Pharmacological modification of iron transport via DMT1

Disorders of iron homeostasis are a major global health concern. While iron deficiency and iron restricted anemia affect more then 10% of the global population, iron overload disorders on the other hand, such as hereditary hemochromatosis and transfusional iron overload in hematopoietic disorders, are frequent clinical conditions associated with progressive iron accumulation in the liver and other parenchyma leading to eventual organ failure. In recent years numerous transmembrane molecules, cellular receptors and circulating proteins have been identified which regulate iron absorption, tissue iron distribution and cellular iron homeostasis. Among these molecules the divalent metal transporter 1 (DMT-1) has attracted great interest since it is essential for duodenal iron absorption, iron delivery for erythroid cells and may also harbor yet not fully elucidated functions in tissue iron distribution. Four DMT-1 different splicing variants have been described, and as first part of this project we want to investigate their subcellular distribution and functional importance for the transport of iron in various tissues. Based on pre- liminary data we will then be aimed to search for pharmacological modifiers of DMT-1 mediated iron transport- as we have already identified a stimulator and an inhibitor of DMT-1 function- and to further characterize their mode of action using path clamp techniques. In vivo models of primary and secondary iron overload and iron deficiency will then study the effects of pharmacological modification of DMT-1 function on body iron homeostasis and the expression of critical iron regulating/regulated genes and proteins in the duodenum, liver and kidney. Dynamic studies will further examine the effects of such interventions on circulating amounts of iron, iron absorption and tissue iron distribution. As the importance of DMT-1 at the levels of the individual cell and the whole organism are only beginning to be understood our data will not only to contribute to elucidation of the multiples functions of DMT-1 in the regulation of body iron homeostasis but also emerge a novel pharmacological concept to treat disorders of iron homeostasis.

Hereditary hemochromatosis (HH) is one of the most frequent genetic disorders in people of Middle and Northern European origin. The underlying uncontrolled dietary iron absorption leads to tissue iron overload and eventual organ failure (liver, heart, endocrine glands) over time. Previous work of our group (supported by FWF grants 14215/15943) has demonstrated that this increased duodenal iron absorption is associated with an increased functional activity of the iron transport protein divalent metal transporter 1 (DMT1). In the current project we now aimed to identify substances which are able to modify DMT1 activity to combat iron overload. We could provide evidence for a completely novel mechanism by which the calcium antagonist nifedipine, a drug being in clinical use for the treatment of hypertension, could almost completely reverse tissue iron overload in mice suffering from hemochomatosis (Hfe-/-) or secondary iron overload. Based on the known interaction between iron and immunity and our contributions to this knowledge, we then asked whether modulation of cellular iron homeostasis would impact on the efficacy of the immune response towards pathogens and how the interaction of macrophages with specific microbes (bacteria and fungi) would impact on host iron metabolism. As part of these studies we could demonstrate that macrophages produce minute amounts of the anti-microbial peptide hepcidin which blocks transcellular iron export and retains iron from extracellular pathogens. Opposite, when macrophages are exposed to intracellular bacteria, such as Salmonella, they stimulate cellular iron export in order to limit the availability of the essential microbial nutrient iron for these microbes. Along this line the reduction of intracellular iron levels promotes anti-microbial immune effector pathways directed against such intracellular bacteria. Using the mouse model of genetic hemochromatosis (Hfe-/-) we could then show that such mice are protected from infection with Salmonella. The reason for this is that macrophage from Hfe-/- mice are iron depleted which could be traced back to increased expression of the peptide lipocalin-2 which targets iron loaded bacterial siderophores thus limiting the availability of iron for microbes such as Salmonella. In addition, we could then show that the pharmacological activation of DMT1 mediated iron transport with nifedipine resulted in an improved control of infections with Salmonella by macrophages and mice providing proof of principal for a new concept of anti-bacterial therapy by which the limitation of microbial iron availability results in a cure or improved control of an infection.