title: Identification of retinol binding protein 1 promoter hypermethylation in isocitrate dehydrogenase 1 and 2 mutant gliomas.
abstract: BACKGROUND
  Mutations in isocitrate dehydrogenase 1 (IDH1) and associated CpG island hypermethylation represent early events in the development of low-grade gliomas and secondary glioblastomas. To identify candidate tumor suppressor genes whose promoter methylation may contribute to gliomagenesis, we compared methylation profiles of IDH1 mutant (MUT) and IDH1 wild-type (WT) tumors using massively parallel reduced representation bisulfite sequencing.
  
  
  METHODS
  Reduced representation bisulfite sequencing was performed on ten pathologically matched WT and MUT glioma samples and compared with data from a methylation-sensitive restriction enzyme technique and data from The Cancer Genome Atlas (TCGA). Methylation in the gene retinol-binding protein 1 (RBP1) was identified in IDH1 mutant tumors and further analyzed with primer-based bisulfite sequencing. Correlation between IDH1/IDH2 mutation status and RBP1 methylation was evaluated with Spearman correlation. Survival data were collected retrospectively and analyzed with Kaplan-Meier and Cox proportional hazards analysis. All statistical tests were two-sided.
  
  
  RESULTS
  Methylome analysis identified coordinated CpG island hypermethylation in IDH1 MUT gliomas, consistent with previous reports. RBP1, important in retinoic acid metabolism, was found to be hypermethylated in 76 of 79 IDH1 MUT, 3 of 3 IDH2 MUT, and 0 of 116 IDH1/IDH2 WT tumors. IDH1/IDH2 mutation was highly correlated with RBP1 hypermethylation (n = 198; Spearman R = 0.94, 95% confidence interval = 0.92 to 0.95, P < .001). The Cancer Genome Atlas showed IDH1 MUT tumors (n = 23) to be RBP1-hypermethylated with decreased RBP1 expression compared with WT tumors (n = 124). Among patients with primary glioblastoma, patients with RBP1-unmethylated tumors (n = 102) had decreased median overall survival compared with patients with RBP1-methylated tumors (n = 22) (20.3 months vs 36.8 months, respectively; hazard ratio of death = 2.48, 95% confidence interval = 1.30 to 4.75, P = .006).
  
  
  CONCLUSIONS
  RBP1 promoter hypermethylation is found in nearly all IDH1 and IDH2 mutant gliomas and is associated with improved patient survival. Because RBP1 is involved in retinoic acid synthesis, our results suggest that dysregulation of retinoic acid metabolism may contribute to glioma formation along the IDH1/IDH2-mutant pathway.
authors.affiliation: Department of Neurology, David Geffen School of Medicine at the University of California Los Angeles, 90095, USA.
authors.forename: Arthur P
authors.initials: AP
authors.lastname: Chou
authors.forename: Reshmi
authors.initials: R
authors.lastname: Chowdhury
authors.forename: Sichen
authors.initials: S
authors.lastname: Li
authors.forename: Weidong
authors.initials: W
authors.lastname: Chen
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Kim
authors.forename: David E
authors.initials: DE
authors.lastname: Piccioni
authors.forename: Julia M
authors.initials: JM
authors.lastname: Selfridge
authors.forename: Reema R
authors.initials: RR
authors.lastname: Mody
authors.forename: Stephen
authors.initials: S
authors.lastname: Chang
authors.forename: Shadi
authors.initials: S
authors.lastname: Lalezari
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Lin
authors.forename: Desiree E
authors.initials: DE
authors.lastname: Sanchez
authors.forename: Ryan W
authors.initials: RW
authors.lastname: Wilson
authors.forename: Matthew C
authors.initials: MC
authors.lastname: Garrett
authors.forename: Bret
authors.initials: B
authors.lastname: Harry
authors.forename: Jack
authors.initials: J
authors.lastname: Mottahedeh
authors.forename: Phioanh L
authors.initials: PL
authors.lastname: Nghiemphu
authors.forename: Harley I
authors.initials: HI
authors.lastname: Kornblum
authors.forename: Paul S
authors.initials: PS
authors.lastname: Mischel
authors.forename: Robert M
authors.initials: RM
authors.lastname: Prins
authors.forename: William H
authors.initials: WH
authors.lastname: Yong
authors.forename: Timothy
authors.initials: T
authors.lastname: Cloughesy
authors.forename: Stanley F
authors.initials: SF
authors.lastname: Nelson
authors.forename: Linda M
authors.initials: LM
authors.lastname: Liau
authors.forename: Albert
authors.initials: A
authors.lastname: Lai
mesh_terms: D000328:Adult; D000368:Aged; D014408:Biomarkers, Tumor / Q000235:genetics*; D015153:Blotting, Western; D001932:Brain Neoplasms / Q000737:chemistry / Q000235:genetics*; D018899:CpG Islands / Q000235:genetics; D019175:DNA Methylation; D004252:DNA Mutational Analysis / Q000379:methods; D005260:Female; D020869:Gene Expression Profiling; D005910:Glioma / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D007521:Isocitrate Dehydrogenase / Q000235:genetics*; D053208:Kaplan-Meier Estimate; D008297:Male; D008875:Middle Aged; D009154:Mutation; D011401:Promoter Regions, Genetic / Q000235:genetics; D016016:Proportional Hazards Models; D060888:Real-Time Polymerase Chain Reaction; D015183:Restriction Mapping; D054840:Retinol-Binding Proteins, Cellular / Q000032:analysis / Q000235:genetics*; D012189:Retrospective Studies; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013447:Sulfites / Q000378:metabolism; D014212:Tretinoin / Q000378:metabolism
chemical_list: D014408:Biomarkers, Tumor; C518456:RBP1 protein, human; D054840:Retinol-Binding Proteins, Cellular; D013447:Sulfites; D014212:Tretinoin; C538784:IDH2 protein, human; D007521:Isocitrate Dehydrogenase; C543588:IDH1 protein, human; C042345:hydrogen sulfite