Abstract 2204: Probiotic <i>Lactobacilli</i> and their secreted metabolites exert anti-inflammatory and anti-genotoxic effects in Barrett’s esophagus associated tumorigenesis <i>in vitro</i> and <i>in vivo</i>

Abstract Gastroesophageal reflux disease (GERD) leads to the accumulation of bile-induced reactive oxygen species and oxidative stress in esophageal tissues, causing inflammation and DNA damage. As GERD progresses to Barrett’s esophagus (BE), the only known precursor lesion for esophageal adenocarcinoma, probiotic bacteria with anti-inflammatory and anti-genotoxic function are of significant interest in the prevention and treatment of Barrett’s esophagus (BE) and esophageal adenocarcinoma. To assess the outcome of probiotic Lactobacillus administration in vivo, we used the model of L2-driven expression of IL1b for inflammation-associated Barrett’s tumorigenesis. We administered 109 CFU/ml (colony-forming unit) L. acidophilus and L. plantarum by oral gavage starting at 6 weeks of age until the IL-1β mice were 6 months old (prevention model) or initiated treatments when the mice were 4 months old until 6 months of age (treatment group). At the end of the study, mice were sacrificed, and esophageal and forestomach tissues were harvested for DNA and RNA extraction and FFPE. Histopathological analysis following Lactobacillus spp. administration in the murine study showed a significant reduction in inflammation (p&amp;lt;0.07) and tumor incidence (p&amp;lt;0.05) compared to untreated sham mice in the prevention and treatment groups. Assessing mucin production at the GEJ visualized by Alcian Blue/PAS, we observed 80% of the sham mice had extended glands compared to significantly less when Lactobacillus spp. was administered (p&amp;lt;0.03).Multiplex immunofluorescence staining was used to evaluate immune cell recruitment and BE marker expression. In vitro assays utilized immortalized normal esophageal cells (STR), and the BE cell lines, CP-B and BAR-T and cell-free Lactobacillus supernatant. Upon DNA damage induction using deoxycholic acid or ox bile, cell lines were incubated with live Lactobacilli or cell-free supernatant secreted by L. fermentum, L. acidophilus and L. plantarum. To identify if inflammation and DNA damage can also be reduced using potential Lactobacillus-secreted bioactive metabolites, we detected 8-oxo-dG, pH2AX and RAD51 in vitro using immunofluorescence. We show a reduction in DNA damage and NF-KB-mediated inflammation in the presence of Lactobacillus cell-free supernatants similar to the effect of live bacteria.Our data demonstrate the feasibility of utilizing probiotic Lactobacillus supplementation in mouse models of BE, as well as support the hypothesis that Lactobacillus spp. aid in the prevention of disease progression and its treatment in experimental models of gastroesophageal reflux disease. Citation Format: Maanya Pradeep, Divya Joshi, Joshua N. Bernard, Thomas Andl, Claudia D. Andl. Probiotic Lactobacilli and their secreted metabolites exert anti-inflammatory and anti-genotoxic effects in Barrett’s esophagus associated tumorigenesis in vitro and in vivo [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 2204.