Journal of Endocrinological Investigation

, Volume 41, Issue 6, pp 655–661 | Cite as

Expression of p27Kip1 and p18Ink4c in human multiple endocrine neoplasia type 1-related pancreatic neuroendocrine tumors

  • E. B. Conemans
  • G. M. Raicu-Ionita
  • C. R. C. Pieterman
  • K. M. A. Dreijerink
  • O. M. Dekkers
  • A. R. Hermus
  • W. W. de Herder
  • M. L. Drent
  • A. N. A. van der Horst-Schrivers
  • B. Havekes
  • P. H. Bisschop
  • G. J. Offerhaus
  • I. H. M. Borel Rinkes
  • G. D. Valk
  • H. Th. M. Timmers
  • M. R. VriensEmail author
Original Article



Pancreatic neuroendocrine tumors are a major manifestation of multiple endocrine neoplasia type 1 (MEN1). This tumor syndrome is caused by germline mutations in MEN1, encoding menin. Insight into pathogenesis of these tumors might lead to new biomarkers and therapeutic targets for these patients. Several lines of evidence point towards a role for p27Kip1 and p18Ink4c in MEN1-related tumor development in animal models for MEN1, but their contribution to human MEN1-related pancreatic neuroendocrine tumor development is not known.


In this study, we characterized protein expression of p27Kip1 and p18Ink4c in human MEN1-related PanNETs by immunohistochemistry. From the nationwide DutchMEN1 Study Group database including > 90% of the Dutch MEN1 population, MEN1-patients, who underwent pancreatic surgery, were selected. A tissue micro-array was constructed with available paraffin tissue blocks, and PanNETs from 61 MEN1 patients were eligible for analysis.


Expression of p27Kip1 was high in 57 (93%) PanNETs and 67% of the tumors showed low expression of p18Ink4c (67.3%). No association was found between expression of either p27Kip1 or p18Ink4c and clinic-pathological characteristics.


These findings indicate that loss of p18Ink4c, but not p27Kip1, is a common event in the development of MEN1-related PanNETs. Restoration of p18Ink4c function through CDK4/6 inhibitors could be a therapeutic option for MEN1-related PanNETs.


Multiple endocrine neoplasia type 1 Pancreatic neuroendocrine tumors Menin p27Kip1 p18Ink4c 



We thank Annette H. Bruggink for her assistance in the tissue collection. We thank Domenico Castigliego and Lutske Lodewijk for their help with the construction of the TMA. We thank Folkert H. Morsink, Roel Broekhuizen and Radhika A. Varier for their advice on the optimization of procedures for immunohistochemistry.

Complicance with ethical standards


Gerlof D. Valk and Menno R. Vriens are the receivers of an unrestricted grant from Ipsen. Koen M. A. Dreijerink is supported by a fellowship from the Dutch Cancer Society (UU 2012 5370).

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

All procedures performed in this study were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.

Informed consent

For this type of study formal consent is not required.

Supplementary material

40618_2017_783_MOESM1_ESM.docx (22 kb)
Supplementary material 1 (DOCX 22 kb)


  1. 1.
    Thakker RV, Newey PJ, Walls GV, Bilezikian J, Dralle H, Ebeling PR et al (2012) Clinical practice guidelines for multiple endocrine neoplasia type 1 (MEN1). J Clin Endocrinol Metab 97(9):2990–3011CrossRefPubMedGoogle Scholar
  2. 2.
    Pieterman CR, Conemans EB, Dreijerink KM, de Laat JM, Timmers HT, Vriens MR et al (2014) Thoracic and duodenopancreatic neuroendocrine tumors in multiple endocrine neoplasia type 1: natural history and function of menin in tumorigenesis. Endocr Relat Cancer 21(3):R121–R142CrossRefPubMedGoogle Scholar
  3. 3.
    Knudson AG Jr (1971) Mutation and cancer: statistical study of retinoblastoma. Proc Natl Acad Sci USA 68(4):820–823CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Bartsch DK, Langer P, Wild A, Schilling T, Celik I, Rothmund M et al (2000) Pancreaticoduodenal endocrine tumors in multiple endocrine neoplasia type 1: surgery or surveillance? Surgery 128(6):958–966CrossRefPubMedGoogle Scholar
  5. 5.
    Hasani-Ranjbar S, Amoli MM, Ebrahim-Habibi A, Gozashti MH, Khalili N, Sayyahpour FA et al (2011) A new frameshift MEN1 gene mutation associated with familial malignant insulinomas. Fam Cancer 10(2):343–348CrossRefPubMedGoogle Scholar
  6. 6.
    Kouvaraki MA, Lee JE, Shapiro SE, Gagel RF, Sherman SI, Sellin RV et al (2002) Genotype–phenotype analysis in multiple endocrine neoplasia type 1. Arch Surg (Chicago, Ill : 1960) 137(6):641–647CrossRefGoogle Scholar
  7. 7.
    Raef H, Zou M, Baitei EY, Al-Rijjal RA, Kaya N, Al-Hamed M et al (2011) A novel deletion of the MEN1 gene in a large family of multiple endocrine neoplasia type 1 (MEN1) with aggressive phenotype. Clin Endocrinol 75(6):791–800CrossRefGoogle Scholar
  8. 8.
    Schaaf L, Pickel J, Zinner K, Hering U, Hofler M, Goretzki PE et al (2007) Developing effective screening strategies in multiple endocrine neoplasia type 1 (MEN 1) on the basis of clinical and sequencing data of German patients with MEN 1. Exp Clin Endocrinol Diabetes 115(8):509–517CrossRefPubMedGoogle Scholar
  9. 9.
    Jiao Y, Shi C, Edil BH, de Wilde RF, Klimstra DS, Maitra A et al (2011) DAXX/ATRX, MEN1, and mTOR pathway genes are frequently altered in pancreatic neuroendocrine tumors. Science (New York) 331(6021):1199–1203CrossRefGoogle Scholar
  10. 10.
    Dreijerink KM, Hoppener JW, Timmers HM, Lips CJ (2006) Mechanisms of disease: multiple endocrine neoplasia type 1-relation to chromatin modifications and transcription regulation. Nat Clin Pract Endocrinol Metab 2(10):562–570CrossRefPubMedGoogle Scholar
  11. 11.
    Hughes CM, Rozenblatt-Rosen O, Milne TA, Copeland TD, Levine SS, Lee JC et al (2004) Menin associates with a trithorax family histone methyltransferase complex and with the hoxc8 locus. Mol Cell 13(4):587–597CrossRefPubMedGoogle Scholar
  12. 12.
    Yokoyama A, Wang Z, Wysocka J, Sanyal M, Aufiero DJ, Kitabayashi I et al (2004) Leukemia proto-oncoprotein MLL forms a SET1-like histone methyltransferase complex with menin to regulate Hox gene expression. Mol Cell Biol 24(13):5639–5649CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    van Nuland R, Smits AH, Pallaki P, Jansen PW, Vermeulen M, Timmers HT (2013) Quantitative dissection and stoichiometry determination of the human SET1/MLL histone methyltransferase complexes. Mol Cell Biol 33(10):2067–2077CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Shilatifard A (2012) The COMPASS family of histone H3K4 methylases: mechanisms of regulation in development and disease pathogenesis. Annu Rev Biochem 81:65–95CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Milne TA, Hughes CM, Lloyd R, Yang Z, Rozenblatt-Rosen O, Dou Y et al (2005) Menin and MLL cooperatively regulate expression of cyclin-dependent kinase inhibitors. Proc Natl Acad Sci USA 102(3):749–754CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Chu IM, Hengst L, Slingerland JM (2008) The Cdk inhibitor p27 in human cancer: prognostic potential and relevance to anticancer therapy. Nat Rev Cancer 8(4):253–267CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Fontaniere S, Casse H, Bertolino P, Zhang CX (2006) Analysis of p27(Kip1) expression in insulinomas developed in pancreatic beta-cell specific MEN1 mutant mice. Fam Cancer 5(1):49–54CrossRefPubMedGoogle Scholar
  18. 18.
    Franklin DS, Godfrey VL, O’Brien DA, Deng C, Xiong Y (2000) Functional collaboration between different cyclin-dependent kinase inhibitors suppresses tumor growth with distinct tissue specificity. Mol Cell Biol 20(16):6147–6158CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Karnik SK, Hughes CM, Gu X, Rozenblatt-Rosen O, McLean GW, Xiong Y et al (2005) Menin regulates pancreatic islet growth by promoting histone methylation and expression of genes encoding p27Kip1 and p18INK4c. Proc Natl Acad Sci USA 102(41):14659–14664CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Agarwal SK, Mateo CM, Marx SJ (2009) Rare germline mutations in cyclin-dependent kinase inhibitor genes in multiple endocrine neoplasia type 1 and related states. J Clin Endocrinol Metab 94(5):1826–1834CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Occhi G, Regazzo D, Trivellin G, Boaretto F, Ciato D, Bobisse S et al (2013) A novel mutation in the upstream open reading frame of the CDKN1B gene causes a MEN4 phenotype. PLoS Genet 9(3):e1003350CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Circelli L, Ramundo V, Marotta V, Sciammarella C, Marciello F, Del Prete M et al (2015) Prognostic role of the CDNK1B V109G polymorphism in multiple endocrine neoplasia type 1. J Cell Mol Med 19(7):1735–1741CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    de Laat JM, Pieterman CR, Weijmans M, Hermus AR, Dekkers OM, de Herder WW et al (2013) Low accuracy of tumor markers for diagnosing pancreatic neuroendocrine tumors in multiple endocrine neoplasia type 1 patients. J Clin Endocrinol Metab 98(10):4143–4151CrossRefPubMedGoogle Scholar
  24. 24.
    Casparie M, Tiebosch AT, Burger G, Blauwgeers H, van de Pol A, van Krieken JH et al (2007) Pathology databanking and biobanking in The Netherlands, a central role for PALGA, the nationwide histopathology and cytopathology data network and archive. Cell Oncol 29(1):19–24PubMedPubMedCentralGoogle Scholar
  25. 25.
    Dreijerink KM, Varier RA, van Nuland R, Broekhuizen R, Valk GD, van der Wal JE et al (2009) Regulation of vitamin D receptor function in MEN1-related parathyroid adenomas. Mol Cell Endocrinol 313(1–2):1–8CrossRefPubMedGoogle Scholar
  26. 26.
    Horree N, Gort EH, van der Groep P, Heintz AP, Vooijs M, van Diest PJ (2008) Hypoxia-inducible factor 1 alpha is essential for hypoxic p27 induction in endometrioid endometrial carcinoma. J Pathol 214(1):38–45CrossRefPubMedGoogle Scholar
  27. 27.
    Lee HS, Chen M, Kim JH, Kim WH, Ahn S, Maeng K et al (2014) Analysis of 320 gastroenteropancreatic neuroendocrine tumors identifies TS expression as independent biomarker for survival. Int J Cancer 135(1):128–137CrossRefPubMedGoogle Scholar
  28. 28.
    Klimstra DS, Modlin IR, Coppola D, Lloyd RV, Suster S (2010) The pathologic classification of neuroendocrine tumors: a review of nomenclature, grading, and staging systems. Pancreas 39(6):707–712CrossRefPubMedGoogle Scholar
  29. 29.
    Bosman FTCF, Hruban RH, Theisse ND (2010) WHO classification of tumours of the digestive system. International Agency for Research on Cancer (IARC), LyonGoogle Scholar
  30. 30.
    Grabowski P, Schrader J, Wagner J, Horsch D, Arnold R, Arnold CN et al (2008) Loss of nuclear p27 expression and its prognostic role in relation to cyclin E and p53 mutation in gastroenteropancreatic neuroendocrine tumors. Clin Cancer Res 14(22):7378–7384CrossRefPubMedGoogle Scholar
  31. 31.
    Korshunov A, Shishkina L, Golanov A (2003) Immunohistochemical analysis of p16INK4a, p14ARF, p18INK4c, p21CIP1, p27KIP1 and p73 expression in 271 meningiomas correlation with tumor grade and clinical outcome. Int J Cancer 104(6):728–734CrossRefPubMedGoogle Scholar
  32. 32.
    Bai F, Pei XH, Nishikawa T, Smith MD, Xiong Y (2007) p18Ink4c, but not p27Kip1, collaborates with Men1 to suppress neuroendocrine organ tumors. Mol Cell Biol 27(4):1495–1504CrossRefPubMedGoogle Scholar
  33. 33.
    Lindberg D, Akerstrom G, Westin G (2008) Evaluation of CDKN2C/p18, CDKN1B/p27 and CDKN2B/p15 mRNA expression, and CpG methylation status in sporadic and MEN1-associated pancreatic endocrine tumours. Clin Endocrinol 68(2):271–277Google Scholar
  34. 34.
    Lapenna S, Giordano A (2009) Cell cycle kinases as therapeutic targets for cancer. Nat Rev Drug Discovery 8(7):547–566CrossRefPubMedGoogle Scholar
  35. 35.
    Lemos MC, Thakker RV (2008) Multiple endocrine neoplasia type 1 (MEN1): analysis of 1336 mutations reported in the first decade following identification of the gene. Hum Mutat 29(1):22–32CrossRefPubMedGoogle Scholar

Copyright information

© Italian Society of Endocrinology (SIE) 2017

Authors and Affiliations

  • E. B. Conemans
    • 1
    • 2
  • G. M. Raicu-Ionita
    • 3
  • C. R. C. Pieterman
    • 2
  • K. M. A. Dreijerink
    • 2
  • O. M. Dekkers
    • 4
  • A. R. Hermus
    • 5
  • W. W. de Herder
    • 6
  • M. L. Drent
    • 7
  • A. N. A. van der Horst-Schrivers
    • 8
  • B. Havekes
    • 9
  • P. H. Bisschop
    • 10
  • G. J. Offerhaus
    • 3
  • I. H. M. Borel Rinkes
    • 1
  • G. D. Valk
    • 2
  • H. Th. M. Timmers
    • 11
  • M. R. Vriens
    • 1
    Email author
  1. 1.Department of SurgeryUniversity Medical Center UtrechtUtrechtThe Netherlands
  2. 2.Department of Internal MedicineUniversity Medical Center UtrechtUtrechtThe Netherlands
  3. 3.Department of PathologyUniversity Medical Center UtrechtUtrechtThe Netherlands
  4. 4.Department of Endocrinology and Metabolism and Department of Clinical EpidemiologyLeiden University Medical CenterLeidenThe Netherlands
  5. 5.Department of EndocrinologyRadboud University Medical CenterNijmegenThe Netherlands
  6. 6.Department of Internal MedicineErasmus Medical CenterRotterdamThe Netherlands
  7. 7.Department of Internal Medicine, Section EndocrinologyVU University Medical CenterAmsterdamThe Netherlands
  8. 8.Department of EndocrinologyUniversity Medical Center GroningenGroningenThe Netherlands
  9. 9.Department of Internal Medicine, Division of EndocrinologyMaastricht University Medical CenterMaastrichtThe Netherlands
  10. 10.Department of Endocrinology and MetabolismAcademic Medical CenterAmsterdamThe Netherlands
  11. 11.Section Stem Cells, Regenerative Medicine Center and Center for Molecular MedicineUniversity Medical Center UtrechtUtrechtThe Netherlands

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