Engineered mesenchymal stem cell-derived small extracellular vesicles delivering miR-124 suppress colorectal cancer progression through EZH2 downregulation and attenuation of STAT3 signaling.

Colorectal cancer (CRC) progression is driven by dysregulated molecular pathways and epigenetic alterations, including enhancer of zeste homolog 2 (EZH2)-mediated histone methylation and STAT3 hyperactivation. While microRNA-124 (miR-124) acts as a tumor suppressor by targeting EZH2, its clinical translation is hindered by delivery challenges. Here, we engineered mesenchymal stem cell-derived small extracellular vesicles (MSC-sEVs) to deliver miR-124, exploiting their tumor-homing capacity and biocompatibility to suppress CRC progression. Bioinformatic analysis of clinical datasets revealed miR-124 downregulation in CRC tissues, correlating with advanced stages and poor prognosis. Lentiviral transduction enabled efficient miR-124 packaging into MSC-sEVs, which were internalized by CRC cells, restoring functional miR-124 levels. Mechanistically, miR-124 directly silenced EZH2, thereby suppressing its oncogenic activity. This led to attenuated STAT3 phosphorylation-a dual effect that inhibited proliferation, migration, and epithelial-mesenchymal transition (EMT) in CRC cells. In vitro, MSC-sEV-miR-124 induced apoptosis, sensitized cells to 5-fluorouracil, and reversed chemoresistance via EZH2 downregulation. In vivo, MSC-sEV-miR-124 significantly reduced tumor growth in BALB/c mice, downregulated EZH2/STAT3 signaling, and suppressed Ki-67 and EMT markers in xenografts. This study provides proof-of-concept that engineered MSC-sEVs can serve as a delivery platform for miR-124 in murine CRC models, offering a dual-targeting strategy against EZH2 and STAT3 to disrupt CRC progression. Our findings suggest that EV-based miRNA delivery warrants further investigation as a strategy to modulate the tumor microenvironment, providing a foundation for development aimed at overcoming epigenetic and oncogenic signaling barriers in CRC therapy.