Abstract A051: Bitter agonists increase macrophage phagocytosis of head and neck squamous cell carcinoma cells

Abstract Head and neck squamous cell carcinoma (HNSCC) is the 7th most commonly diagnosed cancer worldwide and is expected to increase in incidence by 30% by 2030. The HNSCC microenvironment is characterized by substantial macrophage infiltration. However, tumor-associated macrophages often cultivate an immunosuppressive environment that aids tumor progression. Previous work has shown that macrophages express bitter taste receptors (T2Rs), a class of GPCRs that can be activated with bitter compounds. Previous work has also demonstrated that stimulation of T2Rs in macrophages with bitter agonists, such as denatonium benzoate, increases phagocytosis of bacteria, but the impact of bitterants on macrophage phagocytosis of tumor cells is unknown. The goal of this study was to assess the impact of bitter compounds on macrophage phagocytosis of HNSCC cells in vitro. Human peripheral blood mononuclear cells obtained from healthy donors were used to generate monocyte-derived macrophages (MDMs). To investigate the effects of bitter agonist stimulation on macrophage phagocytosis, CFSE-labeled MDMs and DAPI-labeled HNSCC cell line SCC47, were co-stimulated with denatonium benzoate, and macrophage phagocytosis was quantified. This revealed that treatment with denatonium benzoate led to a 70% increase in macrophage phagocytosis compared to controls. Additional analysis revealed a 1.8-fold increase in F-actin in MDMs in response to denatonium benzoate measured via CellMask™ Actin staining, and that macrophages had generated F-actin positive tunneling nanotubes (TNTs). Further characterization with live cell imaging showed that these TNTs were being utilized as a means of mitochondrial transfer as observed by MitoTracker™ and CellMask™ Actin staining. Furthermore, there was a 2.5-fold decrease in mitochondrial ATP production in MDMs stimulated with denatonium benzoate, possibly indicating metabolic reprogramming. Timelapse imaging of MDMs demonstrated a reduction in mitochondrial ATP production, with a concomitant increase in F-actin production. These observations were confirmed in Phorbol 12-myristate 13-acetate differentiated THP-1 cells. Taken together, these results indicate that bitter taste receptor stimulation may contribute to a more activated macrophage phenotype denoted by a decrease in mitochondrial ATP production, increased F-actin staining, and increased phagocytosis of tumor cells. Citation Format: Brianna L Hill, Sarah M Sywanycz, Zoey A Miller, Robert J Lee, Ryan M Carey. Bitter agonists increase macrophage phagocytosis of head and neck squamous cell carcinoma cells [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2024 Oct 18-21; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2024;12(10 Suppl):Abstract nr A051.