Identifying DNA methylation biomarkers for non-endoscopic detection of Barrett’s esophagus

We report a biomarker-based non-endoscopic method for detecting Barrett’s esophagus (BE), based on detecting methylated DNAs retrieved via a swallowable balloon-based esophageal sampling device. BE is the precursor of, and a major recognized risk factor for, developing esophageal adenocarcinoma (EAC). Endoscopy, the current standard for BE detection, is not cost-effective for population screening. We performed genome-wide screening to ascertain regions targeted for recurrent aberrant cytosine methylation in BE, identifying high-frequency methylation within the CCNA1 locus. We tested CCNA1 DNA methylation as a BE biomarker in cytology brushings of the distal esophagus from 173 individuals with or without BE. CCNA1 DNA methylation demonstrated an area under the curve (AUC)=0.95 for discriminating BE-related metaplasia and neoplasia cases versus normal individuals, performing identically to methylation of VIM DNA, an established BE biomarker. When combined, the resulting two biomarker panel was 95% sensitive and 91% specific. These results were replicated in an independent validation cohort of 149 individuals, who were assayed using the same cutoff values for test positivity established in the training population. To progress toward non-endoscopic esophageal screening, we engineered a well-tolerated, swallowable, encapsulated balloon device able to selectively sample the distal esophagus within 5 minutes. In balloon samples from 86 individuals, tests of CCNA1 plus VIM DNA methylation detected BE metaplasia with 90.3% sensitivity and 91.7% specificity. Combining the balloon sampling device with molecular assays of CCNA1 plus VIM DNA methylation enables an efficient, well-tolerated, sensitive, and specific method of screening at-risk populations for BE

GNAS Codon 201 Mutations.

<p>Representative chromatograms (left) and pyrograms (right) of GNAS wild-type, GNAS R201C, and GNAS R201H mutations detected in colon cancer. Arrows in chromatograms show mutant, heterozygous peaks. Boxed peaks in pyrograms highlight mutant alleles. Percentages indicate allele frequencies calculated from pyrogram peak intensities.</p

GNAS tumors are associated with a villous morphology.

<p>H&E Photomicrograph of a GNAS mutant, villous adenocarcinoma. The tumor has typical villous morphology with protruding papillae containing a fibrovascular core and lined with adenomatous epithelium.</p

Characteristics of GNAS mutant adenomas.

<p>Abbreviations: TVA, tubulovillous adenoma; VA, villous adenom.</p