ROLE AND REGULATION OF EXPRESSION OF 92-KDA TYPE-IV COLLAGENASE (MMP-9) IN 2 INVASIVE SQUAMOUS-CELL-CARCINOMA CELL-LINES OF THE ORAL CAVITY

The present study was undertaken to determine the role of the metallop roteinase MMP-9 in the invasive phenotype of squamous-cell carcinoma o f the oral cavity and the regulation of its expression. Zymographic an alysis of conditioned medium from 2 highly invasive squamous-cell-carc inoma cell lines indicated large amounts of an enzyme which was indist inguishable, in size (92 kDa) from the MMP-9 pro-enzyme. Conversion of the 92-kDa gelatinase into a lower-molecular-weight species (84 kDa), identical in size to the activated gelatinase, was evident when both cell lines, which are avid secretors of urokinase, were cultured in th e presence of plasminogen. Penetration of an extracellular-matrix-coat ed filter was dramatically reduced in the presence of the collagenase inhibitor, tissue inhibitor of metalloproteinase-2, suggesting a criti cal role for MMP-9 in the invasive process. Immunohistochemical studie s demonstrating the presence of MMP-9 in tumor cells of resected squam ous-cell cancers suggested that secretion of this collagenase by cells in vitro was reflective of the in vivo setting. Since several phorbol -ester response elements are present in the MMP-9 promoter, we determi ned the role of protein-kinase-C pathways in the regulation of MMP-9 e xpression in cultured SCC. Treatment of cells with PMA resulted in a m ore-than-20-fold increase in the level of protein and mRNA. Conversely , culturing of cells in the presence of the protein-kinase-C inhibitor , calphostin-C, led to a dose-dependent decrease in the amount of MMP- 9 mRNA and protein, suggesting that the constitutive expression of thi s collagenase reflects activation of this signal transduction pathway. In summary, our data suggest that, for a sub-population of squamous-c ell carcinomas, secreted MMP-9 is an important determinant of the inva sive phenotype, and that the expression of this metalloproteinase is r egulated by protein-kinase-C pathways. (C) 1993 Wiley-Liss, Inc.