Abstract A051: Melanoma displays an evolutionarily conserved resistance to upregulation of prophagocytic signals and to CD47 blockade

Abstract

Therapeutic activation of macrophage phagocytosis has the ability to restrain tumor growth through phagocytic clearance of tumor cells and activation of the adaptive immune response. Our objective for this study was to evaluate the effects of modulating pro- and anti-phagocytic pathways in malignant melanoma. In order to identify evolutionarily conserved mechanisms of resistance that may be important for melanoma cell survival, we utilized a multispecies approach and examined the phagocytosis of human, mouse, and dog melanoma cells in vitro. We blocked the interaction between CD47 on tumor cells and SIRPα on macrophages using anti-CD47 monoclonal antibodies, SIRPα mimotopes, and a soluble fusion protein of the SIRPα extracellular domain. In all cases, we confirmed that these reagents blocked >85% of the maximum binding of CD47 to recombinant SIRPα. We observed that melanoma cells from all three species displayed unexpected resistance to phagocytosis that could not be fully mitigated by blockade of the “don’t eat me” signal CD47 or by chemotherapeutic enhancement of known “eat me” signals. In vitro, CD47 blockade minimally enhanced phagocytosis of melanoma cells, and loss of CD47 expression did not increase sensitivity to phagocytosis. In vivo, CD47 blockade enhanced the proliferation of tumor-specific CD8+ T-cells, but this response failed to inhibit tumor growth. Resistance to phagocytosis was not mediated by soluble factors, as phagocytosis of melanoma cells was not enhanced by inhibition of secretory pathways, and phagocytosis of sensitive lymphoma tumor cells was not impaired in the presence of melanoma cells or melanoma culture supernatants. siRNA-mediated knockdown of 47 prospective “don’t eat me” signals similarly did not enhance melanoma cell phagocytosis. Interestingly, in lymphoma cells, CD47 knockout alone did not enhance phagocytosis, suggesting that at least part of the pro-phagocytic effect of CD47 blockade is due to Fc receptor engagement. Restoration of CD47 expression in lymphoma cells re-established sensitivity to CD47 blockade. We conclude that melanoma cells possess an evolutionarily conserved resistance to macrophage phagocytosis. Further investigation will be needed to define and overcome the mechanisms that mediate melanoma cell resistance to innate immunity. Citation Format: Katie L. Anderson, Kristin M. Snyder, Daisuke Ito, Debra C. Lins, Lauren J. Mills, Kipp Weiskopf, Nan G. Ring, Aaron M Ring, Yoji Shimizu, Matthew F. Mescher, Irving L. Weissman, Jaime F. Modiano. Melanoma displays an evolutionarily conserved resistance to upregulation of prophagocytic signals and to CD47 blockade [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A051.