoogle Scholar
  21. 21.
    Mologni L (2011) Development of RET kinase inhibitors for targeted cancer therapy. Curr Med Chem 18:162–175PubMedCrossRefGoogle Scholar
  22. 22.
    Lahiry L, Saha B, Chakraborty J, Bhattacharyya S, Chattopadhyay S, Banerjee S et al (2008) Contribution of p53-mediated Bax transactivation in theaflavin-induced mammary epithelial carcinoma cell apoptosis. Apoptosis 13:771–781PubMedCrossRefGoogle Scholar
  23. 23.
    Lahiry L, Saha B, Chakraborty J, Adhikary A, Mohanty S, Hossain DM et al (2010) Theaflavins target Fas/caspase-8 and Akt/pBad pathways to induce apoptosis in p53-mutated human breast cancer cells. Carcinogenesis 31:259–268PubMedCrossRefGoogle Scholar
  24. 24.
    Adhikary A, Mohanty S, Lahiry L, Hossain DM, Chakraborty S, Das T (2010) Theaflavins retard human breast cancer cell migration by inhibiting NF-kappaB via p53-ROS cross-talk. FEBS Lett 584:7–14PubMedCrossRefGoogle Scholar
  25. 25.
    Li YC, Park MJ, Ye SK, Kim CW, Kim YN (2006) Elevated levels of cholesterol-rich lipid rafts in cancer cells are correlated with apoptosis sensitivity induced by cholesterol-depleting agents. Am J Pathol 168:1107–1118PubMedCrossRefGoogle Scholar
  26. 26.
    Bhattacharyya A, Mandal D, Lahiry L, Bhattacharyya S, Chattopadhyay S, Ghosh UK et al (2007) Black tea-induced amelioration of hepatic oxidative stress through antioxidative activity in EAC-bearing mice. J Environ Pathol Toxicol Oncol 26:245–254PubMedCrossRefGoogle Scholar
  27. 27.
    Choudhuri T, Pal S, Das T, Sa G (2005) Curcumin selectively induces apoptosis in deregulated cyclin D1-expressed cells at G2 phase of cell cycle in a p53-dependent manner. J Biol Chem 280:20059–20068PubMedCrossRefGoogle Scholar
  28. 28.
    Jaqaman K, Kuwata H, Touret N, Collins R, Trimble WS, Danuser G et al (2011) Cytoskeletal control of CD36 diffusion promotes its receptor and signaling function. Cell 146:593–606PubMedCrossRefGoogle Scholar
  29. 29.
    Balasubramanian N, Scott DW, Castle JD, Casanova JE, Schwartz MA (2007) Arf6 and microtubules in adhesion-dependent trafficking of lipid rafts. Nat Cell Biol 9(12):1381–1391PubMedCrossRefGoogle Scholar
  30. 30.
    Chakraborty J, Banerjee S, Ray P, Hossain DM, Bhattacharyya S, Adhikary A et al (2010) Gain of cellular adaptation due to prolonged p53 impairment leads to functional switchover from p53 to p73 during DNA damage in acute myeloid leukemia cells. J Biol Chem 285:33104–33112PubMedCrossRefGoogle Scholar
  31. 31.
    Stennicke HR, Jürgensmeier JM, Shin H, Deveraux Q, Wolf BB, Yang X et al (1998) Pro-caspase-3 is a major physiologic target of caspase-8. J Biol Chem 273:27084–27090PubMedCrossRefGoogle Scholar
  32. 32.
    Mitsiades N, Poulaki V, Tseleni-Balafouta S, Koutras DA, Stamenkovic I (2000) Thyroid carcinoma cells are resistant to FAS-mediated apoptosis but sensitive to tumor necrosis factor-related apoptosis-inducing ligand. Cancer Res 60:4122–4129PubMedGoogle Scholar
  33. 33.
    Nicolini V, Cassinelli G, Cuccuru G, Bongarzone I, Petrangolini G, Tortoreto M et al (2011) Interplay between Ret and Fap-1 regulates CD95-mediated apoptosis in medullary thyroid cancer cells. Biochem Pharmacol 82(7):778–788PubMedCrossRefGoogle Scholar
  34. 34.
    Rzeszutko M, Rzeszutko W, Dziegiel P, Balcerzak W, Kaliszewski K, Bolanowski M (2007) Expression of FAS/APO 1/CD 95 in thyroid tumors. Folia Histochem Cytobiol 45:87–91PubMedGoogle Scholar
  35. 35.
    Park MT, Choi JA, Kim MJ, Um HD, Bae S, Kang CM et al (2003) Suppression of extracellular signal-related kinase and activation of p38 MAPK are two critical events leading to caspase-8- and mitochondria-mediated cell death in phytosphingosine-treated human cancer cells. J Biol Chem 278:50624–50634PubMedCrossRefGoogle Scholar
  36. 36.
    Xiao YQ, Malcolm K, Worthen GS, Gardai S, Schiemann WP, Fadok VA et al (2002) Cross-talk between ERK and p38 MAPK mediates selective suppression of pro-inflammatory cytokines by transforming growth factor-beta. J Biol Chem 277:14884–14893PubMedCrossRefGoogle Scholar
  37. 37.
    Soderstrom TS, Poukkula M, Holmstrom TH, Heiskanen KM, Eriksson JE (2002) Mitogen-activated protein kinase/extracellular signal-regulated kinase signaling in activated T cells abrogates TRAIL-induced apoptosis upstream of the mitochondrial amplification loop and caspase-8. J Immunol 169:2851–2860PubMedGoogle Scholar
  38. 38.
    Boutros T, Chevet E, Metrakos P (2008) Mitogen-activated protein (MAP) kinase/MAP kinase phosphatase regulation: roles in cell growth, death, and cancer. Pharmacol Rev 60:261–310PubMedCrossRefGoogle Scholar
  39. 39.
    Peyssonnaux C, Eychène A (2001) The Raf/MEK/ERK pathway: new concepts of activation. Biol Cell 93:53–62PubMedCrossRefGoogle Scholar
  40. 40.
    Giehl K, Skripczynski B, Mansard A, Menke A, Gierschik P (2000) Growth factor-dependent activation of the Ras-Raf-MEK-MAPK pathway in the human pancreatic carcinoma cell line PANC-1 carrying activated K-ras: implications for cell proliferation and cell migration. Oncogene 19:2930–2942PubMedCrossRefGoogle Scholar
  41. 41.
    Datta SR, Dudek H, Tao X, Masters S, Fu H, Gotoh Y et al (1997) Akt phosphorylation of BAD couples survival signals to the cell-intrinsic death machinery. Cell 91:231–241PubMedCrossRefGoogle Scholar
  42. 42.
    Richardson DS, Lai AZ, Mulligan LM (2006) RET ligand-induced internalization and its consequences for downstream signaling. Oncogene 25(22):3206–3211PubMedCrossRefGoogle Scholar
  43. 43.
    Shin I, Yakes FM, Rojo F, Shin NY, Bakin AV, Baselga J et al (2002) PKB/Akt mediates cell-cycle progression by phosphorylation of p27(Kip1) at threonine 157 and modulation of its cellular localization. Nat Med 8:1145–1152PubMedCrossRefGoogle Scholar
  44. 44.
    Rodríguez-Antona C, Pallares J, Montero-Conde C, Inglada-Pérez L, Castelblanco E, Landa I et al (2010) Overexpression and activation of EGFR and VEGFR2 in medullary thyroid carcinomas is related to metastasis. Endocr Relat Cancer 17(1):7–16PubMedCrossRefGoogle Scholar
  45. 45.
    Hayashi H, Ichihara M, Iwashita T, Murakami H, Shimono Y, Kawai K et al (2000) Characterization of intracellular signals via tyrosine 1062 in RET activated by glial cell line-derived neurotrophic factor. Oncogene 19:4469–4475PubMedCrossRefGoogle Scholar
  46. 46.
    Viglietto G, Chiappetta G, Martinez-Tello FJ, Fukunaga FH, Tallini G, Rigopoulou D et al (1995) RET/PTC oncogene activation is an early event in thyroid carcinogenesis. Oncogene 11:1207–1210PubMedGoogle Scholar
  47. 47.
    Hodgson JM, Croft KD (2010) Tea flavonoids and cardiovascular health. Mol Aspects Med 31(6):495–502PubMedCrossRefGoogle Scholar
  48. 48.
    Mujtaba T, Duo QP (2012) Black tea polyphenols inhibit tumor proteasome activity. In Vivo 26(2):197–202PubMedGoogle Scholar
  49. 49.
    Wiseman S, Mulder T, Rietveld A (2001) Tea flavonoids: bioavailability in vivo and effects on cell signaling pathways in vitro. Antioxid Redox Signal 3:1009–1021PubMedCrossRefGoogle Scholar
  50. 50.
    Mulder TP, van Platerink CJ, Wijnand Schuyl PJ, van Amelsvoort JM (2001) Analysis of theaflavins in biological fluids using liquid chromatography-electrospray mass spectrometry. J Chromatogr B Biomed Sci Appl 760:271–279PubMedCrossRefGoogle Scholar
  51. 51.
    Henning SM, Aronson W, Niu Y, Conde F, Lee NH, Seeram NP et al (2006) Tea polyphenols and theaflavins are present in prostate tissue of humans and mice after green and black tea consumption. J Nutr 136:1839–1843PubMedGoogle Scholar
  52. 52.
    Saha B, Adhikary A, Ray P, Saha S, Chakraborty S, Mohanty S et al (2011) Restoration of tumor suppressor p53 by differentially regulating pro- and anti-p53 networks in HPV-18-infected cervical cancer cells. Oncogene 31:173–186PubMedCrossRefGoogle Scholar
  53. 53.
    Herfarth KK, Wick MR, Marshall HN, Gartner E, Lum S, Moley JF (1997) Absence of TP53 alterations in pheochromocytomas and medullary thyroid carcinomas. Genes Chromosomes Cancer 20:24–29PubMedCrossRefGoogle Scholar
  54. 54.
    Micheau O, Solary E, Hammann A, Dimanche-Boitrel MT (1999) Fas ligand-independent, FADD-mediated activation of the Fas death pathway by anticancer drugs. J Biol Chem 274:7987–7992PubMedCrossRefGoogle Scholar
  55. 55.
    Shammas MA, Neri P, Koley H, Batchu RB, Bertheau RC, Munshi V et al (2006) Specific killing of multiple myeloma cells by (−)-epigallocatechin-3-gallate extracted from green tea: biologic activity and therapeutic implications. Blood 108:2804–2810PubMedCrossRefGoogle Scholar
  56. 56.
    Shao RG, Cao CX, Nieves-Neira W, Dimanche-Boitrel MT, Solary E, Pommier Y (2001) Activation of the Fas pathway independently of Fas ligand during apoptosis induced by camptothecin in p53 mutant human colon carcinoma cells. Oncogene 20:1852–1859PubMedCrossRefGoogle Scholar
  57. 57.
    Hayakawa S, Saeki K, Sazuka M, Suzuki Y, Shoji Y, Ohta T et al (2001) Apoptosis induction by epigallocatechin gallate involves its binding to Fas. Biochem Biophys Res Commun 285:1102–1106PubMedCrossRefGoogle Scholar
  58. 58.
    Coxon AB, Ward JM, Geradts J, Otterson GA, Zajac-Kaye M, Kaye FJ (1998) RET cooperates with RB/p53 inactivation in a somatic multi-step model for murine thyroid cancer. Oncogene 17:1625–1628PubMedCrossRefGoogle Scholar
  59. 59.
    Zhang HY, Meng X, Du ZX, Fang CQ, Liu GL, Wang HQ et al (2009) Significance of survivin, caspase-3, and VEGF expression in thyroid carcinoma. Clin Exp Med 9:207–213PubMedCrossRefGoogle Scholar
  60. 60.
    Rinner B, Siegl V, Pürstner P, Efferth T, Brem B, Greger H et al (2004) Activity of novel plant extracts against medullary thyroid carcinoma cells. Anticancer Res 24:495–500PubMedGoogle Scholar
  61. 61.
    Segouffin-Cariou C, Billaud M (2000) Transforming ability of MEN2A-RET requires activation of the phosphatidylinositol 3-kinase/AKT signaling pathway. J Biol Chem 275:3568–3576PubMedCrossRefGoogle Scholar
  62. 62.
    Santoro M, Melillo RM, Carlomagno F, Fusco A, Vecchio G (2002) Molecular mechanisms of RET activation in human cancer. Ann N Y Acad Sci 963:116–121PubMedCrossRefGoogle Scholar
  63. 63.
    Drosten M, Hilken G, Bokmann M, Rodicker F, Mise N, Cranston AN et al (2004) Role of MEN2A-derived RET in maintenance and proliferation of medullary thyroid carcinoma. J Natl Cancer Inst 96:1231–1239PubMedCrossRefGoogle Scholar
  64. 64.
    Kunnimalaiyaan M, Ndiaye M, Chen H (2006) Apoptosis-mediated medullary thyroid cancer growth suppression by the PI3K inhibitor LY294002. Surgery 140:1009–1014PubMedCrossRefGoogle Scholar
  65. 65.
    Furuya F, Lu C, Willingham MC, Cheng SY (2007) Inhibition of phosphatidylinositol 3-kinase delays tumor progression and blocks metastatic spread in a mouse model of thyroid cancer. Carcinogenesis 28:2451–2458PubMedCrossRefGoogle Scholar
  66. 66.
    Liu Z, Hou P, Ji M, Guan H, Studeman K, Jensen K et al (2008) Highly prevalent genetic alterations in receptor tyrosine kinases and phosphatidylinositol 3-kinase/akt and mitogen-activated protein kinase pathways in anaplastic and follicular thyroid cancers. J Clin Endocrinol Metab 93:3106–3116PubMedCrossRefGoogle Scholar
  67. 67.
    She M, Yang H, Sun L, Yeung SC (2006) Redox control of manumycin A-induced apoptosis in anaplastic thyroid cancer cells: involvement of the xenobiotic apoptotic pathway. Cancer Biol Ther 5:275–280PubMedCrossRefGoogle Scholar
  68. 68.
    Schlumberger M, Carlomagno F, Baudin E, Bidart JM, Santoro M (2008) New therapeutic approaches to treat medullary thyroid carcinoma. Nat Clin Pract Endocrinol Metab 4:22–32PubMedCrossRefGoogle Scholar
  69. 69.
    Adachi S, Nagao T, Ingolfsson HI, Maxfield FR, Andersen OS, Kopelovich L et al (2007) The inhibitory effect of (−)-epigallocatechin gallate on activation of the epidermal growth factor receptor is associated with altered lipid order in HT29 colon cancer cells. Cancer Res 67:6493–6501PubMedCrossRefGoogle Scholar
  70. 70.
    Vitagliano D, De Falco V, Tamburrino A, Coluzzi S, Troncone G, Chiappetta G et al (2011) The tyrosine kinase inhibitor ZD6474 blocks proliferation of RET mutant medullary thyroid carcinoma cells. Endocr Relat Cancer 18(1):1–11PubMedCrossRefGoogle Scholar
  71. 71.
    Bucci C, Thomsen P, Nicoziani P, McCarthy J, Deurs B (2000) Rab7: a key to lysosome biogenesis. Mol Biol Cell 11:467–480PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Minakshi Mazumdar
    • 1
  • Arghya Adhikary
    • 1
  • Samik Chakraborty
    • 1
  • Shravanti Mukherjee
    • 1
  • Argha Manna
    • 1
  • Shilpi Saha
    • 1
  • Suchismita Mohanty
    • 1
  • Amrita Dutta
    • 1
  • Pushpak Bhattacharjee
    • 1
  • Pallab Ray
    • 1
  • Sreya Chattopadhyay
    • 1
  • Shuvomoy Banerjee
    • 1
  • Juni Chakraborty
    • 1
  • Arun K. Ray
    • 1
  • Gaurisankar Sa
    • 1
  • Tanya Das
    • 1
  1. 1.Division of Molecular MedicineBose InstituteKolkataIndia

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