Single-cell and Spatial Transcriptomic Mapping Reveals MSEC-myoCAF Interactions Driving Lymph Node Metastasis in Colorectal Cancer

Lymph node metastasis (LNM) is a major determinant of poor prognosis in colorectal cancer (CRC), yet the cellular and spatial mechanisms driving metastatic initiation remain poorly defined. Here, we performed integrative single-cell RNA sequencing (scRNA-seq; 84,735 cells) and spatial transcriptomics (ST; 20,859 spots) on paired primary tumors and metastatic lymph nodes from five CRC patients. We identified a metastatic stem-like epithelial cell population (MSECs), marked by LGR5, AXIN2, UBE2H, and LAMC2, enriched at invasive fronts and metastatic niches. MSECs co-localized with myofibroblastic cancer-associated fibroblasts (myoCAFs) and communicated via a COL1A1-SDC4 axis that activated NOTCH signaling and promoted epithelial–mesenchymal transition (EMT). Functional assays demonstrated that UBE2H or LAMC2 knockdown impaired CRC cell migration, invasion, and EMT, while their high expression predicted poor disease-free survival. Spatial analysis further revealed that myoCAFs formed a stromal barrier facilitating immune exclusion and metastatic expansion. These findings highlight the MSEC–myoCAF crosstalk as a key driver of CRC metastasis and nominate UBE2H, LAMC2, and COL1A1-SDC4 as potential therapeutic targets.