Tumor-to-endothelium mitochondrial transfer licenses endothelial cells for CD8(+) T cell recognition via mitochondrial neoantigen presentation.

Renal cell carcinoma (RCC) frequently exhibits resistance to immune checkpoint blockade, highlighting the need for strategies that enhance tumor-specific T cell priming and improve immune access to the tumor microenvironment. Here we show that vaccination targeting tumor-associated mitochondrial antigens (TAMAs), derived from tumor-specific mitochondrial DNA (mtDNA) missense mutations, synergizes with PD-1/PD-L1 blockade to overcome checkpoint refractoriness in the RENCA RCC model. TAMAs vaccination elicits antigen-specific T cell responses, increases intratumoral CD8(+) T cell infiltration, and reduces immunosuppressive myeloid populations, resulting in delayed tumor progression and improved survival when combined with checkpoint inhibition. In parallel, TAMAs + checkpoint blockade induces vascular remodeling characterized by increased pericyte coverage, reduced vascular leakage, improved perfusion and reduced hypoxia. Mechanistically, vascular remodeling is driven by CD8(+) T cell-dependent, IFNγ-associated immune activity and is associated with endothelial apoptosis and diminished intratumoral CD31 signal. We further identify tumor-to-endothelium mitochondrial transfer as a mechanism linking mitochondrial neoantigens to the tumor vascular compartment: tumor-derived mitochondria enter human and mouse endothelial cells in vitro and in vivo, and tumor-associated mtDNA mutations are detectable in endothelial fractions from murine tumors and human RCC specimens. Human endothelial cells can present mitochondrial neoantigens via MHC class I and become targets of TAMAs-specific CD8(+) T cell cytotoxicity, including following mitochondrial acquisition from tumor cells. Together, these findings establish mitochondrial neoantigen immunity as a tractable approach to enhance checkpoint responses and reveal mitochondrial transfer as an antigenic bridge that expands immune targeting to the tumor vasculature.