Abstract 6032: Intratumoral Tregs restrain the immune response against primary and metastatic cancer

Abstract Advancements in cancer immunotherapies have significantly improved patient outcomes in hematologic malignancies but are dramatically limited by the suppressive function of Regulatory T cells (Tregs) in solid cancers. Antibody-based therapeutics, such as anti-CTLA-4 and anti-CCR8, are being tested to selectively deplete intratumoral Tregs, with promising preclinical efficacy. However, these therapies have multiple effects that may do more than just deplete intratumoral Tregs, such as activate Fc receptors, bind to other types of immune cells, or bind to cells outside of the tumor microenvironment. While Tregs highly infiltrate most tumor types, it is unclear whether they function to suppress cancer immunity only in tumor tissue itself or at peripheral sites. To test whether Tregs function directly within the tumor microenvironment to suppress antitumor immune responses, we developed a model in which intratumoral (IT) delivery of low-dose diphtheria toxin (DT) in Foxp3DTR mice is used to only deplete IT Tregs. We show that mice that receive IT DT have reduced Tregs frequencies only in the tumor and do not succumb to lymphoproliferative disease, as observed with systemic Treg depletion. Critically, tumor Treg ablation via IT DT produced more effective and durable tumor control than systemic Treg ablation. Mechanistically, we show that robust tumor control is achieved through enhanced function of effector CD4+ and CD8+ T cells and requires MHC II+ CD11b+ Batf3- type 2 conventional dendritic cells. Through the use of disseminated tumor models, we find that disrupting Treg suppression within a primary tumor unleashes antitumor immunity against metastatic lesions that are not directly treated. Interestingly, this multifocal tumor control is dependent upon the cooperation between CD4+ and CD8+ T cells and requires matched primary and metastatic tumors. Through in vivo cellular trafficking experiments and blockade of T cell egress from lymphoid tissues we show a high frequency of Tregs circulate between local and metastatic tumors to mediate systemic immune effects. Upon disruption of tumor-Treg circulation between lesions, we demonstrate enhanced effector function and decreased disease burden at both tumor sites. These findings identify a tumor-resident Treg niche that limits CD4+-cDC2 mediated antitumor immune responses. Importantly, our results uncover a novel Treg suppressive mechanism that promotes systemic cancer progression. Citation Format: Alissa Bockman, Mariela Moreno Ayala, Brenna Gittins, Chenyu Zhang, Skye Benyon, Michel DuPage. Intratumoral Tregs restrain the immune response against primary and metastatic cancer [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 6032.