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Background Ovarian cancer (OC) is the most lethal gynecological malignancy. The majority of patients recur following surgical resection and chemotherapy, 1 and only 10-15% of patients respond to immune checkpoint blockade therapy.2 Personalized vaccination with tumor lysate-pulsed dendritic cells amplifies T cell responses, including neoepitope-specific T cells, but is not curative.3 Thus, amplifying the pre-existing tumor-specific T cell response is insufficient. While known OC antigens such as WT-1, mesothelin, and PRAME comprise the bulk of tumor-associated antigens that have been targeted in clinical trials, many of these are expressed in non-malignant tissues or exhibit limited presentation on HLA.4 Strategies to target multiple antigens may limit tumor immune escape. We developed a novel method to expand T cells ex vivo for OC specificity using Prussian Blue nanoparticle-photothermal therapy (PBNP-PTT), a method of tumor ablation that elicits immunogenic cell death (ICD).5 Methods Immunogenic OC cell lysate was generated with PBNP-PTT and used to stimulate bone marrow-derived dendritic cells (DCs) isolated from healthy female C57Bl6 mice. Autologous splenic CD3+ T cells were cocultured ex vivo with DCs and then assessed for tumor-specific activation and killing using IFNγ ELISpot and MTS cytotoxicity assays. OC-specific T cells were phenotyped with spectral flow cytometry and evaluated in vivo using a syngeneic mouse model.Results We confirmed PBNP-PTT elicits ICD in murine OC cells and confirmed activation of DCs during priming with OC cell lysate via flow cytometry. Ex vivo expansion of T cells with DCs presenting antigens from PBNP-PTT-generated OC cell lysate produced a pool of CD8+ T cells with enhanced IFNγ and TNFα secretion in tumor coculture compared to non-specifically expanded T cells. OC-expanded T cells exhibited greater tumor cell killing at increasing effector-to-target ratios, but did not demonstrate cytotoxicity in culture with healthy murine immune cells and did not secrete IFNγ in the presence of actin. Tumor-bearing female C57Bl6 mice receiving OC-expanded T cells lived significantly longer and exhibited decreased tumor burden compared to mice receiving non-specifically expanded T cells. These mice had greater proportions of B cells in the ascites compared to untreated mice, and had significantly more adoptively transferred T cells expressing TNFα in vivo compared to mice receiving non-specific T cells.Conclusions Ex vivo expansion of T cells using PBNP-PTT to generate cancer cell lysate produces OC-specific CD8+ T cells with potent anti-tumor activity that increases survival in vivo. These data demonstrate a novel method of generating potent T cell products against OC.References Pignata S, Cecere SC, Du Bois A, Harter P, Heitz, F. Treatment of recurrent ovarian cancer. Ann Oncol. 2017;28:51-56.Pawłowsk A, Rekowska A, Kuryło W, Pańczyszyn A, Kotarski J, Wertel I. Current understanding on why ovarian cancer is resistant to immune checkpoint inhibitors. Int J Mol Sci. 2023;24.Rojas LA, Sethna Z, Soares KC, et al. Personalized RNA neoantigen vaccines stimulate T cells in pancreatic cancer. Nat. 2023;618:144-150.Vázquez-García I, Uhlitz F, Ceglia N, et al. Ovarian cancer mutational processes drive site-specific immune evasion. Nat. 2022;612:778–786.Sweeney EE, Cano-Mejia J, Fernandes R. Photothermal therapy generates a thermal window of immunogenic cell death in neuroblastoma. Small. 2018;14.Ethics Approval The animal studies discussed in the abstract received ethics approval by the Institutional Animal Care and Use Committee (IACUC) under the IACUC-approved protocols #A2024-111 and #A2023-083 at the George Washington University.