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Subtypes of Barrett’s oesophagus and oesophageal adenocarcinoma based on genome-wide methylation analysis
  1. Ming Yu1,
  2. Sean K Maden1,
  3. Matthew Stachler2,3,
  4. Andrew M Kaz1,4,5,
  5. Jessica Ayers1,
  6. Yuna Guo1,
  7. Kelly T Carter1,
  8. Amber Willbanks1,
  9. Tai J Heinzerling1,
  10. Rachele M O’Leary1,
  11. Xinsen Xu3,
  12. Adam Bass3,6,
  13. Apoorva K Chandar7,8,
  14. Amitabh Chak7,9,10,
  15. Robin Elliott10,11,
  16. Joseph E Willis10,11,
  17. Sanford D Markowitz8,10,
  18. William M Grady1,4
  1. 1 Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, Washington, USA
  2. 2 Department of Pathology, Brigham and Women’s Hospital and Harvard Medical School, Boston, Massachusetts, USA
  3. 3 Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA
  4. 4 Department of Medicine, University of Washington School of Medicine, Seattle, Washington, USA
  5. 5 Gastroenterology Section, VA Puget Sound Health Care System, Seattle, Washington, USA
  6. 6 Eli and Edythe L Broad Institute, Cambridge, Massachusetts, USA
  7. 7 Division of Gastroenterology, Hospitals Cleveland Medical Center, Cleveland, Ohio, USA
  8. 8 Department of Medicine, Case Western Reserve University, Cleveland, Ohio, USA
  9. 9 Division of Oncology, University Hospitals Cleveland Medical Center, Cleveland, Ohio, USA
  10. 10 Case Comprehensive Cancer Center, Case Western Reserve University, Cleveland, Ohio, USA
  11. 11 Department of Pathology, University Hospitals Cleveland Medical Center, Cleveland, Ohio, USA
  1. Correspondence to Dr Ming Yu and Dr William M Grady, Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, Washington, USA; myu{at}, wgrady{at}


Objective To identify and characterise DNA methylation subtypes in oesophageal adenocarcinoma (EAC) and its precursor Barrett’s oesophagus (BE).

Design We performed genome-wide DNA methylation profiling on samples of non-dysplastic BE from cancer-free patients (n=59), EAC (n=23), normal squamous oesophagus (n=33) and normal fundus (n=9), and identified methylation subtypes using a recursively partitioned mixture model. We assessed genomic alterations for 9 BE and 22 EAC samples with massively parallel sequencing of 243 EAC-associated genes, and we conducted integrative analyses with transcriptome data to identify epigenetically repressed genes. We also carried out in vitro experiments treating EAC cell lines with 5-Aza-2'-Deoxycytidine (5-Aza-dC), short hairpin RNA knockdown and anticancer therapies.

Results We identified and validated four methylation subtypes of EAC and BE. The high methylator subtype (HM) of EAC had the greatest number of activating events in ERBB2 (p<0.05, Student’s t-test) and the highest global mutation load (p<0.05, Fisher’s exact test). PTPN13 was silenced by aberrant methylation in the HM subtype preferentially and in 57% of EACs overall. In EAC cell lines, 5-Aza-dC treatment restored PTPN13 expression and significantly decreased its promoter methylation in HM cell lines (p<0.05, Welch’s t-test). Inhibition of PTPN13 expression in the SK-GT-4 EAC cell line promoted proliferation, colony formation and migration, and increased phosphorylation in ERBB2/EGFR/Src kinase pathways. Finally, EAC cell lines showed subtype-specific responses to topotecan, SN-38 and palbociclib treatment.

Conclusions We identified and characterised methylator subtypes in BE and EAC. We further demonstrated the biological and clinical relevance of EAC methylator subtypes, which may ultimately help guide clinical management of patients with EAC.

  • dysplasia
  • gastrointestinal cancer
  • methylation

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  • MY, SKM and MS contributed equally.

  • Contributors Study concept and design: MY, WMG, AMK, AB, JEW, SDM. Data acquisition: MY, MS, AMK, JA, YG, KTC, AW, TJH, RMO, RE, JEW. Data analysis and interpretation: MY, SM, MS, WMG. Manuscript drafting: MY, SM, MS, WMG. Critical revision of the manuscript for important intellectual content: MY, SDM, MS, AB, AC, WMG. Funding support: MY, SDM, WMG. Reagents/materials/analysis support: XX, JEW, AC, SDM. Study supervision: MY, WMG.

  • Funding These studies were supported by funding from the NIH: UO1CA152756, 5P30CA015704, U54CA163060, UO1CA086402 and UO1CA182940 to WMG; P50CA150964 to SDM; and T32DK007742 to MY. Funding is also provided by the DeGregorio Family Foundation and the Price Family Foundation to WMG. This material is the result of work supported in part by resources from the VA Puget Sound Health Care System. The views expressed in this article are those of the authors and do not necessarily represent the views of the Department of Veterans Affairs.

  • Competing interests None declared.

  • Patient consent Next of kin consent obtained.

  • Ethics approval University Hospitals Cleveland Medical Center (Cleveland, Ohio) Review Board approval.

  • Provenance and peer review Not commissioned; externally peer reviewed.

  • Data sharing statement The HM450 array data will be deposited in GEO and accessible there. Anyone wanting other data is welcome to contact the corresponding author and make a request.

  • Author note Accession codes: HumanMethylation450 array data for 89 samples (n=23 EAC; n=59 BE from cancer-free patients; n=7 BE from EAC patients) have been deposited at the Gene Expression Omnibus under accession numbers GSE81334.