Abstract 5188: Recapitulating the initiation, progression and functional heterogeneity of Barrett’s esophagus using a comprehensive patient-derived organoid model platform

Abstract Barrett's esophagus (BE) is a common condition that increases the risk of esophageal adenocarcinoma (EAC). It originates from gastric cells at the gastroesophageal junction (GEJ) and can progress stepwise from BE to dysplasia and eventually to EAC. However, most BE cases do not progress, suggesting involvement of the tissue microenvironment and other extrinsic factors. BE is a mosaic tissue consisting of diverse clonal units of gastric and/or intestinal phenotypes, but the association between this heterogeneity and progression risk remains unclear. Further investigation is challenging due to a lack of human-based models. Here, we established a comprehensive organoid platform covering the full disease spectrum to study clonal and phenotypic heterogeneity, extrinsic factors, and other cellular components. Methods: We derived organoid cultures from endoscopic biopsies and organ donor tissues and developed a microscopic method to generate single-clonal organoids (sc-organoids) lines, serving as functional proxies of single clones in the original tissue. We comprehensively characterized the molecular features of all organoids and their parental tissues. We also established two models: an aggregated model that co-cultures esophageal fibroblasts with gastric/BE organoids under various pathological conditions; and a labeling-tracking model where BE/EAC organoids were genetically modified with fluorescent nuclear tags for live imaging and tracking. A mathematical model was developed to quantify organoid growth and morphological kinetics. Results: We established 48 organoid lines: 13 non-dysplastic BE, 7 dysplasia, 2 early EAC, and 26 gastric covering the full disease spectrum, recapitulating key molecular hallmarks of stepwise initiation, development, and progression. Transcriptomic analysis indicated that gastric cells at the GEJ are predisposed to BE development due to higher oncogenic stress. We developed 31 sc-organoids from 2 non-dysplastic BE cases and 1 dysplasia case, which captured phenotypic diversity and clonal heterogeneity with some displaying advanced-stage genomic features. The aggregated co-culture model revealed that BE cells promote polarization of fibroblasts into cancer-associated fibroblast-like phenotypes when exposed to refluxate, further enhancing BE's expansion. The labelling-tracking model showed increased BE organoid size and number when co-cultured with EAC, suggesting that cell competition and/or secreted factors alter the balance between precancerous lesions and established tumor. Conclusion: The organoid models provide a valuable platform to study mechanisms underlying precancerous initiation, progression, and neoplastic susceptibility in normal tissues, potentially informing risk stratification and new therapeutic strategies. Citation Format: Lizhe (John) Zhuang, Daniel H. Jacobson, Emily L. Black, Ginny Devonshire, Ahmad Miremadi, Massimiliano Di Pietro, Rebecca C. Fitzgerald. Recapitulating the initiation, progression and functional heterogeneity of Barrett’s esophagus using a comprehensive patient-derived organoid model platform [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5188.