CD47 blockade synergizes with doxorubicin in enhancing macrophage phagocytosis of Ewing sarcoma cells and deceasing xenograft tumor growth and metastasis in an orthotopic model of Ewing sarcoma
Macrophages are immune cells whose major functions include phagocytosing tumor cells. However, tumors escape macrophage phagocytosis through the expression of anti-phagocytic signals, most commonly CD47. Moreover, in Ewing sarcoma (ES), we found that tumor cells utilize dual mechanisms to evade macrophage clearance by simultaneously over-expressing CD47 and down-regulating cell surface calreticulin (csCRT), the pro-phagocytic signal. Here we utilize a CD47 blockade (magrolimab, MAG) to inhibit the anti-phagocytic signal and a chemotherapy regimen (doxorubicin, DOX) to enhance the pro-phagocytic signal and evaluated the effects of MAG or DOX alone and in combination on macrophage phagocytosis of ES cells in vitro and tumor formation and development in vivo. Furthermore, we performed RNA-Seq to identify potential mechanisms of resistance to the CD47 blockade combinatorial therapy. Overall design: Doxorubicin was utilized to enhance csCRT on ES cells. Magrolimab, a humanized CD47 monoclonal antibody was used to block CD47 signal. An ES orthotopic mouse model was utilized to assess the effect of doxorubicin and magrolimab on ES tumor growth and metastasis and animal survival. Tumors were harvested at the end of the study. Total RNA was extracted from the tumors and bulk RNA-Seq was carried out on NovaSeq 6000.