AJUBA: The Master Regulator Bridging EMT and Immune Evasion in Colorectal Cancer.

[BACKGROUND] Epithelial-mesenchymal transition (EMT) represents a critical process that facilitates metastatic dissemination and immune evasion in colorectal cancer (CRC); however, the molecular factors that connect EMT to modifications in the immune microenvironment remain poorly elucidated. In this investigation, we identify AJUBA as an essential regulator that mediates the association between EMT and immune modulation in CRC.

[METHODS] By integrating multi-cohort transcriptomic datasets (The Cancer Genome Atlas (TCGA)-CRC, GSE18105, GSE22598, GSE89076, and GSE110224) with single-cell RNA-seq data (GSE132465), we applied machine learning and deep learning methodologies to comprehensively identify EMT-associated genes demonstrating prognostic significance. AJUBA validation was performed at mRNA and protein levels in a cohort of 90 CRC patient samples using quantitative PCR, Western blotting, and immunohistochemical (IHC) approaches. Functional analyses involved siRNA-mediated knockdown experiments, coupled with evaluations of cell proliferation (CCK-8 assay), migration and invasion (transwell assay), clonogenic capacity (colony formation assay), and in vivo tumor growth in xenograft models. Immune infiltration was assessed via ssGSEA and CIBERSORT algorithms, and spatial transcriptomics data (GSE225857) were used to delineate AJUBA expression within tumor microdomains.

[RESULTS] Across multiple CRC cohorts, AJUBA exhibited marked upregulation and showed distinct enrichment in epithelial cells with activated EMT characteristics. Spatial transcriptomic profiling demonstrated AJUBA colocalization with cancer-associated fibroblasts (CAFs) within immune-excluded niches. Enhanced AJUBA expression exhibited a positive correlation with heightened infiltration of M2 macrophages and activation of VEGF/NOTCH signaling cascades. In vivo, AJUBA knockdown led to suppressed tumor growth, reduced Ki-67 proliferation indices, and diminished M2 macrophage abundance. Clinically, elevated AJUBA expression correlated with advanced nodal metastasis and served as an independent predictor of poor overall survival (OS; HR = 4.809, 95% CI: 2.385-9.695, p < 0.001).

[CONCLUSIONS] AJUBA functions as a key regulator that links EMT to immune modulation, promoting macrophage polarization and facilitating immune evasion in CRC. Through its coupling of EMT activation with proangiogenic signaling, AJUBA represents both a prognostic biomarker and a promising therapeutic target for alleviating immune exclusion in metastatic CRC.