Inhibition of COX-2 and EGFR by Melafolone Improves Anti-PD-1 Therapy through Vascular Normalization and PD-L1 Downregulation in Lung Cancer

Checkpoint blockade therapy has been proven efficacious in lung cancer patients. However, primary/acquired resistance hampered its efficacy. Therefore, there is an urgent need to develop novel strategies to improve the checkpoint blockade therapy. Here we tested whether dual inhibition of cyclooxygenase-2 (COX-2) and epidermal growth factor receptor (EGFR) by flavonoid melafolone improves program death 1 (PD-1) checkpoint blockade therapy through normalizing tumor vasculature and PD-L1 downregulation. Virtual screening assay, cellular thermal shift assay and enzyme inhibition assay identified melafolone as a potential inhibitor of COX-2 and EGFR. In the Lewis lung carcinoma (LLC) and CMT167 models, dual inhibition of COX-2 and EGFR by melafolone promoted survival, tumor growth inhibition and vascular normalization and ameliorated CD8+ T cell supression, accompanied by the downregulated TGF-β, VEGF and PD-L1 in the tumor cells. Mechanistically, dual inhibition of COX-2 and EGFR in the lung cancer cells by melafolone increased the migration of pericyte, decreased the proliferation and migration of endothelial cells, and enhanced the proliferation and effector function of CD8+ T cells through VEGF, TGF-β or PD-L1 downregulation and PI3K/AKT inactivation. Notably, melafolone improved PD-1 immunotherapy against the LLC and CMT167 tumors. Together, dual inhibition of COX-2 and EGFR by melafolone improves checkpoint blockade therapy through vascular normalization and PD-L1 downregulation, and may be a promising combination strategy for the treatment of human lung cancer by the effects on vessels and immune cells.