Abstract
Purpose
Cushing’s disease is primarily caused by adrenocorticotropic hormone (ACTH)-producing pituitary adenomas. If excision of the tumor from the pituitary, which is the primary treatment for Cushing’s disease, is unsuccessful, further medical therapy is needed to treat the resultant hypercortisolism. Some of the drugs used to treat this condition have shown potential therapeutic benefits, but a more effective treatment should be explored for the treatment of Cushing’s disease. In the present study, we determined the effect of heat shock protein 90 inhibitors on ACTH production and cell proliferation of AtT-20 corticotroph tumor cells.
Methods
AtT-20 pituitary corticotroph tumor cells were cultured. The expression levels of mouse proopiomelanocortin (POMC) and pituitary tumor transforming gene 1 (PTTG1) mRNA were evaluated using quantitative real-time PCR. Cellular DNA content was analyzed with fluorescence-activated cell sorting (FACS) analysis. The protein levels were determined by Western blot analysis.
Results
Both 17-allylamino-17-demethoxygeldanamycin and CCT018159 decreased POMC mRNA levels in AtT-20 cells and ACTH levels in the culture medium of these cells, suggesting that both drugs suppress ACTH synthesis and secretion in corticotroph tumor cells. Both drugs also decreased cell proliferation and induced apoptosis. FACS analyses revealed that both agents increased the percentage of AtT-20 cells in the G2/M phase. These drugs decreased cell proliferation, presumably due to the induction of cell death and arrest of the cell cycle in AtT-20 cells. Tumor weight in mice xenografted with AtT-20 cells and treated with CCT018159 was lower than in AtT-20-xenografted control mice. CCT018159 also decreased plasma ACTH levels, and POMC and PTTG1 mRNA levels in the tumor cells.
Conclusions
CCT018159 inhibits ACTH production and corticotroph tumor cell proliferation in vitro and in vivo.
Similar content being viewed by others
References
Nieman LK, Biller BM, Findling JW, Newell-Price J, Savage MO, Stewart PM, Montori VM (2008) The diagnosis of Cushing’s syndrome: an endocrine society clinical practice guideline. J Clin Endocrinol Metab 93:1526–1540
Kageyama K, Oki T, Sakihara S, Nigawara T, Terui K, Suda T (2013) Evaluation of the diagnostic criteria for Cushing’s disease in Japan. Endocr J 6:127–135
Biller BM, Grossman AB, Stewart PM, Melmed S, Bertagna X, Bertherat J, Buchfelder M, Colao A, Hermus AR, Hofland LJ, Klibanski A, Lacroix A, Lindsay JR, Newell-Price J, Nieman LK, Petersenn S, Sonino N, Stalla GK, Swearingen B, Vance ML, Wass JA, Boscaro M (2008) Treatment of adrenocorticotropin-dependent Cushing’s syndrome; a consensus statement. J Clin Endocrinol Metab 93:2454–3462
Bertagna X, Guignat L (2013) Approach to the Cushing’s disease patient with persistent/recurrent hypercortisolism after pituitary surgery. J Clin Endocrinol Metab 98:1307–1318
Schteingart DE (2009) Drugs in the medical treatment of Cushing’s syndrome. Expert Opin Emerg Drugs 14:661–671
Feelders RA, de Bruin C, Pereira AM, Romijn JA, Netea-Maier RT, Hermus AR, Zelissen PM, van Heerebeek R, de Jong FH, van der Lely AJ, de Herder WW, Hofland LJ, Lamberts SW (2010) Pasireotide alone or with cabergoline and ketoconazole in Cushing’s disease. N Eng J Med 362:1846–1848
Hofland LJ (2008) Somatostatin and somatostatin receptors in Cushing’s disease. Mol Cell Endocrinol 286:199–205
de Bruin C, Feelders RA, Lamberts SW, Hofland LJ (2009) Somatostatin and dopamine receptors as targets for medical treatment of Cushing’s syndrome. Rev Endocr Metab Disord 10:91–102
Ferone D, Gatto F, Arvigo M, Resmini E, Boschetti M, Teti C, Esposito D, Minuto F (2009) The clinical-molecular interface of somatostatin, dopamine and their receptors in pituitary pathophysiology. J Mol Endocrinol 42:361–370
Páez-Pereda M, Kovalovsky D, Hopfner U, Theodoropoulou M, Pagotto U, Uhl E, Losa M, Stalla J, Grübler Y, Missale C, Arzt E, Stalla GK (2001) Retinoic acid prevents experimental Cushing syndrome. J Clin Invest 108:1123–1131
Bangaru ML, Woodliff J, Raff H, Kansra S (2010) Growth suppression of mouse pituitary corticotroph tumor AtT20 cells by curcumin: a model for treating Cushing’s disease. PLoS ONE 5:e9893
Pei L, Melmed S (1997) Isolation and characterization of a pituitary tumor-transforming gene (PTTG). Mol Endocrinol 11:433–441
Vlotides G, Eigler T, Melmed S (2007) Pituitary tumor-transforming gene: physiology and implications for tumorigenesis. Endocr Rev 28:165–186
Zhang X, Horwitz GA, Heaney AP, Nakashima M, Prezant TR, Bronstein MD, Melmed S (1999) Pituitary tumor transforming gene (PTTG) expression in pituitary adenomas. J Clin Endocrinol Metab 84:761–767
Chesnokova V, Zonis S, Zhou C, Ben-Shlomo A, Wawrowsky K, Toledano Y, Tong Y, Kovacs K, Scheithauer B, Melmed S (2011) Lineage-specific restraint of pituitary gonadotroph cell adenoma growth. PLoS ONE 6:e17924
Chesnokova V, Zonis S, Wawrowsky K, Tani Y, Ben-Shlomo A, Ljubimov V, Mamelak A, Bannykh S, Melmed S (2012) Clusterin and FOXL2 act concordantly to regulate pituitary gonadotroph adenoma growth. Mol Endocrinol 26:2092–2103
Hernández A, López-Lluch G, Bernal JA, Navas P, Pintor-Toro JA (2008) Dicoumarol down-regulates human PTTG1/Securin mRNA expression through inhibition of Hsp90. Mol Cancer Ther 7:474–482
Hernández A, López-Lluch G, Navas P, Pintor-Toro JA (2009) HDAC and Hsp90 inhibitors down-regulate PTTG1/securin but do not induce aneuploidy. Genes Chromosomes Cancer 48:194–201
Sharp SY, Boxall K, Rowlands M, Prodromou C, Roe SM, Maloney A, Powers M, Clarke PA, Box G, Sanderson S, Patterson L, Matthews TP, Cheung KM, Ball K, Hayes A, Raynaud F, Marais R, Pearl L, Eccles S, Aherne W, McDonald E, Workman P (2007) In vitro biological characterization of a novel, synthetic diaryl pyrazole resorcinol class of heat shock protein 90 inhibitors. Cancer Res 67:2206–2216
Smith NF, Hayes A, James K, Nutley BP, McDonald E, Dymock B, Drysdale MJ, Raynaud FI, Workman P (2006) Preclinical pharmacokinetics and metabolism of a novel diaryl pyrazole resorcinol series of heat shock protein 90 inhibitors. Mol Cancer Ther 5:1628–1637
Kageyama K, Hanada K, Suda T (2009) Differential regulation of urocortins1-3 mRNA in human umbilical vein endothelial cells. Regul Pept 155:131–138
Kageyama K, Hanada K, Suda T (2010) Differential regulation and roles of urocortins in human adrenal H295R cells. Regul Pept 162:18–25
Born EJ, Hartman SV, Holstein SA (2013) Targeting HSP90 and monoclonal protein trafficking modulates the unfolded protein response, chaperone regulation and apoptosis in myeloma cells. Blood Cancer J 3:e167
Ui T, Morishima K, Saito S, Sakuma Y, Fujii H, Hosoya Y, Ishikawa S, Aburatani H, Fukayama M, Niki T, Yasuda Y (2014) The HSP90 inhibitor 17-N-allylamino-17-demethoxy geldanamycin (17-AAG) synergizes with cisplatin and induces apoptosis in cisplatin-resistant esophageal squamous cell carcinoma cell lines via the Akt/XIAP pathway. Oncol Rep 31:619–624
Zagouri F, Sergentanis TN, Chrysikos D, Papadimitriou CA, Dimopoulos MA, Psaltopoulou T (2013) Hsp90 inhibitors in breast cancer: a systematic review. Breast 22:569–578
Solit DB, Zheng FF, Drobnjak M, Münster PN, Higgins B, Verbel D, Heller G, Tong W, Cordon-Cardo C, Agus DB, Scher HI, Rosen N (2002) 17-Allylamino-17-demethoxygeldanamycin induces the degradation of androgen receptor and HER-2/neu and inhibits the growth of prostate cancer xenografts. Clin Cancer Res 8:986–993
Senju M, Sueoka N, Sato A, Iwanaga K, Sakao Y, Tominaga M, Irie K, Hayashi S, Sueoka E (2006) Hsp90 inhibitors cause G2/M arrest associated with the reduction of Cdc25C and Cdc2 in lung cancer cell lines. J Cancer Res Clin Oncol 132:150–158
Kageyama K, Hanada K, Moriyama T, Imaizumi T, Satoh K, Suda T (2007) Differential regulation of CREB and ERK phosphorylation through corticotropin-releasing factor receptors type 1 and 2 in AtT-20 and A7r5 cells. Mol Cell Endocrinol 263:90–102
Kageyama K, Hanada K, Moriyama T, Nigawara T, Sakihara S, Suda T (2006) G Protein-coupled receptor kinase 2 involvement in desensitization of corticotropin-releasing factor (CRF) receptor type 1 by CRF in murine corticotrophs. Endocrinology 147:441–450
Tsukamoto N, Otsuka F, Miyoshi T, Yamanaka R, Inagaki K, Yamashita M, Otani H, Takeda M, Suzuki J, Ogura T, Iwasaki Y, Makino H (2010) Effects of bone morphogenetic protein (BMP) on adrenocorticotropin production by pituitary corticotrope cells: involvement of up-regulation of BMP receptor signaling by somatostatin analogs. Endocrinology 151:1129–1141
Theodoropoulou M, Zhang J, Laupheimer S, Paez-Pereda M, Erneux C, Florio T, Pagotto U, Stalla GK (2006) Octreotide, a somatostatin analogue, mediates its antiproliferative action in pituitary tumor cells by altering phosphatidylinositol 3-kinase signaling and inducing Zac1 expression. Cancer Res 66:1576–1582
Solit DB, Basso AD, Olshen AB, Scher HI, Rosen N (2003) Inhibition of heat shock protein 90 function down-regulates Akt kinase and sensitizes tumors to Taxol. Cancer Res 63:2139–2144
Muşat M, Vax VV, Borboli N, Gueorguiev M, Bonner S, Korbonits M, Grossman AB (2004) Cell cycle dysregulation in pituitary oncogenesis. Front Horm Res 32:34–62
Lu C, Willingham MC, Furuya F, Cheng SY (2008) Activation of phosphatidylinositol 3-kinase signaling promotes aberrant pituitary growth in a mouse model of thyroid-stimulating hormone-secreting pituitary tumors. Endocrinology 149:3339–3345
Cerovac V, Monteserin-Garcia J, Rubinfeld H, Buchfelder M, Losa M, Florio T, Paez-Pereda M, Stalla GK, Theodoropoulou M (2010) The somatostatin analogue octreotide confers sensitivity to rapamycin treatment on pituitary tumor cells. Cancer Res 70:666–674
Taguchi T, Takao T, Iwasaki Y, Nishiyama M, Asaba K, Hashimoto K (2006) Suppressive effects of dehydroepiandrosterone and the nuclear factor-kappaB inhibitor parthenolide on corticotroph tumor cell growth and function in vitro and in vivo. J Endocrinol 188:321–331
Fukuoka H, Cooper O, Ben-Shlomo A, Mamelak A, Ren SG, Bruyette D, Melmed S (2011) EGFR as a therapeutic target for human, canine, and mouse ACTH-secreting pituitary adenomas. J Clin Invest 121:4712–4721
Kageyama K, Oki Y, Nigawara T, Suda T, Daimon M (2014) Pathophysiology and treatment of subclinical Cushing’s disease and pituitary silent corticotroph adenomas. Endocr J 61:941–948
Acknowledgments
We thank the Department of Social Medicine, Hirosaki University Graduate School of Medicine, for generously providing us with FACS analysis. This research did not receive any specific grant from any funding agency in the public, commercial or not-for-profit sector.
Conflict of interest
None of the authors have any potential conflicts of interest associated with this research.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Sugiyama, A., Kageyama, K., Murasawa, S. et al. Inhibition of heat shock protein 90 decreases ACTH production and cell proliferation in AtT-20 cells. Pituitary 18, 542–553 (2015). https://doi.org/10.1007/s11102-014-0607-4
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11102-014-0607-4