2207 - Riluzole as a Dual-Targeted Radiosensitizer for Osteosarcoma: Targeting Tumor Cells and Angiogenic Vasculature to Enhance Single High Dose Radiotherapy Efficacy

Purpose/Objective(s): Osteosarcoma (OS), a highly vascular and aggressive bone cancer prevalent among young adults, presents significant treatment challenges due to its resistance to conventional radiation therapy. Consequently, effective radiosensitizers are crucial for improving therapeutic outcomes in this malignancy. Single high-dose radiation therapy (SDRT) has emerged as a promising approach for overcoming radioresistance in highly vascularized sarcomas. However, anti-angiogenic monotherapies targeting tumor vasculature alone have shown limited clinical success as radiosensitizers. Thus, there is a critical need for compounds capable of simultaneously targeting both tumor cells and angiogenic endothelial cells to maximize the therapeutic efficacy of SDRT. Riluzole, an FDA-approved drug for Amyotrophic Lateral Sclerosis, is being investigated for repurposing in OS treatment. Its multifaceted mechanism of action encompassing glutamate release inhibition, suppression of antioxidant defenses, and angiogenesis inhibition, highlights its potential as a radiosensitizer. We hypothesize that Riluzole enhances the radiosensitivity of OS tumors to SDRT by concurrently targeting tumor cells and their vasculature.

Materials/Methods: To assess the radiosensitizing effects of Riluzole, OS cells (LM7 (metastatic, human); OS482 (metastatic, mouse)) were pre-treated with Riluzole prior to SDRT exposure. Clonogenic survival was evaluated using a colony assay, while apoptosis was quantified through Hoechst 33342 staining and confirmed via western blot analysis. Intracellular reactive oxygen species (ROS) levels were measured using a fluorometric ROS assay in OS and bovine aortic endothelial cells. The impact of Riluzole on DNA damage and repair was further assessed using western blot and flow cytometric analysis. Anti-angiogenic effect in OS cells was assessed by quantifying VEGFA expression using western blot.

Results: Our preliminary findings indicate that Riluzole sensitizes osteosarcoma cells to SDRT-induced apoptosis and significantly reduces the survival fraction of irradiated cells compared to radiation alone. We identified that Riluzole-induced radiosensitization is mediated through oxidative stress augmentation and impaired DNA repair. Riluzole enhances radiation-induced ROS generation, inhibits the DNA repair enzyme PARP1, and exacerbates radiation-induced DNA damage. Additionally, Riluzole suppresses SDRT-induced VEGFA expression, signifying its potential antiangiogenic effects. Furthermore, Riluzole induces ROS production and reduces endothelial cell viability, supporting its dual mechanism of radiosensitization.

Conclusion: Our study demonstrates Riluzole as a potent radiosensitizer for OS, effectively targeting both tumor cells and its vasculature. We aim to translate these findings into preclinical studies using an orthotopic osteosarcoma mouse model, with important implications for metastatic patients undergoing SDRT.