Suppression of in vivo tumor growth and induction of suspension cell death by tissue inhibitor of metalloproteinases (TIMP)-3

Abstract

Tissue inhibitor of metalloproteinases-3(TEMP-3), a novel member of TEMP family genes, has been recently cloned and shown to be expressed in preneoplastic but not in neoplastic mouse JB6 epidermal cells (Sun et al. 1994 Cancer Res., 54, 11139). This down regulation of the gene appears to be attributable at least in part to alteration of gene methylation (Sun et al. 1995 J. Biol Chem., 270, 19312). Little is known, however, about the role of TEMP-3 in human cancers. We screened several human tumor cell lines for TEMP-3 expression and found that a colon carcinoma line, DLD-1, did not express TEMP-3. If down regulation of TIMP-3 is causally related to carcino-genesis, re-expression by transfection may reverse the tumor cell phenotype. We therefore overexpressed human TEMP-3 in DLD-1 cells. TEMP-3 transfectants showed a serum-dependent growth inhibition in monolayer culture and a decreased growth potential in nude mice in a manner dependent on the level of TEMP-3 expression. A transfectant expressing a high level of active hTEMP-3 completely lost the ability to form tumors following s.c. injection into nude mice. We also tested TEMP-3 expressing cells and neocontrol TEMP-3 negative cells for their ability to grow in liquid suspension culture, since both cells grew in semi-solid soft agar. As compared to neocontrol cells, TIMP-3 over-expressors formed large aggregates, followed by cell death. This effect was not mimicked by BB94, a broad MMP inhibitor. We conclude from this study that (i) TEMP-3 overexpression in human colon carcinoma cells induces growth arrest in low serum conditions and inhibits in vivo tumor growth and (ii) the TEMP-3-induced large aggregate formation and subsequent cell death under suspension growth cannot be explained by its MMP inhibitory activity.