Abstract
Icaritin, a small molecule currently being investigated in phase III clinical trials in China (NCT03236636 and NCT03236649) for treatment of advanced hepatocellular carcinoma (HCC), is a prenylflavonoid derivative obtained from the Epimedium genus. Previously, it was found that Icaritin decreased the expression of PD-L1, but its direct molecular targets and the underlying mechanisms have not been identified. In this study, we report the identification of IKK-α as the protein target of Icaritin by biotin-based affinity binding assay. The further mutagenesis assay has provided evidence that C46 and C178 in IKK-α were essential amino acids for Icaritin binding to IKK-α, revealing the binding sites of Icaritin to IKK-α for the first time. Functionally, Icaritin inhibited the NF-κB signalling pathway by blocking IKK complex formation, which led to decreased nuclear translocation of NF-κB p65, and subsequent downregulation of PD-L1 expression in a dose-dependent manner. More importantly, PD-L1-positive patients exhibited longer overall survival upon Icaritin therapy. Finally, Icaritin in combination with checkpoints antibodies, such as α-PD-1, has demonstrated much better efficacy than any single therapy in animal models. This is the first report that anticancer effects of Icaritin are mediated, at least in part, by impairing functions of IKK-α.