Introduction
Esophageal adenocarcinoma (EAC) typically develops through a metaplasia-dysplasia-carcinoma sequence driven by chronic bile/acid reflux. SerpinB3, a serine protease inhibitor, is expressed in nearly half of the lesions along this carcinogenic cascade [1] and has been implicated in tumor-promoting mechanisms in several organs [2-4]. However, its role at the esophageal squamocolumnar junction (SCJ) remains unclear.
Aims & Methods
We investigated the functional role of SerpinB3 in reflux-induced carcinogenesis. A validated murine model of gastro-jejunal anastomosis (GJA) was used to induce chronic bile/acid reflux in wild-type (WT; n=21) and SerpinB3-knockout (KO; n=15) Balb/C mice. A subgroup of WT mice (n=6) received 1-piperidine propionic acid (1-PPA; months 6–9) to inhibit PAR2, previously shown to downregulate SerpinB3 [5]. Animals were euthanised at 6 and 9 months and tissues analysed histologically. Transcriptomic profiling was performed on gastroesophageal tissues (6 GJA-WT vs. 6 GJA-KO) using the nCounter® PanCancer IO 360TM panel (NanoString Technologies), including time of sacrifice as covariate. Differential expression, gene set analysis (GSA), pathway and cell type scoring were applied. Additionally, esophageal mucosa samples from 18 treatment-naïve EAC patients were analysed for SerpinB3 expression (Real-Time RT-PCR) and immune markers (CD8, CD107a) by immunohistochemistry. Statistical significance was set at p<0.05; transcriptomic data were corrected for multiple testing using the Benjamini–Hochberg false discovery rate (FDR <0.05).
Results
At 9 months, SerpinB3 deficiency and pharmacological PAR2 inhibition (associated with SerpinB3 downregulation) were associated with a higher incidence of adenocarcinoma (p=0.022). At 6 months, KO mice showed increased lesion extension (p=0.036) and a trend toward more advanced histological stage (p=0.056). Transcriptomic analysis identified 33 nominally deregulated genes (p<0.05), without FDR-significant differences. GSA demonstrated coordinated downregulation of apoptosis, autophagy, antigen presentation, Notch and Hedgehog signalling. Pathway scoring revealed a marked reduction in mucosal activity in KO mice, with the strongest decreases in metabolic stress, MAPK and PI3K-Akt pathways. Immune cell profiling suggested a trend toward reduced T- and NK-cell infiltration in KO tissues. In human samples, low SerpinB3 expression was associated with decreased CD8+ T-cell infiltration and reduced cytotoxic activity (CD107a+).
Conclusion
SerpinB3 deficiency promotes reflux-induced esophageal carcinogenesis. Its loss is associated with a broadly suppressed mucosal transcriptional response, characterised by impaired metabolic stress responses, reduced activation of epithelial homeostatic pathways, and diminished immune effector function. These findings support a protective role of SerpinB3 in maintaining epithelial resilience and immunosurveillance under chronic reflux conditions. In this context, SerpinB3 deficiency may generate a permissive microenvironment for neoplastic progression. Notably, these data suggest a stage- and context-dependent role of SerpinB3, with protective effects during early reflux-induced mucosal injury, contrasting with previously reported tumor-promoting functions in established cancers [2-4]. Reduced SerpinB3 expression may represent a biomarker of increased risk for EAC progression.
References
- Fassan M, et al. Squamous cell carcinoma antigen (SCCA) is up-regulated during Barrett's carcinogenesis and predicts esophageal adenocarcinoma resistance to neoadjuvant chemotherapy. Oncotarget. 2017;8(15):24372-24379. doi: 10.18632/oncotarget.14108.
- Suminami Y, et al. Inhibition of apoptosis in human tumour cells by the tumour-associated serpin, SCC antigen-1. Br J Cancer. 2000;82(4):981-9.
- Quarta S, et al. SERPINB3 induces epithelial-mesenchymal transition. J Pathol. 2010;221(3):343-56. doi: 10.1002/path.2708.
- Turato C, et al. Squamous cell carcinoma antigen 1 is associated to poor prognosis in esophageal cancer through immune surveillance impairment and reduced chemosensitivity. Cancer Sci. 2019;110(5):1552-1563. doi: 10.1111/cas.13986.
- Villano G, et al. The Protease Activated Receptor 2 - CCAAT/Enhancer-Binding Protein beta - SerpinB3 axis inhibition as a novel strategy for the treatment of Non-Alcoholic Steatohepatitis. Mol Metab. (2024) 81:101889. doi: 10.1016/j.molmet.2024.101889.
Disclosure
No conflict of interest to declare