Abstract 3226: Intratumoral manipulation of melanoma using novel hyperthermic treatment to enhance anti-PD-L1 therapy

Abstract Despite tremendous advancement in the understanding and manipulation of immune checkpoint molecules in immunotherapeutic design, there are still vast limitations in treatment efficacy. Immunotherapy manipulates the immune checkpoint blockade (ICB), blocking negative regulators of the immune system-preventing tumor immune evasion. Combination therapy enhances the mechanistic actions of each individual component, enhancing tumoricidal behavior. Thermal ablation to induce cancer cell death as a therapeutic adjuvant is an understudied area. B16-F10 melanoma cells were grown at 37°C or 41°C. B16-F10 cells grown at 41°C decreased cell migration by 70% in 24 hours. B16-F10 cell proliferation was decreased at 41°C by 62% at 48 hours and 94% at 72 hours. Protein expression profiles were evaluated under hyperthermic conditions in vitro. Expression of pERK and ERK decreased by 86% and 50% and Caspase-3 increased by 31% at 41°C. Cell stress can induce death pathways and the heat shock protein response simultaneously, therefore we investigated inducible Hsp70 expression. Hsp70 expression was undetectable at 37°C and 188% at 41°C. Cytokine analysis of conditioned media showed that at 41°C, TNF-α expression was increased and IL-4 expression was decreased, suggesting a proinflammatory shift in cytokine profiles at higher temperatures. In support of our data, hyperthermia-induced TNF-α apoptotic responses have been reported. Programmed cell death ligand (PD-L1) is expressed by tumors to suppress immune cells and induce tolerogenic phenotypes within the tumor microenvironment. We observed that hyperthermic potentiation decreased the expression of this checkpoint molecule in B16-F10 by 35%. Therefore, we hypothesize that hyperthermia modulates the tumor microenvironment to enhance their responsiveness to checkpoint immunotherapies. Based on these promising in vitro results, we plan to further explore the clinical aspect of our findings through a translation in vivo. B16-F10 cells will be implanted onto C57BL/6 mice and treated with infrared radiation in combination with either anti-PD-L1, anti-PD-1, or IL-15. Citation Format: Nicole DeSouza, Danielle Quaranto, Kaci Kopec, Jan Geliebter, Raj K. Tiwari, Mark D. Hurwitz. Intratumoral manipulation of melanoma using novel hyperthermic treatment to enhance anti-PD-L1 therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3226.