Effects of a Combined Therapy of Bortezomib and Ionizing Radiation on Chondrosarcoma Three-dimensional Spheroid Cultures

Chondrosarcomas represent a heterogeneous group of primary bone cancers that are characterized by hyaline cartilaginous neoplastic tissue and are predominantly resistant to radiation and chemotherapy. However, adjuvant radiotherapy is often recommended in inoperable cases or after incomplete resections. To improve the efficiency of treatment, the present study tested a combination therapy with ionizing radiation (IR) and the proteasome inhibitor bortezomib. Using a three-dimensional (3D) spheroid model, 0-20 Gy of IR was applied to chondrosarcoma cells and healthy human chondrocytes. Following combined treatment with IR and bortezomib, the cell cycle distribution, apoptotic induction, the survivin pathway, autophagy and DNA damage were evaluated. Both cell types exhibited a slight decrease in viability following increasing doses of IR; the chondrosarcoma cells demonstrated a significant dose-dependent increase in the expression levels of the DNA damage marker histone H2AX phosphorylation at serine 139 (γH2AX). The combination treatment with bortezomib significantly decreased the cell viability after 48 h compared with that in irradiated cells. High-dose IR induced a G/M phase arrest, which was accompanied by a decrease in the number of cells at the G and S phase. Co-treatment with bortezomib changed the distribution of the cell cycle phases. The mRNA expression levels of the proapoptotic genes Bcl-2-associated X protein (Bax) and Bak were significantly increased by bortezomib treatment and combination therapy with IR. In addition, the combination therapy resulted in a synergistic decrease of the expression levels of survivin and its corresponding downstream pathway molecules, including heat shock protein 90, X-linked inhibitor of apoptosis protein, smad 2 and smad 3. Comparative analyses of γH2AX at 1 and 24 h post-IR revealed efficient DNA repair in human chondrosarcoma cells. Therefore, additional bortezomib treatment may only temporarily improve the radiation sensitivity of chondrosarcoma cells. However, the inhibition of the survivin pathway by the combined treatment with IR and bortezomib, observed in the present study, revealed a novel aspect in the tumor biology of chondrosarcoma 3D spheroid cultures and may represent a potential target for therapy.