The initial steps of ovarian cancer cell metastasis are mediated by MMP-2 cleavage of vitronectin and fibronectin
Hilary A. Kenny, … , Lisa M. Coussens, Ernst Lengyel
Hilary A. Kenny, … , Lisa M. Coussens, Ernst Lengyel
Published March 13, 2008
Citation Information: J Clin Invest. 2008;118(4):1367-1379. https://doi.org/10.1172/JCI33775.
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Research Article Oncology

The initial steps of ovarian cancer cell metastasis are mediated by MMP-2 cleavage of vitronectin and fibronectin

Abstract

Most patients (80%) with ovarian cancer (OvCa) present with metastatic disease. Attachment of OvCa cells to peritoneum and omentum represents the first rate-limiting step for metastatic spread. Therefore, identifying factors regulating cell attachment in the abdominal cavity is critical to the development of therapeutic agents. We show here that MMP-2 expression was upregulated in OvCa cells upon attachment to their microenvironment. Downregulation of MMP-2 mRNA or pharmacological inhibition of MMP-2 proteolytic function, in both human OvCa primary cells and cell lines, reduced attachment of OvCa cells to a 3D organotypic model of metastatic OvCa, full human omentum or peritoneum, and in vivo to mouse peritoneum and omentum. Absence of MMP-2 in the host did not alter OvCa adhesion, as determined utilizing mice harboring homozygous null mutations in either the Mmp2 or Mmp9 genes. Conversely, adhesion induced upregulation of MMP-2 mRNA in OvCa cells. MMP-2 inhibition in OvCa cells through pharmacological or antibody treatment prior to i.p. dissemination in nude mice significantly decreased tumor growth and metastasis and extended survival. MMP-2 enhanced peritoneal adhesion of OvCa cells through cleavage of ECM proteins fibronectin (FN) and vitronectin (Vn) into small fragments and increased binding of OvCa cells to these FN and Vn fragments and their receptors, α5β1 and αVβ3 integrin. These findings indicate that MMP-2 expressed by metastatic OvCa cells functionally regulates their attachment to peritoneal surfaces.

Authors

Hilary A. Kenny, Swayamjot Kaur, Lisa M. Coussens, Ernst Lengyel

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Figure 5

MMP-2 is transcriptionally upregulated in OvCa cells upon interaction with host stroma.

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MMP-2 is transcriptionally upregulated in OvCa cells upon interaction wi...
SKOV3ip1 cells were added to the 3D culture (A), full human omentum (B), or full human peritoneum (C), and then cells were sorted by FACS. The relative expression of MMP-2 normalized to GAPDH, and huGUS was measured by TaqMan quantitative real-time RT-PCR. (D) SKOV3ip1 cells were transfected with 5 μg of the –1,659-bp (WT) or the –1,629-bp MMP-2 promoter and adhered to 3D model, and luciferase activity was measured. (E) The relative expression of p53 normalized to GAPDH, and huGUS was measured by TaqMan quantitative real-time RT-PCR (top panel). Protein lysates from SKOV3ip1 cells cultured on plastic or 3D culture (after sorting) were immunoprecipitated with control mouse IgG or monoclonal p53 antibody, and western blot analysis for p53 was conducted on lysates using a pantropic sheep anti-p53 antibody (bottom panel). Positive control was RKO mRNA or cell lysates. (F) SKOV3ip1 cells were cotransfected with 5 μg of the WT MMP-2 promoter and a WT or mutated p53 expression plasmid and adhered to the 3D model, and luciferase activity was measured. Luciferase activity was normalized to number of bound and unbound cells, and all assays were run in duplicate. *P < 0.01, **P < 0.001. Each graph is representative of 3 independent experiments.