CCSG Pilot: Radiation-Derived Extracellular Vesicles Promote Prostate Cancer Colonization in Bone Through Activation of the mtROS–RANKL–TFAM Axis

Abstract Bone metastasis remains a major cause of morbidity and mortality in advanced prostate cancer. Although radiation therapy (RT) is effective for local tumor control, its unintended systemic effects on metastatic progression remain poorly understood. We propose that irradiated prostate tumors release biologically altered extracellular vesicles (RT-EVs) that remotely condition bone as a tumor-promoting premetastatic niche. Our central hypothesis is that RT-EVs establish permissive, immunosuppressive bone microenvironments through two coordinated mechanisms: first, by activating a previously unrecognized mitochondrial ROS (mtROS)- RANKL-TFAM signaling axis in osteoclasts that drives bone remodeling and facilitates metastatic seeding; and second, by inducing mtROS/Complex III-dependent IL-10 production in myeloid cells, thereby suppressing immune surveillance and supporting metastatic colonization. This model shifts the field from a tumor-intrinsic view of radiation response to a tumor-extrinsic framework in which RT systemically reprograms bone through EV-mediated mitochondrial signaling. To test this hypothesis, we will use complementary in vivo and ex vivo models, including tumor-conditioning, a high-resolution bone-in-culture system to study early bone seeding events, and mechanistic studies in osteoclast and myeloid cell populations. We will define how RT-EVs regulate mtROS, RANKL, TFAM, Complex III activity, and IL-10 production; determine whether inhibition of these pathways disrupts niche formation and reduces metastatic burden; and validate key findings using human datasets and tissue-based approaches, including single-cell and spatial transcriptomic analyses of prostate cancer bone and visceral metastatic specimens. The expected outcome is a first-in-field mechanistic model linking RT-induced EV signaling to osteoclast-driven remodeling and myeloid-cell-mediated immunosuppression in bone. These studies will identify actionable therapeutic targets and candidate circulating biomarkers, including EV-associated mitochondrial signatures such as TFAM, for future strategies aimed at preventing metastatic relapse after RT in prostate cancer. This project has strong translational and clinical relevance because it addresses how a standard-of-care local therapy may unintentionally promote metastatic relapse, and it is designed to generate targetable pathways and circulating EV/mitochondrial biomarkers that could be developed for men at high risk of recurrence after RT as well as for patients with established bone-metastatic disease. Inter- programmatic collaboration will integrate expertise in prostate cancer biology, redox biology, radiation oncology, and tumor immunology, while Shared Resource use will include Biospecimen Procurement and Translational Pathology resources for FFPE processing and IHC staining; Flow Cytometry resources for immune phenotyping; and Redox Metabolism resources for mitochondrial metabolism assays.