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MAP kinase activity supported by BRAF V600E mutation rather than gene amplification is associated with ETV1 expression in melanoma brain metastases

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Abstract

In primary melanoma, ETV1 transcription factor was suggested to be activated mainly by gene amplification and to promote tumor growth in cooperation with BRAF V600E. Aim of this study was to investigate ETV1 expression in human melanoma with a focus on brain metastases. We investigated ETV1 in 68 human melanoma brain metastases using FISH for ETV1 gene (located at chromosome 7p21) and centromere chromosome 7 and immunohistochemistry for ETV1, BRAF V600E, and ETV1/BRAF associated proteins pMSK1, pRSK1, pp38, pMEK1/2, MAPKAP kinase 2, CIC, HIF-1alpha and Ki-67. We further studied ETV1 copy number variations in 32 melanoma cell lines from primary and metastatic lesions using array CGH. The influence of the MAP kinase pathway activity on ETV1 mRNA and protein expression under BRAF wild-type and BRAF V600E conditions were determined in melanoma cell lines using qRT-PCR and Western Blot. No ETV1 high grade amplifications were observed in tissue samples, but low grade ETV1 gene amplifications were found in 7 (10.3 %) melanoma brain metastases. ETV1 protein expression in tissue samples (15 %) correlated with BRAF V600E status (p = 0.007) and HIF-1alpha expression (p = 0.049), but not with ETV1 gene dose. Application of the BRAFV600E-specific inhibitor vemurafenib and the BRAFV6ooE/V600K-inhibitor dabrafenib revealed predominant regulation of ETV-1 mRNA and protein via MAPK-pathway. ETV1 expression is a rare event in human melanoma and seems to be rather based on hyperactivation of MAPK signals, by BRAF V600E mutation, than on ETV1 gene amplification. Consequently, therapeutic inhibition of BRAF and the downstream MAPK pathway also down-regulates oncogenic ETV1 expression.

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References

  1. Monte D, Coutte L, Baert JL, Angeli I, Stehelin D, de Launoit Y (1995) Molecular characterization of the ets-related human transcription factor ER81. Oncogene 11(4):771–779

    CAS  PubMed  Google Scholar 

  2. Charlot C, Dubois-Pot H, Serchov T, Tourrette Y, Wasylyk B (2010) A review of post-translational modifications and subcellular localization of Ets transcription factors: possible connection with cancer and involvement in the hypoxic response. Methods Mol Biol 647:3–30

    Article  CAS  PubMed  Google Scholar 

  3. Hollenhorst PC, McIntosh LP, Graves BJ (2011) Genomic and biochemical insights into the specificity of ETS transcription factors. Annu Rev Biochem 80:437–471

    Article  CAS  PubMed  Google Scholar 

  4. Hermans KG, van der Korput HA, van Marion R, van de Wijngaart DJ, Ziel-van der Made A, Dits NF, Boormans JL, van der Kwast TH, van Dekken H, Bangma CH, Korsten H, Kraaij R, Jenster G, Trapman J (2008) Truncated ETV1, fused to novel tissue-specific genes, and full-length ETV1 in prostate cancer. Cancer Res 68(18):7541–7549

    Article  CAS  PubMed  Google Scholar 

  5. Tomlins SA, Rhodes DR, Perner S, Dhanasekaran SM, Mehra R, Sun XW, Varambally S, Cao X, Tchinda J, Kuefer R, Lee C, Montie JE, Shah RB, Pienta KJ, Rubin MA, Chinnaiyan AM (2005) Recurrent fusion of TMPRSS2 and ETS transcription factor genes in prostate cancer. Science 310(5748):644–648

    Article  CAS  PubMed  Google Scholar 

  6. Jeon IS, Davis JN, Braun BS, Sublett JE, Roussel MF, Denny CT, Shapiro DN (1995) A variant Ewing’s sarcoma translocation (7;22) fuses the EWS gene to the ETS gene ETV1. Oncogene 10(6):1229–1234

    CAS  PubMed  Google Scholar 

  7. Chi P, Chen Y, Zhang L, Guo X, Wongvipat J, Shamu T, Fletcher JA, Dewell S, Maki RG, Zheng D, Antonescu CR, Allis CD, Sawyers CL (2010) ETV1 is a lineage survival factor that cooperates with KIT in gastrointestinal stromal tumours. Nature 467(7317):849–853

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  8. Vitari AC, Leong KG, Newton K, Yee C, O’Rourke K, Liu J, Phu L, Vij R, Ferrando R, Couto SS, Mohan S, Pandita A, Hongo JA, Arnott D, Wertz IE, Gao WQ, French DM, Dixit VM (2011) COP1 is a tumour suppressor that causes degradation of ETS transcription factors. Nature 474(7351):403–406

    Article  CAS  PubMed  Google Scholar 

  9. Wu J, Janknecht R (2002) Regulation of the ETS transcription factor ER81 by the 90-kDa ribosomal S6 kinase 1 and protein kinase A. J Biol Chem 277(45):42669–42679

    Article  CAS  PubMed  Google Scholar 

  10. Dakour J, Vinayagamoorthy T, Jimbow K, Chen H, Luo D, Dixon W, Munoz V (1993) Identification of a cDNA coding for a Ca(2+)-binding phosphoprotein (p90), calnexin, on melanosomes in normal and malignant human melanocytes. Exp Cell Res 209(2):288–300

    Article  CAS  PubMed  Google Scholar 

  11. Willmore-Payne C, Holden JA, Tripp S, Layfield LJ (2005) Human malignant melanoma: detection of BRAF- and c-kit-activating mutations by high-resolution amplicon melting analysis. Hum Pathol 36(5):486–493

    Article  CAS  PubMed  Google Scholar 

  12. Jane-Valbuena J, Widlund HR, Perner S, Johnson LA, Dibner AC, Lin WM, Baker AC, Nazarian RM, Vijayendran KG, Sellers WR, Hahn WC, Duncan LM, Rubin MA, Fisher DE, Garraway LA (2010) An oncogenic role for ETV1 in melanoma. Cancer Res 70(5):2075–2084

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  13. Birner P, Beer A, Vinatzer U, Stary S, Hoftberger R, Nirtl N, Wrba F, Streubel B, Schoppmann SF (2012) MAPKAP kinase 2 overexpression influences prognosis in gastrointestinal stromal tumors and associates with copy number variations on chromosome 1 and expression of p38 MAP kinase and ETV1. Clin Cancer Res 18(7):1879–1887

    Article  CAS  PubMed  Google Scholar 

  14. Davies H, Bignell GR, Cox C, Stephens P, Edkins S, Clegg S, Teague J, Woffendin H, Garnett MJ, Bottomley W, Davis N, Dicks E, Ewing R, Floyd Y, Gray K, Hall S, Hawes R, Hughes J, Kosmidou V, Menzies A, Mould C, Parker A, Stevens C, Watt S, Hooper S, Wilson R, Jayatilake H, Gusterson BA, Cooper C, Shipley J, Hargrave D, Pritchard-Jones K, Maitland N, Chenevix-Trench G, Riggins GJ, Bigner DD, Palmieri G, Cossu A, Flanagan A, Nicholson A, Ho JW, Leung SY, Yuen ST, Weber BL, Seigler HF, Darrow TL, Paterson H, Marais R, Marshall CJ, Wooster R, Stratton MR, Futreal PA (2002) Mutations of the BRAF gene in human cancer. Nature 417(6892):949–954

    Article  CAS  PubMed  Google Scholar 

  15. Huang FW, Hodis E, Xu MJ, Kryukov GV, Chin L, Garraway LA (2013) Highly recurrent TERT promoter mutations in human melanoma. Science 339(6122):957–959

    Article  CAS  PubMed  Google Scholar 

  16. Schouten LJ, Rutten J, Huveneers HA, Twijnstra A (2002) Incidence of brain metastases in a cohort of patients with carcinoma of the breast, colon, kidney, and lung and melanoma. Cancer 94(10):2698–2705

    Article  PubMed  Google Scholar 

  17. Sosman JA, Kim KB, Schuchter L, Gonzalez R, Pavlick AC, Weber JS, McArthur GA, Hutson TE, Moschos SJ, Flaherty KT, Hersey P, Kefford R, Lawrence D, Puzanov I, Lewis KD, Amaravadi RK, Chmielowski B, Lawrence HJ, Shyr Y, Ye F, Li J, Nolop KB, Lee RJ, Joe AK, Ribas A (2012) Survival in BRAF V600-mutant advanced melanoma treated with vemurafenib. N Engl J Med 366(8):707–714

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  18. Falchook GS, Long GV, Kurzrock R, Kim KB, Arkenau TH, Brown MP, Hamid O, Infante JR, Millward M, Pavlick AC, O’Day SJ, Blackman SC, Curtis CM, Lebowitz P, Ma B, Ouellet D, Kefford RF (2012) Dabrafenib in patients with melanoma, untreated brain metastases, and other solid tumours: a phase 1 dose-escalation trial. Lancet 379(9829):1893–1901

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  19. Kumar SM, Yu H, Edwards R, Chen L, Kazianis S, Brafford P, Acs G, Herlyn M, Xu X (2007) Mutant V600E BRAF increases hypoxia inducible factor-1alpha expression in melanoma. Cancer Res 67(7):3177–3184

    Article  CAS  PubMed  Google Scholar 

  20. Birner P, Piribauer M, Fischer I, Gatterbauer B, Marosi C, Ambros PF, Ambros IM, Bredel M, Oberhuber G, Rossler K, Budka H, Harris AL, Hainfellner JA (2003) Vascular patterns in glioblastoma influence clinical outcome and associate with variable expression of angiogenic proteins: evidence for distinct angiogenic subtypes. Brain Pathol 13(2):133–143

    Article  CAS  PubMed  Google Scholar 

  21. Hirsch FR, Varella-Garcia M, Bunn PA Jr, Di Maria MV, Veve R, Bremmes RM, Baron AE, Zeng C, Franklin WA (2003) Epidermal growth factor receptor in non-small-cell lung carcinomas: correlation between gene copy number and protein expression and impact on prognosis. J Clin Oncol 21(20):3798–3807

    Article  CAS  PubMed  Google Scholar 

  22. Capper D, Berghoff AS, Magerle M, Ilhan A, Wohrer A, Hackl M, Pichler J, Pusch S, Meyer J, Habel A, Petzelbauer P, Birner P, von Deimling A, Preusser M (2012) Immunohistochemical testing of BRAF V600E status in 1,120 tumor tissue samples of patients with brain metastases. Acta Neuropathol 123(2):223–233

    Article  CAS  PubMed  Google Scholar 

  23. Capper D, Preusser M, Habel A, Sahm F, Ackermann U, Schindler G, Pusch S, Mechtersheimer G, Zentgraf H, von Deimling A (2011) Assessment of BRAF V600E mutation status by immunohistochemistry with a mutation-specific monoclonal antibody. Acta Neuropathol 122(1):11–19

    Article  CAS  PubMed  Google Scholar 

  24. Berger W, Elbling L, Minai-Pour M, Vetterlein M, Pirker R, Kokoschka EM, Micksche M (1994) Intrinsic MDR-1 gene and P-glycoprotein expression in human melanoma cell lines. Int J Cancer 59(5):717–723

    Article  CAS  PubMed  Google Scholar 

  25. Berger W, Hauptmann E, Elbling L, Vetterlein M, Kokoschka EM, Micksche M (1997) Possible role of the multidrug resistance-associated protein (MRP) in chemoresistance of human melanoma cells. Int J Cancer 71(1):108–115

    Article  CAS  PubMed  Google Scholar 

  26. Mathieu V, Pirker C, Schmidt WM, Spiegl-Kreinecker S, Lotsch D, Heffeter P, Hegedus B, Grusch M, Kiss R, Berger W (2012) Aggressiveness of human melanoma xenograft models is promoted by aneuploidy-driven gene expression deregulation. Oncotarget 3(4):399–413

    PubMed Central  PubMed  Google Scholar 

  27. Lotsch D, Steiner E, Holzmann K, Spiegl-Kreinecker S, Pirker C, Hlavaty J, Petznek H, Hegedus B, Garay T, Mohr T, Sommergruber W, Grusch M, Berger W (2013) Major vault protein supports glioblastoma survival and migration by upregulating the EGFR/PI3K signalling axis. Oncotarget 4(11):1904–1918

    PubMed Central  PubMed  Google Scholar 

  28. Heffeter P, Jakupec MA, Korner W, Chiba P, Pirker C, Dornetshuber R, Elbling L, Sutterluty H, Micksche M, Keppler BK, Berger W (2007) Multidrug-resistant cancer cells are preferential targets of the new antineoplastic lanthanum compound KP772 (FFC24). Biochem Pharmacol 73(12):1873–1886

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  29. Hatzivassiliou G, Song K, Yen I, Brandhuber BJ, Anderson DJ, Alvarado R, Ludlam MJ, Stokoe D, Gloor SL, Vigers G, Morales T, Aliagas I, Liu B, Sideris S, Hoeflich KP, Jaiswal BS, Seshagiri S, Koeppen H, Belvin M, Friedman LS, Malek S (2010) RAF inhibitors prime wild-type RAF to activate the MAPK pathway and enhance growth. Nature 464(7287):431–435

    Article  CAS  PubMed  Google Scholar 

  30. Lee JT, Li L, Brafford PA, van den Eijnden M, Halloran MB, Sproesser K, Haass NK, Smalley KS, Tsai J, Bollag G, Herlyn M (2010) PLX4032, a potent inhibitor of the B-Raf V600E oncogene, selectively inhibits V600E-positive melanomas. Pigment Cell Melanoma Res 23(6):820–827

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  31. Mehra R, Dhanasekaran SM, Palanisamy N, Vats P, Cao X, Kim JH, Kim DS, Johnson T, Fullen DR, Chinnaiyan AM (2013) Comprehensive analysis of ets family members in melanoma by fluorescence in situ hybridization reveals recurrent ETV1 amplification. Transl Oncol 6(4):405–412

    Article  PubMed Central  PubMed  Google Scholar 

  32. Pratilas CA, Taylor BS, Ye Q, Viale A, Sander C, Solit DB, Rosen N (2009) (V600E)BRAF is associated with disabled feedback inhibition of RAF-MEK signaling and elevated transcriptional output of the pathway. Proc Natl Acad Sci USA 106(11):4519–4524

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  33. Yang XM, Wang YS, Zhang J, Li Y, Xu JF, Zhu J, Zhao W, Chu DK, Wiedemann P (2009) Role of PI3K/Akt and MEK/ERK in mediating hypoxia-induced expression of HIF-1alpha and VEGF in laser-induced rat choroidal neovascularization. Invest Ophthalmol Vis Sci 50(4):1873–1879

    Article  PubMed  Google Scholar 

  34. Aebersold DM, Burri P, Beer KT, Laissue J, Djonov V, Greiner RH, Semenza GL (2001) Expression of hypoxia-inducible factor-1alpha: a novel predictive and prognostic parameter in the radiotherapy of oropharyngeal cancer. Cancer Res 61(7):2911–2916

    CAS  PubMed  Google Scholar 

  35. Bachtiary B, Schindl M, Potter R, Dreier B, Knocke TH, Hainfellner JA, Horvat R, Birner P (2003) Overexpression of hypoxia-inducible factor 1alpha indicates diminished response to radiotherapy and unfavorable prognosis in patients receiving radical radiotherapy for cervical cancer. Clin Cancer Res 9(6):2234–2240

    CAS  PubMed  Google Scholar 

  36. Birner P, Preusser M, Gelpi E, Berger J, Gatterbauer B, Ambros IM, Ambros PF, Acker T, Plate KH, Harris AL, Hainfellner JA (2004) Expression of hypoxia-related tissue factors correlates with diminished survival of adjuvantly treated patients with chromosome 1p aberrant oligodendroglial neoplasms and therapeutic implications. Clin Cancer Res 10(19):6567–6571

    Article  CAS  PubMed  Google Scholar 

  37. Koukourakis MI, Giatromanolaki A, Skarlatos J, Corti L, Blandamura S, Piazza M, Gatter KC, Harris AL (2001) Hypoxia inducible factor (HIF-1a and HIF-2a) expression in early esophageal cancer and response to photodynamic therapy and radiotherapy. Cancer Res 61(5):1830–1832

    CAS  PubMed  Google Scholar 

  38. Valencak J, Kittler H, Schmid K, Schreiber M, Raderer M, Gonzalez-Inchaurraga M, Birner P, Pehamberger H (2009) Prognostic relevance of hypoxia inducible factor-1alpha expression in patients with melanoma. Clin Exp Dermatol 34(8):e962–e964

    Article  CAS  PubMed  Google Scholar 

  39. Nazarian R, Shi H, Wang Q, Kong X, Koya RC, Lee H, Chen Z, Lee MK, Attar N, Sazegar H, Chodon T, Nelson SF, McArthur G, Sosman JA, Ribas A, Lo RS (2010) Melanomas acquire resistance to B-RAF(V600E) inhibition by RTK or N-RAS upregulation. Nature 468(7326):973–977

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  40. Dissanayake K, Toth R, Blakey J, Olsson O, Campbell DG, Prescott AR, MacKintosh C (2011) ERK/p90(RSK)/14-3-3 signalling has an impact on expression of PEA3 Ets transcription factors via the transcriptional repressor capicua. Biochem J 433(3):515–525

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  41. Astigarraga S, Grossman R, Diaz-Delfin J, Caelles C, Paroush Z, Jimenez G (2007) A MAPK docking site is critical for down-regulation of Capicua by Torso and EGFR RTK signaling. EMBO J 26(3):668–677

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  42. Andreu MJ, Ajuria L, Samper N, Gonzalez-Perez E, Campuzano S, Gonzalez-Crespo S, Jimenez G (2012) EGFR-dependent down-regulation of Capicua and the establishment of Drosophila dorsoventral polarity. Fly (Austin) 6(4):234–239

    Article  CAS  Google Scholar 

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Acknowledgments

We thank Mrs. Gerda Ricken for excellent technical assistance. This work was supported by the research budget of the Medical University of Vienna, by the “Medizinisch-wissenschaftlicher Fonds des Bürgermeisters der Bundeshauptstadt Wien” (project# 13056) and unrestricted research grants by Roche and Glaxo Smith Kline (GSK).

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The authors have declared no conflicts of interest.

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Authors and Affiliations

  1. Department of Neuropathology, Institute of Pathology, Ruprecht-Karls-University, Heidelberg, Germany

    Peter Birner, David Capper & Andreas von Deimling

  2. Clinical Institute of Pathology, Medical University of Vienna, Vienna, Austria

    Peter Birner

  3. Comprehensive Cancer Center—CNS Tumors Unit, Medical University of Vienna, Vienna, Austria

    Peter Birner, Anna S. Berghoff, Carina Dinhof, Christine Pirker, Walter Berger & Matthias Preusser

  4. Institute of Neurology, Medical University of Vienna, Vienna, Austria

    Anna S. Berghoff

  5. Clinical Cooperation Unit Neuropathology, German Cancer Research Center, Heidelberg, Germany

    David Capper & Andreas von Deimling

  6. Department of Surgery, Medical University of Vienna, Vienna, Austria

    Sebastian F. Schoppmann

  7. Department of Dermatology, Medical University of Vienna, Vienna, Austria

    Peter Petzelbauer

  8. Department of Medicine I, Medical University of Vienna, Währinger Gürtel 18-20, 1090, Vienna, Austria

    Carina Dinhof, Christine Pirker, Walter Berger & Matthias Preusser

  9. Institute of Cancer Research, Medical University of Vienna, Vienna, Austria

    Christine Pirker & Walter Berger

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  1. Peter Birner
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Correspondence to Matthias Preusser.

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Birner, P., Berghoff, A.S., Dinhof, C. et al. MAP kinase activity supported by BRAF V600E mutation rather than gene amplification is associated with ETV1 expression in melanoma brain metastases. Arch Dermatol Res 306, 873–884 (2014). https://doi.org/10.1007/s00403-014-1490-6

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  • DOI: https://doi.org/10.1007/s00403-014-1490-6

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