Abstract 3320: Combined effect of zoledronic acid and telomerase-specific oncolytic virotherapy for human osteosarcoma cells.
Authors: Yamakawa Yasuaki , Hasei Joe , Tazawa Hiroshi , Osaki Shuhei , Sasaki Tsuyoshi , Kunisada Toshiyuki , Yoshida Aki , Urata Yasuo , Ozaki Toshifumi , Fujiwara Toshiyoshi 
Published: August 2014

Abstract
Osteosarcoma is the most frequent type of primary malignant bone tumor and common between the ages of 10 to 25. Based on preoperative and postoperative chemotherapy, the prognosis of osteosarcoma patients has improved. However, 30% of osteosarcoma patients who become refractory to chemotherapy still die from tumor or metastasis. We previously reported that telomerase-specific, replication-competent oncolytic adenovirus (Telomelysin, OBP-301), efficiently killed human bone and soft tissue sarcoma cells but not normal human somatic cells. Recently, third-generation bisphosphonates, zoledronic acid (ZOL), is widely used to inhibit bone destruction in metastatic bone tumor through induction of apoptosis in osteoclast cells in the clinical settings. Moreover, ZOL has been shown to induce the antitumor effect synergistically in combination with chemotherapeutic agents in human sarcoma cells. In this study, we investigated the combined antitumor effect of ZOL and OBP-301 in human osteosarcoma cells. Three human osteosarcoma cell lines, HOS, SaOS-2, U2OS, were used. We used XTT assay to examine the antitumor effect of ZOL and OBP-301 individually and combinatory on days 2, 3, 5 after treatment. ZOL was treated at concentration of 0 to 10μM, and OBP-301 was infected at multiplicity of infections (MOI) of 0 to 50 plaque forming units (PFU)/cell. Synergy between ZOL and OBP-301 was analyzed with the CalcuSyn software (BioSoft). XTT assay showed that treatment of ZOL or OBP-301 decreased the cell viability in a time- and dose-dependent manner. Combination treatment of ZOL and OBP-301 showed synergy and additive effects. After calculation of the half maximal (50%) inhibitory concentration (IC50) of ZOL and dose (ID50) of OBP-301 for each cell, cells were treated with ZOL and OBP-301 for 3 days at concentration of IC50 and ID50, respectively. Whole cell lysates were subjected to western blot analysis for the cleaved PARP (C-PARP) and PARP, LC3, p62 and β-actin. Western blot analysis revealed that ZOL induced apoptosis, which was confirmed by increasing of C- PARP, and OBP-301 induced not only apoptosis but also autophagy, which is confirmed by conversion of LC3- I to LC3- II and p62 down-regulation. Combination treatment showed increasing of these effects synergistically or additively in each cell. These results suggest that combination treatment of ZOL and OBP-301 is a promising antitumor strategy for patients with osteosarcomas. We are now planning to investigate the antitumor and bone-protective effects of combination treatment in an orthotopic xenograft tumor model using 3D-CT imaging system.
Citation Format: Yasuaki Yamakawa, Joe Hasei, Hiroshi Tazawa, Shuhei Osaki, Tsuyoshi Sasaki, Toshiyuki Kunisada, Aki Yoshida, Yasuo Urata, Toshifumi Ozaki, Toshiyoshi Fujiwara. Combined effect of zoledronic acid and telomerase-specific oncolytic virotherapy for human osteosarcoma cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3320. doi:10.1158/1538-7445.AM2013-3320