Supplementary Material for: Tissue-Based Multiomic Exploratory Analysis of the uPa/uPAR System and Matrix Metalloproteinases in SARIFA-Positive Gastrointestinal Cancers

Introduction: We recently proposed SARIFA (Stroma AReactive Invasion Front Areas), defined as direct tumour-adipocyte interaction, as an H&E-based histopathologic biomarker in gastrointestinal cancers, particularly gastric (GC) and colorectal cancer (CRC). Despite SARIFA’s well-validated prognostic value, its mechanistic underpinnings remain unclear. We hypothesized that extracellular matrix remodelling, specifically the plasmin/plasminogen activator system, may contribute to SARIFA formation. Methods: To test this, we compared the prognostic value of H&E-based SARIFA-status with ELISA-based protein levels of the serine proteases urokinase-type plasminogen activator (uPA, encoded by PLAU) and plasminogen activator inhibitor-1 (PAI-1, encoded by SERPINE1) in CRC. We further examined associations between SARIFA-status and the plasmin/plasminogen activator system as well as downstream metalloproteinases using both protein (ELISA, immunohistochemistry) and bulk gene expression data (TCGA-COAD/READ and TCGA-STAD), as well as spatial gene expression profiling in CRC (n=8) and GC (n=12). Results: Our findings show that high expression of the plasmin/plasminogen activator system and downstream metalloproteinases correlates with SARIFA-positivity. Digital spatial profiling revealed PLAU upregulation in tumour cells and PLAUR (encoding uPAR) upregulation in adjacent stromal cells at SARIFAs, suggesting a potential receptor-ligand interaction. Notably, SARIFA-positive tumours showed significantly higher numbers of tumour buds. Conclusion: These results provide new insights into the biological basis of SARIFAs and suggest therapeutic vulnerabilities related to the plasmin/plasminogen activator system.