Malignant peripheral nerve sheath tumors contain loss of histone H3K27 trimethylation (H3K27me3) due to driver mutations affecting the polycomb repressive complex 2 (PRC2). Consequently, loss of H3K27me3 staining has served as a diagnostic marker for this tumor type. However, recent reports demonstrate H3K27me3 loss in numerous other tumors, including some in the differential diagnosis of malignant peripheral nerve sheath tumor. Since these tumors lose H3K27me3 through mechanisms distinct from PRC2 loss, we set out to determine whether loss of dimethylation of H3K27, which is also catalyzed by PRC2, might be a more specific marker of PRC2 loss and malignant peripheral nerve sheath tumor. Using mass spectrometry, we identify a near complete loss of H3K27me2 in malignant peripheral nerve sheath tumors and cell lines. Immunohistochemical analysis of 72 malignant peripheral nerve sheath tumors, seven K27M-mutant gliomas, 43 ependymomas, and 10 Merkel cell carcinomas demonstrates that while H3K27me3 loss is common across these tumor types, H3K27me2 loss is limited to malignant peripheral nerve sheath tumors and is highly concordant with H3K27me3 loss (33/34 cases). Thus, increased specificity does not come at the cost of greatly reduced sensitivity. To further compare H3K27me2 and H3K27me3 immunohistochemistry, we investigated 42 melanomas and 54 synovial sarcomas, histologic mimics of malignant peripheral nerve sheath tumor with varying degrees of H3K27me3 loss in prior reports. While global H3K27me3 loss was not seen in these tumors, weak and limited H3K27me3 staining was common. By contrast, H3K27me2 staining was more clearly retained in all cases, making it a superior binary classifier. This was confirmed by digital image analysis of stained slides. Our findings indicate that H3K27me2 loss is highly specific for PRC2 loss and that PRC2 loss is a rarer phenomenon than H3K27me3 loss. Consequently, H3K27me2 loss is a superior diagnostic marker for malignant peripheral nerve sheath tumor.
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Rodriguez FJ, Folpe AL, Giannini C, Perry A. Pathology of peripheral nerve sheath tumors: diagnostic overview and update on selected diagnostic problems. Acta Neuropathol. 2012;123:295–319.
Hornick JL Limited biopsies of soft tissue tumors: the contemporary role of immunohistochemistry and molecular diagnostics. Mod Pathol. 2019. https://doi.org/10.1038/s41379-018-0139-y.
Zou C, Smith KD, Liu J, Lahat G, Myers S, Wang WL, et al. Clinical, pathological, and molecular variables predictive of malignant peripheral nerve sheath tumor outcome. Ann Surg. 2009;249:1014–22.
Le Guellec S, Decouvelaere A-V, Filleron T, Valo I, Charon-Barra C, Robin YM, et al. Malignant peripheral nerve sheath tumor is a challenging diagnosis: a systematic pathology review, immunohistochemistry, and molecular analysis in 160 patients from the french sarcoma group database. Am J Surg Pathol. 2016;40:896–908.
Lee W, Teckie S, Wiesner T, Ran L, Prieto Granada CN, Lin M, et al. PRC2 is recurrently inactivated through EED or SUZ12 loss in malignant peripheral nerve sheath tumors. Nat Genet. 2014;46:1227–32.
De Raedt T, Beert E, Pasmant E, Luscan A, Brems H, Ortonne N, et al. PRC2 loss amplifies Ras-driven transcription and confers sensitivity to BRD4-based therapies. Nature. 2014;514:247–51.
Zhang M, Wang Y, Jones S, Sausen M, McMahon K, Sharma R, et al. Somatic mutations of SUZ12 in malignant peripheral nerve sheath tumors. Nat Genet. 2014;46:1170–2.
Prieto-Granada CN, Wiesner T, Messina JL, Jungbluth AA, Chi P, Antonescu CR. Loss of H3K27me3 expression is a highly sensitive marker for sporadic and radiation-induced MPNST. Am J Surg Pathol. 2016;40:479–89.
Schaefer I-M, Fletcher CD, Hornick JL. Loss of H3K27 trimethylation distinguishes malignant peripheral nerve sheath tumors from histologic mimics. Mod Pathol. 2016;29:4–13.
Cleven AHG, Sannaa GAA, Briaire-de Bruijn I, Ingram DR, van de Rijn M, Rubin BP, et al. Loss of H3K27 tri-methylation is a diagnostic marker for malignant peripheral nerve sheath tumors and an indicator for an inferior survival. Mod Pathol. 2016;29:582–90.
Otsuka H, Kohashi K, Yoshimoto M, Ishihara S, Toda Y, Yamada Y, et al. Immunohistochemical evaluation of H3K27 trimethylation in malignant peripheral nerve sheath tumors. Pathol Res Pr. 2018;214:417–25.
Le Guellec S, Macagno N, Velasco V, Lamant L, Lae M, Filleron T, et al. Loss of H3K27 trimethylation is not suitable for distinguishing malignant peripheral nerve sheath tumor from melanoma: a study of 387 cases including mimicking lesions. Mod Pathol. 2017;30:1677–87.
Asano N, Yoshida A, Ichikawa H, Mori T, Nakamura M, Kawai A, et al. Immunohistochemistry for trimethylated H3K27 in the diagnosis of malignant peripheral nerve sheath tumours. Histopathology. 2017;70:385–93.
Pekmezci M, Cuevas-Ocampo AK, Perry A, Horvai AE. Significance of H3K27me3 loss in the diagnosis of malignant peripheral nerve sheath tumors. Mod Pathol. 2017;30:1710–9.
Lewis PW, Müller MM, Koletsky MS, Cordero F, Lin S, Banaszynski LA, et al. Inhibition of PRC2 activity by a gain-of-function H3 mutation found in pediatric glioblastoma. Science. 2013;340:857–61.
Bender S, Tang Y, Lindroth AM, Hovestadt V, Jones DT, Kool M, et al. Reduced H3K27me3 and DNA hypomethylation are major drivers of gene expression in K27M mutant pediatric high-grade gliomas. Cancer Cell. 2013;24:660–72.
Chan K-M, Fang D, Gan H, Hashizume R, Yu C, Schroeder M, et al. The histone H3.3K27M mutation in pediatric glioma reprograms H3K27 methylation and gene expression. Genes Dev. 2013;27:985–90.
Bayliss J, Mukherjee P, Lu C, Jain SU, Chung C, Martinez D, et al. Lowered H3K27me3 and DNA hypomethylation define poorly prognostic pediatric posterior fossa ependymomas. Sci Transl Med. 2016;8:366ra161.
Panwalkar P, Clark J, Ramaswamy V, Hawes D, Yang F, Dunham C, et al. Immunohistochemical analysis of H3K27me3 demonstrates global reduction in group-A childhood posterior fossa ependymoma and is a powerful predictor of outcome. Acta Neuropathol. 2017;134:705–14.
Busam KJ, Pulitzer MP, Coit DC, Arcila M, Leng D, Jungbluth AA, et al. Reduced H3K27me3 expression in Merkel cell polyoma virus-positive tumors. Mod Pathol. 2017;30:877–83.
Katz LM, Hielscher T, Liechty B, Silverman J, Zagzag D, Sen R, et al. Loss of histone H3K27me3 identifies a subset of meningiomas with increased risk of recurrence. Acta Neuropathol. 2018;135:955–63.
Makise N, Sekimizu M, Konishi E, Motoi T, Kubo T, Ikoma H, et al. H3K27me3 deficiency defines a subset of dedifferentiated chondrosarcomas with characteristic clinicopathological features. Mod Pathol. 2018. https://doi.org/10.1038/s41379-018-0140-5.
Piunti A, Shilatifard A. Epigenetic balance of gene expression by Polycomb and COMPASS families. Science. 2016;352:aad9780.
Pasini D, Bracken AP, Jensen MR, Lazzerini Denchi E, Helin K. Suz12 is essential for mouse development and for EZH2 histone methyltransferase activity. EMBO J. 2004;23:4061–71.
Ferrari KJ, Scelfo A, Jammula S, Cuomo A, Barozzi I, Stützer A, et al. Polycomb-dependent H3K27me1 and H3K27me2 regulate active transcription and enhancer fidelity. Mol Cell. 2014;53:49–62.
Wessel D, Flügge UI. A method for the quantitative recovery of protein in dilute solution in the presence of detergents and lipids. Anal Biochem. 1984;138:141–3.
Sidoli S, Garcia BA. Characterization of individual histone posttranslational modifications and their combinatorial patterns by mass spectrometry-based proteomics strategies. Methods Mol Biol. 2017;1528:121–48.
Yuan Z-F, Sidoli S, Marchione DM, Simithy J, Janssen KA, Szurgot MR, et al. EpiProfile 2.0: a computational platform for processing epi-proteomics mass spectrometry data. J Proteome Res. 2018;17:2533–41.
Leroy G, Dimaggio PA, Chan EY, Zee BM, Blanco MA, Bryant B, et al. A quantitative atlas of histone modification signatures from human cancer cells. Epigenetics Chromatin. 2013;6:20.
Stafford JM, Lee C-H, Voigt P, Descostes N, Saldaña-Meyer R, Yu JR, et al. Multiple modes of PRC2 inhibition elicit global chromatin alterations in H3K27M pediatric glioma. Sci Adv. 2018;4:eaau5935.
Bankhead P, Loughrey MB, Fernández JA, Dombrowski Y, McArt DG, Dunne PD, et al. QuPath: open source software for digital pathology image analysis. Sci Rep. 2017;7:16878.
Noberini R, Osti D, Miccolo C, Richichi C, Lupia M, Corleone G, et al. Extensive and systematic rewiring of histone post-translational modifications in cancer model systems. Nucleic Acids Res. 2018;46:3817–32.
Zhang Y, Muller M, Xu B, Yoshida Y, Horlacher O, Nikitin K, et al. Unrestricted modification search reveals lysine methylation as major modification induced by tissue formalin fixation and paraffin embedding. Proteomics. 2015;15:2568–79.
Schoeftner S, Sengupta AK, Kubicek S, Mechtler K, Spahn L, Koseki H, et al. Recruitment of PRC1 function at the initiation of X inactivation independent of PRC2 and silencing. EMBO J. 2006;25:3110–22.
Shen X, Liu Y, Hsu Y-J, Fujiwara Y, Kim J, Mao X, et al. EZH1 mediates methylation on histone H3 lysine 27 and complements EZH2 in maintaining stem cell identity and executing pluripotency. Mol Cell. 2008;32:491–502.
Peters AHFM, Kubicek S, Mechtler K, O'Sullivan RJ, Derijck AA, Perez-Burgos L, et al. Partitioning and plasticity of repressive histone methylation states in mammalian chromatin. Mol Cell. 2003;12:1577–89.
Busam KJ, Shah KN, Gerami P, Sitzman T, Jungbluth AA, Kinsler V. Reduced H3K27me3 expression is common in nodular melanomas of childhood associated with congenital melanocytic nevi but not in proliferative nodules. Am J Surg Pathol. 2017;41:396–404.
Pajtler KW, Wen J, Sill M, Lin T, Orisme W, Tang B, et al. Molecular heterogeneity and CXorf67 alterations in posterior fossa group A (PFA) ependymomas. Acta Neuropathol. 2018;136:211–26.
Mohammad F, Weissmann S, Leblanc B, Pandey DP, Højfeldt JW, Comet I, et al. EZH2 is a potential therapeutic target for H3K27M-mutant pediatric gliomas. Nat Med. 2017;23:483–92.
Kadoch C, Williams RT, Calarco JP, Miller EL, Weber CM, Braun SM, et al. Dynamics of BAF-Polycomb complex opposition on heterochromatin in normal and oncogenic states. Nat Genet. 2017;49:213–22.
Perdigoto CN, Dauber KL, Bar C, Tsai PC, Valdes VJ, Cohen I, et al. Polycomb-mediated repression and sonic hedgehog signaling interact to regulate merkel cell specification during skin development. PLoS Genet. 2016;12:e1006151.
Piunti A, Hashizume R, Morgan MA, Bartom ET, Horbinski CM, Marshall SA, et al. Therapeutic targeting of polycomb and BET bromodomain proteins in diffuse intrinsic pontine gliomas. Nat Med. 2017;23:493–500.
Amirnasr A, Verdijk RM, van Kuijk PF, Taal W, Sleijfer S, Wiemer EAC. Expression and inhibition of BRD4, EZH2 and TOP2A in neurofibromas and malignant peripheral nerve sheath tumors. PLoS ONE. 2017;12:e0183155.
Wassef M, Luscan A, Aflaki S, Zielinski D, Jansen PWTC, Baymaz HI, et al. EZH1/2 function mostly within canonical PRC2 and exhibit proliferation-dependent redundancy that shapes mutational signatures in cancer. Proc Natl Acad Sci USA. 2019. https://doi.org/10.1073/pnas.1814634116.
Grasso CS, Tang Y, Truffaux N, Berlow NE, Liu L, Debily MA, et al. Functionally defined therapeutic targets in diffuse intrinsic pontine glioma. Nat Med. 2015;21:555–9.
Nagaraja S, Vitanza NA, Woo PJ, Taylor KR, Liu F, Zhang L, et al. Transcriptional dependencies in diffuse intrinsic pontine glioma. Cancer Cell. 2017;31:635–.e6.
Wojcik JB, Marchione DM, Sidoli S, Djedid A, Lisby A, Majewski J, et al. Epigenomic reordering induced by Polycomb loss drives oncogenesis but leads to therapeutic vulnerabilities in malignant peripheral nerve sheath tumors. Cancer Res. 2019;3704:2018. canres
Makise N, Sekimizu M, Kubo T, Wakai S, Watanabe SI, Kato T, et al. Extraskeletal osteosarcoma: MDM2 and H3K27me3 analysis of 19 cases suggest disease heterogeneity. Histopathology. 2018;73:147–56.
Makise N, Sekimizu M, Kubo T, Wakai S, Hiraoka N, Komiyama M, et al. Clarifying the distinction between malignant peripheral nerve sheath tumor and dedifferentiated liposarcoma: a critical reappraisal of the diagnostic utility of MDM2 and H3K27me3 status. Am J Surg Pathol. 2018;42:656–64.
This research was supported by US National Institutes of Health (NIH) grants (GM110174, AI118891, and CA196539 to BAG and P01CA196539 to BAG; 5K12CA076931 to JBW; T32GM008275 and TL1TR001880 to DMM). JBW is also supported by the University of Pennsylvania Department of Pathology and Laboratory Medicine startup funds. BAG is also supported by a Robert Arceci Scholar award from the Leukemia and Lymphoma Society. The authors would also like to acknowledge Dr. Kumarasen Cooper in the Department of Pathology at the Hospital of the University of Pennsylvania for his support and helpful discussions about the content of the manuscript.
Conflict of interest
The authors declare that they have no conflict of interest.
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