Abstract 2780: Genome-wide methylation analysis reveals methylator subtypes of Barrett's esophagus and esophageal adenocarcinoma
Authors: Yu Ming , Maden Sean , Kaz Andrew , Heinzerling Tai , O’Leary Rachele , Stachler Matthew , Bass Adam , Chak Amitabh , Willis Joseph , Markowitz Sanford , Grady William 
Published: July 2016

Abstract
Esophageal adenocarcinoma (EAC) arises from Barrett's Esophagus (BE) through a progression sequence driven by the accumulation of epi/genetic alterations. Genetic alterations and aberrantly methylated genes/loci, the best-characterized epigenetic alteration, vary among individual BE and EAC cases. To better understand the molecular heterogeneity in BE/ EAC, we analyzed genome-wide DNA methylation patterns in BE (N = 98), EAC (N = 23), and normal esophagus and stomach (N = 78) collected via Esophagus Translational Research Network (BETRNet) using Human Methylation 450K (HM450K) arrays. We identified methylation-based subtypes using a model-based recursively partitioned mixture model and unsupervised resampling-based hierarchical cluster analysis with the most variable probes based on standard deviation. All analyses were performed in R. Four methylator subtypes were identified in the BE/EAC dataset using our clustering approach: high (HM); intermediate (IM); low (LM); and minimal (MM). The clusters showed widespread methylation differences in CpGs primarily located in CpG islands (72% of 414 total probes). Each methylation subtype showed distinct genetic and clinical features suggesting underlying biological differences (Table 1). HM exhibited methylation patterns reminiscent of a CpG Island Methylator Phenotype (CIMP). Among BE/EAC cases with known TP53 mutation status, the majority of mutants were among HM and IM clusters (15/19, 78.9%), Finally, identification of methylation subtypes in an independent HM450K dataset of EAC samples (N = 89) from the Cancer Genome Atlas validated the existence of the BE/EAC methylation subtypes. In summary, our data suggest genome wide alterations in DNA methylation occur in the BE stage of the BE-to-EAC progression. We also identified four distinct methylation subtypes of BE/EAC, including a CIMP-like HM subtype, and found more TP53 mutants in the HM and IM subtypes, revealing molecular heterogeneity in different methylation subtypes. Table 1.Characteristics of primary BE/EAC dataset and subtypes from cluster analyses.Cluster CharacteristicSubtypesHigh N (%)Intermediate N (%)Low N (%)Minimal N (%)Total N (%)Case CountsTotal31 (26%)44 (36%)28 (23%)18 (23%)121 (100%)BE23 (23%)34 (35%)24 (24%)17 (17%)98 (100%)EAC8 (35%)10 (43%)4 (17%)1 (4%)23 (100%)Demographic VariablesGenderM28 (28%)35 (35%)23 (23%)14 (14%)100 (100%)GenderF3 (14%)9 (43%)5 (24%)4 (19%)21 (100%)Age (mean ± sd)68 ± 1163 ± 1361 ± 1461 ± 1064 ± 13TP53 Mutations***Mutation*6 (32%)9 (47%)4 (21%)0 (0%)19 (100%)WT6 (30%)8 (40%)5 (25%)1 (5%)20 (100%)Unknown1927191782*** Mutation categories included: Frame Shift Deletion; In Frame Insertion; In Frame Deletion; Missense Mutation; Nonsense Mutation; and Splice Site
Citation Format: Ming Yu, Sean Maden, Andrew Kaz, Tai Heinzerling, Rachele O’Leary, Matthew Stachler, Adam Bass, Amitabh Chak, Joseph Willis, Sanford Markowitz, William Grady. Genome-wide methylation analysis reveals methylator subtypes of Barrett's esophagus and esophageal adenocarcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2780.