Abstract The central purpose of this study is to uncover and mechanistically describe the role of Hedgehog (Hh) signaling in CD4+ regulatory T cells (Tregs) in the context of murine triple negative breast cancer (TNBC). We employ a novel genetically engineered mouse model, murine TNBC models, flow cytometry, and targeted molecular assays to provide compelling evidence that inhibiting Hh activity in Tregs mitigates their suppressive function and subsequently slows tumor growth. TNBC is an aggressive form of breast cancer with limited treatment options. Any therapeutic benefits can be stifled by an immunosuppressive tumor microenvironment (TME) often found in TNBC due to an abundance of Tregs. Tregs are under transcriptional control of Foxp3, suppress other tumor-infiltrating lymphocytes and innate immune cells, and are associated with worse patient outcomes.Hh signaling normally plays a key role in embryonic development, tissue regeneration, and stem-cell renewal. Importantly, Hh signaling is aberrantly upregulated in a quarter of all cancers, and in TNBC, it has been implicated in drug resistance, tumor growth, migration, and invasion. Recent work from our lab has demonstrated that systemic pharmacological inhibition of Hh signaling led to a decrease in Treg abundance and function in the TME of TNBC.In this study, we report that inducible ablation of the Hh transcription factor, Gli2, in a Foxp3-dependent manner, impedes mammary tumor growth. The resultant TME is characterized by decreased abundance of Tregs and an increase in inflammatory Th17s. To uncover the mechanism behind these changes, we utilized RNA-seq and chromatin immunoprecipitation qPCR (ChIP-qPCR). Our data demonstrate that Gli2 ablation alters the transcriptional controls in Tregs leading them to pivot towards inflammatory Th17 cells. These changes in Tregs functionally reduce the immunosuppressive nature of the TME of TNBC and permit an inflammatory TME, thereby reducing tumor growth. Cumulatively, these findings introduce a novel strategy for immunotherapy to target and impair Tregs in the TME of TNBC. Citation Format: Ian Miranda, Courtney Swain, Brandon J. Metge, Lalita Shevde. Treg-specific inhibition of hedgehog signaling alters their function and impedes the growth of murine triple-negative breast cancer 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 5003.
Miranda et al. (Fri,) studied this question.