title: Genome-wide methylation profiling identifies an essential role of reactive oxygen species in pediatric glioblastoma multiforme and validates a methylome specific for H3 histone family 3A with absence of G-CIMP/isocitrate dehydrogenase 1 mutation.
abstract: BACKGROUND
  Pediatric glioblastoma multiforme (GBM) is rare, and there is a single study, a seminal discovery showing association of histone H3.3 and isocitrate dehydrogenase (IDH)1 mutation with a DNA methylation signature. The present study aims to validate these findings in an independent cohort of pediatric GBM, compare it with adult GBM, and evaluate the involvement of important functionally altered pathways.
  
  
  METHODS
  Genome-wide methylation profiling of 21 pediatric GBM cases was done and compared with adult GBM data (GSE22867). We performed gene mutation analysis of IDH1 and H3 histone family 3A (H3F3A), status evaluation of glioma cytosine-phosphate-guanine island methylator phenotype (G-CIMP), and Gene Ontology analysis. Experimental evaluation of reactive oxygen species (ROS) association was also done.
  
  
  RESULTS
  Distinct differences were noted between methylomes of pediatric and adult GBM. Pediatric GBM was characterized by 94 hypermethylated and 1206 hypomethylated cytosine-phosphate-guanine (CpG) islands, with 3 distinct clusters, having a trend to prognostic correlation. Interestingly, none of the pediatric GBM cases showed G-CIMP/IDH1 mutation. Gene Ontology analysis identified ROS association in pediatric GBM, which was experimentally validated. H3F3A mutants (36.4%; all K27M) harbored distinct methylomes and showed enrichment of processes related to neuronal development, differentiation, and cell-fate commitment.
  
  
  CONCLUSIONS
  Our study confirms that pediatric GBM has a distinct methylome compared with that of adults. Presence of distinct clusters and an H3F3A mutation-specific methylome indicate existence of epigenetic subgroups within pediatric GBM. Absence of IDH1/G-CIMP status further indicates that findings in adult GBM cannot be simply extrapolated to pediatric GBM and that there is a strong need for identification of separate prognostic markers. A possible role of ROS in pediatric GBM pathogenesis is demonstrated for the first time and needs further evaluation.
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences, New Delhi, India (P.J., P.P., VI.S., VA.S., M.C.S., C.S.); Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India (I.R.P.P., S.S., J.P., S.T., K.S.); Department of Neuropathology, National Institute of Mental Health and Neuro Sciences, Bangalore, India (VAI.S.); Department of Neurosurgery, National Institute of Mental Health and Neurosciences, Bangalore, India (A.A.); Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, India (A.S., D.G.).
authors.forename: Prerana
authors.initials: P
authors.lastname: Jha
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences, New Delhi, India (P.J., P.P., VI.S., VA.S., M.C.S., C.S.); Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India (I.R.P.P., S.S., J.P., S.T., K.S.); Department of Neuropathology, National Institute of Mental Health and Neuro Sciences, Bangalore, India (VAI.S.); Department of Neurosurgery, National Institute of Mental Health and Neurosciences, Bangalore, India (A.A.); Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, India (A.S., D.G.).
authors.forename: Irene Rosita
authors.initials: IR
authors.lastname: Pia Patric
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences, New Delhi, India (P.J., P.P., VI.S., VA.S., M.C.S., C.S.); Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India (I.R.P.P., S.S., J.P., S.T., K.S.); Department of Neuropathology, National Institute of Mental Health and Neuro Sciences, Bangalore, India (VAI.S.); Department of Neurosurgery, National Institute of Mental Health and Neurosciences, Bangalore, India (A.A.); Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, India (A.S., D.G.).
authors.forename: Sudhanshu
authors.initials: S
authors.lastname: Shukla
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences, New Delhi, India (P.J., P.P., VI.S., VA.S., M.C.S., C.S.); Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India (I.R.P.P., S.S., J.P., S.T., K.S.); Department of Neuropathology, National Institute of Mental Health and Neuro Sciences, Bangalore, India (VAI.S.); Department of Neurosurgery, National Institute of Mental Health and Neurosciences, Bangalore, India (A.A.); Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, India (A.S., D.G.).
authors.forename: Pankaj
authors.initials: P
authors.lastname: Pathak
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences, New Delhi, India (P.J., P.P., VI.S., VA.S., M.C.S., C.S.); Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India (I.R.P.P., S.S., J.P., S.T., K.S.); Department of Neuropathology, National Institute of Mental Health and Neuro Sciences, Bangalore, India (VAI.S.); Department of Neurosurgery, National Institute of Mental Health and Neurosciences, Bangalore, India (A.A.); Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, India (A.S., D.G.).
authors.forename: Jagriti
authors.initials: J
authors.lastname: Pal
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences, New Delhi, India (P.J., P.P., VI.S., VA.S., M.C.S., C.S.); Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India (I.R.P.P., S.S., J.P., S.T., K.S.); Department of Neuropathology, National Institute of Mental Health and Neuro Sciences, Bangalore, India (VAI.S.); Department of Neurosurgery, National Institute of Mental Health and Neurosciences, Bangalore, India (A.A.); Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, India (A.S., D.G.).
authors.forename: Vikas
authors.initials: V
authors.lastname: Sharma
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences, New Delhi, India (P.J., P.P., VI.S., VA.S., M.C.S., C.S.); Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India (I.R.P.P., S.S., J.P., S.T., K.S.); Department of Neuropathology, National Institute of Mental Health and Neuro Sciences, Bangalore, India (VAI.S.); Department of Neurosurgery, National Institute of Mental Health and Neurosciences, Bangalore, India (A.A.); Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, India (A.S., D.G.).
authors.forename: Sivaarumugam
authors.initials: S
authors.lastname: Thinagararanjan
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences, New Delhi, India (P.J., P.P., VI.S., VA.S., M.C.S., C.S.); Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India (I.R.P.P., S.S., J.P., S.T., K.S.); Department of Neuropathology, National Institute of Mental Health and Neuro Sciences, Bangalore, India (VAI.S.); Department of Neurosurgery, National Institute of Mental Health and Neurosciences, Bangalore, India (A.A.); Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, India (A.S., D.G.).
authors.forename: Vani
authors.initials: V
authors.lastname: Santosh
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences, New Delhi, India (P.J., P.P., VI.S., VA.S., M.C.S., C.S.); Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India (I.R.P.P., S.S., J.P., S.T., K.S.); Department of Neuropathology, National Institute of Mental Health and Neuro Sciences, Bangalore, India (VAI.S.); Department of Neurosurgery, National Institute of Mental Health and Neurosciences, Bangalore, India (A.A.); Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, India (A.S., D.G.).
authors.forename: Vaishali
authors.initials: V
authors.lastname: Suri
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences, New Delhi, India (P.J., P.P., VI.S., VA.S., M.C.S., C.S.); Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India (I.R.P.P., S.S., J.P., S.T., K.S.); Department of Neuropathology, National Institute of Mental Health and Neuro Sciences, Bangalore, India (VAI.S.); Department of Neurosurgery, National Institute of Mental Health and Neurosciences, Bangalore, India (A.A.); Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, India (A.S., D.G.).
authors.forename: Mehar Chand
authors.initials: MC
authors.lastname: Sharma
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences, New Delhi, India (P.J., P.P., VI.S., VA.S., M.C.S., C.S.); Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India (I.R.P.P., S.S., J.P., S.T., K.S.); Department of Neuropathology, National Institute of Mental Health and Neuro Sciences, Bangalore, India (VAI.S.); Department of Neurosurgery, National Institute of Mental Health and Neurosciences, Bangalore, India (A.A.); Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, India (A.S., D.G.).
authors.forename: Arimappamagan
authors.initials: A
authors.lastname: Arivazhagan
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences, New Delhi, India (P.J., P.P., VI.S., VA.S., M.C.S., C.S.); Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India (I.R.P.P., S.S., J.P., S.T., K.S.); Department of Neuropathology, National Institute of Mental Health and Neuro Sciences, Bangalore, India (VAI.S.); Department of Neurosurgery, National Institute of Mental Health and Neurosciences, Bangalore, India (A.A.); Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, India (A.S., D.G.).
authors.forename: Ashish
authors.initials: A
authors.lastname: Suri
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences, New Delhi, India (P.J., P.P., VI.S., VA.S., M.C.S., C.S.); Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India (I.R.P.P., S.S., J.P., S.T., K.S.); Department of Neuropathology, National Institute of Mental Health and Neuro Sciences, Bangalore, India (VAI.S.); Department of Neurosurgery, National Institute of Mental Health and Neurosciences, Bangalore, India (A.A.); Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, India (A.S., D.G.).
authors.forename: Deepak
authors.initials: D
authors.lastname: Gupta
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences, New Delhi, India (P.J., P.P., VI.S., VA.S., M.C.S., C.S.); Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India (I.R.P.P., S.S., J.P., S.T., K.S.); Department of Neuropathology, National Institute of Mental Health and Neuro Sciences, Bangalore, India (VAI.S.); Department of Neurosurgery, National Institute of Mental Health and Neurosciences, Bangalore, India (A.A.); Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, India (A.S., D.G.).
authors.forename: Kumaravel
authors.initials: K
authors.lastname: Somasundaram
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences, New Delhi, India (P.J., P.P., VI.S., VA.S., M.C.S., C.S.); Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India (I.R.P.P., S.S., J.P., S.T., K.S.); Department of Neuropathology, National Institute of Mental Health and Neuro Sciences, Bangalore, India (VAI.S.); Department of Neurosurgery, National Institute of Mental Health and Neurosciences, Bangalore, India (A.A.); Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, India (A.S., D.G.).
authors.forename: Chitra
authors.initials: C
authors.lastname: Sarkar
mesh_terms: D000328:Adult; D001932:Brain Neoplasms / Q000235:genetics*; D045744:Cell Line, Tumor; D002648:Child; D002675:Child, Preschool; D018899:CpG Islands; D019175:DNA Methylation; D005260:Female; D005909:Glioblastoma / Q000235:genetics*; D006657:Histones / Q000235:genetics*; D006801:Humans; D007521:Isocitrate Dehydrogenase / Q000235:genetics*; D008297:Male; D009154:Mutation; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D006657:Histones; D017382:Reactive Oxygen Species; D007521:Isocitrate Dehydrogenase; C543588:IDH1 protein, human
keywords: G-CIMP; H3F3A; IDH1; methylation; pediatric GBM; reactive oxygen species (ROS)