InVivoMAb anti-mouse CD47 (IAP)

Macrophage phagocytosis of tumor cells mediated by CD47-specific blocking antibodies has been proposed to be the major effector mechanism in xenograft models. Here, using syngeneic immunocompetent mouse tumor models, we reveal that the therapeutic effects of CD47 blockade depend on dendritic cell but not macrophage cross-priming of T cell responses. The therapeutic effects of anti-CD47 antibody therapy were abrogated in T cell-deficient mice. In addition, the antitumor effects of CD47 blockade required expression of the cytosolic DNA sensor STING, but neither MyD88 nor TRIF, in CD11c(+) cells, suggesting that cytosolic sensing of DNA from tumor cells is enhanced by anti-CD47 treatment, further bridging the innate and adaptive responses. Notably, the timing of administration of standard chemotherapy markedly impacted the induction of antitumor T cell responses by CD47 blockade. Together, our findings indicate that CD47 blockade drives T cell-mediated elimination of immunogenic tumors.

Macrophages enforce antitumor immunity by engulfing and killing tumor cells. Although these functions are determined by a balance of stimulatory and inhibitory signals, the role of macrophage metabolism is unknown. Here, we study the capacity of macrophages to circumvent inhibitory activity mediated by CD47 on cancer cells. We show that stimulation with a CpG oligodeoxynucleotide, a Toll-like receptor 9 agonist, evokes changes in the central carbon metabolism of macrophages that enable antitumor activity, including engulfment of CD47(+) cancer cells. CpG activation engenders a metabolic state that requires fatty acid oxidation and shunting of tricarboxylic acid cycle intermediates for de novo lipid biosynthesis. This integration of metabolic inputs is underpinned by carnitine palmitoyltransferase 1A and adenosine tri-phosphate citrate lyase, which, together, impart macrophages with antitumor potential capable of overcoming inhibitory CD47 on cancer cells. Our findings identify central carbon metabolism to be a novel determinant and potential therapeutic target for stimulating antitumor activity by macrophages.

BACKGROUND: Human pancreatic ductal adenocarcinoma (PDAC) responds poorly to immune checkpoint inhibitor (ICPi). While the mechanism is not completely clear, it has been recognized that tumor microenvironment (TME) plays key roles. We investigated if targeting CD47 with a monoclonal antibody could enhance the response of PDAC to ICPi by altering the TME. METHODS: Using immunohistochemistry, we examined tumor-infiltrating CD68(+) pan-macrophages (CD68(+) M) and CD163(+) M2 macrophages (CD163(+) M2) and tumor expression of CD47 and PD-L1 proteins in 106 cases of PDAC. The efficacy of CD47 blockade was examined in xenograft models. CD45(+) immune cells from syngeneic tumor models were subjected to single-cell RNA-sequencing (scRNA-seq) by using the 10x Genomics pipeline. RESULTS: We found that CD47 expression correlated with the level of CD68(+) M but not CD163(+) M2. High levels of tumor-infiltrating CD68(+) M, CD163(+) M2, and CD47 expression were significantly associated with worse survival. CD47(high)/CD68(+) M(high) and CD47(high)/CD163(+) M2(high) correlated significantly with shorter survival, whereas CD47(low)/CD68(+) M(low) and CD47(low)/CD163(+) M2(low) correlated with longer survival. Intriguingly, CD47 blockade decreased the tumor burden in the Panc02 but not in the MPC-83 syngeneic mouse model. Using scRNA-seq, we showed that anti-CD47 treatment significantly remodeled the intratumoral lymphocyte and macrophage compartments in Panc02 tumor-bearing mice by increasing the pro-inflammatory macrophages that exhibit anti-tumor function, while reducing the anti-inflammatory macrophages. Moreover, CD47 blockade not only increased the number of intratumoral CD8(+) T cells, but also remodeled the T cell cluster toward a more activated one. Further, combination therapy targeting both CD47 and PD-L1 resulted in synergistic inhibition of PDAC growth in the MPC-83 but not in Panc02 model. MPC-83 but not Panc02 mice treated with both anti-CD47 and anti-PD-L1 showed increased number of PD-1(+)CD8(+) T cells and enhanced expression of key immune activating genes. CONCLUSION: Our data indicate that CD47 targeting induces compartmental remodeling of tumor-infiltrating immune cells of the TME in PDAC. Different PDAC mouse models exhibited differential response to the anti-CD47 and anti-PD-L1 blockade due to the differential effect of this combination treatment on the infiltrating immune cells and key immune activating genes in the TME established by the different PDAC cell lines.

CD47 is a ubiquitously expressed transmembrane glycoprotein that regulates inflammatory responses and tissue repair. Here, we show that normal mice treated with anti-CD47 antibodies, and Cd47-null mice have impaired intestinal mucosal wound healing. Furthermore, intestinal epithelial cell (IEC)-specific loss of CD47 does not induce spontaneous immune-mediated intestinal barrier disruption but results in defective mucosal repair after biopsy-induced colonic wounding or Dextran Sulfate Sodium (DSS)-induced mucosal damage. In vitro analyses using primary cultures of CD47-deficient murine colonic IEC or human colonoid-derived IEC treated with CD47-blocking antibodies demonstrate impaired epithelial cell migration in wound healing assays. Defective wound repair after CD47 loss is linked to decreased epithelial β1 integrin and focal adhesion signaling, as well as reduced thrombospondin-1 and TGF-β1. These results demonstrate a critical role for IEC-expressed CD47 in regulating mucosal repair and raise important considerations for possible alterations in wound healing secondary to therapeutic targeting of CD47.