Abstract 1170: Proton-induced exosomal signatures reveal radiation response pathways in pediatric diffuse midline glioma.

Abstract Diffuse intrinsic pontine glioma (DIPG) primarily affects children between five and ten years of age and carries a poor prognosis, with median survival of nine to twelve months. Standard treatment options are limited and typically include radiation therapy, which provides only temporary improvement, while chemotherapy and targeted therapy trials remain ongoing. DIPG remains one of the most lethal pediatric brain tumors, and emerging evidence suggests that extracellular vesicles such as exosomes may contribute to treatment resistance by carrying cargo that regulates apoptosis, DNA damage signaling, and intercellular survival pathways. Despite growing interest in exosome-mediated communication, the radiation-dependent dynamics of exosome release in DIPG remain understudied and poorly defined. This study aims to characterize radiation response features in DIPG1 and DIPG16A by comparing proliferation behavior, cell cycle distribution, and apoptosis and necrosis patterns after proton exposure, while establishing early and late exosomal dynamics through quantification of exosome release and size distribution. Here, we provide early evidence that proton radiation induces distinct cellular stress phenotypes accompanied by measurable and time-dependent exosomal output, suggesting an active radiation-responsive vesicle program. DIPG16A was derived from a treatment-naïve pretreatment biopsy specimen, whereas DIPG1 was obtained from a post-treatment autopsy specimen following exposure to radiation and multiple chemotherapeutic regimens. Both cell lines were cultured under standard conditions, exposed to graded proton doses, and exosomes were isolated and analyzed using nanoparticle tracking. A summary analysis of global treatment response revealed model-specific differences that may reflect prior therapeutic exposure and intrinsic tumor biology. Proliferation assays demonstrated that DIPG16A was more sensitive to radiation than DIPG1. DIPG16A showed a higher apoptotic population after irradiation, while both models displayed minimal necrosis and neither line showed detectable cell cycle arrest. Nanoparticle tracking analysis confirmed measurable exosome release at both early and late intervals in each line, establishing the foundation for future work investigating how apoptosis inhibitor proteins within exosomes may contribute to exosome-mediated radiation resistance in diffuse midline glioma. Collectively, these findings highlight proton-induced exosomal signaling as a potentially important and previously underappreciated component of treatment response in pediatric diffuse midline glioma and support further mechanistic exploration of exosome-carried survival factors. Generative artificial intelligence was used only for text editing and clarity, and all scientific content was produced and verified by the authors. Citation Format: Ann Morcos, Yeonkyu Jung, Nathan R. Wall. Proton-induced exosomal signatures reveal radiation response pathways in pediatric diffuse midline glioma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 1170.