Suppression of Invasion by Inducible Expression of Tissue Inhibitor of Metalloproteinase-1 (TIMP-1) in B 16-F10 Melanoma Cells

Abstract

Background: We have previously shown that down-modulation (i.e., by antisense expression vector) of tissue inhibitor of metalloproteinase-1 (TIMP-1) in a noninvasive, nontumorigenic cell line led to an acquisition of an invasive, tumorigenic, and metastatic ability in these cells. Purpose: Our purpose was to examine whether increased levels of murine TIMP-1 can directly suppress the invasive ability of malignant cells. Methods: Murine B16-F10 melanoma cells were transfected with an expression vector to overproduce TIMP-1. Among these transfectants, we isolated five clonal cell lines (2–5, 2–8, 2–10, 6–5, and 6–9) that showed upregulation (i.e., overexpression) of TIMP-1. Results: These cell lines had an increased basal level of TIMP-1 messenger RNA. TIMP-1 expression was under the control of the mouse metallothionein-I promoter, and four of these five clones (2–5, 2–8, 6–5, and 6–9) showed a threefold to 10-fold induction of TIMP-1 message when they were treated with 20 [sM cadmium for 4 hours. An increase in TIMP-1 message led to an increase in TIMP-1 protein activity measured in the conditioned medium of clones 2–10 and 6–5. The invasive ability of the TIMP-1-upregulated cells was tested in a matrigel transwell invasion assay. All of the upregu-lated clones showed a significant reduction in their invasive ability, relative to the invasive ability of parental B16-F10 and the control 1–2 cell line; this reduction correlated with the level of TIMP-1 overexpression. Cadmium induction of TIMP-1 messenger RNA resulted in a further suppression of the invasive ability of the two inducible cell lines tested (i.e., 2–8 and 6–5). Conclusions: Our data demonstrate that a specific upregulation of murine TIMP-1 expression in B16-F10 melanoma cells directly suppressses their invasive ability. [J Natl Cancer Inst 84:1017–1022, 1992]