Download PDF
Horm Metab Res 2015; 47(03): 200-208
DOI: 10.1055/s-0034-1390484
Endocrine Research
© Georg Thieme Verlag KG Stuttgart · New York

Expression of the cAMP Binding Protein EPAC1 in Thyroid Tumors and Growth Regulation of Thyroid Cells and Thyroid Carcinoma Cells by EPAC Proteins

M. Broecker-Preuss
1   Department of Endocrinology and Metabolism, and Division of Laboratory Research, University Hospital Essen, Essen, Germany
2   Current Address: University Hospital Essen, Department of Clinical Chemistry, Core-Lab, Essen, Germany
,
J. Baten
1   Department of Endocrinology and Metabolism, and Division of Laboratory Research, University Hospital Essen, Essen, Germany
,
S.-Y. Sheu-Grabellus
3   Institute of Pathology, University Hospital Essen, Essen, Germany
,
R. Görges
4   Clinic of Nuclear Medicine, University Hospital Essen, Essen, Germany
,
A. Bockisch
4   Clinic of Nuclear Medicine, University Hospital Essen, Essen, Germany
,
K. W. Schmid
3   Institute of Pathology, University Hospital Essen, Essen, Germany
,
D. Führer
1   Department of Endocrinology and Metabolism, and Division of Laboratory Research, University Hospital Essen, Essen, Germany
,
K. Mann
1   Department of Endocrinology and Metabolism, and Division of Laboratory Research, University Hospital Essen, Essen, Germany
› Author Affiliations
Further Information

Publication History

received 27 March 2014

accepted 15 September 2014

Publication Date:
05 November 2014 (online)

Also available at
    Permissions and Reprints

Abstract

The thyrotropin receptor-cAMP pathway is central in growth regulation of thyroid cells and thyroid tumorigenesis, and it regulates expression of thyroid specific genes. Recently, 2 new protein kinase A-independent cAMP effectors named EPAC1 and 2 were described that activate additional intracellular pathways. The aim of our study was to investigate the role of EPAC proteins in growth regulation of thyroid cells and thyroid carcinomas. EPAC1 expression was investigated immunohistochemically in tissues of various thyroid tumors. Utilizing MTT assay, the effect of EPAC stimulation on proliferation in thyroid carcinoma cells and in non-transformed rat FRTL5 cells was investigated. The activation of intracellular signaling pathways was examined by RAP pull-down assay and Western blots. EPAC1 expression was strong in non-oxyphilic follicular thyroid adenomas and carcinomas and in follicular papillary thyroid carcinomas. It was moderate in oxyphilic follicular tumors and classical and tall cell papillary carcinomas. In contrast, EPAC1 expression was low in poorly differentiated carcinomas and very low in anaplastic carcinomas. Thyroid carcinoma cell lines showed no or very weak EPAC1 expression and exhibited no growth-promoting effect after EPAC stimulation. Non-transformed rat FRTL5 cells were growth-stimulated by an EPAC-specific cAMP-analogue and showed EPAC-dependent activation of RAP, ERK, and p70S6 kinase. EPAC1 expression and cellular response to EPAC activation in rat FRTL5 cells reflect cellular responses to cAMP and TSH stimulation in non-transformed thyroid cells. In undifferentiated thyroid carcinomas, loss of EPAC1 expression may be in accordance with the loss of thyroid-specific functions and the loss of responsiveness of the TSHR-cAMP pathway.

Key words

cAMP signaling - TSH receptor - thyroid carcinoma - exchange protein activated by cAMP
 

  • References

  • 1 Davies L, Welch HG. Increasing incidence of thyroid cancer in the United States, 1973–2002. JAMA 2006; 295: 2164-2167
    CrossrefPubMedGoogle Scholar
  • 2 Jemal A, Bray F, Center MM, Ferlay J, Ward E, Forman D. Global cancer statistics. CA Cancer J Clin 2011; 61: 69-90
    CrossrefPubMedGoogle Scholar
  • 3 Kapiteijn E, Schneider TC, Morreau H, Gelderblom H, Nortier JWR, Smit JW. New treatment modalities in advanced thyroid cancer. Ann Oncol 2012; 23: 10-18
    CrossrefPubMedGoogle Scholar
  • 4 Sherman SI. Thyroid carcinoma. The Lancet 2003; 361: 501-511
    CrossrefPubMedGoogle Scholar
  • 5 Baudin E, Schlumberger M. New therapeutic approaches for metastatic thyroid carcinoma. Lancet Oncol 2007; 8: 148-156
    CrossrefPubMedGoogle Scholar
  • 6 Kondo T, Ezzat S, Asa SL. Pathogenetic mechanisms in thyroid follicular-cell neoplasia. Nat Rev Cancer 2006; 6: 292-306
    CrossrefPubMedGoogle Scholar
  • 7 Görges R. The changing epidemiology of thyroid cancer. In: Biersack HJ, Grünwald F. (eds.) Thyroid Cancer. 2nd Edition Berlin: Springer; 2005: 3-27
    CrossrefGoogle Scholar
  • 8 Xing M. Molecular pathogenesis and mechanisms of thyroid cancer. Nat Rev Cancer 2013; 13: 184-199
    CrossrefPubMedGoogle Scholar
  • 9 Pitt SC, Moley J. Medullary, anaplastic, and metastatic cancers of the thyroid. Semin Oncol 2010; 37: 567-579
    CrossrefPubMedGoogle Scholar
  • 10 Filetti S, Bidart JM, Arturi F, Caillou B, Russo D, Schlumberger M. Sodium/iodide symporter: a key transport system in thyroid cancer cell metabolism. Eur J Endocrinol 1999; 141: 443-457
    CrossrefPubMedGoogle Scholar
  • 11 Schlumberger M, Sherman SI. Approach to the patient with advanced differentiated thyroid cancer. Eur J Endocrinol 2012; 166: 5-11
    CrossrefPubMedGoogle Scholar
  • 12 Rodriguez JM, Parrilla P, Moreno A, Sola J, Pinero A, Ortiz S, Soria T. Insular carcinoma: an infrequent subtype of thyroid cancer. J Am Coll Surg 1998; 187: 503-508
    CrossrefPubMedGoogle Scholar
  • 13 Kebebew E, Greenspan FS, Clark OH, Woeber KA, McMillan A. Anaplastic thyroid carcinoma. Treatment outcome and prognostic factors. Cancer 2005; 103: 1330-1335
    CrossrefPubMedGoogle Scholar
  • 14 Hiltzik D, Carlson DL, Tuttle RM, Chuai S, Ishill N, Shaha A, Shah JP, Singh B, Ghossein RA. Poorly differentiated thyroid carcinomas defined on the basis of mitosis and necrosis: a clinicopathologic study of 58 patients. Cancer 2006; 106: 1286-1295
    CrossrefPubMedGoogle Scholar
  • 15 Sanders Jr EM, LiVolsi VA, Brierley J, Shin J, Randolph GW. An evidence-based review of poorly differentiated thyroid cancer. World J Surg 2007; 31: 934-945
    CrossrefPubMedGoogle Scholar
  • 16 Volante M, Rapa I, Papotti M. Poorly differentiated thyroid carcinoma: diagnostic features and controversial issues. Endocr Pathol 2008; 19: 150-155
    CrossrefPubMedGoogle Scholar
  • 17 Smallridge RC, Marlow LA, Copland JA. Anaplastic thyroid cancer: molecular pathogenesis and emerging therapies. Endocr Relat Cancer 2009; 16: 17-44
    PubMedGoogle Scholar
  • 18 Dumont JE, Lamy F, Roger P, Maenhaut C. Physiological and pathological regulation of thyroid cell proliferation and differentiation by thyrotropin and other factors. Physiol Rev 1992; 72: 667-697
    CrossrefPubMedGoogle Scholar
  • 19 Dremier S, Coulonval K, Perpete S, Vandeput N, Fortemaison N, Van Keymeulen A, Deleu S, Ledent C, Clément S, Schurmans S, Dumont JE, Lamy F, Roger PP, Maenhaut C. The role of cyclic AMP and its effect on protein kinase A in the mitogenic action of thyrotropin on the thyroid cell. Ann N Y Acad Sci 2002; 968: 106-121
    CrossrefPubMedGoogle Scholar
  • 20 Robbins RJ, Schlumberger MJ. The evolving role of (131)I for the treatment of differentiated thyroid carcinoma. J Nucl Med 2005; 46: 28S-37S
    PubMedGoogle Scholar
  • 21 American Thyroid Association Surgery Working Group; American Association of Endocrine Surgeons; American Academy of Otolaryngology-Head and Neck Surgery; American Head and Neck Society . Carty SE, Cooper DS, Doherty GM, Haugen BR, Kloos RT, Lee SL, Mandel SJ, Mazzaferri EL, McIver B, Pacini F, Schlumberger M, Sherman SI, Steward DL, Tuttle RM. Revised American Thyroid Association management guidelines for patients with thyroid nodules and differentiated cancer. Thyroid 2009; 19: 1167-1214
    CrossrefPubMedGoogle Scholar
  • 22 De Rooij J, Zwartkruis FJ, Verheijen MH, Cool RH, Nijman SM, Wittinghofer A, Bos JL. Epac is a Rap1 guanine-nucleotide-exchange factor directly activated by cyclic AMP. Nature 1998; 396: 474-477
    CrossrefPubMedGoogle Scholar
  • 23 Kawasaki H, Springett GM, Mochizuki N, Toki S, Nakaya M, Matsuda M, Housman DE, Graybiel AM. A family of cAMP-binding proteins that directly activate Rap1. Science 1998; 282: 2275-2279
    CrossrefPubMedGoogle Scholar
  • 24 Gloerich M, Bos JL. Epac: defining a new mechanism for cAMP action. Annu Rev Pharmacol Toxicol 2010; 50: 355-375
    CrossrefPubMedGoogle Scholar
  • 25 Breckler M, Berthouze M, Laurent AC, Crozatier B, Morel E, Lezoualc’h F. Rap-linked cAMP signaling Epac proteins: compartmentation, functioning and disease implications. Cell Signal 2011; 23: 1257-1266
    CrossrefPubMedGoogle Scholar
  • 26 Dremier S, Milenkovic M, Blancquaert S, Dumont JE, Døskeland SO, Maenhaut C, Roger PP. Cyclic adenosine 3′,5′-monophosphate (cAMP)-dependent protein kinases, but not exchange proteins directly activated by cAMP (Epac), mediate thyrotropin/cAMP-dependent regulation of thyroid cells. Endocrinology 2007; 148: 4612-4622
    CrossrefPubMedGoogle Scholar
  • 27 Grandoch M, Roscioni SS, Schmidt M. The role of Epac proteins, novel cAMP mediators, in the regulation of immune, lung and neuronal function. Br J Pharmacol 2010; 159: 265-284
    CrossrefPubMedGoogle Scholar
  • 28 Fujita T, Meguro T, Fukuyama R, Nakamuta H, Koida M. New signaling pathway for parathyroid hormone and cyclic AMP action on extracellular-regulated kinase and cell proliferation in bone cells. Checkpoint of modulation by cyclic AMP. J Biol Chem 2002; 277: 22191-22200
    CrossrefPubMedGoogle Scholar
  • 29 Misra UK, Pizzo SV. Coordinate regulation of forskolin-induced cellular proliferation in macrophages by protein kinase A/cAMP-response element-binding protein (CREB) and Epac1-Rap1 signaling: effects of silencing CREB gene expression on Akt activation. J Biol Chem 2005; 280: 38276-38289
    CrossrefPubMedGoogle Scholar
  • 30 Kupperman E, Wen W, Meinkoth JL. Inhibition of thyrotropin-stimulated DNA synthesis by microinjection of inhibitors of cellular Ras and cyclic AMP-dependent protein kinase. Mol Cell Biol 1993; 13: 4477-4484
    PubMedGoogle Scholar
  • 31 Dremier S, Pohl V, Poteet-Smith C, Roger PP, Corbin J, Doskeland SO, Dumont JE, Maenhaut C. Activation of cyclic AMP-dependent kinase is required but may not be sufficient to mimic cyclic AMP-dependent DNA synthesis and thyroglobulin expression in dog thyroid cells. Mol Cell Biol 1997; 17: 6717-6726
    CrossrefPubMedGoogle Scholar
  • 32 Cass LA, Summers SA, Prendergast GV, Backer JM, Birnbaum MJ, Meinkoth JL. Protein kinase A-dependent and -independent signaling pathways contribute to cyclic AMP-stimulated proliferation. Mol Cell Biol 1999; 19: 5882-5891
    CrossrefPubMedGoogle Scholar
  • 33 Tsygankova OM, Kupperman E, Wen W, Meinkoth JL. Cyclic AMP activates Ras. Oncogene 2000; 19: 3609-3615
    CrossrefPubMedGoogle Scholar
  • 34 Cass LA, Meinkoth JL. Ras signaling through PI3K confers hormone-independent proliferation that is compatible with differentiation. Oncogene 2000; 19: 924-932
    CrossrefPubMedGoogle Scholar
  • 35 Gonzalez-Robayna IJ, Falender AE, Ochsner S, Firestone GL, Richards JS. Follicle-Stimulating hormone (FSH) stimulates phosphorylation and activation of protein kinase B (PKB/Akt) and serum and glucocorticoid-Induced kinase (Sgk): evidence for A kinase-independent signaling by FSH in granulosa cells. Mol Endocrinol 2000; 14: 1283-1300
    CrossrefPubMedGoogle Scholar
  • 36 Buscà R, Abbe P, Mantoux F, Aberdam E, Peyssonnaux F, Eychène A, Ortonne JP, Ballotti R. Ras mediates the cAMP-dependent activation of extracellular signal-regulated kinases (ERKs) in melanocytes. EMBO J 2000; 19: 2900-2910
    CrossrefPubMedGoogle Scholar
  • 37 Hochbaum D, Hong K, Barila G, Ribeiro-Neto F, Altschuler DL. Epac in synergy with cAMP-dependent protein kinase (PKA), is required for cAMP-mediated mitogenesis. J Biol Chem 2008; 283: 4464-4468
    CrossrefPubMedGoogle Scholar
  • 38 Remmele W, Hildebrand U, Hienz HA, Klein PJ, Vierbuchen M, Behnken LJ, Heicke B, Scheidt E. Comparative histological, histochemical, immunohistochemical and biochemical studies on oestrogen receptors, lectin receptors, and Barr bodies in human breast cancer. Virchows Arch A Pathol Anat Histopathol 1986; 409: 127-147
    CrossrefPubMedGoogle Scholar
  • 39 Ambesi-Impiombato FS, Parks LA, Coon HG. Culture of hormone-dependent functional epithelial cells from rat thyroids. Proc Natl Acad Sci USA 1980; 77: 3455-3459
    CrossrefPubMedGoogle Scholar
  • 40 Mark J, Ekedahl C, Dahlenfors R, Westermark B. Cytogenetical observations in five human anaplastic thyroid carcinomas. Hereditas 1987; 107: 163-174
    PubMedGoogle Scholar
  • 41 Carlsson J, Nilsson K, Westermark B, Pontén J, Sundström C, Larsson E, Bergh J, Påhlman S, Busch C, Collins VP. Formation and growth of multicellular spheroids of human origin. Int J Cancer 1983; 31: 523-533
    CrossrefPubMedGoogle Scholar
  • 42 Heldin NE, Cvejić D, Smeds S, Westermark B. Coexpression of functionally active receptors for thyrotropin and platelet-derived growth factor in human thyroid carcinoma cells. Endocrinology 1991; 129: 2187-2193
    CrossrefPubMedGoogle Scholar
  • 43 Dahlman T, Lammerts E, Wik M, Bergström D, Grimelius L, Westermark K, Rubin K, Heldin NE. Fibrosis in undifferentiated (anaplastic) thyroid carcinomas: evidence for a dual action of tumour cells in collagen type I synthesis. J Pathol 2000; 191: 376-386
    CrossrefPubMedGoogle Scholar
  • 44 Heldin NE, Gustavsson B, Claesson-Welsh L, Hammacher A, Mark J, Heldin CH, Westermark B. Aberrant expression of receptors for platelet-derived growth factor in an anaplastic thyroid carcinoma cell line. Proc Natl Acad Sci USA 1988; 85: 9302-9306
    CrossrefPubMedGoogle Scholar
  • 45 Fabien N, Fusco A, Santoro M, Barbier Y, Dubois PM, Paulin C. Description of a human papillary thyroid carcinoma cell line. Morphologic study and expression of tumoral markers. Cancer 1994; 15: 2206-2212
    PubMedGoogle Scholar
  • 46 Schönberger J, Bauer J, Spruss T, Weber G, Chahoud I, Eilles C, Grimm D. Establishment and characterization of the follicular thyroid carcinoma cell line ML-1. J Mol Med (Berl) 2000; 78: 102-110
    CrossrefPubMedGoogle Scholar
  • 47 Goretzki PE, Frilling A, Simon D, Rastegar M, Ohmann C. Growth regulation of human thyrocytes by thyrotropin, cyclic adenosine monophosphate, epidermal growth factor and insulin-like growth factor. Front Horm Res Basel, Karger 1989; 18: 56-80
    PubMedGoogle Scholar
  • 48 Enserink JM, Christensen AE, de Rooij J, van Triest M, Schwede F, Genieser HG, Døskeland SO, Blank JL, Bos JL. A novel Epac-specific cAMP analogue demonstrates independent regulation of Rap1 and ERK. Nat Cell Biol 2002; 4: 901-906
    CrossrefPubMedGoogle Scholar
  • 49 Christensen AE, Selheim F, de Rooij J, Dremier S, Schwede F, Dao KK, Martinez A, Maenhaut C, Bos JL, Genieser HG, Døskeland SO. cAMP analog mapping of Epac1 and cAMP kinase. Discriminating analogs demonstrate that Epac and cAMP kinase act synergistically to promote PC-12 cell neurite extension. J Biol Chem 2003; 278: 35394-35402
    CrossrefPubMedGoogle Scholar
  • 50 Holz GG, Chepurny OG, Schwede F. Epac-selective cAMP analogs: new tools with which to evaluate the signal transduction properties of cAMP-regulated guanine nucleotide exchange factors. Cell Signal 2008; 20: 10-20
    CrossrefPubMedGoogle Scholar
  • 51 Van Triest M, de Rooij J, Bos JL. Measurement of GTP-bound Ras-like GTPases by activation-specific probes. Methods Enzymol 2001; 333: 343-348
    PubMedGoogle Scholar
  • 52 Sands WA, Palmer TM. Regulating gene transcription in response to cyclic AMP elevation. Cell Signal 2008; 20: 460-466
    CrossrefPubMedGoogle Scholar
  • 53 Kimura T, Van Keymeulen A, Golstein J, Fusco A, Dumont JE, Roger PP. Regulation of thyroid cell proliferation by TSH and other factors: a critical evaluation of in vitro models. Endocr Rev 2001; 22: 631-656
    CrossrefPubMedGoogle Scholar
  • 54 Parma J, Duprez L, Van Sande J, Cochaux P, Gervy C, Mockel J, Dumont J, Vassart G. Somatic mutations in the thyrotropin receptor gene cause hyperfunctioning thyroid adenomas. Nature 1993; 365: 649-651
    Google Scholar
  • 55 Lyons J, Landis CA, Harsh G, Vallar L, Grünewald K, Feichtinger H, Duh QY, Clark OH, Kawasaki E, Bourne HR. Two G protein oncogenes in human endocrine tumors. Science 1990; 249: 655-659
    Google Scholar
  • 56 Ledent C, Dumont JE, Vassart G, Parmentier M. Thyroid expression of an A2 adenosine receptor transgene induces thyroid hyperplasia and hyperthyroidism. EMBO J 1992; 11: 537-542
    PubMedGoogle Scholar
  • 57 Michiels FM, Caillou B, Talbot M, Dessarps-Freichey F, Maunoury MT, Schlumberger M, Mercken L, Monier R, Feunteun J. Oncogenic potential of guanine nucleotide stimulatory factor alpha subunit in thyroid glands of transgenic mice. Proc Natl Acad Sci USA 1994; 91: 10488-10492
    CrossrefPubMedGoogle Scholar
  • 58 Zeiger MA, Saji M, Gusev Y, Westra WH, Takiyama Y, Dooley WC, Kohn LD, Levine MA. Thyroid-specific expression of cholera toxin A1 subunit causes thyroid hyperplasia and hyperthyroidism in transgenic mice. Endocrinology 1997; 138: 3133-3140
    CrossrefPubMedGoogle Scholar
  • 59 Franco AT, Malaguarnera R, Refetoff S, Liao XH, Lundsmith E, Kimura S, Pritchard C, Marais R, Davies TF, Weinstein LS, Chen M, Rosen N, Ghossein R, Knauf JA, Fagin JA. Thyrotropin receptor signaling dependence of Braf-induced thyroid tumor initiation in mice. Proc Natl Acad Sci USA 2011; 108: 1615-1620
    CrossrefPubMedGoogle Scholar
  • 60 Kaneshige M, Kaneshige K, Zhu X, Dace A, Garrett L, Carter TA, Kazlauskaite R, Pankratz DG, Wynshaw-Boris A, Refetoff S, Weintraub B, Willingham MC, Barlow C, Cheng S. Mice with a targeted mutation in the thyroid hormone beta receptor gene exhibit impaired growth and resistance to thyroid hormone. Proc Natl Acad Sci USA 2000; 97: 13209-13214
    CrossrefPubMedGoogle Scholar
  • 61 Lu C, Zhao L, Ying H, Willingham MC, Cheng SY. Growth activation alone is not sufficient to cause metastatic thyroid cancer in a mouse model of follicular thyroid carcinoma. Endocrinology 2010; 151: 1929-1939
    CrossrefPubMedGoogle Scholar
  • 62 Fang Y, Olah ME. Cyclic AMP-dependent, protein kinase A-independent activation of extracellular signal-regulated kinase 1/2 following adenosine receptor stimulation in human umbilical vein endothelial cells: role of exchange protein activated by cAMP 1 (Epac1). J Pharmacol Exp Ther 2007; 322: 1189-1200
    CrossrefPubMedGoogle Scholar
  • 63 Misra UK, Pizzo SV. Epac1-induced cellular proliferation in prostate cancer cells is mediated by B-Raf/ERK and mTOR signaling cascades. J Cell Biochem 2009; 108: 998-1011
    CrossrefPubMedGoogle Scholar
  • 64 Van Staveren WC, Beeckman S, Tomás G, Dom G, Hébrant A, Delys L, Vliem MJ, Trésallet C, Andry G, Franc B, Libert F, Dumont JE, Maenhaut C. Role of Epac and protein kinase A thyrotropin-induced gene expression in primary thyrocytes. Exp Cell Res 2012; 318: 444-452
    CrossrefPubMedGoogle Scholar
  • 65 Wright PA, Lemoine NR, Goretzki PE, Wyllie FS, Bond J, Hughes C, Röher HD, Williams ED, Wynford-Thomas D. Mutation of the p53 gene in a differentiated human thyroid carcinoma cell line, but not in primary thyroid tumours. Oncogene 1991; 6: 1693-1697
    PubMedGoogle Scholar
  • 66 Adeniran AJ, Zhu Z, Gandhi M, Steward DL, Fidler JP, Giordano TJ, Biddinger PW, Nikiforov YE. Correlation between genetic alterations and microscopic features, clinical manifestations, and prognostic characteristics of thyroid papillary carcinomas. Am J Surg Pathol 2006; 30: 216-222
    CrossrefPubMedGoogle Scholar
  • 67 Xing M. BRAF mutation in papillary thyroid cancer: pathogenic role, molecular bases, and clinical implications. Endocr Rev 2007; 28: 742-762
    CrossrefPubMedGoogle Scholar
  • 68 Ros P, Rossi DL, Acebrón P, Santisteban P. Thyroid-specific gene expression in the multi-step process of thyroid carcinogenesis. Biochimie 1999; 81: 389-396
    CrossrefPubMedGoogle Scholar