Caveolin-3 Null Mutation in Family with Barretts Esophagus and Esophageal Adenocarcinoma

INTRODUCTION: Aggregation of Barretts esophagus (BE) and esophageal adenocarcinoma (EAC) in families has been termed familial Barretts esophagus (FBE). Analysis of single, large FBE families can enable the identification of genetic susceptibility to complex diseases such as BE and EAC. METHODS: Phenotypes of BE and EAC were ascertained in a large FBE family with 7 affected members: 4 men with EAC, 1 man with BE and high grade dysplasia, and 2 women with non-dysplastic BE by review of endoscopy and surgical pathology reports. Whole exome sequencing was performed on germline DNA from 3 affected members to identify variants in coding genes that segregated with disease. Formalin fixed paraffin embedded tissue from an affected family member as well as non-familial subjects with BE and EAC was examined with regular histology and immunohistochemistry. The CAV3 gene with a variant segregating in the family was further characterized in a porcine model of esophageal injury using immunofluorescence. RESULTS: Using a whole exome sequencing approach on an exceptional FBE family we identified a segregating nonsense mutation in the gene Caveolin-3 (CAV3). Histologic examination of a formalin fixed paraffin embedded (FFPE) esophagectomy specimen from an individual carrying the CAV3 null mutation revealed esophageal submucosal glands (ESMG) that showed acinar metaplasia with marked atypia and absence of myoepithelial cells, distinctly different from acinar metaplasia seen in ESMG of non-familial subjects with BE and high grade dysplasia. Immunofluorescence studies of ESMG in porcine esophagus revealed the presence of CAV3 in selected cells in a distribution that was consistent with myoepithelial cells. Experimental injury of the porcine esophagus using radiofrequency ablation revealed that CAV3 expression increased markedly within ESMGs, ESMG ductal epithelium, and overlying healing neosquamous epithelium 10 days after injury. CONCLUSIONS: We theorize that CAV3 expression, perhaps through myoepithelial cells within ESMGs, controls the differentiation and proliferation of squamous epithelial precursor cells in response to injury. Furthermore, the truncating nonsense CAV3 mutation discovered in a family disrupts normal squamous healing and the organization of ESMGs, making affected family members susceptible to the proliferation and development of metaplastic columnar BE and EAC.

Origins of cancer genome complexity revealed by haplotype-resolved genomic analysis of evolution of Barretts esophagus to esophageal adenocarcinoma

Complex chromosomal alterations are a hallmark of advanced cancers but rarely seen in normal tissue. The progression of precancerous lesions to malignancy is often accompanied by increasing complexity of chromosomal alterations that can drive their transformation through focal oncogenic amplifications. However, the etiology and evolution dynamics of these alterations are poorly understood. Here we study chromosomal copy-number evolution in the progression of Barretts esophagus (BE) to esophageal adenocarcinoma (EAC) by multi-regional whole-genome sequencing analysis of BE samples with dysplasia and microscopic EAC foci. Through haplotype-specific copy-number analysis of BE genome evolution, we identified distinct patterns of episodic copy-number evolution consistent with the outcomes of abnormal mitosis and dicentric chromosome breakage. While abnormal mitosis, including whole-genome duplication, accounts for most chromosome or arm-level copy-number changes, segmental copy-number alterations display signatures of multi-generational evolution of unstable dicentric chromosomes. Continuous evolution of dicentric chromosomes through breakage-fusion-bridge cycles and chromothripsis rapidly increases genomic complexity and diversity among BE cells, culminating in the generation of distinct focal amplifications. These mutational processes enable multiple subclones within small dysplastic areas to undergo parallel transformation to cancer following acquisition of distinct oncogenic amplifications. Our results demonstrate how chromosomal instability drives clonal diversification in precancer evolution and promotes tumorigenesis in primary human samples.

Triage-driven diagnosis for early detection of esophageal cancer using deep learning

Deep learning methods have been shown to achieve excellent performance on diagnostic tasks, but it is still an open challenge how to optimally combine them with expert knowledge and existing clinical decision pathways. This question is particularly important for the early detection of cancer, where high volume workflows might potentially benefit substantially from automated analysis. Here, we present a deep learning framework to analyse samples of the Cytosponge(R)-TFF3 test, a minimally invasive alternative to endoscopy, for detecting Barretts Esophagus, the main precursor of esophageal cancer. We trained and independently validated the framework on data from two clinical trials, analysing a combined total of 4,662 pathology slides from 2,331 patients. Our approach exploits screening patterns of expert gastrointestinal16pathologists and established decision pathways to define eight triage classes of varying priority for manual expert review. By substituting manual review with automated review in low-priority classes, we can reduce pathologist workload by up to 66% while matching the diagnostic performance of expert pathologists. These results lay the foundation for tailored, semi-automated decision support systems embedded in clinical workflows.

Evaluation of a novel infra-red endoscopy system in the assessment of early neoplasia in Barretts esophagus: pilot study from a single center

SUMMARY Infrared endoscopy (IRE) has been shown to be useful in detecting submucosal (SM) invasion in early gastric cancer. Its role in the endoscopic assessment of Barrett's neoplasia has not been reported to date. We aimed in this study to evaluate the role of IRE in the detection and characterization of early neoplastic lesions within Barrett's esophagus (BE). The secondary aim was to explore its usefulness for the assessment of the presence of submucosal invasion in these early neoplastic Barrett's lesions. We included in the study patients with dysplastic BE who were referred to our institution for endoscopic therapy of a previously diagnosed early Barrett's neoplasia. An examination with white light high resolution endoscopy (HRE) and near IRE after intravenous injection of indocyanine green was performed for all patients using an infrared endoscope prototype. Staining on IRE and correlation with final histological staging by endoscopic mucosal resection/surgery or histological diagnosis on mapping biopsies was analyzed. A total of 23 patients were enrolled in our study: 17 of them with 19 visible lesions and 6 patients with flat BE and no lesions. Staining on IRE was noted in 18 cases: 17 (94%) had at least high grade dysplasia (HGD). No stain was noted in 7 cases: final histology was <HGD in 5 (71%) and ≥HGD in 2 (29%). There was statistically significant difference between cases with no stain and any staining on IRE with regard to the presence of ≥HGD [2/7 (29%) vs. 17/18 (94%) P = 0.0022]. Stain was reported as faint in 12 and dense in 6. All 6 cases with dense staining had at least HGD. We concluded that IRE can provide additional information to the currently available white light endoscopy for detecting early neoplastic lesions within BE. IRE also allows detecting HGD and most advanced histology in BE. Usefulness of IRE to detect submucosal involvement in early Barrett's neoplastic lesions needs to be assessed further in larger cohort studies.