Abstract 5072: Inhibition of EZH2 as a therapeutic strategy for osteosarcoma

Abstract

Abstract Introduction: Osteosarcoma is the most common primary malignant bone tumor. About 30% of osteosarcoma patients experience relapse and die of their disease. We previously demonstrated that IL-11Rα is overexpressed in primary and metastatic osteosarcoma and plays a role in the development of metastases. Our recent microarray data from KRIB, SJSA1, and LM7 IL-11Rα shRNA cells show decreased expression of several components of polycomb repressive complex 2 (PRC2). EZH2 is the catalytic subunit of PRC2 and blocks transcription of numerous tumor suppressors’ genes. EZH2 has been widely implicated in cancer and inhibition of its catalytic activity has recently emerged as a novel approach to treat cancers. Purpose: To explore the effects and mechanisms of inhibition of EZH2 on osteosarcoma metastasis. Inhibition was tested using GSK126, a novel small-molecule EZH2 inhibitor that competes with methyl group donor S-adenosyl-methionine in a highly selective fashion. Methods: Proliferation assays were performed on a panel of osteosarcoma cell lines subjected to increasing concentrations (0.0625 μm to 20 μm) of GSK126 for 3 days. The effects of IL-11 in PRC1 complex histone H3 methylation (H3K27) were studied by Western blotting. For in vivo experiments, we used an orthotopic metastatic model. Luciferase-labeled SJSA1 osteosarcoma cells were injected into the tibias of 4- to 5-week-old nude mice. After 2 weeks, mice were treated with vehicle or GSK126 three times per week intraperitoneally at 0.1 ml per 20 g of body weight in 20% captisol for 48 days. The tumor-containing legs were amputated when tumor volume reached 2 cm. Primary tumors were measured with calipers, and tumorigenesis and metastasis were evaluated by luciferase imaging. After 4 more weeks of treatment, all the mice were killed and their lungs collected. Immunohistochemical study for EZH2 (1:200, D2C9, Cell Signaling) was performed on tissue microarrays composed of 200 formalin-fixed decalcified human OS specimens: 141 primary and 59 metastatic. Intensity and extent of tumoral labeling were evaluated. Results: EZH2 expression was seen in 79/200 OS specimens (40%) with 54 specimens (27%) with moderate to strong labeling. In the majority of cases, when staining was present, more than 50% of labeling was seen. More metastatic specimens (24/59, 41%) had moderate to strong labeling than primary resection specimens (30/141, 21%). In vitro, SJSA1, HOS and CCH-OS-D osteosarcoma cell lines were sensitive to EZH2 inhibition (IC50= 3.576- 6.450uM). In vivo, mice treated with GSK126 developed significantly fewer osteosarcoma lung metastases than control mice injected with vehicle alone (P<0.01). Conclusion: GSK126, a potent small-molecule EZH2 inhibitor, inhibits the development of osteosarcoma lung metastases in vivo. Thus, GSK126 may be considered as a novel anticancer drug candidate for osteosarcoma. Citation Format: Eswaran Devarajan, Wei-lin Wang, Jen-Wei Tsai, Andrew Futreal, Valerae O. Lewis. Inhibition of EZH2 as a therapeutic strategy for osteosarcoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5072. doi:10.1158/1538-7445.AM2017-5072