Collagenase-3 (MMP-13) is a human matrix metalloproteinase specifically expressed by invading tumor cells in squamous cell carcinomas (SCCs) of the head and neck. Here, we have further elucidated the role of MMP-13 in tumor invasion by examining its expression in invasive malignant tumors of the female genital tract. Using in situ hybridization, expression of MMP-13 mRNA was detected in 9 of 12 vulvar SCCs, primarily in tumor cells, but not in intact vulvar epithelium, in cervical SCCs (n = 12), or in endometrial (n = 11) or ovarian adenocarcinomas (n = 8). MMP-13 expression was especially abundant in vulvar carcinomas showing metastasis to lymph nodes and was associated with expression of membrane type 1 MMP by tumor cells and gelatinase-A (MMP-2) by stromal cells, as detected by immunohistochemistry. MMP-13 mRNAs were detected in 9 of 11 cell lines established from vulvar carcinomas and in 4 of 6 cell lines from cervical carcinomas, whereas endometrial (n = 10) and ovarian (n = 9) carcinoma cell lines were negative for MMP-13 mRNA. No correlation was detected between MMP-13 expression and p53 gene mutations in vulvar SCC cell lines. However, MMP-13 expression was detected in 5 of 6 vulvar and cervical SCC cell lines harboring HPV 16 or 68 DNA. These results show that MMP-13 is specifically expressed by malignantly transformed squamous epithelial cells, including vulvar SCC cells, and appears to serve as a marker for their invasive capacity.