The notch-miR-188-5p-TIMP2/3 axis orchestrates exosome-driven pre-metastatic niche formation in colorectal cancer

Colorectal cancer (CRC) metastasis is critically mediated by pre-metastatic niche (PMN) formation, a process driven by tumor-derived exosomes. This study establishes a highly invasive CT-26 subline (HI CT-26) through iterative in vivo selection, demonstrating enhanced metastatic potential via epithelial-mesenchymal transition (EMT) activation and transendothelial migration. HI CT-26-derived exosomes were found to enrich miR-188-5p, which directly suppresses TIMP2 and TIMP3, key regulators of extracellular matrix (ECM) homeostasis and immune modulation. This suppression is associated with the establishment of a PMN, characterized by an increased relative abundance of M2-like macrophages and myeloid-derived suppressor cell (MDSC) accumulation in target organs. Mechanistically, Notch signaling activates miR-188-5p transcription through RBP-J binding to its promoter, forming a regulatory axis that links Notch pathway activation to metastatic progression. Clinical validation using TCGA data confirmed elevated RBP-J and miR-188-5p expression, alongside reduced TIMP2/3 levels, as prognostic biomarkers for poor CRC outcomes. These findings reveal a novel Notch-miR-188-5p-TIMP2/3 signaling cascade driving exosome-mediated PMN formation, offering insights into therapeutic strategies targeting the metastatic microenvironment.