Abstract 2832: Multiplex fluorescent imaging of the metastatic melanoma microenvironment reveals spatially-dependent macrophage interactions modulating tumor-infiltrating T-cells

Abstract Tumor-infiltrating lymphocytes (TIL) hold strong prognostic value in determining patient survival as well as response to immune checkpoint blockade across many tumor types, especially metastatic melanoma. Tumors often evolve mechanisms to prevent TIL access and/or function to allow for continued growth. One such mechanism is the recruitment and reprogramming of tumor-associated macrophages (TAM) which suppress TILs through a variety of pathways. The interactions between TAMs and TILs that result in a suppressive tumor microenvironment (TME) have been studied in vitro and with spatially-agnostic sequencing, leaving the location-specific contributions underexplored. Interrogating the TAM-TIL interactions in an intact TME will uncover novel mechanisms responsible for TAM function and may be key to reversing TIL immunosuppression, improving anti-tumor immune responses, survival, and potentially therapy response. We use histocytometry, a multiplex quantitative tissue imaging method capable of capturing whole-slide tumor sections, to explore the human metastatic melanoma TME in situ and resolve the spatial contributions of TAM-TIL interactions. We demonstrate a T-cell preference for physical contact with phagocytic TAMs in the tumor but non-phagocytic TAMs in the stroma. We further model immune synapses from histocytometry to probe the function underlying the TAM-TIL preference based on the directionality of signaling components. The model suggests T-cells in contact with macrophages in the tumor are communicating more frequently with the phagocytic TAMs as compared to their non-phagocytic counterparts, indicating that this interaction extends beyond a simple proximity preference. In addition, proliferating T-cells communicate more frequently with macrophages than non-proliferating T-cells but this effect limited to the intratumor space. Together we demonstrate the functional consequences of physical TAM-TIL interactions with unprecedented spatial resolution. This work unravels important TAM interactions that shape the TME which have not been previously appreciated, providing novel insight into the forces modifying T-cell function in the TME and revealing new TAM biology to explore further. Citation Format: Victor G. Wang, Jan Martinek, Ananya Gulati, Hannah Boruchov, Kelly Ray, Karolina Palucka, Jeffrey H. Chuang. Multiplex fluorescent imaging of the metastatic melanoma microenvironment reveals spatially-dependent macrophage interactions modulating tumor-infiltrating T-cells [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2832.