Hepcidin, a peptide hormone, is a key regulator in mammalian iron homeostasis. Increased level of hepcidin due to inflammatory conditions stimulates the ferroportin (FPN) transporter internalization, impairing the iron absorption; clinically manifested as anemia of inflammation (AI). Inhibiting hepcidin-mediated FPN degradation is proposed as an important strategy to combat AI. A systematic approach involving in silico, in vitro, ex vivo and in vivo studies is employed to identify hepcidin-binding agents. The virtual screening of 68,752 natural compounds via molecular docking resulted into identification of guanosine 5'-diphosphate (GDP) as a promising hepcidin-binding agent. The molecular dynamics simulations helped to identify the important hepcidin residues involved in stabilization of hepcidin-GDP complex. The results gave a preliminary indication that GDP may possibly inhibit the hepcidin-FPN interactions. The in vitro studies revealed that GDP caused FPN stabilization (FPN-GFP cell lines) and increased the FPN-mediated cellular iron efflux (HepG2 and Caco-2 cells). Interestingly, the co-administration of GDP and ferrous sulphate (FeSO4) ameliorated the turpentine-induced AI in mice (indicated by increased haemoglobin level, serum iron, FPN expression and decreased ferritin level). These results suggest that GDP a promising natural small-molecule inhibitor that targets Hepcidin-FPN complex may be incorporated with iron supplement regimens to ameliorate AI.

Hepcidin, master regulator of iron homeostasis, causes anemia under infectious and inflammatory conditions by reducing intestinal absorption of iron with decreased release of iron from macrophages and liver despite adequate iron stores leading to Anemia of Inflammation (AI). Many therapeutic trials have been carried out but none have been effective due to its adverse effects. In present study, we discover that Guanosine 5'-diphosphate (GDP) encapsulated in lipid vesicle (NH+) was found to inhibit NF-ҝB activation by limiting phosphorylation and degradation of IҝBα, thus, attenuating IL-6 secretion from macrophage cells. Moreover, the suppressed IL-6 levels down regulated JAK2/STAT3 pathway with decrease inflammation-mediated Hamp mRNA transcription (HepG2) and increase iron absorption (Caco2) in HepG2/Caco2 co-culture model. Analogous results were obtained in acute and chronic AI mice model thus, correcting haemoglobin level. These results proved NH + GDP as novel therapeutic agent to overcome limitations and suggests it as potential drug to ameliorate AI.