ca-074-methyl-ester and Neoplasm-Metastasis

Cathepsin B is a cysteine proteinase that primarily functions as an endopeptidase within endolysosomal compartments in normal cells. However, during tumoral expansion, the regulation of cathepsin B can be altered at multiple levels, thereby resulting in its overexpression and export outside of the cell. This may suggest a possible role of cathepsin B in alterations leading to cancer progression. The aim of this study was to determine the contribution of intracellular and extracellular cathepsin B in growth, tumorigenesis, and invasion of colorectal cancer (CRC) cells. Results show that mRNA and activated levels of cathepsin B were both increased in human adenomas and in CRCs of all stages. Treatment of CRC cells with the highly selective and non-permeant cathepsin B inhibitor Ca074 revealed that extracellular cathepsin B actively contributed to the invasiveness of human CRC cells while not essential for their growth in soft agar. Cathepsin B silencing by RNAi in human CRC cells inhibited their growth in soft agar, as well as their invasion capacity, tumoral expansion, and metastatic spread in immunodeficient mice. Higher levels of the cell cycle inhibitor p27(Kip1) were observed in cathepsin B-deficient tumors as well as an increase in cyclin B1. Finally, cathepsin B colocalized with p27(Kip1) within the lysosomes and efficiently degraded the inhibitor. In conclusion, the present data demonstrate that cathepsin B is a significant factor in colorectal tumor development, invasion, and metastatic spreading and may, therefore, represent a potential pharmacological target for colorectal tumor therapy.

Topics: Animals; Caco-2 Cells; Carcinogenesis; Cathepsin B; Cell Line, Tumor; Cell Movement; Cell Proliferation; Colorectal Neoplasms; Dipeptides; Gene Expression Regulation, Neoplastic; HCT116 Cells; HEK293 Cells; HT29 Cells; Humans; Mice; Neoplasm Invasiveness; Neoplasm Metastasis; Neoplasm Transplantation

The receptor for the type 1 insulin-like growth factor (IGF-I) has been implicated in cellular transformation and the acquisition of an invasive/metastatic phenotype in various tumors. Following ligand binding, the IGF-I receptor is internalized, and the receptor.ligand complex dissociates as the ligand is degraded by endosomal proteinases. In the present study we show that the inhibition of endosomal IGF-I-degrading enzymes in human breast and murine lung carcinoma cells by the cysteine proteinase inhibitors, E-64 and CA074-methyl ester, profoundly altered receptor trafficking and signaling. In treated cells, intracellular ligand degradation was blocked, and although the receptor and two substrates, Shc and Insulin receptor substrate, were hyperphosphorylated on tyrosine, IGF-I-induced DNA synthesis, anchorage-independent growth, and matrix metalloproteinase synthesis were inhibited. The results suggest that ligand processing by endosomal proteinases is a key step in receptor signaling and function and a potential target for therapy.

Topics: Adaptor Proteins, Signal Transducing; Adaptor Proteins, Vesicular Transport; Animals; Blotting, Western; Cell Membrane; Chromatography, High Pressure Liquid; Cysteine Proteinase Inhibitors; Dipeptides; DNA; Dose-Response Relationship, Drug; Endosomes; Female; Flow Cytometry; Humans; Insulin-Like Growth Factor I; Kinetics; Leucine; Ligands; Liver; Male; Mice; Models, Biological; Neoplasm Metastasis; Phosphorylation; Precipitin Tests; Protein Binding; Proteins; Rats; Rats, Sprague-Dawley; Receptor, IGF Type 1; Shc Signaling Adaptor Proteins; Signal Transduction; Src Homology 2 Domain-Containing, Transforming Protein 1; Time Factors; Tumor Cells, Cultured; Tyrosine