Abstract: BACKGROUND: Traditional cell culture models exhibit substantial limitations in accurately recapitulating the complex biological characteristics of tumors. In contrast, organoid, which maintains the essential biological features of primary tumors, offers a highly effective platform for comprehensive investigation into the pathogenesis of adenocarcinoma of the esophagogastric junction and the advancement of precision diagnostics and targeted therapeutic strategies.
OBJECTIVE: To Establish patient-derived organoid models of gastroesophageal junction adenocarcinoma, evaluate their fidelity in representing the biological characteristics of primary tumors, and provide a standardized in vitro model for investigating the pathogenesis of gastroesophageal junction adenocarcinoma and advancing precision medicine.
METHODS: Fifty surgical resection specimens were obtained from patients with adenocarcinoma of the esophagogastric junction and used for tissue dissociation, culture, and the establishment of organoid models. Cell viability in the constructed organoid models was evaluated using fluorescence co-localization staining. Hematoxylin-eosin staining was performed to compare structural morphological features between patient-derived organoid models of esophagogastric junction adenocarcinoma and their corresponding primary tumor tissues. Periodic Acid-Schiff staining was carried out to detect mucin-secreting phenotypes in the patient-derived organoid models. Immunohistochemistry was conducted to assess histopathological consistency between the organoid models and the original tumor tissues. The chi-square test and univariate logistic regression analysis were utilized to explore potential associations between the success rate of organoid model establishment and the clinicopathological characteristics as well as tissue features of the patients.
RESULTS AND CONCLUSION: In the patient-derived organoid models of esophagogastric junction adenocarcinoma established in this study, the primary culture success rate was 86.0% (43/50). The organoids exhibited a typical three-dimensional structure, progressively developing from cell aggregates into glandular lumen-like spherical formations. Stable subculturing was achieved within 7-10 days after primary culture, with no significant phenotypic changes observed across multiple passages. Furthermore, the organoids maintained a stable proliferative capacity after cryopreservation and subsequent thawing. Fluorescence co-localization staining confirmed that the patient-derived organoid models of esophagogastric junction adenocarcinoma maintained high cell viability across all stages of culture. Hematoxylin-eosin staining revealed that both the patient-derived organoid models and the corresponding primary tumor tissues exhibited dysplastic glandular architecture and an increased nuclear-to-cytoplasmic ratio. Periodic Acid-Schiff staining demonstrated consistent mucin secretion positivity. Immunohistochemistry analysis revealed a high degree of concordance in the positive expression rates of Ki67, CEA, CK7, and Cadherin17 between the organoids and the corresponding primary tumor tissues. Moreover, chi-square test and logistic regression analyses indicated that poorly differentiated tumor tissues (P=0.02), major pathological response to neoadjuvant chemotherapy in patients with adenocarcinoma of the esophagogastric junction (P=0.007), ex vivo duration of tumor tissues (P=0.006), and tumor tissue weight (P=0.006) were significantly associated with the success rate of organoid model establishment.

Key words: adenocarcinoma of the esophagogastric junction, primary culture, three-dimensional culture, organoids, model establishment, cell viability, histopathological analysis, tumor model