Identification of Potential Biomarkers for Giant Cell Tumor of Bone Using Comparative Proteomics Analysis

Abstract

Giant cell tumor of bone can be locally aggressive and occasionally can metastasize in the lungs. To identify new markers predictive of aggressive behavior, we analyzed five patients who developed lung metastasis and five who remained disease free for a minimum of 5 years. Using two-dimensional electrophoresis, we detected 28 differentially expressed spots. Fourteen spots were identified using mass spectrometry, including seven up-regulated and seven down-regulated in metastatic samples and classified according to functional categories. We then selected five proteins involved in cell cycle or apoptosis. Thioredoxin peroxidase, allograft inflammatory factor 1, and ubiquitin E2N had more than threefold up-regulation; glutathione peroxidase 1 had 1.9-fold up-regulation; and heat shock protein 27 showed down-regulation in metastatic samples with a very low P value. After validation and analysis of protein levels, evaluation of clinical impact was assessed in a much wider cohort of primary archival specimens. Immunodetection showed a higher frequency of thioredoxin peroxidase, allograft inflammatory factor 1, ubiquitin E2N, and glutathione peroxidase 1 overexpression in primary tumors that developed into lung metastases or that locally relapsed than in the disease-free group, with variable stain intensity and distribution. KaplanMeier analysis showed that high expression of glutathi- Giant cell tumor (GCT) is a benign bone tumor with fairly high local aggressiveness, and development of lung metastases is rare, occurring in 2% to 5% of cases.1 Histologically, the tumor pattern is formed by a network of spindle-shaped mononuclear stroma cells, round mononuclear histiocytic cells, and multinuclear giant cells similar to osteoclasts.2 Cellular components interact with various factors playing a role in osteoclast function regulation. In fact, precursors of osteoclasts express receptor activator of NF-B that in the presence of macrophage colonystimulating factor and its ligand, receptor activator of NF-B ligand, mediates osteoclast formation by increasing the expression of enzymes that dissolve organic and inorganic components of bone.3,4 At the same time, the endogenous osteoprotegerin counteracts these effects by competing for receptor activator of NF-B ligand and neutralizing it. These interactions may provide information to help develop new approaches to biological therapy of this tumor. Drugs that target the osteolytic process lower recurrence rates associated with morbidity and mortality and are considered useful for new clinical treatments. 5,6

Potential role of heat-shock proteins in giant cell tumors

“…New proteins have been identified that are involved in tumor cell survival and are associated with increased aggressiveness of GCTs that leads, in most cases, to local recurrences and, more rarely, to metastasis (Conti et al, 2011). PTHrP excreted by GCT stromal cells increases bone tumor cell local invasiveness and migration (Mak et al, 2013).…”

“…Histologically, the tumor pattern is formed by a network of multinucleated osteoclastlike giant cells, spindle-shaped fibroblast-like stromal cells, round mononuclear histiocytic cells, and CD68-positive phagocytic histiocytes (Werner, 2006). A few proteomics analysis studies have been reported (Conti et al, 2011;Zhang et al, 2013). However, the use of isobaric tags for relative and absolute quantitation (iTRAQ) technology to identify the differentially expressed proteins associated with GCT has not been reported to date.…”

Heat Shock Protein 70 Expression in Giant Cell Tumor of Bone and Its Clinical Application

“…The process of the identification of Hsp70 was carried out by the use of a comparative proteinanalysis [22]. Identification of Hsp70 expression was carried out in this study because these proteins related to natural history of most tumor and might be the target of therapy in the future [23] [24].…”

Orthopaedic Research in Italy: State of the Art

“…They were unable to promote osteoclast differentiation,although they exerted a strong chemotactic effect on osteoclast precursors (47). Tenascin C (48) and other biomarkers (49) were identified as predictors of aggressive behaviour. The best treatment of giant cell tumor of the sacrum is controversial.…”