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Stromal cells in phyllodes tumors of the breast are enriched for EZH2 and stem cell marker expression

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Abstract

Phyllodes tumors (PTs) of the breast are fibroepithelial neoplasms with stromal hypercellularity, which is the basis for their classification as benign, borderline, and malignant. The histologic diagnosis of PTs is often difficult, and the pathological features may not always predict clinical behavior. The pathobiology of PT remains poorly understood. Enhancer of Zeste 2 (EZH2) epigenetically regulates cell-type identity, cellular differentiation, and breast cancer stem cells. EZH2 exerts oncogenic functions in breast cancer and is associated with metastasis. We hypothesized that in PTs, EZH2 and the stem cell marker ALDH1 may be expressed in stromal cells and may be associated with their degree of differentiation. Forty PTs were histologically characterized at our institution following the World Health Organization criteria. We investigated the expression of EZH2 and ALDH1 by immunohistochemistry and recorded as percentage of positive epithelial and stromal cells. EZH2 was positive when over 10 % of cells exhibited nuclear staining; ALDH1 was positive when over 5 % of cells had cytoplasmic staining. Of the 40 PTs, 24 (60 %) were histologically benign, 8 (20 %) borderline, and 8 (20 %) malignant. Stromal EZH2 was significantly associated with the diagnosis of malignant PT, as it was detected in 1 of 24 (4 %) benign, 3 of 8 (37.5 %) borderline, and 5 of 8 (62.5 %) malignant tumors. Stromal EZH2 was significantly associated with stromal overgrowth (p = 0.01), atypia (p = 0.01), hypercellularity (p = 0.01), and mitoses (p = 0.02), all features of malignant PT. Stromal EZH2 and ALDH1 were significantly associated with grade of PT (p = 0.01 and p < 0.05 respectively). In conclusion, EZH2 and ALDH1 expression in the stroma of PT may mark malignant progression and may be helpful to distinguish histologically benign from borderline and malignant tumors in challenging cases. Our study also suggests that PTs contain mesenchymal stem cells, shedding light into the pathogenesis of these tumors.

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References

  1. Rosen PP, Oberman HA (1993) Tumors of the mammary gland, vol 7., Atlas of tumor pathology Third seriesArmed Forces Institute of Pathology, Washington

    Google Scholar 

  2. Lakhani SR, Ellis. IO, Schnitt, SJ, Tan, PH, van de Vijver, M.J. eds (2012) World Health Organization Classification of Tumours of the Breast. IARC Lyon

  3. Sawyer EJ, Hanby AM, Ellis P, Lakhani SR, Ellis IO, Boyle S, Tomlinson IP (2000) Molecular analysis of phyllodes tumors reveals distinct changes in the epithelial and stromal components. Am J Pathol 156(3):1093–1098

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Sawyer EJ, Hanby AM, Rowan AJ, Gillett CE, Thomas RE, Poulsom R, Lakhani SR, Ellis IO, Ellis P, Tomlinson IP (2002) The Wnt pathway, epithelial-stromal interactions, and malignant progression in phyllodes tumours. J Pathol 196(4):437–444. doi:10.1002/path.1067

    Article  CAS  PubMed  Google Scholar 

  5. Kwon JE, Jung WH, Koo JS (2012) Molecules involved in epithelial-mesenchymal transition and epithelial-stromal interaction in phyllodes tumors: implications for histologic grade and prognosis. Tumour Biol 33(3):787–798. doi:10.1007/s13277-011-0296-9

    Article  PubMed  Google Scholar 

  6. Laible G, Wolf A, Dorn R, Reuter G, Nislow C, Lebersorger A, Popkin D, Pillus L, Jenuwein T (1997) Mammalian homologues of the Polycomb-group gene Enhancer of zeste mediate gene silencing in Drosophila heterochromatin and at S. cerevisiae telomeres. EMBO J 16(11):3219–3232

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Ringrose L, Paro R (2004) Epigenetic regulation of cellular memory by the Polycomb and Trithorax group proteins. Annu Rev Genet 38:413–443

    Article  CAS  PubMed  Google Scholar 

  8. Boyer LA, Plath K, Zeitlinger J, Brambrink T, Medeiros LA, Lee TI, Levine SS, Wernig M, Tajonar A, Ray MK, Bell GW, Otte AP, Vidal M, Gifford DK, Young RA, Jaenisch R (2006) Polycomb complexes repress developmental regulators in murine embryonic stem cells. Nature 441(7091):349–353

    Article  CAS  PubMed  Google Scholar 

  9. Alford SH, Toy K, Merajver SD, Kleer CG (2012) Increased risk for distant metastasis in patients with familial early-stage breast cancer and high EZH2 expression. Breast Cancer Res Treat 132(2):429–437. doi:10.1007/s10549-011-1591-2

    Article  CAS  PubMed  Google Scholar 

  10. Gonzalez ME, Moore HM, Li X, Toy KA, Huang W, Sabel MS, Kidwell KM, Kleer CG (2014) EZH2 expands breast stem cells through activation of NOTCH1 signaling. Proc Natl Acad Sci USA 111(8):3098–3103. doi:10.1073/pnas.1308953111

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Kleer CG, Cao Q, Varambally S, Shen R, Ota I, Tomlins SA, Ghosh D, Sewalt RG, Otte AP, Hayes DF, Sabel MS, Livant D, Weiss SJ, Rubin MA, Chinnaiyan AM (2003) EZH2 is a marker of aggressive breast cancer and promotes neoplastic transformation of breast epithelial cells. Proc Natl Acad Sci USA 100(20):11606–11611. doi:10.1073/pnas.1933744100

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Kunju LP, Cookingham C, Toy KA, Chen W, Sabel MS, Kleer CG (2011) EZH2 and ALDH-1 mark breast epithelium at risk for breast cancer development. Mod Pathol 24(6):786–793. doi:10.1038/modpathol.2011.8

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Chang CJ, Yang JY, Xia W, Chen CT, Xie X, Chao CH, Woodward WA, Hsu JM, Hortobagyi GN, Hung MC (2011) EZH2 promotes expansion of breast tumor initiating cells through activation of RAF1-beta-catenin signaling. Cancer Cell 19(1):86–100. doi:10.1016/j.ccr.2010.10.035

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Dontu G, Abdallah WM, Foley JM, Jackson KW, Clarke MF, Kawamura MJ, Wicha MS (2003) In vitro propagation and transcriptional profiling of human mammary stem/progenitor cells. Genes Dev 17(10):1253–1270

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Ginestier C, Hur MH, Charafe-Jauffret E, Monville F, Dutcher J, Brown M, Jacquemier J, Viens P, Kleer CG, Liu S, Schott A, Hayes D, Birnbaum D, Wicha MS, Dontu G (2007) ALDH1 Is a marker of normal and malignant human mammary stem cells and a predictor of poor clinical outcome. Cell Stem Cell 1(5):555–567

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Lin JJ, Huang CS, Yu J, Liao GS, Lien HC, Hung JT, Lin RJ, Chou FP, Yeh KT, Yu AL (2014) Malignant phyllodes tumors display mesenchymal stem cell features and aldehyde dehydrogenase/disialoganglioside identify their tumor stem cells. Breast Cancer Res 16(2):R29. doi:10.1186/bcr3631

    Article  PubMed  PubMed Central  Google Scholar 

  17. Bracken AP, Pasini D, Capra M, Prosperini E, Colli E, Helin K (2003) EZH2 is downstream of the pRB-E2F pathway, essential for proliferation and amplified in cancer. EMBO J 22(20):5323–5335

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Kamminga LM, Bystrykh LV, de Boer A, Houwer S, Douma J, Weersing E, Dontje B, de Haan G (2006) The Polycomb group gene Ezh2 prevents hematopoietic stem cell exhaustion. Blood 107(5):2170–2179

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Varambally S, Dhanasekaran SM, Zhou M, Barrette TR, Kumar-Sinha C, Sanda MG, Ghosh D, Pienta KJ, Sewalt RG, Otte AP, Rubin MA, Chinnaiyan AM (2002) The polycomb group protein EZH2 is involved in progression of prostate cancer. Nature 419(6907):624–629

    Article  CAS  PubMed  Google Scholar 

  20. Ciarapica R, Miele L, Giordano A, Locatelli F, Rota R (2011) Enhancer of zeste homolog 2 (EZH2) in pediatric soft tissue sarcomas: first implications. BMC Med 9:63. doi:10.1186/1741-7015-9-63

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Ciarapica R, Russo G, Verginelli F, Raimondi L, Donfrancesco A, Rota R, Giordano A (2009) Deregulated expression of miR-26a and Ezh2 in rhabdomyosarcoma. Cell Cycle 8(1):172–175

    Article  CAS  PubMed  Google Scholar 

  22. Richter GH, Plehm S, Fasan A, Rossler S, Unland R, Bennani-Baiti IM, Hotfilder M, Lowel D, von Luettichau I, Mossbrugger I, Quintanilla-Martinez L, Kovar H, Staege MS, Muller-Tidow C, Burdach S (2009) EZH2 is a mediator of EWS/FLI1 driven tumor growth and metastasis blocking endothelial and neuro-ectodermal differentiation. Proc Natl Acad Sci USA 106(13):5324–5329. doi:10.1073/pnas.0810759106

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Charytonowicz E, Cordon-Cardo C, Matushansky I, Ziman M (2009) Alveolar rhabdomyosarcoma: is the cell of origin a mesenchymal stem cell? Cancer Lett 279(2):126–136. doi:10.1016/j.canlet.2008.09.039

    Article  CAS  PubMed  Google Scholar 

  24. Juan AH, Kumar RM, Marx JG, Young RA, Sartorelli V (2009) Mir-214-dependent regulation of the polycomb protein Ezh2 in skeletal muscle and embryonic stem cells. Mol Cell 36(1):61–74. doi:10.1016/j.molcel.2009.08.008

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Lenhard MS, Kahlert S, Himsl I, Ditsch N, Untch M, Bauerfeind I (2008) Phyllodes tumour of the breast: clinical follow-up of 33 cases of this rare disease. Eur J Obstet Gynecol Reprod Biol 138(2):217–221

    Article  PubMed  Google Scholar 

  26. Karim RZ, Gerega SK, Yang YH, Spillane A, Carmalt H, Scolyer RA, Lee CS (2009) Phyllodes tumours of the breast: a clinicopathological analysis of 65 cases from a single institution. Breast 18(3):165–170. doi:10.1016/j.breast.2009.03.001

    Article  CAS  PubMed  Google Scholar 

  27. Tan PH, Thike AA, Tan WJ, Thu MM, Busmanis I, Li H, Chay WY, Tan MH (2012) Predicting clinical behaviour of breast phyllodes tumours: a nomogram based on histological criteria and surgical margins. J Clin Pathol 65(1):69–76. doi:10.1136/jclinpath-2011-200368

    Article  PubMed  Google Scholar 

  28. Belkacemi Y, Bousquet G, Marsiglia H, Ray-Coquard I, Magne N, Malard Y, Lacroix M, Gutierrez C, Senkus E, Christie D, Drumea K, Lagneau E, Kadish SP, Scandolaro L, Azria D, Ozsahin M (2008) Phyllodes tumor of the breast. Int J Radiat Oncol Biol Phys 70(2):492–500

    Article  PubMed  Google Scholar 

  29. Barth RJ Jr (1999) Histologic features predict local recurrence after breast conserving therapy of phyllodes tumors. Breast Cancer Res Treat 57(3):291–295

    Article  PubMed  Google Scholar 

  30. Pezner RD, Schultheiss TE, Paz IB (2008) Malignant phyllodes tumor of the breast: local control rates with surgery alone. Int J Radiat Oncol Biol Phys 71(3):710–713. doi:10.1016/j.ijrobp.2007.10.051

    Article  PubMed  Google Scholar 

  31. Grimes MM (1992) Cystosarcoma phyllodes of the breast: histologic features, flow cytometric analysis, and clinical correlations. Mod Pathol 5(3):232–239

    CAS  PubMed  Google Scholar 

  32. Noronha Y, Raza A, Hutchins B, Chase D, Garberoglio C, Chu P, Weiss L, Wang J (2011) CD34, CD117, and Ki-67 expression in phyllodes tumor of the breast: an immunohistochemical study of 33 cases. Int J Surg Pathol 19(2):152–158. doi:10.1177/1066896910382009

    Article  PubMed  Google Scholar 

  33. Tan PH, Jayabaskar T, Yip G, Tan Y, Hilmy M, Selvarajan S, Bay BH (2005) p53 and c-kit (CD117) protein expression as prognostic indicators in breast phyllodes tumors: a tissue microarray study. Mod Pathol 18(12):1527–1534

    CAS  PubMed  Google Scholar 

  34. Tse GM, Lui PC, Vong JS, Lau KM, Putti TC, Karim R, Scolyer RA, Lee CS, Yu AM, Ng DC, Tse AK, Tan PH (2009) Increased epidermal growth factor receptor (EGFR) expression in malignant mammary phyllodes tumors. Breast Cancer Res Treat 114(3):441–448

    Article  CAS  PubMed  Google Scholar 

  35. Karim RZ, Gerega SK, Yang YH, Spillane A, Carmalt H, Scolyer RA, Lee CS (2010) p16 and pRb immunohistochemical expression increases with increasing tumour grade in mammary phyllodes tumours. Histopathology 56(7):868–875. doi:10.1111/j.1365-2559.2010.03562.x

    Article  PubMed  Google Scholar 

  36. Kleer CG, Giordano TJ, Braun T, Oberman HA (2001) Pathologic, immunohistochemical, and molecular features of benign and malignant phyllodes tumors of the breast. Mod Pathol 14(3):185–190

    Article  CAS  PubMed  Google Scholar 

  37. Tan WJ, Thike AA, Bay BH, Tan PH (2014) Immunohistochemical expression of homeoproteins Six1 and Pax3 in breast phyllodes tumours correlates with histological grade and clinical outcome. Histopathology 64(6):807–817. doi:10.1111/his.12329

    Article  PubMed  Google Scholar 

  38. Khosravi-Shahi P (2011) Management of non metastatic phyllodes tumors of the breast: review of the literature. Surg Oncol 20(4):e143–e148. doi:10.1016/j.suronc.2011.04.007S0960-7404(11)00038-7

    Article  PubMed  Google Scholar 

  39. Chen WH, Cheng SP, Tzen CY, Yang TL, Jeng KS, Liu CL, Liu TP (2005) Surgical treatment of phyllodes tumors of the breast: retrospective review of 172 cases. J Surg Oncol 91(3):185–194. doi:10.1002/jso.20334

    Article  PubMed  Google Scholar 

  40. Mitus J, Reinfuss M, Mitus JW, Jakubowicz J, Blecharz P, Wysocki WM, Skotnicki P (2014) Malignant phyllodes tumor of the breast: treatment and prognosis. Breast J 20(6):639–644. doi:10.1111/tbj.12333

    Article  PubMed  Google Scholar 

  41. August DA, Kearney T (2000) Cystosarcoma phyllodes: mastectomy, lumpectomy, or lumpectomy plus irradiation. Surg Oncol 9(2):49–52

    Article  CAS  PubMed  Google Scholar 

  42. Kondo Y (2014) Targeting histone methyltransferase EZH2 as cancer treatment. J Biochem 156(5):249–257. doi:10.1093/jb/mvu054

    Article  PubMed  Google Scholar 

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Funding

This work was supported by NIH Grants R01 CA107469 and CA125577, and the National Institutes of Health through the University of Michigan’s Cancer Center Support Grant (5 P30 CA46592).

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Authors and Affiliations

  1. Department of Pathology, University of California Davis, Sacramento, CA, USA

    Yanhong Zhang

  2. Departments of Radiation Oncology, University of Michigan, Ann Arbor, MI, USA

    Adam L. Liss, Eugene Chung & Lori J. Pierce

  3. Department of Pathology, University of Michigan, 4217 Comprehensive Cancer Center, 1500 E. Medical Center Drive, Ann Arbor, MI, 48109, USA

    Celina G. Kleer

  4. Comprehensive Cancer Center, University of Michigan, Ann Arbor, MI, USA

    Lori J. Pierce & Celina G. Kleer

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  1. Yanhong Zhang
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  2. Adam L. Liss
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  5. Celina G. Kleer
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Correspondence to Celina G. Kleer.

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Yanhong Zhang and Adam L. Liss are co-first authors.

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Zhang, Y., Liss, A.L., Chung, E. et al. Stromal cells in phyllodes tumors of the breast are enriched for EZH2 and stem cell marker expression. Breast Cancer Res Treat 158, 21–28 (2016). https://doi.org/10.1007/s10549-016-3853-5

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