Background: The incidence of esophageal adenocarcinoma (EAC) is rapidly increasing in Western countries. Most EACs originate from Barrett’s esophagus (BE), a precancerous lesion initiated by chronic reflux of gastrointestinal contents, especially acidic bile salts (ABS). Several chemotherapeutics induce high levels of reactive oxygen species (ROS) to eliminate tumor growth. Ferroptosis is a recently identified cell death mode triggered by ROS and subsequent lipid peroxidation. However, cancer cells develop adaptive abilities to recalibrate redox balance via hijacking the antioxidant systems, resulting in drug resistance. PRDX2 protein is a member of the peroxiredoxin family of antioxidant enzymes. We hypothesized that PRDX2 may serve as a critical mediator in redox-associated chemoresistance under reflux conditions.
Methods: In this study, we analyzed RNA sequencing data and public databases to unveil the aberrant overexpression and potential chemoresistance function of PRDX2 in EAC. To simulate both acute and chronic clinical conditions of gastroesophageal reflux disease (GERD) in vitro, we employed transient ABS exposure as well as repeated ABS exposure (rABS). EAC cells and 3D organotypic culture (OTC) served as platforms to explore the anti-ferroptosis function and regulatory mechanisms of PRDX2. Our initial findings were validated using patient-derived xenografts (PDX) and human EAC tissue microarrays (TMA).
Results: Aberrant expression of PRDX2 was detected in both human EAC tissues and cell lines exhibiting resistance to oxaliplatin treatment. The knockdown of PRDX2 impaired the recovery of EAC cells from ABS-induced ROS-dependent lipid peroxidation. Moreover, silencing PRDX2 sensitized the chemo-resistant EAC cells to oxaliplatin. Chromatin immunoprecipitation (ChIP) assay identified that NF-kB activated PRDX2 transcription by directly binding to its promoter region. We also found that APE1-redox-mediated NF-kB activation is crucial for ABS-induced PRDX2. We further discovered that PRDX2 contributes to chemoresistance by upregulating GPX4 and suppressing ferroptosis. Furthermore, immunofluorescent staining in the 3D model (OTC) and immunohistochemistry staining in human TMA confirmed co-overexpression of APE1 and PRDX2. In addition, APX2009, an APE1-redox-specific inhibitor, significantly sensitized EAC cells to oxaliplatin by inhibiting PRDX2 expression and inducing ferroptosis. Notably, we observed synergistic effects of the combination of oxaliplatin and APX2009 in EAC-PDX tumors.
Conclusion: Our findings revealed a novel link between reflux-induced redox unbalance and ferroptosis-related chemoresistance in EAC via APE1/redox/NF-kB/PRDX2/GPX4 axis. Targeting PRDX2 by APE1-redox-specific inhibitors is a potential novel strategy for drug combination in refractory EAC.

Heng Lu

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