Piperine Inhibits the Immune Checkpoint CD47 to Enhance Immunity Against Pancreatic Cancer

ABSTRACT Pancreatic cancer is a common malignant tumor of the digestive tract, with insidious onset and rapid metastasis. Most patients have already developed to an advanced stage at the time of diagnosis, and the fatality rate is very high. In recent years, immunotherapy has emerged as a promising potential treatment for pancreatic cancer. Some studies have shown that piperine has the potential to inhibit pancreatic cancer, but the role of piperine in tumor progression and tumor immunity in vivo remains unclear. The effects and mechanisms of piperine on the immunity of pancreatic cancer were studied. The effects of piperine on the proliferation, migration and apoptosis of pancreatic cancer Panc02 and PANC‐1 cells were detected by CCK‐8, plate colony formation, CFSE staining, wound healing, transwell and flow cytometry. The effects of piperine on the growth, metastasis and immune microenvironment of pancreatic cancer were studied by constructing subcutaneous and orthotopic pancreatic cancer models in mice and using live animal imaging and flow cytometry. The intersection targets of piperine and pancreatic cancer were analyzed by network pharmacology and RNA‐seq, and the key genes of immune cell changes were identified. The specific effects of piperine on CD47 in pancreatic cancer tissues and cells were studied by immunohistochemistry, flow cytometry, qRT‐PCR, and Western blot. Tumor cells were cocultured with peritoneal/bone marrow‐derived macrophages or CD8 + T cells to explore the cellular mechanism, and the phagocytosis of macrophages and the activation of CD8 + T cells were detected by immunofluorescence, magnetic bead sorting and flow cytometry. In the in vitro experiments, piperine inhibited the proliferation and migration of the pancreatic cancer cell lines Panc02 and PANC‐1, and induced its apoptosis. In the in vivo experiments, piperine inhibited the growth and metastasis of subcutaneous and orthotopic pancreatic tumors in mice. In the immune microenvironment, piperine significantly increased the proportion of macrophages, especially M1‐type macrophages, as well as the proportion of CD8 + T cells in the tumor and spleen. Moreover, piperine reduced the expression of CD47 at the mRNA and protein levels of pancreatic cancer cells, thus enhancing the phagocytosis of macrophages toward tumor cells, and increasing the activation level of CD8 + T cells. Our results suggest that piperine enhances its antitumor immunity in pancreatic cancer by blocking the immune checkpoint CD47. Therefore, piperine is expected to provide a new approach for the immunotherapy of pancreatic cancer.