Breast Cancer Research and Treatment

, Volume 117, Issue 2, pp 443–451

Inhibition of histone deacetylase suppresses EGF signaling pathways by destabilizing EGFR mRNA in ER-negative human breast cancer cells

Brief Report


Estrogen receptor alpha (ER)-negative human breast cancer cells frequently overexpress epidermal growth factor receptor (EGFR) and respond poorly to endocrine therapies. Our previous studies demonstrate that histone deacetylation plays a key role in ER gene silencing, and ER expression can be restored with histone deacetylase (HDAC) inhibitors in ER-negative human breast cancer cells. Whether inhibition of HDAC also alters epidermal growth factor (EGF) signaling pathways is not defined. Here we present evidence that reexpression of ER protein by a clinically available HDAC inhibitor, suberoylanilide hydroxamic acid (SAHA or vorinostat), is coupled with loss of EGFR in ER-negative human breast cancer cells. Consistent with this observation, MDA-MB-231 cells, which are ER-negative and overexpress EGFR, that are engineered to express ER show a decrease in EGFR protein expression. Down-regulation of EGFR by SAHA results from attenuation of its mRNA stability. We also confirm that new protein synthesis is required for maintaining EGFR mRNA stability. Further experiments indicate that a decrease in EGFR abolished EGF-initiated signaling pathways including phosphorylated PAK1, p38MAPK and AKT. Thus, SAHA may not only reactivate silenced ER, but also simultaneously deplete EGFR expression. These data suggest that inhibition of HDAC is a promising epigenetic therapy for ER-negative human breast cancer.


HDAC inhibitor SAHA ER EGFR 


  1. 1.
    Nicholson RI, McClelland RA, Gee JM, Manning DL, Cannon P, Robertson JF et al (1994) Epidermal growth factor receptor expression in breast cancer: association with response to endocrine therapy. Breast Cancer Res Treat 29:117–125. doi:10.1007/BF00666187 PubMedCrossRefGoogle Scholar
  2. 2.
    Nicholson S, Wright C, Sainsbury JR, Halcrow P, Kelly P, Angus B et al (1990) Epidermal growth factor receptor (EGFr) as a marker for poor prognosis in node-negative breast cancer patients: neu and tamoxifen failure. J Steroid Biochem Mol Biol 37:811–814. doi:10.1016/0960-0760(90)90424-J PubMedCrossRefGoogle Scholar
  3. 3.
    Knowlden JM, Hutcheson IR, Jones HE, Madden T, Gee JM, Harper ME et al (2003) Elevated levels of epidermal growth factor receptor/c-erbB2 heterodimers mediate an autocrine growth regulatory pathway in tamoxifen-resistant MCF-7 cells. Endocrinology 144:1032–1044. doi:10.1210/en.2002-220620 PubMedCrossRefGoogle Scholar
  4. 4.
    Jordan NJ, Gee JM, Barrow D, Wakeling AE, Nicholson RI (2004) Increased constitutive activity of PKB/Akt in tamoxifen resistant breast cancer MCF-7 cells. Breast Cancer Res Treat 87:167–180. doi:10.1023/B:BREA.0000041623.21338.47 PubMedCrossRefGoogle Scholar
  5. 5.
    Citri A, Yarden Y (2006) EGF-ERBB signaling: towards the systems level. Nat Rev Mol Cell Biol 7:505–516. doi:10.1038/nrm1962 PubMedCrossRefGoogle Scholar
  6. 6.
    Davis RJ (2000) Signal transduction by the JNK group of MAP kinases. Cell 103:239–252. doi:10.1016/S0092-8674(00)00116-1 PubMedCrossRefGoogle Scholar
  7. 7.
    Garcia R, Franklin RA, McCubrey JA (2006) Cell death of MCF-7 human breast cancer cells induced by EGFR activation in the absence of other growth factors. Cell Cycle 5:1840–1846PubMedGoogle Scholar
  8. 8.
    Kumar R, Gururaj AE, Barnes CJ (2006) p21-activated kinases in cancer. Nat Rev Cancer 6:459–471. doi:10.1038/nrc1892 PubMedCrossRefGoogle Scholar
  9. 9.
    Chrysogelos SA, Yarden RI, Lauber AH, Murphy JM (1994) Mechanisms of EGF receptor regulation in breast cancer cells. Breast Cancer Res Treat 31:227–236. doi:10.1007/BF00666156 PubMedCrossRefGoogle Scholar
  10. 10.
    Fan P, Wang J, Santen RJ, Yue W (2007) Long-term treatment with tamoxifen facilitates translocation of estrogen receptor alpha out of the nucleus and enhances its interaction with EGFR in MCF-7 breast cancer cells. Cancer Res 67:1352–1360. doi:10.1158/0008-5472.CAN-06-1020 PubMedCrossRefGoogle Scholar
  11. 11.
    Davidson NE, Gelmann EP, Lippman ME, Dickson RB (1987) Epidermal growth factor receptor gene expression in estrogen receptor-positive and negative human breast cancer cell lines. Mol Endocrinol 1:216–223PubMedCrossRefGoogle Scholar
  12. 12.
    Chrysogelos SA (1993) Chromatin structure of the EGFR gene suggests a role for intron 1 sequences in its regulation in breast cancer cells. Nucleic Acids Res 21:5736–5741. doi:10.1093/nar/21.24.5736 PubMedCrossRefGoogle Scholar
  13. 13.
    McInerney JM, Wilson MA, Strand KJ, Chrysogelos SA (2001) A strong intronic enhancer element of the EGFR gene is preferentially active in high EGFR expressing breast cancer cells. J Cell Biochem 80:538–549. doi :10.1002/1097-4644(20010315)80:4<538::AID-JCB1008>3.0.CO;2-2PubMedCrossRefGoogle Scholar
  14. 14.
    Sheikh MS, Shao ZM, Chen JC, Li XS, Hussain A, Fontana JA (1994) Expression of estrogen receptors in estrogen receptor-negative human breast carcinoma cells: modulation of epidermal growth factor-receptor (EGF-R) and transforming growth factor alpha (TGF alpha) gene expression. J Cell Biochem 54:289–298. doi:10.1002/jcb.240540305 PubMedCrossRefGoogle Scholar
  15. 15.
    Wilson MA, Chrysogelos SA (2002) Identification and characterization of a negative regulatory element within the epidermal growth factor receptor gene first intron in hormone-dependent breast cancer cells. J Cell Biochem 85:601–614. doi:10.1002/jcb.10168 PubMedCrossRefGoogle Scholar
  16. 16.
    Boerner JL, Gibson MA, Fox EM, Posner ED, Parsons SJ, Silva CM et al (2005) Estrogen negatively regulates epidermal growth factor (EGF)-mediated signal transducer and activator of transcription 5 signaling in human EGF family receptor-overexpressing breast cancer cells. Mol Endocrinol 19:2660–2670. doi:10.1210/me.2004-0439 PubMedCrossRefGoogle Scholar
  17. 17.
    Levin ER (2002) Bidirectional signaling between the estrogen receptor and the epidermal growth factor receptor. Mol Endocrinol 17:309–317. doi:10.1210/me.2002-0368 PubMedCrossRefGoogle Scholar
  18. 18.
    Ottaviano YL, Issa JP, Parl FF, Smith HS, Baylin SB, Davidson NE (1994) Methylation of the estrogen receptor gene CpG island marks loss of estrogen receptor expression in human breast cancer cells. Cancer Res 54:2552–2555PubMedGoogle Scholar
  19. 19.
    Ferguson AT, Lapidus R, Baylin S, Davidson NE (1995) Demethylation of the estrogen receptor gene in estrogen receptor-negative breast cancer cells can reactivate estrogen receptor gene expression. Cancer Res 55:2279–2283PubMedGoogle Scholar
  20. 20.
    Yang X, Phillips DL, Ferguson AT, Nelson WG, Herman JG, Davidson NE (2001) Synergistic activation of functional estrogen receptor (ER)-alpha by DNA methyltransferase and histone deacetylase inhibition in human ER-alpha-negative breast cancer cells. Cancer Res 61:7025–7029PubMedGoogle Scholar
  21. 21.
    Sharma D, Blum J, Yang X, Beaulieu N, Macleod AR, Davidson NE (2005) Release of methyl CpG binding proteins and histone deacetylase 1 from the Estrogen receptor alpha (ER) promoter upon reactivation in ER-negative human breast cancer cells. Mol Endocrinol 19:1740–1751. doi:10.1210/me.2004-0011 PubMedCrossRefGoogle Scholar
  22. 22.
    Sharma D, Saxena NK, Davidson NE, Vertino PM (2006) Restoration of tamoxifen sensitivity in estrogen receptor-negative breast cancer cells: tamoxifen-bound reactivated ER recruits distinctive corepressor complexes. Cancer Res 66:6370–6378. doi:10.1158/0008-5472.CAN-06-0402 PubMedCrossRefGoogle Scholar
  23. 23.
    Zhou Q, Atadja P, Davidson NE (2007) Histone deacetylase inhibitor LBH589 reactivates silenced estrogen receptor alpha (ER) gene expression without loss of DNA hypermethylation. Cancer Biol Ther 6:64–69PubMedGoogle Scholar
  24. 24.
    Leu YW, Yan PS, Fan M, Jin VX, Liu JC, Curran EM et al (2004) Loss of estrogen receptor signaling triggers epigenetic silencing of downstream targets in breast cancer. Cancer Res 64:8184–8192. doi:10.1158/0008-5472.CAN-04-2045 PubMedCrossRefGoogle Scholar
  25. 25.
    Bolden JE, Peart MJ, Johnstone RW (2006) Anticancer activities of histone deacetylase inhibitors. Nat Rev Drug Discov 5:769–784. doi:10.1038/nrd2133 PubMedCrossRefGoogle Scholar
  26. 26.
    Keen JC, Zhou Q, Park BH, Pettit C, Mack KM, Blair B et al (2005) Protein phosphatase 2A regulates estrogen receptor alpha (ER) expression through modulation of ER mRNA stability. J Biol Chem 280:29519–29524. doi:10.1074/jbc.M505317200 PubMedCrossRefGoogle Scholar
  27. 27.
    Adam L, Vadlamudi R, Mandal M, Chernoff J, Kumar R (2000) Regulation of microfilament reorganization and invasiveness of breast cancer cells by kinase dead p21-activated kinase-1. J Biol Chem 275:12041–12050. doi:10.1074/jbc.275.16.12041 PubMedCrossRefGoogle Scholar
  28. 28.
    Vadlamudi RK, Adam L, Wang RA, Mandal M, Nguyen D, Sahin A et al (2000) Regulatable expression of p21-activated kinase-1 promotes anchorage-independent growth and abnormal organization of mitotic spindles in human epithelial breast cancer cells. J Biol Chem 275:36238–36244. doi:10.1074/jbc.M002138200 PubMedCrossRefGoogle Scholar
  29. 29.
    Rayala SK, Molli PR, Kumar R (2006) Nuclear p21-activated kinase 1 in breast cancer packs off tamoxifen sensitivity. Cancer Res 66:5985–5988. doi:10.1158/0008-5472.CAN-06-0978 PubMedCrossRefGoogle Scholar
  30. 30.
    Balasenthil S, Sahin AA, Barnes CJ, Wang RA, Pestell RG, Vadlamudi RK et al (2004) p21-Activated kinase-1 signaling mediates cyclin D1 expression in mammary epithelial and cancer cells. J Biol Chem 279:1422–1428. doi:10.1074/jbc.M309937200 PubMedCrossRefGoogle Scholar
  31. 31.
    Massarweh S, Osborne CK, Jiang S, Wakeling AE, Rimawi M, Mohsin SK et al (2006) Mechanisms of tumor regression and resistance to estrogen deprivation and fulvestrant in a model of estrogen receptor-positive, HER-2/neu-positive breast cancer. Cancer Res 66:8266–8273. doi:10.1158/0008-5472.CAN-05-4045 PubMedCrossRefGoogle Scholar
  32. 32.
    Rayala SK, Talukder AH, Balasenthil S, Tharakan R, Barnes CJ, Wang RA et al (2006) P21-Activated kinase 1 regulation of estrogen receptor-alpha activation involves serine 305 activation linked with serine 118 phosphorylation. Cancer Res 66:1694–1701. doi:10.1158/0008-5472.CAN-05-2922 PubMedCrossRefGoogle Scholar
  33. 33.
    Hirokawa Y, Arnold M, Nakajima H, Zalcberg J, Maruta H (2005) Signal therapy of breast cancers by the HDAC inhibitor FK228 that blocks the activation of PAK1 and abrogates the tamoxifen-resistance. Cancer Biol Ther 4:956–960. doi:10.1158/1535-7163.MCT-04-0321 PubMedCrossRefGoogle Scholar
  34. 34.
    Banerjee M, Worth D, Prowse DM, Nikolic M (2002) Pak1 phosphorylation on t212 affects microtubules in cells undergoing mitosis. Curr Biol 12:1233–1239. doi:10.1016/S0960-9822(02)00956-9 PubMedCrossRefGoogle Scholar
  35. 35.
    Zhao ZS, Lim JP, Ng YW, Lim L, Manser E (2005) The GIT-associated kinase PAK targets to the centrosome and regulates Aurora-A. Mol Cell 20:237–249. doi:10.1016/j.molcel.2005.08.035 PubMedCrossRefGoogle Scholar
  36. 36.
    Holm C, Rayala S, Jirstrom K, Stal O, Kumar R, Landberg G (2006) Association between Pak1 expression and subcellular localization and tamoxifen resistance in breast cancer patients. J Natl Cancer Inst 98:671–680PubMedCrossRefGoogle Scholar
  37. 37.
    Puto LA, Pestonjamasp K, King CC, Bokoch GM (2003) p21-Activated kinase 1 (PAK1) interacts with the Grb2 adapter protein to couple to growth factor signaling. J Biol Chem 278:9388–9393. doi:10.1074/jbc.M208414200 PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC. 2008

Authors and Affiliations

  • Qun Zhou
    • 1
  • Patrick G. Shaw
    • 1
  • Nancy E. Davidson
    • 1
  1. 1.The Sidney Kimmel Comprehensive Cancer CenterJohns Hopkins UniversityBaltimoreUSA

Personalised recommendations