18β-Glycyrrhetinic acid ameliorates renal anemia via inactivating PHD2 and activating HIF-2α-EPO axis

18β-Glycyrrhetinic acid (GA) is a major metabolite of glycyrrhizin (GL) that is a key ingredient of licorice, a popular edible and medicinal plant, and is often used in the food and pharmaceutical industry as a flavoring agent. Previous studies suggest that licorice possess hematopoietic activity. However, the bioactive ingredients in licorice that produce hematopoietic effect remain elusive. This study aimed to test if GA is the key active molecule that contributed to the anti-anemia activity of licorice. Both in vivo and in vitro models were utilized to determine the efficacy of GA against renal anemia and to decipher its related mechanisms. We found for the first time that oral administration of GA at clinically safe doses is effective against renal anemia, illustrated by the improvement in key hematological parameters. Mechanistically, GA was found to be able to directly bind to and inhibit PHD2, analyzed by molecular docking simulation, surface plasmon resonance (SPR), cellular thermal shift assay (CETSA), and a time-resolved fluorescence resonance energy transfer (TR-FRET)-based enzyme activity assay. PHD2 inactivation by GA led to the activation of HIF-2α-erythropoietin (EPO) pathway, accompanied by improved iron availability and inhibition of inflammation. Further, the tissue distribution analysis revealed that the concentrations of GA in target organ kidney and liver are 120.39 and 83.86 μg/g respectively, exceeding IC50 of 18.83 μg/mL for inhibition of PHD2. The findings therefore identified GA as a novel HIF-PHD2 inhibitor with favorable target tissue distribution that contributed to the anti-anemia effect of licorice and warrants further investigation.