Abstract C100: Therapeutic targeting of microRNA-31 promotes chemosensitivity via ATOX1 in pancreatic ductal adenocarcinoma

Abstract Introduction: Pancreatic ductal adenocarcinoma (PDAC) has a dismal long-term patient survival rate. This poor prognosis is due in part to poor patient responses to current standard-of-care chemotherapy regimens. Patients receiving platinum-based chemotherapy often succumb to treatment resistance. MicroRNAs (miRNAs) are small, non-coding RNAs that play an important role in modulating cellular processes involved in chemoresistance. We have previously demonstrated that loss of miR-31 influences cellular sensitivity to chemotherapy and radiotherapy in several cancers, including PDAC. Here, we identify a molecular mechanism underlying miR-31–mediated alterations in platinum agent chemosensitivity, supporting miR-31 as an important therapeutic target. Materials and Methods: A pCMV-miR vector containing a miR-31 mimic was stably expressed into a miR-31–deficient PDAC cell line, BxPC-3. Additionally, a pmiRZip lentivector suppressing miR-31 was stably expressed into a miR-31–abundant PDAC cell line, Panc-1. To assess ATOX1 alterations, a pCMV6-AC-ATOX1 overexpression vector was stably expressed in Panc-1 cells, and a pRS ATOX1 shRNA suppression vector was stably expressed in BxPC-3 cells. Clonogenic assays were performed to assess the impact of miR-31 and ATOX1 modulation on chemosensitivity. Expression levels of candidate miR-31 targets were evaluated using ELISA and Western blot analysis. Following cisplatin treatment, cytoplasmic and nuclear levels of Pt195 were quantified by ICP-MS in the miR-31– and ATOX1–manipulated PDAC cells. Results: MiR-31 overexpression in BxPC-3 cells significantly promoted resistance to cisplatin. Conversely, miR-31 suppression in Panc-1 cells significantly enhanced cisplatin sensitivity. Kaplan-Meier survival analysis of PDAC patients showed that low miR-31 expression inversely correlates with high expression of ATOX1, a cytoplasmic copper (Cu) chaperone and transcription factor, and that these patients have significantly improved overall survival. In silico analysis identified ATOX1 as a predicted target of miR-31. Overexpression of miR-31 significantly reduced ATOX1 expression in PDAC cells. There is evidence that cisplatin binds to the Cu(I)-ATOX1 complex, potentially altering intracellular cisplatin transport. Consequently, we found that miR-31 expression inversely correlated with the nuclear accumulation of cisplatin in PDAC cells. Moreover, direct overexpression of ATOX1 in Panc-1 cells significantly enhanced cisplatin sensitivity. Conversely, direct suppression of ATOX1 in BxPC-3 cells significantly promoted resistance to cisplatin. However, direct suppression of ATOX1 did not alter intranuclear platinum levels, suggesting that ATOX1 modulates chemosensitivity through mechanisms independent of cisplatin nuclear trafficking. Conclusions: Our study demonstrates, for the first time, that miR-31 regulates ATOX1 expression, thereby modulating cisplatin sensitivity in PDAC. These findings reveal a novel miR-31–ATOX1 axis as a potential therapeutic target to overcome chemoresistance in PDAC. Citation Format: David Hackett, Jason McGrath, Xuehua Lin, David Van Acken, Ralf A. Hilger, Christina Cahill, Stephen G. Maher. Therapeutic targeting of microRNA-31 promotes chemosensitivity via ATOX1 in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2025 Oct 22-26; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2025;24(10 Suppl):Abstract nr C100.