title: Expression of DNA methyltransferases 1 and 3B correlates with EZH2 and this 3-marker epigenetic signature predicts outcome in glioblastomas.
abstract: This study aims to analyze expression of EZH2 and DNA-methyltransferases (DNMT1, 3A and 3B) in astrocytic tumors and investigate their link as well as their correlation with survival, especially in GBMs. Expression of EZH2 and DNMTs (DNMT1, DNMT3A and DNMT3B) in different grades of astrocytomas (n=93) was assessed by qRT-PCR and immunohistochemistry. GBM-U87MG cell line was used for functional studies. Strong immunopositivity (LI≥25%) for EZH2, DNMT1 and DNMT3B was detected in 52%, 56% and 64% cases of GBMs respectively, which was significantly higher as compared to Grade II/III cases. Similarly, their median fold change of mRNA expression was also significantly higher in GBMs. There was also a significant positive correlation between DNMT1/DNMT3B and EZH2 mRNA and protein expression, which was in concordance with TCGA data set. Inhibition of DNMTs in cell line by Azacytidine resulted in down-regulation of EZH2, while knock-down of EZH2 by siRNA was not associated with any significant alteration of DNMTs, indicating that EZH2 expression in GBMs is possibly regulated by DNMTs, but not the reverse. Strong immunopositivity for EZH2, DNMT1 and DNMT3B were individually associated with significantly shorter survival and showed no correlation with IDH1 mutation status. In addition, the combination of these 3 markers represented an independent prognostic signature with cases having weak/negative expression of all 3 markers being associated with best prognosis. For the first time, the present study describes an epigenetic prognostic signature in GBMs based on immunohistochemical expression of EZH2, DNMT1 and 3B which can be used easily in routine neuropathology practice.
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences (AIIMS), New Delhi, India.
authors.forename: Suvendu
authors.initials: S
authors.lastname: Purkait
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences (AIIMS), New Delhi, India.
authors.forename: Vikas
authors.initials: V
authors.lastname: Sharma
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences (AIIMS), New Delhi, India.
authors.forename: Anupam
authors.initials: A
authors.lastname: Kumar
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences (AIIMS), New Delhi, India.
authors.forename: Pankaj
authors.initials: P
authors.lastname: Pathak
authors.affiliation: Radiation Oncology, All India Institute of Medical Sciences (AIIMS), New Delhi, India.
authors.forename: Supriya
authors.initials: S
authors.lastname: Mallick
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences (AIIMS), New Delhi, India.
authors.forename: Prerana
authors.initials: P
authors.lastname: Jha
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences (AIIMS), New Delhi, India.
authors.forename: Mehar Chand
authors.initials: MC
authors.lastname: Sharma
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences (AIIMS), New Delhi, India.
authors.forename: Vaishali
authors.initials: V
authors.lastname: Suri
authors.affiliation: Radiation Oncology, All India Institute of Medical Sciences (AIIMS), New Delhi, India.
authors.forename: Pramod Kumar
authors.initials: PK
authors.lastname: Julka
authors.affiliation: Neurosurgery, All India Institute of Medical Sciences (AIIMS), New Delhi, India.
authors.forename: Ashish
authors.initials: A
authors.lastname: Suri
authors.affiliation: Neurosurgery, All India Institute of Medical Sciences (AIIMS), New Delhi, India.
authors.forename: B S
authors.initials: BS
authors.lastname: Sharma
authors.affiliation: Department of Pathology, All India Institute of Medical Sciences (AIIMS), New Delhi, India. Electronic address: sarkar.chitra@gmail.com.
authors.forename: Chitra
authors.initials: C
authors.lastname: Sarkar
mesh_terms: D001374:Azacitidine / Q000494:pharmacology; D045744:Cell Line, Tumor; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000235:genetics* / Q000378:metabolism; D000071221:Enhancer of Zeste Homolog 2 Protein; D004791:Enzyme Inhibitors / Q000494:pharmacology; D044127:Epigenesis, Genetic / Q000187:drug effects / Q000235:genetics; D005260:Female; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D005909:Glioblastoma / Q000235:genetics* / Q000378:metabolism / Q000628:therapy; D006801:Humans; D007150:Immunohistochemistry; D053208:Kaplan-Meier Estimate; D008297:Male; D015999:Multivariate Analysis; D017063:Outcome Assessment, Health Care / Q000379:methods / Q000706:statistics & numerical data; D063151:Polycomb Repressive Complex 2 / Q000235:genetics*; D011379:Prognosis; D016016:Proportional Hazards Models; D034622:RNA Interference; D012189:Retrospective Studies; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000097584:DNA Methyltransferase 3B
chemical_list: D001374:Azacitidine; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; D000071221:Enhancer of Zeste Homolog 2 Protein; D004791:Enzyme Inhibitors; D063151:Polycomb Repressive Complex 2; D000097584:DNA Methyltransferase 3B; C000619380:DNMT1 protein, human; C498409:EZH2 protein, human
keywords: DNMT1; DNMT3A; DNMT3B; EZH2; GBM; Survival