Both the matrix metalloproteinase matrilysin and the prostaglandin H synthase cyclooxygenase‐2 (Cox‐2), are thought to play key roles in colorectal carcinogenesis. These enzymes are overexpressed in 85–90% of human colorectal cancers. Furthermore, mice carrying an adenomatous polyposis coli germline mutation that are also nullizygous for either matrilysin or Cox‐2 display a significant reduction in tumor multiplicity. To determine if there is a direct link between matrilysin and Cox‐2, their expression was characterized in two mouse models of intestinal carcinogenesis and in human colorectal tumor samples. Both matrilysin and Cox‐2 expression was increased in the mouse models and in the human colorectal cancers; however, immunohistochemistry and in situ hybridization indicated that their localization within the tumors was different. In the mouse models, Cox‐2 was expressed in the superficial stroma, whereas matrilysin expression was localized exclusively to the neoplastic epithelium. In contrast, in human colorectal cancers, both Cox‐2 and matrilysin were expressed in the neoplastic epithelium. Although over 80% of the specimens expressed both matrilysin and Cox‐2, the levels and localization of matrilysin and Cox‐2 expression were distinct. Cox‐2 expression was strongest in well‐differentiated areas, and matrilysin immunostaining was strongest in the more dysplastic and invasive regions of the tumor. These results indicate that these two important modulators of colorectal tumorigenesis are differentially expressed and imply that the therapeutic benefit may be improved by combination therapy utilizing selective Cox‐2 and matrilysin inhibitors. Mol. Carcinog. 24:177–187, 1999. © 1999 Wiley‐Liss, Inc.