Abstract A65: Mechanisms of melanoma cell resistance to phagocytosis

Abstract

Abstract Melanoma is notoriously resistant to apoptosis and to other cytolytic mechanisms used by immune cells; however, the mechanisms by which melanoma cells evade phagocytosis by the innate immune system are incompletely understood. Previous studies in our lab revealed that melanoma cells from human, mouse, and dog tumors display an evolutionarily conserved resistance to phagocytosis that was not observed in lymphoma cells or in other solid tumors from these three species. Blockade of CD47 to disable “don't eat me” signals and doxorubicin treatment to enhance the “eat me” signals calreticulin and phosphatidylserine lead to small increases in melanoma cell phagocytosis in vitro. However, this increase did not appear to be biologically relevant, as combination chemo-immunotherapy did not consistently promote enhanced activation of antigen-specific T cells in the tumor draining lymph node or reduce tumor burden in the B16 melanoma model. From these studies, we concluded that melanoma cells display an evolutionarily conserved resistance to phagocytosis, which cannot be fully mitigated by modulation of known pro- and anti-phagocytic pathways. Therefore, we hypothesize that melanoma cells possess an uncharacterized mechanism of resistance to phagocytosis, such as expression of a soluble or membrane bound “don't eat me” signal. To test for the presence of a soluble anti-phagocytic factor, we performed a series of assays in which human lymphoma (Raji) cells, shown to be readily phagocytosed by mouse macrophages, were placed in supernatant harvested from cultured human melanoma (M21) cells. Blockade of the known “don't eat me” signal CD47 was achieved using a SIRPα; mimotope, CV1-G4, or an isotype control. Lymphoma cell phagocytosis by mouse macrophages and response to CD47 blockade were unaffected by the presence of melanoma cell supernatant, suggesting that a secreted factor is not responsible for resistance. To test for the presence of an uncharacterized, membrane-bound “don't eat me” signal, we designed a siRNA panel containing 48 genes whose protein products are expressed on the cell surface of melanoma cells. In ongoing experiments, we are testing how knockdown of these proteins through RNA interference affects macrophage-mediated phagocytosis of melanoma cells and response to CD47 blockade. Citation Format: Katie L. Anderson, Kristin M. Snyder, Debra Lins, Kipp Weiskopf, Yoji Shimizu, Irving Weissman, Matthew Mescher, Jaime Modiano. Mechanisms of melanoma cell resistance to phagocytosis. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr A65.