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Mucin 2 (MUC2) modulates the aggressiveness of breast cancer

  • Anna Astashchanka
  • Thomas M. Shroka
  • Britta M. Jacobsen
Preclinical study
  • 77 Downloads

Abstract

Purpose

Tumors that secrete large volumes of mucus are chemotherapy resistant, however, mechanisms underlying this resistance are unknown. One protein highly expressed in mucin secreting breast cancers is the secreted mucin, Mucin 2 (MUC2). While MUC2 is expressed in some breast cancers it is absent in normal breast tissue, implicating it in breast cancer. However, the effects of MUC2 on breast cancer are largely unknown. This study examined the role of MUC2 in modulating breast cancer proliferation, response to chemotherapy and metastasis.

Methods

Using patient derived xenografts we developed two novel cell lines, called BCK4 and PT12, which express high levels of MUC2. To modulate MUC2 levels, BCK4 and PT12 cells were engineered to express shRNA targeted to MUC2 (shMUC2, low MUC2) or a non-targeting control (shCONT, high MUC2) and proliferation and apoptosis were measured in vitro and in vivo. BCK4 cells with shCONT or shMUC2 were labeled with GFP-luciferase and examined in an experimental metastasis model; disease burden and site specific dissemination were monitored by intravital imaging and fluorescence guided dissection, respectively.

Results

Proliferation decreased in BCK4 and PT12 shMUC2 cells versus control cells both in vitro and in vivo. Chemotherapy induced minimal apoptosis in control cells expressing high MUC2 but increased apoptosis in shMUC2 cells containing low MUC2. An experimental metastasis model showed disease burden decreased when breast cancer cells contained low versus high MUC2. Treatment with Epidermal Growth Factor (EGF) increased MUC2 expression in BCK4 cells; this induction was abolished by the EGF-receptor inhibitor, Erlotinib.

Conclusions

MUC2 plays an important role in mediating proliferation, apoptosis and metastasis of breast cancer cells. MUC2 may be important in guiding treatment and predicting outcomes in breast cancer patients.

Keywords

Breast cancer Mucin 2 Metastasis Mucinous Estrogen receptor 

Notes

Acknowledgements

We thank Dr. Carol Sartorius for helpful comments on the manuscript. We also thank the following University of Colorado Cancer Center Core laboratories: Tissue Culture Core, Functional Genomics Facility, the University of Colorado Cancer Center (P30CA046984), and the University of Colorado Anschutz Medical Campus Biorepository Core Facility.

Funding

This study was supported by the University of Colorado Cancer Center Core Laboratories (P30CA046984). Funding provided by the University of Colorado Department of Pathology and the Breast Cancer Research Foundation. Studies also supported (in part) by a research grant from the Cancer League of Colorado, Inc (to BMJ).

Compliance with ethical standards

Conflict of interest

Anna Astashchanka declares that she has no conflict of interest. Thomas M. Shroka declares that he has no conflict of interest. Britta M. Jacobsen declares that she has no conflict of interest.

Ethical approval

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. This article does not contain any studies with human participants performed by any of the authors.

Supplementary material

10549_2018_4989_MOESM1_ESM.pdf (1.9 mb)
Supplementary material 1 (PDF 1916 KB)
10549_2018_4989_MOESM2_ESM.doc (52 kb)
Supplementary material 2 (DOC 51 KB)

References

  1. 1.
    Siegel RL, Miller KD, Jemal A (2016) Cancer statistics, 2016. CA Cancer J Clin 66(1):7–30.  https://doi.org/10.3322/caac.21332 CrossRefGoogle Scholar
  2. 2.
    Kufe DW (2009) Mucins in cancer: function, prognosis and therapy. Nat Rev Cancer 9(12):874–885.  https://doi.org/10.1038/nrc2761 CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Mukhopadhyay P, Chakraborty S, Ponnusamy MP, Lakshmanan I, Jain M, Batra SK (2011) Mucins in the pathogenesis of breast cancer: implications in diagnosis, prognosis and therapy. Biochim Biophys Acta 1815(2):224–240.  https://doi.org/10.1016/j.bbcan.2011.01.001 CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Yang C, Murray JL, Ibrahim NK (2018) MUC1 and cancer immunotherapy, vol 1. Elsevier Inc., Amsterdam, pp 225–240Google Scholar
  5. 5.
    Macha MA, Krishn SR, Jahan R, Banerjee K, Batra SK, Jain M (2015) Emerging potential of natural products for targeting mucins for therapy against inflammation and cancer. Cancer Treat Rev 41(3):277–288.  https://doi.org/10.1016/j.ctrv.2015.01.001 CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Diaz LK, Wiley EL, Morrow M (2001) Expression of epithelial mucins Muc1, Muc2, and Muc3 in ductal carcinoma in situ of the breast. Breast J 7(1):40–45CrossRefGoogle Scholar
  7. 7.
    Do SI, Kim K, Kim DH, Chae SW, Park YL, Park CH, Sohn JH (2013) Associations between the expression of mucins (MUC1, MUC2, MUC5AC, and MUC6) and clinicopathologic parameters of human breast ductal carcinomas. J Breast Cancer 16(2):152–158.  https://doi.org/10.4048/jbc.2013.16.2.152 CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Matsukita S, Nomoto M, Kitajima S, Tanaka S, Goto M, Irimura T, Kim YS, Sato E, Yonezawa S (2003) Expression of mucins (MUC1, MUC2, MUC5AC and MUC6) in mucinous carcinoma of the breast: comparison with invasive ductal carcinoma. Histopathology 42(1):26–36CrossRefGoogle Scholar
  9. 9.
    Rakha EA, Boyce RW, Abd El-Rehim D, Kurien T, Green AR, Paish EC, Robertson JF, Ellis IO (2005) Expression of mucins (MUC1, MUC2, MUC3, MUC4, MUC5AC and MUC6) and their prognostic significance in human breast cancer. Mod Pathol 18(10):1295–1304.  https://doi.org/10.1038/modpathol.3800445 CrossRefPubMedGoogle Scholar
  10. 10.
    Chu JS, Chang KJ (1999) Mucin expression in mucinous carcinoma and other invasive carcinomas of the breast. Cancer Lett 142(1):121–127. doi:S0304-3835(99)00161-5 [pii]CrossRefGoogle Scholar
  11. 11.
    Walsh MD, McGuckin MA, Devine PL, Hohn BG, Wright RG (1993) Expression of MUC2 epithelial mucin in breast carcinoma. J Clin Pathol 46(10):922–925CrossRefGoogle Scholar
  12. 12.
    Xu Y, Kimura N, Yoshida R, Lin H, Yoshinaga K (2001) Immunohistochemical study of Muc1, Muc2 and human gastric mucin in breast carcinoma: relationship with prognostic factors. Oncol Rep 8(5):1177–1182PubMedGoogle Scholar
  13. 13.
    Adsay NV, Merati K, Nassar H, Shia J, Sarkar F, Pierson CR, Cheng JD, Visscher DW, Hruban RH, Klimstra DS (2003) Pathogenesis of colloid (pure mucinous) carcinoma of exocrine organs: Coupling of gel-forming mucin (MUC2) production with altered cell polarity and abnormal cell-stroma interaction may be the key factor in the morphogenesis and indolent behavior of colloid carcinoma in the breast and pancreas. Am J Surg Pathol 27(5):571–578CrossRefGoogle Scholar
  14. 14.
    Patel DS, Khandeparkar SGS, Joshi AR, Kulkarni MM, Dhande B, Lengare P, Phegade LA, Narkhede K (2017) Immunohistochemical study of MUC1, MUC2 and MUC5AC expression in primary breast carcinoma. J Clin Diagn Res 11(4):EC30–EC34.  https://doi.org/10.7860/JCDR/2017/26533.9707 CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Kasashima S, Kawashima A, Zen Y, Ozaki S, Kobayashi M, Tsujibata A, Minato H (2007) Expression of aberrant mucins in lobular carcinoma with histiocytoid feature of the breast. Virchows Arch 450(4):397–403.  https://doi.org/10.1007/s00428-007-0381-z CrossRefPubMedGoogle Scholar
  16. 16.
    Lesuffleur T, Porchet N, Aubert JP, Swallow D, Gum JR, Kim YS, Real FX, Zweibaum A (1993) Differential expression of the human mucin genes MUC1 to MUC5 in relation to growth and differentiation of different mucus-secreting HT-29 cell subpopulations. J Cell Sci 106(Pt 3):771–783PubMedGoogle Scholar
  17. 17.
    Nagao T, Kinoshita T, Hojo T, Tsuda H, Tamura K, Fujiwara Y (2012) The differences in the histological types of breast cancer and the response to neoadjuvant chemotherapy: the relationship between the outcome and the clinicopathological characteristics. Breast 21(3):289–295.  https://doi.org/10.1016/j.breast.2011.12.011 CrossRefPubMedGoogle Scholar
  18. 18.
    Munzone E, Giobbie-Hurder A, Gusterson BA, Mallon E, Viale G, Thurlimann B, Ejlertsen B, MacGrogan G, Bibeau F, Lelkaitis G, Price KN, Gelber RD, Coates AS, Goldhirsch A, Colleoni M, International Breast Cancer Study G, the BIGCG (2015) Outcomes of special histotypes of breast cancer after adjuvant endocrine therapy with letrozole or tamoxifen in the monotherapy cohort of the BIG 1–98 trial. Ann Oncol 26(12):2442–2449.  https://doi.org/10.1093/annonc/mdv391 CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Bomeisl PE, Thompson CL, Harris LN, Gilmore HL (2015) Comparison of oncotype DX recurrence score by histologic types of breast carcinoma. Arch Pathol Lab Med 139(12):1546–1549.  https://doi.org/10.5858/arpa.2014-0557-OA CrossRefPubMedGoogle Scholar
  20. 20.
    Siegelmann-Danieli N, Silverman B, Zick A, Beit-Or A, Katzir I, Porath A (2013) The impact of the Oncotype DX Recurrence Score on treatment decisions and clinical outcomes in patients with early breast cancer: the Maccabi Healthcare Services experience with a unified testing policy. Ecancermedicalscience 7:380.  https://doi.org/10.3332/ecancer.2013.380 CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Jambal P, Badtke MM, Harrell JC, Borges VF, Post MD, Sollender GE, Spillman MA, Horwitz KB, Jacobsen BM (2013) Estrogen switches pure mucinous breast cancer to invasive lobular carcinoma with mucinous features. Breast Cancer Res Treat 137(2):431–448.  https://doi.org/10.1007/s10549-012-2377-x CrossRefPubMedGoogle Scholar
  22. 22.
    Kabos P, Finlay-Schultz J, Li C, Kline E, Finlayson C, Wisell J, Manuel CA, Edgerton SM, Harrell JC, Elias A, Sartorius CA (2012) Patient-derived luminal breast cancer xenografts retain hormone receptor heterogeneity and help define unique estrogen-dependent gene signatures. Breast Cancer Res Treat 135(2):415–432.  https://doi.org/10.1007/s10549-012-2164-8 CrossRefPubMedGoogle Scholar
  23. 23.
    D’Amato NC, Gordon MA, Babbs B, Spoelstra NS, Carson Butterfield KT, Torkko KC, Phan VT, Barton VN, Rogers TJ, Sartorius CA, Elias A, Gertz J, Jacobsen BM, Richer JK (2016) Cooperative dynamics of AR and ER activity in breast cancer. Mol Cancer Res 14(11):1054–1067.  https://doi.org/10.1158/1541-7786.MCR-16-0167 CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Korch C, Spillman MA, Jackson TA, Jacobsen BM, Murphy SK, Lessey BA, Jordan VC, Bradford AP (2012) DNA profiling analysis of endometrial and ovarian cell lines reveals misidentification, redundancy and contamination. Gynecol Oncol 127(1):241–248.  https://doi.org/10.1016/j.ygyno.2012.06.017 CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    Harrell JC, Shroka TM, Jacobsen BM (2017) Estrogen induces c-Kit and an aggressive phenotype in a model of invasive lobular breast cancer. Oncogenesis 6(11):396.  https://doi.org/10.1038/s41389-017-0002-x CrossRefPubMedPubMedCentralGoogle Scholar
  26. 26.
    Badtke MM, Jambal P, Dye WW, Spillman MA, Post MD, Horwitz KB, Jacobsen BM (2012) Unliganded progesterone receptors attenuate taxane-induced breast cancer cell death by modulating the spindle assembly checkpoint. Breast Cancer Res Treat 131(1):75–87.  https://doi.org/10.1007/s10549-011-1399-0 CrossRefPubMedGoogle Scholar
  27. 27.
    Harrell JC, Dye WW, Allred DC, Jedlicka P, Spoelstra NS, Sartorius CA, Horwitz KB (2006) Estrogen receptor positive breast cancer metastasis: altered hormonal sensitivity and tumor aggressiveness in lymphatic vessels and lymph nodes. Cancer Res 66(18):9308–9315CrossRefGoogle Scholar
  28. 28.
    Goswami CP, Nakshatri H (2013) PROGgene: gene expression based survival analysis web application for multiple cancers. J Clin Bioinform 3(1):22.  https://doi.org/10.1186/2043-9113-3-22 CrossRefGoogle Scholar
  29. 29.
    Pawitan Y, Bjohle J, Amler L, Borg AL, Egyhazi S, Hall P, Han X, Holmberg L, Huang F, Klaar S, Liu ET, Miller L, Nordgren H, Ploner A, Sandelin K, Shaw PM, Smeds J, Skoog L, Wedren S, Bergh J (2005) Gene expression profiling spares early breast cancer patients from adjuvant therapy: derived and validated in two population-based cohorts. Breast Cancer Res 7(6):R953–R964.  https://doi.org/10.1186/bcr1325 CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Yousef EM, Furrer D, Laperriere DL, Tahir MR, Mader S, Diorio C, Gaboury LA (2017) MCM2: An alternative to Ki-67 for measuring breast cancer cell proliferation. Mod Pathol 30(5):682–697.  https://doi.org/10.1038/modpathol.2016.231 CrossRefPubMedGoogle Scholar
  31. 31.
    Aksoy N, Thornton DJ, Corfield A, Paraskeva C, Sheehan JK (1999) A study of the intracellular and secreted forms of the MUC2 mucin from the PC/AA intestinal cell line. Glycobiology 9(7):739–746CrossRefGoogle Scholar
  32. 32.
    Rose MC, Voynow JA (2006) Respiratory tract mucin genes and mucin glycoproteins in health and disease. Physiol Rev 86(1):245–278.  https://doi.org/10.1152/physrev.00010.2005 CrossRefPubMedGoogle Scholar
  33. 33.
    Wittel UA, Goel A, Varshney GC, Batra SK (2001) Mucin antibodies - new tools in diagnosis and therapy of cancer. Front Biosci 6:D1296–D1310CrossRefGoogle Scholar
  34. 34.
    Vokuda RS, Verma SK, Srinivas BH (2018) Tissue Microarray Technology-A Brief Review. Natl J Lab Med 7(1):PR01–PR04Google Scholar
  35. 35.
    Sonora C, Mazal D, Berois N, Buisine MP, Ubillos L, Varangot M, Barrios E, Carzoglio J, Aubert JP, Osinaga E (2006) Immunohistochemical analysis of MUC5B apomucin expression in breast cancer and non-malignant breast tissues. J Histochem Cytochem 54(3):289–299.  https://doi.org/10.1369/jhc.5A6763.2005 CrossRefPubMedGoogle Scholar
  36. 36.
    Ookawa K, Kudo T, Aizawa S, Saito H, Tsuchida S (2002) Transcriptional activation of the MUC2 gene by p53. J Biol Chem 277(50):48270–48275.  https://doi.org/10.1074/jbc.M207986200 CrossRefPubMedGoogle Scholar
  37. 37.
    Valque H, Gouyer V, Gottrand F, Desseyn JL (2012) MUC5B leads to aggressive behavior of breast cancer MCF7 cells. PLoS ONE 7(10):e46699.  https://doi.org/10.1371/journal.pone.0046699 CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Valque H, Gouyer V, Husson MO, Gottrand F, Desseyn JL (2011) Abnormal expression of Muc5b in Cftr-null mice and in mammary tumors of MMTV-ras mice. Histochem Cell Biol 136(6):699–708.  https://doi.org/10.1007/s00418-011-0872-5 CrossRefPubMedGoogle Scholar
  39. 39.
    Garcia EP, Tiscornia I, Libisch G, Trajtenberg F, Bollati-Fogolin M, Rodriguez E, Noya V, Chiale C, Brossard N, Robello C, Santinaque F, Folle G, Osinaga E, Freire T (2016) MUC5B silencing reduces chemo-resistance of MCF-7 breast tumor cells and impairs maturation of dendritic cells. Int J Oncol 48(5):2113–2123.  https://doi.org/10.3892/ijo.2016.3434 CrossRefPubMedGoogle Scholar
  40. 40.
    Tadesse S, Corner G, Dhima E, Houston M, Guha C, Augenlicht L, Velcich A (2017) MUC2 mucin deficiency alters inflammatory and metabolic pathways in the mouse intestinal mucosa. Oncotarget 8(42):71456–71470.  https://doi.org/10.18632/oncotarget.16886 CrossRefPubMedPubMedCentralGoogle Scholar
  41. 41.
    Shan YS, Hsu HP, Lai MD, Yen MC, Fang JH, Weng TY, Chen YL (2014) Suppression of mucin 2 promotes interleukin-6 secretion and tumor growth in an orthotopic immune-competent colon cancer animal model. Oncol Rep 32(6):2335–2342.  https://doi.org/10.3892/or.2014.3544 CrossRefPubMedPubMedCentralGoogle Scholar
  42. 42.
    He YF, Zhang MY, Wu X, Sun XJ, Xu T, He QZ, Di W (2013) High MUC2 expression in ovarian cancer is inversely associated with the M1/M2 ratio of tumor-associated macrophages and patient survival time. PLoS ONE 8(12):e79769.  https://doi.org/10.1371/journal.pone.0079769 CrossRefPubMedPubMedCentralGoogle Scholar
  43. 43.
    Yonezawa S, Goto M, Yamada N, Higashi M, Nomoto M (2008) Expression profiles of MUC1, MUC2, and MUC4 mucins in human neoplasms and their relationship with biological behavior. Proteomics 8(16):3329–3341.  https://doi.org/10.1002/pmic.200800040 CrossRefPubMedGoogle Scholar
  44. 44.
    Hollingsworth MA, Swanson BJ (2004) Mucins in cancer: protection and control of the cell surface. Nat Rev Cancer 4(1):45–60.  https://doi.org/10.1038/nrc1251 CrossRefPubMedGoogle Scholar
  45. 45.
    Scully OJ, Bay BH, Yip G, Yu Y (2012) Breast cancer metastasis. Cancer Genom Proteom 9(5):311–320Google Scholar
  46. 46.
    Hsu HP, Lai MD, Lee JC, Yen MC, Weng TY, Chen WC, Fang JH, Chen YL (2017) Mucin 2 silencing promotes colon cancer metastasis through interleukin-6 signaling. Sci Rep 7(1):5823.  https://doi.org/10.1038/s41598-017-04952-7 CrossRefPubMedPubMedCentralGoogle Scholar
  47. 47.
    Cardillo MR, Castagna G, Memeo L, De Bernardinis E, Di Silverio F (2000) Epidermal growth factor receptor, MUC-1 and MUC-2 in bladder cancer. J Exp Clin Cancer Res 19(2):225–233PubMedGoogle Scholar
  48. 48.
    Utsunomiya T, Yonezawa S, Sakamoto H, Kitamura H, Hokita S, Aiko T, Tanaka S, Irimura T, Kim YS, Sato E (1998) Expression of MUC1 and MUC2 mucins in gastric carcinomas: its relationship with the prognosis of the patients. Clin Cancer Res 4(11):2605–2614PubMedGoogle Scholar
  49. 49.
    Jonckheere N, Skrypek N, Van Seuningen I (2014) Mucins and tumor resistance to chemotherapeutic drugs. Biochim Biophys Acta 1846(1):142–151.  https://doi.org/10.1016/j.bbcan.2014.04.008 CrossRefPubMedGoogle Scholar
  50. 50.
    Leteurtre E, Gouyer V, Rousseau K, Moreau O, Barbat A, Swallow D, Huet G, Lesuffleur T (2004) Differential mucin expression in colon carcinoma HT-29 clones with variable resistance to 5-fluorouracil and methotrexate. Biol Cell 96(2):145–151.  https://doi.org/10.1016/j.biolcel.2003.12.005 CrossRefPubMedGoogle Scholar

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Authors and Affiliations

  1. 1.Department of PathologyUniversity of Colorado Anschutz Medical CampusAuroraUSA
  2. 2.Division of Endocrinology, Metabolism and Diabetes, Department of MedicineUniversity of Colorado Anschutz Medical CampusAuroraUSA

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