Osteosarcoma is the most frequent primary malignant tumor of bone typically affecting children and young adults. The overall prognosis for patients with localized osteosarcoma has improved significantly (about 65% survive 5 yr) because of progress in chemotherapy and refined surgical techniques. In contrast, for 10–20% of the patients who present with metastasis, with the lung being the most common site, the 5-yr survival rate is 20%, which plateaued over the last 15 yr 1, 2. Therefore, new treatment strategies are required to prevent metastasis to improve the survival of these patients.
The development of an invasive tumor with metastatic potential is a complex, multistep process that includes cell proliferation, angiogenesis, tissue remodeling, and invasion 3. The degradation of the surrounding extracellular matrix (ECM) is one of the first steps initiated by the release of proteases from different cell types, including tumor and stromal cells 4. The proteases participating in these processes belong to all known classes of proteases (metallo-, serine-, aspartyl-, and cysteine proteases). Proteolytic enzymes can act directly by degrading ECM or cell surface proteins or indirectly by activating cascades of proteolytic enzymes. Under physiological conditions these cascades are regulated in complex networks of activators, inhibitors, and receptors. During tumor progression and metastasis this protease network is unbalanced with the consequence of not only destroying the surrounding ECM, but also releasing active molecules stored in the matrix or generating active fragments of the matrix proteins, which promote cell proliferation, invasion, and angiogenesis 5, 6. In addition to matrix metalloproteinases (MMPs) and serine proteases, cathepsin proteases are also involved in the different processes of metastasis 7–9. For cathepsins, 11 cysteine proteases (cathepsins B, C, F, H, K, L, O, S, V, W, and X), 2 aspartyl proteases (cathepsins D and E), and 1 serine protease (cathepsin G) have been identified in human 10. In normal cells, they are required for protein turnover, processing of proteins in secretory granules, antigen presentation, or bone remodeling 11, 12. In addition, cathepsins have also functions outside the lysosomal compartment. They execute programmed cell death upon release into the cytosol, and when released from the cell, they degrade ECM 13, 14. Several cathepsins have been identified as markers of tumor progression and metastasis in different types of cancer 8, 15. Specifically, cathepsin B and cathepsin L were suggested to participate in metastasis of osteosarcoma tumors and are expressed in corresponding cell lines 16–19.
The present study aimed at the identification of individual cathepsins as markers for metastasis in osteosarcoma. For this analysis we took advantage of the human SAOS-2 osteosarcoma cell line with low metastatic potential and of its derivatives LM5 and LM7 with increased metastatic potential in vivo 20. Recently, this model system has been used to study the role of interleukin-12 and Fas/FasL in osteosarcoma lung metastasis in vivo 21, 22. Here, the expression of 14 cathepsins was studied at the mRNA and the protein levels in the SAOS-2 osteosarcoma cell line and the highly metastatic LM5 and LM7 sublines. The results confirmed the observations of previous reports that suggested a role of cathepsin L in metastasis of osteosarcoma. Importantly, the results also imply that cathepsin K may be of diagnostic relevance with respect to prognosis of patients with high-grade tumors and metastasis at diagnosis.