Acta Neuropathologica

, Volume 129, Issue 1, pp 133–146 | Cite as

ATRX and IDH1-R132H immunohistochemistry with subsequent copy number analysis and IDH sequencing as a basis for an “integrated” diagnostic approach for adult astrocytoma, oligodendroglioma and glioblastoma

  • David E. Reuss
  • Felix Sahm
  • Daniel Schrimpf
  • Benedikt Wiestler
  • David Capper
  • Christian Koelsche
  • Leonille Schweizer
  • Andrey Korshunov
  • David T. W. Jones
  • Volker Hovestadt
  • Michel Mittelbronn
  • Jens Schittenhelm
  • Christel Herold-Mende
  • Andreas Unterberg
  • Michael Platten
  • Michael Weller
  • Wolfgang Wick
  • Stefan M. Pfister
  • Andreas von DeimlingEmail author
Original Paper


Diffuse gliomas are represented in the 2007 WHO classification as astrocytomas, oligoastrocytomas and oligodendrogliomas of grades II and III and glioblastomas WHO grade IV. Molecular data on these tumors have a major impact on prognosis and therapy of the patients. Consequently, the inclusion of molecular parameters in the WHO definition of brain tumors is being planned and has been forwarded as the “ISN-Haarlem” consensus. We, here, analyze markers of special interest including ATRX, IDH and 1p/19q codeletion in a series of 405 adult patients. Among the WHO 2007 classified tumors were 152 astrocytomas, 61 oligodendrogliomas, 63 oligoastrocytomas and 129 glioblastomas. Following the concepts of the “ISN-Haarlem”, we rediagnosed the series to obtain “integrated” diagnoses with 155 tumors being astrocytomas, 100 oligodendrogliomas and 150 glioblastomas. In a subset of 100 diffuse gliomas from the NOA-04 trial with long-term follow-up data available, the “integrated” diagnosis had a significantly greater prognostic power for overall and progression-free survival compared to WHO 2007. Based on the “integrated” diagnoses, loss of ATRX expression was close to being mutually exclusive to 1p/19q codeletion, with only 2 of 167 ATRX-negative tumors exhibiting 1p/19q codeletion. All but 4 of 141 patients with loss of ATRX expression and diffuse glioma carried either IDH1 or IDH2 mutations. Interestingly, the majority of glioblastoma patients with loss of ATRX expression but no IDH mutations exhibited an H3F3A mutation. Further, all patients with 1p/19 codeletion carried a mutation in IDH1 or IDH2. We present an algorithm based on stepwise analysis with initial immunohistochemistry for ATRX and IDH1-R132H followed by 1p/19q analysis followed by IDH sequencing which reduces the number of molecular analyses and which has a far better association with patient outcome than WHO 2007.


ATRX IDH Diffuse glioma Glioblastoma Oligodendroglioma Astrocytoma 7p/10q H3F3A 1p/19q 



The authors (WW, MW) conducting this work represent the Neurooncology Working Group (NOA) of the German Cancer Society. We gratefully acknowledge the contributions of Ulrike Ernemann, MD and Christoph Meisner, PhD (Tübingen, Germany), Guido Reifenberger, MD and Michael C. Sabel, MD (Düsseldorf, Germany), Susanne Koeppen, MD (Essen, Germany), Otmar Wiestler, MD and Thorsten Pietsch, MD (Bonn, Germany) and Ralf Ketter, MD to the first publication of the study. The study was supported by the Medical Faculty Heidelberg PostDoc Fellowship and the DKFZ Intramural Funding Program, Priority Topic Intratumoral Heterogeneity, to FS. BW is a scholar of the NCT Heidelberg School of Oncology Postdoc Program. We thank Tanja Göck and Viktoria Zeller for excellent technical assistance.

Supplementary material

401_2014_1370_MOESM1_ESM.xlsx (42 kb)
Supplementary material 1 (XLSX 42 kb)
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Supplementary material 2 (XLSX 10 kb)
401_2014_1370_MOESM3_ESM.pdf (27 kb)
Supplementary material 3 (PDF 26 kb)


  1. 1.
    Bady P, Sciuscio D, Diserens AC et al (2012) MGMT methylation analysis of glioblastoma on the Infinium methylation BeadChip identifies two distinct CpG regions associated with gene silencing and outcome, yielding a prediction model for comparisons across datasets, tumor grades, and CIMP-status. Acta Neuropathol 124:547–560PubMedCentralPubMedCrossRefGoogle Scholar
  2. 2.
    Balss J, Meyer J, Mueller W, Korshunov A, Hartmann C, von Deimling A (2008) Analysis of the IDH1 codon 132 mutation in brain tumors. Acta Neuropathol 116:597–602PubMedCrossRefGoogle Scholar
  3. 3.
    Bello MJ, Leone PE, Vaquero J, de Campos JM, Kusak ME, Sarasa JL, Pestana A, Rey JA (1995) Allelic loss at 1p and 19q frequently occurs in association and may represent early oncogenic events in oligodendroglial tumors. Int J Cancer 64:207–210PubMedCrossRefGoogle Scholar
  4. 4.
    Bello MJ, Vaquero J, de Campos JM, Kusak ME, Sarasa JL, Saez-Castresana J, Pestana A, Rey JA (1994) Molecular analysis of chromosome 1 abnormalities in human gliomas reveals frequent loss of 1p in oligodendroglial tumors. Int J Cancer 57:172–175PubMedCrossRefGoogle Scholar
  5. 5.
    Bigner SH, Mark J, Burger PC, Mahaley SM, Bullard DEJ, Muhlbaier LH, Bigner DD (1988) Specific chromosomal abnormalities in malignant human gliomas. Cancer Res 48:405–411PubMedGoogle Scholar
  6. 6.
    Bleeker FE, Lamba S, Leenstra S et al (2009) IDH1 mutations at residue p. R132 (IDH1(R132)) occur frequently in high-grade gliomas but not in other solid tumors. Hum Mutat 30:7–11PubMedCrossRefGoogle Scholar
  7. 7.
    Burger PC (2002) What is an oligodendroglioma? Brain Pathol 12:257–259PubMedCrossRefGoogle Scholar
  8. 8.
    Cairncross G, Wang M, Shaw E et al (2013) Phase III trial of chemoradiotherapy for anaplastic oligodendroglioma: long-term results of RTOG 9402. J Clin Oncol 31:337–343PubMedCentralPubMedCrossRefGoogle Scholar
  9. 9.
    Cairncross JG, Ueki K, Zlatescu MC et al (1998) Specific genetic predictors of chemotherapeutic response and survival in patients with anaplastic oligodendrogliomas. J Natl Cancer Inst 90:1473–1479PubMedCrossRefGoogle Scholar
  10. 10.
    Capper D, Sahm S, Hartmann C, Meyermann R, von Deimling A, Schittenhelm J (2010) Application of mutant IDH1 antibody to differentiate diffuse glioma from non-neoplastic central nervous system lesions and therapy induced changes. Am J Surg Pathol 34:1199–1204PubMedCrossRefGoogle Scholar
  11. 11.
    Capper D, Weißert S, Balss J et al (2010) Characterization of R132H mutation specific IDH1 antibody binding in brain tumors. Brain Pathol 20:245–254PubMedCrossRefGoogle Scholar
  12. 12.
    Capper D, Zentgraf H, Balss J, Hartmann C, von Deimling A (2009) Monoclonal antibody specific for IDH1 R132H mutation. Acta Neuropathol 118:599–601PubMedCrossRefGoogle Scholar
  13. 13.
    Coons SW, Johnson PC, Scheithauer BW, Yates AJ, Pearl DK (1997) Improving diagnostic accuracy and interobserver concordance in the classification and grading of primary gliomas. Cancer 79:1381–1393PubMedCrossRefGoogle Scholar
  14. 14.
    Fuller CE, Schmidt RE, Roth KA, Burger PC, Scheithauer BW, Banerjee R, Trinkaus K, Lytle R, Perry A (2003) Clinical utility of fluorescence in situ hybridization (FISH) in morphologically ambiguous gliomas with hybrid oligodendroglial/astrocytic features. J Neuropathol Exp Neurol 62:1118–1128PubMedGoogle Scholar
  15. 15.
    Griffin CA, Burger P, Morsberger L, Yonescu R, Swierczynski S, Weingart JD, Murphy KM (2006) Identification of der(1;19)(q10;p10) in five oligodendrogliomas suggests mechanism of concurrent 1p and 19q loss. J Neuropathol Exp Neurol 65:988–994PubMedCrossRefGoogle Scholar
  16. 16.
    Hartmann C, Hentschel B, Wick W et al (2010) Patients with IDH1 wild type anaplastic astrocytomas exhibit worse prognosis than IDH1 mutated glioblastomas and IDH1 mutation status accounts for the unfavorable prognostic effect of higher age: implications for classification of gliomas. Acta Neuropathol 120:707–718PubMedCrossRefGoogle Scholar
  17. 17.
    Hartmann C, Meyer J, Balss J et al (2009) Type and frequency of IDH1 and IDH2 mutations are related to astrocytic and oligodendroglial differentiation and age: a study of 1,010 diffuse gliomas. Acta Neuropathol 118:469–474PubMedCrossRefGoogle Scholar
  18. 18.
    Heaphy CM, de Wilde RF, Jiao Y et al (2011) Altered telomeres in tumors with ATRX and DAXX mutations. Science 333:425PubMedCentralPubMedCrossRefGoogle Scholar
  19. 19.
    Ichimura K, Pearson DM, Kocialkowski S, Backlund LM, Chan R, Jones DT, Collins VP (2009) IDH1 mutations are present in the majority of common adult gliomas but are rare in primary glioblastomas. Neuro Oncol 11:341–347PubMedCentralPubMedCrossRefGoogle Scholar
  20. 20.
    Idbaih A, Marie Y, Pierron G et al (2005) Two types of chromosome 1p losses with opposite significance in gliomas. Ann Neurol 58:483–487PubMedCrossRefGoogle Scholar
  21. 21.
    Jenkins RB, Blair H, Ballman KV et al (2006) A t(1;19)(q10;p10) mediates the combined deletions of 1p and 19q and predicts a better prognosis of patients with oligodendroglioma. Cancer Res 66:9852–9861PubMedCrossRefGoogle Scholar
  22. 22.
    Jiao Y, Killela PJ, Reitman ZJ et al (2012) Frequent ATRX, CIC, FUBP1 and IDH1 mutations refine the classification of malignant gliomas. Oncotarget 3:709–722PubMedCentralPubMedGoogle Scholar
  23. 23.
    Jiao Y, Shi C, Edil BH et al (2011) DAXX/ATRX, MEN1, and mTOR pathway genes are frequently altered in pancreatic neuroendocrine tumors. Science 331:1199–1203PubMedCentralPubMedCrossRefGoogle Scholar
  24. 24.
    Kannan K, Inagaki A, Silber J et al (2012) Whole-exome sequencing identifies ATRX mutation as a key molecular determinant in lower-grade glioma. Oncotarget 3:1194–1203PubMedCentralPubMedGoogle Scholar
  25. 25.
    Killela PJ, Reitman ZJ, Jiao Y et al (2013) TERT promoter mutations occur frequently in gliomas and a subset of tumors derived from cells with low rates of self-renewal. Proc Natl Acad Sci 110:6021–6026PubMedCentralPubMedCrossRefGoogle Scholar
  26. 26.
    Koelsche C, Sahm F, Capper D et al (2013) Distribution of TERT promoter mutations in pediatric and adult tumors of the nervous system. Acta Neuropathol 126:907–915PubMedCrossRefGoogle Scholar
  27. 27.
    Kraus JA, Koopmann J, Kaskel P, Maintz D, Brandner S, Louis DN, Wiestler OD, von Deimling A (1995) Shared allelic losses on chromosomes 1p and 19q suggest a common origin of oligodendroglioma and oligoastrocytoma. J Neuropathol Exp Neurol 54:91–95PubMedCrossRefGoogle Scholar
  28. 28.
    Liu XY, Gerges N, Korshunov A et al (2012) Frequent ATRX mutations and loss of expression in adult diffuse astrocytic tumors carrying IDH1/IDH2 and TP53 mutations. Acta Neuropathol 124:615–625PubMedCrossRefGoogle Scholar
  29. 29.
    Louis D, Ohgaki H, Wiestler O, Cavenee W (2007) World Health Organization Classification of Tumours of the Central Nervous System. In: Bosman F, Jaffe E, Lakhani S, Ohgaki H (eds) World Health Organization Classification of Tumours, 4 edn. IARC, LyonGoogle Scholar
  30. 30.
    Louis DN, Perry A, Burger P et al (2014) International Society of Neuropathology-Haarlem Consensus Guidelines, for Nervous System Tumor Classification and Grading. Brain Pathol 24:429–435PubMedCrossRefGoogle Scholar
  31. 31.
    Louis DN, von Deimling A, Seizinger BR (1992) A (CA)n dinucleotide repeat assay for evaluating loss of allelic heterozygosity in small and archival human brain tumor specimens. Am J Pathol 141:777–782PubMedCentralPubMedGoogle Scholar
  32. 32.
    Miller CR, Dunham CP, Scheithauer BW, Perry A (2006) Significance of necrosis in grading of oligodendroglial neoplasms: a clinicopathologic and genetic study of newly diagnosed high-grade gliomas. J Clin Oncol 24:5419–5426PubMedCrossRefGoogle Scholar
  33. 33.
    Mogensen U, Ishwaran H, Gerds TA (2012) Evaluating random forests for survival analysis using prediction error curves. J Stat Softw 50:1–23PubMedCentralPubMedGoogle Scholar
  34. 34.
    Noushmehr H, Weisenberger DJ, Diefes K et al (2010) Identification of a CpG island methylator phenotype that defines a distinct subgroup of glioma. Cancer Cell 17:510–522PubMedCentralPubMedCrossRefGoogle Scholar
  35. 35.
    Ozawa T, Riester M, Cheng YK, Huse JT, Squatrito M, Helmy K, Charles N, Michor F, Holland EC (2014) Most human non-GCIMP glioblastoma subtypes evolve from a common proneural-like precursor glioma. Cancer Cell 26:288–300PubMedCrossRefGoogle Scholar
  36. 36.
    Parsons DW, Jones S, Zhang X et al (2008) An integrated genomic analysis of human glioblastoma multiforme. Science 321:1807–1812PubMedCentralPubMedCrossRefGoogle Scholar
  37. 37.
    Perry A (2001) Oligodendroglial neoplasms: current concepts, misconceptions, and folklore. Adv Anat Pathol 8:183–199PubMedCrossRefGoogle Scholar
  38. 38.
    R-Core-Team (2014) R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria.
  39. 39.
    Reifenberger J, Reifenberger G, Liu L, James CD, Wechsler W, Collins VP (1994) Molecular genetic analysis of oligodendroglial tumors shows preferential allelic deletions on 19q and 1p. Am J Pathol 145:1175–1190PubMedCentralPubMedGoogle Scholar
  40. 40.
    Sahm F, Reuss D, Koelsche C et al (2014) Farewell to oligoastrocytoma: in situ molecular genetics favor classification as either oligodendroglioma or astrocytoma. Acta Neuropathol 128:551–559PubMedCrossRefGoogle Scholar
  41. 41.
    Sanson M, Marie Y, Paris S et al (2009) Isocitrate dehydrogenase 1 codon 132 mutation is an important prognostic biomarker in gliomas. J Clin Oncol 27:4150–4154PubMedCrossRefGoogle Scholar
  42. 42.
    Schwartzentruber J, Korshunov A, Liu X et al (2012) Driver mutations in histone H3.3 and chromatin remodelling genes in pediatric glioblastoma. Nature 482:226–231PubMedCrossRefGoogle Scholar
  43. 43.
    Smith JS, Perry A, Borell TJ et al (2000) Alterations of chromosome arms 1p and 19q as predictors of survival in oligodendrogliomas, astrocytomas, and mixed oligoastrocytomas. J Clin Oncol 18:636–645PubMedGoogle Scholar
  44. 44.
    Sturm D, Witt H, Hovestadt V et al (2012) Hotspot mutations in H3F3A and IDH1 define distinct epigenetic and biological subgroups of glioblastoma. Cancer Cell 22:425–437PubMedCrossRefGoogle Scholar
  45. 45.
    van den Bent MJ, Brandes AA, Taphoorn MJ et al (2013) Adjuvant procarbazine, lomustine, and vincristine chemotherapy in newly diagnosed anaplastic oligodendroglioma: long-term follow-up of EORTC brain tumor group study 26951. J Clin Oncol 31:344–350PubMedCrossRefGoogle Scholar
  46. 46.
    von Deimling A, Bender B, Jahnke R et al (1994) Loci associated with malignant progression in astrocytomas: a candidate on chromosome 19q. Cancer Res 54:1397–1401Google Scholar
  47. 47.
    von Deimling A, Louis DN, von Ammon K, Petersen I, Wiestler OD, Seizinger BR (1992) Evidence for a tumor suppressor gene on chromosome 19q associated with human astrocytomas, oligodendrogliomas and mixed gliomas. Cancer Res 52:4277–4279Google Scholar
  48. 48.
    Watanabe T, Nobusawa S, Kleihues P, Ohgaki H (2009) IDH1 Mutations are early events in the development of astrocytomas and oligodendrogliomas. Am J Pathol 174:653–656Google Scholar
  49. 49.
    Weller M, Felsberg J, Hartmann C et al (2009) Molecular predictors of progression-free and overall survival in patients with newly diagnosed glioblastoma: a prospective translational study of the German Glioma Network. J Clin Oncol 27:5743–5750PubMedCrossRefGoogle Scholar
  50. 50.
    Wick W, Hartmann C, Engel C et al (2009) NOA-04 randomized phase III trial of sequential radiochemotherapy of anaplastic glioma with procarbazine, lomustine, and vincristine or temozolomide. J Clin Oncol 27:5874–5880PubMedCrossRefGoogle Scholar
  51. 51.
    Wiestler B, Capper D, Holland-Letz T, Korshunov A, von Deimling A, Pfister S, Platten M, Weller M, Wick W (2013) ATRX loss refines the classification of anaplastic gliomas and identifies a subgroup of IDH mutant astrocytic tumors with better prognosis. Acta Neuropathol 126:443–451PubMedCrossRefGoogle Scholar
  52. 52.
    Wiestler B, Capper D, Hovestadt V et al (2014) Assessing CpG island methylator phenotype, 1p/19q codeletion, and MGMT promoter methylation from epigenome-wide data in the biomarker cohort of the NOA-04 trial. Neuro Oncol 16:1630–1638PubMedCrossRefGoogle Scholar
  53. 53.
    Wiestler B, Capper D, Sill M et al (2014) Integrated DNA methylation and copy-number profiling identify three clinically and biologically relevant groups of anaplastic glioma. Acta Neuropathol 128:561–571PubMedCrossRefGoogle Scholar
  54. 54.
    Xu W, Yang H, Liu Y et al (2011) Oncometabolite 2-hydroxyglutarate is a competitive inhibitor of alpha-ketoglutarate-dependent dioxygenases. Cancer Cell 19:17–30PubMedCentralPubMedCrossRefGoogle Scholar
  55. 55.
    Yan H, Parsons DW, Jin G et al (2009) IDH1 and IDH2 mutations in gliomas. N Engl J Med 360:765–773PubMedCentralPubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2014

Authors and Affiliations

  • David E. Reuss
    • 1
    • 2
  • Felix Sahm
    • 1
    • 2
  • Daniel Schrimpf
    • 1
    • 2
  • Benedikt Wiestler
    • 3
    • 4
  • David Capper
    • 1
    • 2
  • Christian Koelsche
    • 1
    • 2
  • Leonille Schweizer
    • 1
    • 2
  • Andrey Korshunov
    • 1
    • 2
  • David T. W. Jones
    • 5
  • Volker Hovestadt
    • 6
  • Michel Mittelbronn
    • 7
    • 8
  • Jens Schittenhelm
    • 9
  • Christel Herold-Mende
    • 10
  • Andreas Unterberg
    • 10
  • Michael Platten
    • 3
    • 11
  • Michael Weller
    • 12
  • Wolfgang Wick
    • 3
    • 4
  • Stefan M. Pfister
    • 5
    • 13
  • Andreas von Deimling
    • 1
    • 2
    Email author
  1. 1.German Cancer Consortium (DKTK), CCU Neuropathology, German Cancer Research Center (DKFZ)HeidelbergGermany
  2. 2.Department Neuropathology, Institute of PathologyUniversity of HeidelbergHeidelbergGermany
  3. 3.Neurology ClinicHeidelberg University HospitalHeidelbergGermany
  4. 4.Clinical Cooperation Unit NeurooncologyGerman Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ)HeidelbergGermany
  5. 5.Division of Pediatric NeurooncologyGerman Cancer Research Center (DKFZ)HeidelbergGermany
  6. 6.Division of Molecular GeneticsGerman Cancer Consortium (DKTK), German Cancer Research Center (DKFZ)HeidelbergGermany
  7. 7.Institute of Neurology (Edinger Institute)Goethe UniversityFrankfurtGermany
  8. 8.German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ)HeidelbergGermany
  9. 9.Department of Neuropathology, Institute of Pathology and NeuropathologyUniversity TuebingenTuebingenGermany
  10. 10.Department of NeurosurgeryUniversity of HeidelbergHeidelbergGermany
  11. 11.DKTK Clinical Cooperation Unit Neuroimmunology and Brain Tumor ImmunologyGerman Cancer Research CenterHeidelbergGermany
  12. 12.Department of NeurologyUniversity Hospital ZurichZurichSwitzerland
  13. 13.Department of Pediatric Oncology, Hematology and ImmunologyUniversity Medical CenterHeidelbergGermany

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