title: Next-generation sequencing of endoscopic biopsies identifies ARID1A as a tumor-suppressor gene in Barrett's esophagus.
abstract: The incidence of Barrett's esophagus (BE)-associated esophageal adenocarcinoma (EAC) is increasing. Next-generation sequencing (NGS) provides an unprecedented opportunity to uncover genomic alterations during BE pathogenesis and progression to EAC, but treatment-naive surgical specimens are scarce. The objective of this study was to establish the feasibility of using widely available endoscopic mucosal biopsies for successful NGS, using samples obtained from a BE 'progressor'. Paired-end whole-genome NGS was performed on the Illumina platform using libraries generated from mucosal biopsies of normal squamous epithelium (NSE), BE and EAC obtained from a patient who progressed to adenocarcinoma during endoscopic surveillance. Selective validation studies, including Sanger sequencing, immunohistochemistry and functional assays, were performed to confirm the NGS findings. NGS identified somatic nonsense mutations of AT-rich interactive domain 1A (SWI like) (ARID1A) and PPIE and an additional 37 missense mutations in BE and/or EAC, which were confirmed by Sanger sequencing. ARID1A mutations were detected in 15% (3/20) high-grade dysplasia (HGD)/EAC patients. Immunohistochemistry performed on an independent archival cohort demonstrated ARID1A protein loss in 0% (0/76), 4.9% (2/40), 14.3% (4/28), 16.0% (8/50) and 12.2% (12/98) of NSE, BE, low-grade dysplasia, HGD and EAC tissues, respectively, and was inversely associated with nuclear p53 accumulation (P=0.028). Enhanced cell growth, proliferation and invasion were observed on ARID1A knockdown in EAC cells. In addition, genes downstream of ARID1A that potentially contribute to the ARID1A knockdown phenotype were identified. Our studies establish the feasibility of using mucosal biopsies for NGS, which should enable the comparative analysis of larger 'progressor' versus 'non-progressor' cohorts. Further, we identify ARID1A as a novel tumor-suppressor gene in BE pathogenesis, reiterating the importance of aberrant chromatin in the metaplasia-dysplasia sequence.
authors: 1] Department of Pathology, John Hopkins Medical Institutions, Baltimore, MD, USA [2] Department of Gastroenterology and Hepatology, University Medical Center Utrecht, Utrecht, The Netherlands [3] Department of Pathology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors: M M
authors: MM
authors: Streppel
authors: Stanley Institute for Cognitive Genomics, Cold Spring Harbor Laboratory, Woodbury, NY, USA.
authors: S
authors: S
authors: Lata
authors: Stanley Institute for Cognitive Genomics, Cold Spring Harbor Laboratory, Woodbury, NY, USA.
authors: M
authors: M
authors: DelaBastide
authors: Department of Pathology, John Hopkins Medical Institutions, Baltimore, MD, USA.
authors: E A
authors: EA
authors: Montgomery
authors: Division of Gastroenterology, Department of Medicine, Washington University School of Medicine, St Louis, MO, USA.
authors: J S
authors: JS
authors: Wang
authors: Department of Medicine (Division of Gastroenterology and Hepatology), John Hopkins Medical Institutions, Baltimore, MD, USA.
authors: M I
authors: MI
authors: Canto
authors: 1] Department of Pathology, John Hopkins Medical Institutions, Baltimore, MD, USA [2] Pathobiology Program, John Hopkins Medical Institutions, Baltimore, MD, USA.
authors: A M
authors: AM
authors: Macgregor-Das
authors: Department of Pathology, John Hopkins Medical Institutions, Baltimore, MD, USA.
authors: S
authors: S
authors: Pai
authors: Department of Pathology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors: F H M
authors: FH
authors: Morsink
authors: Department of Pathology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors: G J
authors: GJ
authors: Offerhaus
authors: Stanley Institute for Cognitive Genomics, Cold Spring Harbor Laboratory, Woodbury, NY, USA.
authors: E
authors: E
authors: Antoniou
authors: 1] Department of Pathology, John Hopkins Medical Institutions, Baltimore, MD, USA [2] Department of Oncology, John Hopkins Medical Institutions, Baltimore, MD, USA [3] Department of Genetic Medicine, John Hopkins Medical Institution, Baltimore, MD, USA.
authors: A
authors: A
authors: Maitra
authors: Stanley Institute for Cognitive Genomics, Cold Spring Harbor Laboratory, Woodbury, NY, USA.
authors: W R
authors: WR
authors: McCombie
mesh_terms: D000230:Adenocarcinoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D000368:Aged; D001471:Barrett Esophagus / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D001706:Biopsy; D015153:Blotting, Western; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D004268:DNA-Binding Proteins; D019723:Endoscopes; D004848:Epithelium / Q000378:metabolism / Q000473:pathology; D004938:Esophageal Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D005260:Female; D053263:Gene Regulatory Networks; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D008875:Middle Aged; D020125:Mutation, Missense; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D020641:Polymorphism, Single Nucleotide; D034622:RNA Interference; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D059467:Transcriptome; D025521:Tumor Suppressor Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: C112476:ARID1A protein, human; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins