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Specific detection of methionine 27 mutation in histone 3 variants (H3K27M) in fixed tissue from high-grade astrocytomas |
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| Authors: | Denise Bechet Gerrit G H Gielen Andrey Korshunov Stefan M Pfister Caterina Rousso Damien Faury Pierre-Olivier Fiset Naciba Benlimane Peter W Lewis Chao Lu C David Allis Mark W Kieran Keith L Ligon Torsten Pietsch Benjamin Ellezam Steffen Albrecht Nada Jabado |
| Institution: | 1. Departments of Experimental Medicine and of Human Genetics, McGill University, 4060 Ste Catherine West, PT239, Montreal, QC, H3Z2Z3, Canada 2. Institute of Neuropathology, University of Bonn Medical Center, Sigmund-Freud-Strasse 25, 53127, Bonn, Germany 3. Clinical Cooperation Unit Neuropathology, German Cancer Research Center, Heidelberg, Germany 4. Division of Pediatric Neurooncology, German Cancer Research Center (DKFZ), Heidelberg, Germany 5. Department of Pediatrics, Montreal Children’s Hospital, McGill University Health Centre, Montreal, QC, Canada 6. Department of Pathology, Montreal Children’s Hospital, McGill University Health Centre, Montreal, QC, Canada 7. Research Pathology Facility, MCETC, Jewish General Hospital, Montreal, QC, Canada 8. Department of Biomolecular Chemistry, School of Medicine and Public Health, University of Wisconsin, Madison, WI, USA 9. Laboratory of Chromatin Biology and Epigenetics, The Rockefeller University, New York, NY, USA 10. Division of Pediatric Hematology/Oncology, Department of Pediatric Oncology, Dana-Farber Cancer Institute, Boston Children’s Hospital, Harvard Medical School, Boston, MA, USA 11. Center for Molecular Oncologic Pathology, Dana-Farber Cancer Institute, Harvard University, Boston, MA, USA 12. Department of Pathology, CHU Ste-Justine, Université de Montréal, Montreal, QC, Canada 14. Hopital Ste Justine, Montreal, QC, Canada 13. Montreal Children’s Hospital, Montreal, QC, H3H1P3, Canada |
| Abstract: | Studies in pediatric high-grade astrocytomas (HGA) by our group and others have uncovered recurrent somatic mutations affecting highly conserved residues in histone 3 (H3) variants. One of these mutations leads to analogous p.Lys27Met (K27M) mutations in both H3.3 and H3.1 variants, is associated with rapid fatal outcome, and occurs specifically in HGA of the midline in children and young adults. This includes diffuse intrinsic pontine gliomas (80 %) and thalamic or spinal HGA (>90 %), which are surgically challenging locations with often limited tumor material available and critical need for specific histopathological markers. Here, we analyzed formalin-fixed paraffin-embedded tissues from 143 pediatric HGA and 297 other primary brain tumors or normal brain. Immunohistochemical staining for H3K27M was compared to tumor genotype, and also compared to H3 tri-methylated lysine 27 (H3K27me3) staining, previously shown to be drastically decreased in samples carrying this mutation. There was a 100 % concordance between genotype and immunohistochemical analysis of H3K27M in tumor samples. Mutant H3K27M was expressed in the majority of tumor cells, indicating limited intra-tumor heterogeneity for this specific mutation within the limits of our dataset. Both H3.1 and H3.3K27M mutants were recognized by this antibody while non-neoplastic elements, such as endothelial and vascular smooth muscle cells or lymphocytes, did not stain. H3K27me3 immunoreactivity was largely mutually exclusive with H3K27M positivity. These results demonstrate that mutant H3K27M can be specifically identified with high specificity and sensitivity using an H3K27M antibody and immunohistochemistry. Use of this antibody in the clinical setting will prove very useful for diagnosis, especially in the context of small biopsies in challenging midline tumors and will help orient care in the context of the extremely poor prognosis associated with this mutation. |
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