Tissue-specific reprogramming leads to angiogenic neutrophil specialization and tumor vascularization in colorectal cancer

Polymorphonuclear neutrophil (PMN) tissue accumulation is an established feature of ulcerative colitis (UC) lesions and colorectal cancer (CRC). Given the emerging evidence of PMN phenotypic and functional heterogeneity, we analyzed the transcriptomic landscape of PMNs in blood and tissue during the spatiotemporal transition from inflammatory ulceration to CRC. Based on their transcriptional programs, PMNs were effectively stratified into distinct spatial compartments of peripheral blood, inflamed colon tissue, and the tumor niche. In silico pathway overrepresentation analysis, protein-network mapping, gene signature identification, and gene-ontology scoring revealed unique enrichment of angiogenic and vasculature development pathways in tumor-associated neutrophils (TANs). Functional studies utilizing ex vivo cultures, colitis-induced murine CRC, and patient-derived xenograft models demonstrated a critical role for TANs in promoting tumor vascularization. Spp1 (OPN) and Mmp14 (MT1-MMP) were identified by unbiased -omics and by mechanistic studies to be highly induced in TANs and function to critically regulate endothelial cell chemotaxis and branched network formation. TCGA dataset and clinical specimens confirmed enrichment of SPP1 and MMP14 in high-grade CRC but not in UC patients. Importantly, pharmacological inhibition of TAN trafficking or MMP14 activity effectively reduced tumor vascular density, leading to CRC regression. The current study identified a niche-directed PMN transcriptional reprograming and functional specialization, highlighting emerging PMN plasticity. Our findings define new TAN contributions to tumor vascularization, delineating a new therapeutic framework for CRC treatment focused on TAN angiogenic properties. Overall design: Mice were subjected to multiple cycle of AOM/DSS. Blood or tumor-associated neutrophils were sorted and RNA-extracted at different disease timepoints (colitis, week 1; early CRC week 8, and advanced CRC week 13-14). We performed bulk RNAsequencing to characterize transcriptomic changes of neutrophils in cancer tissues and perform GO and GSEA analyses to identify angiogenic profiles of neutrophils. DEG analysis by DEseq2 and pairwise comparisons identified SPP1 and MMP14 as top upregulated genes in tumor neutrophils. We thus treated PDX models with MMP14 inhibitor and anti-OPN and extracted tumor cells for bulk RNAsequencing. Gene ontology and GSEA analyses identified collagen pathways and hypoxia pathways being enriched upon treatment with MMP14 inhibitor.