The origin of human metaplastic states and their propensity for cancer is poorly understood. Barrett’s esophagus is a common metaplastic condition that increases the risk for esophageal adenocarcinoma, and its cellular origin is enigmatic. To address this, we harvested tissues spanning the gastroesophageal junction from healthy and diseased donors, including isolation of esophageal submucosal glands. A combination of single-cell transcriptomic profiling, in silico lineage tracing from methylation, open chromatin and somatic mutation analyses, and functional studies in organoid models showed that Barrett’s esophagus originates from gastric cardia through c-MYC and HNF4A-driven transcriptional programs. Furthermore, our data indicate that esophageal adenocarcinoma likely arises from undifferentiated Barrett’s esophagus cell types even in the absence of a pathologically identifiable metaplastic precursor, illuminating early detection strategies.

Background & Aims-Esophageal adenocarcinomas (EAC) are heterogeneous and often preceded by Barrett's esophagus (BE). Many genomic changes have been associated with development of BE and EAC, but little is known about epigenetic alterations. We performed *

BACKGROUND Oncogenes are commonly amplified on extrachromosomal DNA (ecDNA) contributing to poor outcomes for patients. Currently, the chronology of ecDNA development is not known. We studied the origination and evolution of ecDNA in patients with Barrett's esophagus (BE) who progressed to esophageal adenocarcinoma (EAC). METHODS We analyzed whole-genome sequencing (WGS) data from a BE surveillance cohort and EAC patients at Cambridge University UK (n=206 patients). We also analyzed WGS data from biopsies taken at two time points from multiple sites in the esophagus from 80 patients enrolled in a case-control study at the Fred Hutchinson Cancer Center (FHCC) - 40 BE patients who progressed to EAC and 40 who did not. RESULTS ecDNA was detected in 24% and 43% of BE patients with BE-associated early and late-stage EAC, respectively, in the Cambridge cross-sectional cohort. ecDNA was found in 33% of all FHCC BE patients who developed cancer, either prior to, or at EAC diagnosis. ecDNA was strongly associated with patients who developed cancer, in contrast with FHCC BE patients who did not progress (odds ratio, 18.8, CI - 2.3-152, p=3.3x10-4). ecDNAs were enriched for oncogenes and immunomodulatory genes and could be detected early in the transition from high-grade dysplasia to cancer and increased in copy number and complexity over time. CONCLUSIONS ecDNAs can develop before a diagnosis of cancer in BE patients and are strongly selected for during the evolution to EAC. ecDNAs promote diverse oncogene and immunomodulatory gene amplification during EAC development and progression.

Oncogene amplification on extrachromosomal DNA (ecDNA) drives the evolution of tumours and their resistance to treatment, and is associated with poor outcomes for patients with cancer1–6. At present, it is unclear whether ecDNA is a later manifestation of genomic instability, or whether it can be an early event in the transition from dysplasia to cancer. Here, to better understand the development of ecDNA, we analysed whole-genome sequencing (WGS) data from patients with oesophageal ademocarcinoma (EAC) or Barrett’s oesophagus. These data included 206 biopsies in Barrett’s oesophagus surveillance and EAC cohorts from Cambridge University. We also analysed WGS and histology data from biopsies that were collected across multiple regions at 2 time points from 80 patients in a case–control study at the Fred Hutchinson Cancer Center. In the Cambridge cohorts, the frequency of ecDNA increased between Barrett’s-oesophagus-associated early-stage (24%) and late-stage (43%) EAC, suggesting that ecDNA is formed during cancer progression. In the cohort from the Fred Hutchinson Cancer Center, 33% of patients who developed EAC had at least one oesophageal biopsy with ecDNA before or at the diagnosis of EAC. In biopsies that were collected before cancer diagnosis, higher levels of ecDNA were present in samples from patients who later developed EAC than in samples from those who did not. We found that ecDNAs contained diverse collections of oncogenes and immunomodulatory genes. Furthermore, ecDNAs showed increases in copy number and structural complexity at more advanced stages of disease. Our findings show that ecDNA can develop early in the transition from high-grade dysplasia to cancer, and that ecDNAs progressively form and evolve under positive selection.

Barrett’s esophagus is a pre-malignant lesion that can progress to esophageal adenocarcinoma. We perform a multi-omic analysis of pre-cancer samples from 146 patients with a range of outcomes, comprising 642 person years of follow-up. Whole genome sequencing reveals complex structural variants and LINE-1 retrotransposons, as well as known copy number changes, occurring even prior to dysplasia. The structural variant burden captures the most variance across the cohort and genomic profiles do not always match consensus clinical pathology dysplasia grades. Increasing structural variant burden is associated with: high levels of chromothripsis and breakage-fusion-bridge events; increased expression of genes related to cell cycle checkpoint, DNA repair and chromosomal instability; and epigenetic silencing of Wnt signalling and cell cycle genes. Timing analysis reveals molecular events triggering genomic instability with more clonal expansion in dysplastic samples. Overall genomic complexity occurs early in the Barrett’s natural history and may inform the potential for cancer beyond the clinically discernible phenotype.

PurposeEvidence regarding whether or not antibiotic prophylaxis is beneficial in preventing post-operative surgical site infection in adult inguinal hernia repair is conflicting. A recent Cochrane review based on 17 randomised trials did not reach a conclusion on this subject. This study aimed to describe the current practice and determine whether clinical equipoise is prevalent.MethodsSurgeons in training were recruited to administer the Survey of Hernia Antibiotic Prophylaxis usE survey to consultant-level general surgeons in London and the south-east of England on their practices and beliefs regarding antibiotic prophylaxis in adult elective inguinal hernia repair. Local prophylaxis guidelines for the participating hospital sites were also determined.ResultsThe study was conducted at 34 different sites and received completed surveys from 229 out of a possible 245 surgeons, a 93 % response rate. Overall, a large majority of hospital guidelines (22/28) and surgeons’ personal beliefs (192/229, 84 %) supported the use of single-dose pre-operative intravenous antibiotic prophylaxis in inguinal hernia repair, although there was considerable variation in the regimens in use. The most widely used regimen was intravenous co-amoxiclav (1.2 g). Less than half of surgeons were adherent to their own hospital antibiotic guidelines for this procedure, although many incorrectly believed that they were following these.ConclusionIn the south-east of England, there is a strong majority of surgical opinion in favour of the use of antibiotic prophylaxis in this procedure. It is therefore likely to be extremely difficult to conduct further randomised studies in the UK to support or refute the effectiveness of prophylaxis in this commonly performed procedure.

This case highlights the importance of considering other causes of abdominal pain in a young man with normal initial investigations and the role of conservative management.

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.