Morphine Contributes to Epithelial-Mesenchymal Transition in Triple-Negative Breast Cancer Cells by Blocking COX-2 Methylation via Regulating the miR-23a-3p/DNMT3A Feedback.

Abstract

To investigate the effects and mechanisms of morphine on epithelial-mesenchymal transformation (EMT) in triple-negative breast cancer (TNBC). The levels of miR-23a-3p, DNMT3A, and COX-2 in tumor tissues from metastatic TNBC patients treated with morphine were assessed using qRT-PCR. Functional assays assessed morphine's impact on TNBC cell malignancy. Dual luciferase reporter and RNA pull-down assays investigated the interaction between miR-23a-3p and DNMT3A. miR-23a-3p inhibitor and DNMT3A siRNA were transfected into TNBC cells. Protein expression was analyzed by Western blot. Methylation status of miR-23a-3p and COX-2 was assessed via methylation-specific PCR. Rescue experiments were performed to research whether morphine modulates EMT in TNBC through COX-2 methylation regulation via the miR-23a-3p/DNMT3A feedback loop. The effects of morphine on TNBC in nude mice xenotransplantation were studied. In metastatic TNBC patients treated with morphine, miR-23a-3p and COX-2 expression were elevated, and DNMT3A levels were reduced. In TNBC cells, morphine enhanced migration, invasion, and EMT, and suppressed apoptosis. It upregulated miR-23a-3p and COX-2; downregulated DNMT3A; and inhibited methylation of miR-23a-3p and COX-2. miR-23a-3p directly inhibited DNMT3A expression. In morphine-treated TNBC cells, silencing DNMT3A reduced methylation of miR-23a-3p and COX-2. miR-23a-3p inhibitor suppressed migration, invasion, and EMT, and promoted apoptosis; however, these effects were reversed by DNMT3A silencing. In vivo, morphine promoted tumor EMT and metastasis in TNBC; reduced miR-23a-3p and COX-2 methylation; and decreased DNMT3A expression. Morphine accelerated EMT in TNBC by inhibiting COX-2 methylation through the miR-23a-3p/DNMT3A loop.