PROJECT SUMMARY/ABSTRACT Project 1: Shaping the Microenvironment by DPEP1 Facilitates Adenoma Progression Project 1 aims to examine how shaping the microenvironment by DPEP1 facilitates adenoma progression. This will be a basic project. We propose that dipeptidase-1 (DPEP1) marks those adenomas with the potential to progress to colorectal cancer (CRC) through an active bi-directional communication with neutrophils. DPEP1 has two functions: dipeptidase activity and recently identified neutrophil-binding activity. Our hypothesis is that DPEP1, largely through its neutrophil-binding activity, marks adenomas with a predilection for progression. We have found that DPEP1 immunoreactivity is detected in 27% of colorectal adenomas but this increases to 72% of CRCs, consistent with DPEP1 marking the small subset of adenomas that progress to CRC. Utilizing human specimens, a unique Transwell co-culture system of adenoma organoids and freshly isolated neutrophils isolated from healthy volunteers, and an informative mouse model, we will test if DPEP1-expressing adenomas more effectively communicate with neutrophils and create a neutrophil-enriched microenvironment, increasing the likelihood that these adenomas will progress. Exosomes have attracted a great deal of recent attention as a rich source of cargo that may serve as cancer biomarkers. We have found that DPEP1 is released in exosomes from CRC cell lines and that it highly enriched in a subset of exosomes that contain known CRC biomarkers, CEA and EPCAM. Of interest, neutrophils also release small extracellular vesicles (sEVs) that contain neutrophil elastase in a form that cannot be inhibited by elastase inhibitors, and thus it is especially potent in degrading the extracellular matrix, a key step in cancer invasion. We will also use a unique neutrophil reporter mouse to monitor onset and perdurance of neutrophil infiltration, along with the properties of these neutrophils, in an inducible, stem cell-driven mouse model of colonic adenomas. .