Reduction of Experimental Human Fibrosarcoma Lung Metastasis in Mice by Adenovirus-Mediated Cystatin C Overexpression in the Host

Abstract

Tumor cell invasion and metastasis are associated with degradation of components of the extracellular matrix by different proteinases. Among those, papain-like cysteine proteases, such as cathepsin B, seem to play an important role, as they are associated with poor clinical outcome in different cancers. In this study, we tested whether cystatin C, a natural extracellular inhibitor of papain-like cysteine proteases, can inhibit metastasis when overexpressed at the tumor-host interface. Local overexpression of cystatin C in liver and lungs of CD1 nu/nu mice was achieved by gene transfer with a novel adenoviral construct, which also led to the presence of 60 ng/mL of cystatin C in the serum. Three days after gene transfer, these mice were challenged by i.v. inoculation of lacZ-tagged human fibrosarcoma cells (HT1080lacZ-K15), leading to the formation of experimental lung and liver metastases. In this model, formation of experimental metastatic foci correlated with expression of cathepsin B in lungs, whereas there was no correlation with metastasis to the liver. In mice overexpressing cystatin C, the number of lung metastases was significantly reduced by 92%, as compared with mice receiving control adenovirus. The efficacy of extravasation of HT1080lacZ-K15 cells into the liver was not affected, indicating the independence of this process from the activity of cysteine-cathepsins. The present report is the first evidence of successful reduction of metastasis by inhibition of cysteine-cathepsins by cystatin C overexpression in the host microenvironment. Furthermore, organ-specific protease expression during tumor-host cell interactions could affect the success of antiproteolytic intervention against metastasis. (Cancer Res 2005; 65(19): 8608-12)

Tissue Inhibitor of Metalloproteinases-1 Promotes Liver Metastasis by Induction of Hepatocyte Growth Factor Signaling

“…We used lacZ-tagged human fibrosarcoma cells (HT1080lacZ-K15) that typically form large pulmonary metastatic foci and only few single tumor cells as micrometastases, but no macrometastases, in livers (26) to exclude that the TIMP-1 effect was cell type dependent. After challenge of CD1 nu/nu mice with HT1080lacZ-K15, significant reduction of lung metastasis by 93% was observed in TIMP-1-transduced animals compared with mice with virus control (P = 0.018; Fig.…”

“…A DNA fragment consisting of Ign-leader sequence and N-TIMP-1 cDNA or full-length human TIMP-1 cDNA (pGEMhTIMP-1; ref. 25), respectively, was obtained and cloned into the adenoviral shuttle plasmid pKA1 (26). Recombinant adenoviruses were generated, purified, and plaque titered as described previously (26).…”

“…25), respectively, was obtained and cloned into the adenoviral shuttle plasmid pKA1 (26). Recombinant adenoviruses were generated, purified, and plaque titered as described previously (26). As control, Addl70-3, an adenovirus without transgene, was used (27).…”

“…Functionality of adenoviral constructs was tested in vitro (data not shown). Adenoviral infection in vitro was done as described previously (26).…”

Inhibitors of Proteinases as Potential Anti-Cancer Agents

“…In some types of cancers, the changes in cysteine cathepsin epression or activity have diagnostic or prognostic value with imbalance between cathepsins and cystatins being associated with tumor phenotype. Since the latter ones are able to inhibit cathepsins tumor-associated activity many studies have indicated their potential use in therapeutic approaches (Keppler, 2006;Kopitz et al, 2005;Palermo & Joyce, 2007). Indeed, one of these inhibitors, cystatin C (mostly the one isolated from egg white) has been used in preclinical research studies for more than 20 years, however, it has been introduced into clinical practice quite scarcely.…”