Application of a multi‐gene next‐generation sequencing panel to a non‐invasive oesophageal cell‐sampling device to diagnose dysplastic Barrett's oesophagus

Abstract

The early detection and endoscopic treatment of patients with the dysplastic stage of Barrett's oesophagus is a key to preventing progression to oesophageal adenocarcinoma. However, endoscopic surveillance protocols are hampered by the invasiveness of repeat endoscopy, sampling bias, and a subjective histopathological diagnosis of dysplasia. In this case‐control study, we investigated the use of a non‐invasive, pan‐oesophageal cell‐sampling device, the Cytosponge™, coupled with a cancer hot‐spot panel to identify patients with dysplastic Barrett's oesophagus. Formalin‐fixed, paraffin‐embedded (FFPE) Cytosponge™ samples from 31 patients with non‐dysplastic and 28 with dysplastic Barrett's oesophagus with good available clinical annotation were selected for inclusion. Samples were microdissected and amplicon sequencing performed using a panel covering >2800 COSMIC hot‐spot mutations in 50 oncogenes and tumour suppressor genes. Strict mutation criteria were determined and duplicates were run to confirm any mutations with an allele frequency <12%. When compared with endoscopy and biopsy as the gold standard the panel achieved a 71.4% sensitivity (95% CI 51.3–86.8) and 90.3% (95% CI 74.3–98.0) specificity for diagnosing dysplasia. TP53 had the highest rate of mutation in 14/28 dysplastic samples (50%). CDKN2A was mutated in 6/28 (21.4%), ERBB2 in 3/28 (10.7%), and 5 other genes at lower frequency. The only gene from this panel found to be mutated in the non‐dysplastic cases was CDKN2A in 3/31 cases (9.7%) in keeping with its known loss early in the natural history of the disease. Hence, it is possible to apply a multi‐gene cancer hot‐spot panel and next‐generation sequencing to microdissected, FFPE samples collected by the Cytosponge™, in order to distinguish non‐dysplastic from dysplastic Barrett's oesophagus. Further work is required to maximize the panel sensitivity.

Genetic Profiles of Barrett’s Esophagus and Esophageal Adenocarcinoma in Japanese patients

“…TP53 mutations are rarely detected in NDBE without concurrent dysplasia [27][28][29]. However, it is considered a progression risk if detected [30].…”

Expanding beyond endoscopy: A review of non-invasive modalities in Barrett’s esophagus screening and surveillance

“…As the string is pulled back, the Cytosponge collects cells from the lining of the entire esophagus and oropharynx. Although several biomarkers have been used with Cytosponge including a multi-gene next-generation sequencing panel, differentially methylated genes, and microRNAs, the most well-established biomarker used with Cytosponge has been trefoil factor family protein 3 (TFF3) immunohistochemical staining[ 35 - 37 ].…”

Screening for Barrett’s Oesophagus: Are We Ready for it?

“…Although samples collected via the Cytosponge can be assayed for several biomarkers including methylation [ 53 ], multigene next-generation sequencing panels [ 54 ], and microRNAs [ 55 ] useful for screening and surveillance, the current modus operandi for detecting BO from specimens obtained using this device is immunohistochemical staining of Trefoil Factor Protein 3 (TFF3) which is an indicator of intestinal metaplasia (the histopathological feature of BO). The slide-based technique permits the identification of gastric columnar cells as a quality control metric to ensure that the sampling device reached the stomach.…”