EphA2 sustains the adaptive response of colorectal organoids to chemotherapy

Introduction – EphA2 is highly expressed in colorectal cancer (CRC), and high EphA2 expression indicates a worse prognosis. We investigated EphA2 dynamics in a clinically relevant model: CRC patient-derived organoids (PDOs) treated with chemotherapy. Methods – We evaluated the number of EphA2-expressing cells and the Aldehyde dehydrogenase activity by flow cytometry, and analyzed EphA2 protein levels and phosphorylation status using Zn-Phos-tag gels and indirect ELISA. We employed siRNA to deplete the PDO cells of EphA2. Results – EphA2-positive cells form a stable subpopulation in organoid cultures that persists after oxaliplatin treatment. Phosphorylation of EphA2 at Ser897 increases with treatment and correlates with higher EphA2 levels. Silencing EphA2 or reducing Ser897 phosphorylation decreases organoid formation, suggesting chemosensitization. Some EphA2-positive cells show increased ALDH activity after chemotherapy, and EphA2-ALDH1A3 interaction has prognostic value in CRC. Discussion – Here, we discovered that EphA2-positive cells constitute a persistent, ALDH-positive cell subpopulation in CRC-PDOs that withstood exposure to oxaliplatin (OXA) and 5-fluorouracil (5-FU). The enforced suppression of EphA2 or diminished Ser897 stress results in a chemosensitization effect. This sheds further light on the role of EphA2 in the adaptive stress response of CRC.