(T0930-03-16) Oxaliplatin Inhibits Oncogenic Signaling to Potentiate Radiation Response in Preclinical Models of Radioresistant Glioblastoma and Outperforms the Clinical Standard-of-Care Temozolomide

Purpose: Glioblastoma (GBM) is the most aggressive and lethal primary brain tumor in adults and is distinguished by its rapid growth, tremendous infiltration capacity, and persistent resistance to treatment. The current standard-of-care, maximal surgical resection followed by concurrent radiation therapy (RT) and oral temozolomide (TMZ) chemotherapy, achieves only modest survival benefits, with a median survival of less than 18 months from patient diagnosis. A major driver of GBM's radioresistance is the aberrant activation of Signal Transducer and Activator of Transcription 3 (STAT3), a transcription factor that promotes tumor progression, immune evasion, and treatment resistance. Despite advances in RT techniques, survival outcomes remain stagnant, underscoring the urgent need for radiosensitizers that target GBM’s molecular mechanisms of resistance. Platinum-based chemotherapeutics offer a promising approach, not only through their DNA-damaging effects but also by modulating oncogenic signaling pathways. However, their ability to inhibit phosphorylated STAT3 (pSTAT3), a hallmark of GBM pathology, and potentiate radiation efficacy remains underexplored. This study aims to evaluate FDA-approved platinum therapeutics, focusing on oxaliplatin (OXA), for novel applications in GBM radiosensitization. Preliminary in vitro findings in a preclinical model of GBM suggest that OXA inhibits pSTAT3 and significantly augments radiation efficacy. Such findings demonstrate potential for OXA to be repurposed as a radiosensitizer for GBM treatment.

Methods: To assess the efficacy of OXA, a third-generation platinum drug, as a potential radiosensitizer for GBM treatment, multiple in vitro assays were performed on KR158 murine glioma cells, a preclinical model frequently used in GBM research. Western blot analyses were performed to assess temporal changes in pSTAT3 expression following treatment with OXA or TMZ over 3–24 hours. Protein lysates were collected at each timepoint, and pSTAT3 expression levels quantified via immunoblotting. To evaluate the radiation dose response of KR158 cells, the cells were first exposed to radiation at doses ranging from 2 to 10 Gy, and then a clonogenic assay was performed to identify the radiation doses at which they exhibit resistance. Findings from this study are critical in informing the selection of appropriate radiation doses for drug-radiation combination studies aimed at sensitizing radioresistant GBM cells to radiation. From the results of this study, a subsequent clonogenic assay was conducted (at radiation doses that KR158 cells demonstrated resistance to) to evaluate cell survival and self-renewal capacity following treatment with radiation alone, oxaliplatin alone, or their combination, and compared to the TMZ + radiation clinical standard. All experiments were carried out in at least biological triplicates with appropriate controls, and data were analyzed in Graphpad Prism using one-way ANOVA and appropriate post-hoc tests to determine statistical significance.

Results: Western blot analysis revealed that OXA treatment led to a significant, time-dependent reduction in pSTAT3 expression, with the 3-hour treatment showing the most pSTAT3 inhibition. In contrast, the TMZ-treated cells showed no substantial pSTAT3 inhibition at any of the time points tested, especially compared to the OXA-treated cells. An analysis of the radiation dose response study performed on the KR158 cells demonstrated no significant reduction in surviving fractions at 2 Gy and 4 Gy compared to the untreated control, indicating their capacity to survive and proliferate at those doses. This suggests that the cells possess an inherent degree of radioresistance within the 2-4 Gy dose range, consistent with the treatment-refractory nature of GBM. In contrast, exposure to radiation doses above 4 Gy resulted in a significant, dose-dependent decrease in surviving fractions, reflecting increased radiation-induced cytotoxicity at higher doses. By identifying 2 Gy and 4 Gy as sub-lethal radiation doses in this model, we establish a biologically relevant framework for testing drugs that may potentiate radiation effects in otherwise resistant glioma cells. Quantitative analysis of the clonogenic data obtained from the drug-radiation combination experiments revealed that OXA greatly reduced clonogenic survival when combined with radiation, at both 2 Gy and 4 Gy, compared to radiation treatment alone. Notably, the OXA-radiation combination significantly outperformed the clinically used TMZ-radiation regimen, suggesting a mechanistic link between pSTAT3 inhibition and enhanced radiosensitivity. These findings indicate that OXA, but not TMZ, enhances radiation response in radioresistant KR158 glioma cells.

Conclusion: Overall, these results identify OXA as a promising radiosensitizer capable of targeting STAT3 signaling to overcome the intrinsic radioresistance in GBM cells-a major clinical challenge in GBM treatment. Future work will focus on developing a liposomal oxaliplatin formulation to improve drug delivery, tumor accumulation, and tumor retention. Additionally, these in vitro findings will be extended to in vivo GBM models to assess therapeutic efficacy and further validate the potential for clinical translation. Collectively, this study lays the foundation for platinum-based combination therapies as a viable strategy to improve outcomes for GBM patients.<