Second round of anemia Research grants awarded to physicians in the USA and Europe
RoFAR (Roche Foundation for Anemia Research) announced it’s second cycle of research grants totaling 1 524 440 Swiss Francs ($USD 1 250 000) just one year after the organization was established.
The seven successful research projects to receive awards were selected from over 70 initial applications and were chosen for meeting the exacting standards for scientific excellence, feasibility and originality set by the organization's independent Scientific Advisory Board.
Announcing the awards, Dr Nathan Levin, chair of the RoFAR Board of Trustees and a leading nephrologist at the Renal Research Institute, New York, USA, said “Once again we were delighted with the quality of research applications and I would like to congratulate our second wave of award winners. We hope that the research they will now be able to move forward will, over the next two years, yield vital information to improve our understanding of anemia in new and important arenas.”
The award winners and their fields of research are:
 |
Professor Hans-Ulrich Bucher
University Hospital Zurich
Zurich, Switzerland
Erythropoietin reduces brain, eye and lung damage in very preterm infants: proof of concept study |
Erythropoetin (EPO) has been shown to be protective against hypoxic-ischaemic and inflammatory injuries in a broad range of tissues and organs besides promoting red cell formation. Especially protective effects on brain, retina and bowel could be shown in animal models and first human studies.
EPO has been used widely for several weeks in preterm infants to prevent anaemia and is well tolerated. No short and long term adverse effects have been documented with EPO treatment in preterm infants.
Because EPO has been shown to influence several mechanisms associated with these short-term and long-term complications of prematurity and furthermore has been shown to have a positive effect even post hoc, i.e. if given within a period of hours after an hypoxic-ischemic insult, EPO may ameliorate the damage in very premature infants. Very preterm infants may suffer from a variety of short-term complications and long-term sequelae of premature birth. The most critical period are the first days after birth and inflammatory changes as consequence of hypoxia-ischemia or infection seems to have a major impact on short-term as well as permanent damage.
To determine whether early administration of EPO alters the incidence and severity of complications typically associated with preterm birth in infants born between 24 0/7 and 27 6/7 gestational weeks, we want to investigate in a clinical trial for prophylactic therapy with EPO.
Progress report available: 
 |
Dr. Edward Debnam
Royal Free & UniversityCollege
London, UK
Is inflammation an important factor in the anemia of chronic renal failure?
|
Anaemia is a feature of renal failure and it is, in part, due to insufficient production of the hormone erythropoietin (EPO), the action of which is to stimulate the bone marrow to produce mature red blood cells. However, an increased production of red blood cells also requires the absorption of adequate amounts of dietary iron (since iron is a vital component of red blood cells). It is a common clinical observation that many renal failure patients receiving EPO require intravenous iron supplementation, despite the fact that they have adequate dietary iron. This strongly suggests an ab-normality of iron uptake by the intestine.
Recent studies show that another hormone, hepcidin, reduces iron absorption from the gut. Interestingly, hepcidin secretion is increased during infection (also a common finding in renal failure) and this raises the possibility that raised hepcidin levels in renal failure interfere with gut iron uptake, i.e. high blood levels of hepcidin may override the effect of administered EPO in renal failure and limit the supply of dietary iron for red blood cell production. Hence the need for intravenous iron treatment. However, this procedure has risks of iron overload and tissue injury.
To date, there is no information available on the relationship between hepcidin secretion, EPO secretion and infection in renal failure. If an association is found between infection and hepcidin expression in renal failure, the process by which hepcidin influences iron transport may prove to be an important target for therapy of this form of anaemia. Progress report available:
 |
Dr. Diane Gilligan
Puget Sound Blood Centre
Seattle, USA
Regulation of gene expression during erythropoiesis
| The goal of this project is to understand the regulation of appropriate gene expression during red blood cell production. Anemia, the lack of sufficient numbers of red blood cells, is a common disorder that affects patients with many different illnesses. Specifically, it is a side effect of chemotherapy for cancer and it is a secondary sign of kidney failure. Much has been learned about the production of red blood cells since the discovery of erythropoietin, a hormone produced by the kidney that stimulates hematopoietic stem cells to produce red blood cells. At a molecular level, the binding of erythropoietin to its receptor signals the precursor cells to increase expression of genes that are important to red blood cells and to decrease expression of genes that are not important to red blood cells. This signaling pathway is not yet well understood and our experiments are designed to gain more information about the molecular steps that are required to increase production of red blood cells. We have been studying a family of genes, the adducins, that are expressed in all cells, but were first discovered as components of red blood cells. We have demonstrated lineage specific expression of two of the adducin genes. The beta adducin gene is highly expressed in red blood cells, but is not expressed in platelets, while the gamma adducin gene shows the opposite characteristics, highly expressed in platelets, but not expressed in red blood cells. We will study the adducin gene family as a model system for lineage specific gene expression, with particular significance for red blood cell production. We may identify novel factors that are important for red blood cell gene expression and these results may lead to novel therapies for patients with anemia.
Progress report available:
 |
Professor Alexander Maxwell
Queen’s University Belfast
Belfast, UK
Investigation of the role of JUNE-1 in erythropoiesis
| Erythropoietin (EPO) stimulates bone marrow cells to differentiate into mature erythrocytes. Recombinant human erythropoietin therapy is widely used to treat anaemias but there is relatively limited knowledge of how EPO regulates genes in cells and tissues. Investigation of EPO-regulated genes should improve our understanding of how EPO influences erythropoiesis and the function of non-haematopoietic organs which express EPO receptors such as the brain and heart.
We investigated transcriptional events occurring downstream of EPO binding to its receptor. We have now identified a novel gene, JUNE-1, encoding 5 exons expressing a 1.2 kb transcript translated into a 44 kDa protein. Its highly conserved DNA sequence has both nuclear targeting and plant homeodomain motifs, thought to mediate protein-protein or protein-DNA interactions. This suggests that JUNE-1 is involved in chromatin remodeling or transcriptional regulation. RT-PCR confirmed JUNE-1 expression in a variety of tissues and tumour cell lines, implicating JUNE-1 in processes beyond erythropoiesis.
Functional studies of JUNE-1 are proposed using an in vitro model of erythropoiesis (murine erythroleukaemia MEL cell system), where proerythroblasts are cultured and differentiate into mature hemoglobin producing cells. We plan to localize JUNE-1 protein by immunohistochemistry.
The consequences of overexpression and underexpression (knockdown via RNA interference) of JUNE-1 protein on erythroid cell proliferation and differentiation will be as-sessed. MEL cells will be examined for changes in phenotype, such as cell growth rate, apoptosis, and differentiation capacity. Since JUNE-1 may function as a transcription factor, the genes downstream of JUNE-1 will be identified in the MEL cell model using mouse gene microarrays.
To identify proteins that interact with JUNE-1 mammalian expression vectors will be used to produce JUNE-1 fused to GST, and the protein then immobilized on a GST column. Cellular extracts will be loaded on the columns, and interacting proteins eluted and identified by SDS-PAGE and mass spectrometry. JUNE-1 specific antibodies will also be used to identify JUNE-1 interacting proteins via co-immunoprecipitations from the cell lysates. The proposed research should enable the functional characterization of JUNE-1, a novel EPO-regulated gene.
Progress report available:
 |
Dr. Jun-ichi Nishimura
Duke University Medical Centre, USA
Innovative drug design using RNA aptamers for various anaemias.
|
Aptamers are small RNA molecules that bind to target proteins similar to antibodies. Through a selection process, termed SELEX, aptamers have been identified against a wide range of therapeutic targets. Dr. Sullenger (Mentor and Consultant) has established the SELEX system, and Dr. Nishimura (PI) has recently successfully identified RNA aptamers for treatment of paroxysmal nocturnal hemoglobinuria, a rare type hemolytic anemia. Our overall goal is to develop aptamers to treat various anemias using the SELEX system. In this proposal, we exclusively focus on the development of aptamers that inhibit red cell adhesion to prevent or treat vaso-occlusion in sickle cell disease (SCD). Vaso-occlusive crises are the major clinical feature of SCD, and the adhesion of sickle erythrocytes (SS-RBC) to vascular endothelium is important to the generation of vaso-occlusion. SS-RBC express many adhesion molecules, such as Lutheran protein (B-CAM) and LW. Adhesive SS-RBC may bind to endothelial cell P-selectin, integrin aVb3, as well as extracellular matrix proteins, such as laminin. All of these molecules and their ligands are potential targets for reagents to prevent or treat the vaso-occlusive crises of SCD, and we focus 3 mole-cules, including B-CAM, P-selectin, and integrin aVb3. Since RNA aptamers for P-selectin have already been identified, we have synthesized one of these known P-selectin-binding aptamers. We have also begun to identify RNA aptamers that bind specifically to B-CAM and integrin aVb3. Once these aptamers are optimized, we will advance to flow chamber assays, established by Dr. Telen (Co-investigator), to test their ability to inhibit adhesion of SS-RBC to vascular endothelium. Selected high-affinity and high inhibitory aptamers will be further modified in preparation for in vivo studies, using a system established by Dr. Telen. The development of combinatorial blocking aptamers against these 3 adhesion molecules represents a novel potential therapeutic option for patients with SCD.
Progress report available:
 |
Professor Radek Skoda
University Hospital Basel
Basel, Switzerland
The role of Smad4-dependent signaling in anaemia
|
We observed that mice lacking the signaling protein SMAD4 developed a severe anemia. SMAD4 is an essential mediator for signals generated by the transforming growth factor beta (TGFb) family of proteins. This signaling pathway is important for proper embryonic development and regulates differentiation and growth in many adult tissues. However, an association of the TGFb signaling pathway with the appearance of anemia in adults has not been described to date. We used an inducible system to delete the gene for SMAD4 in adult mice and observed anemia 3-4 weeks after induction. Blood cells lacking SMAD4 developed normally when transplanted into healthy recipients, but anemia developed when normal blood cells were transplanted into a SMAD4 deficient recipients. Thus, SMAD4 is required to pro-vide the environment for normal red blood cell homeostasis. Our preliminary results suggest that loss of SMAD4 causes alterations in the iron metabolism that could interfere with red blood cell formation. Iron is essential for the formation of hemoglobin and iron deficiency is the most common cause of anemia. More subtle changes in the availability of iron are thought to play a role in anemia associated with chronic inflammatory diseases and cancer. We propose to elucidate the mechanism of how anemia develops in these mice and to de-fine a novel role for the TGFb signaling in the regulation of red blood cell formation.
Progress report available:
 |
Dr. Carole Soussain
Oregon Health and Sciences University
Portland, USA
Neuroprotective effect of erythropoietin on chemo- and radiotherapy-induced toxicity
|
Combined radiotherapy and chemotherapy approaches have provided significant efficacy against brain tumors such as primary central nervous system lymphoma (PCNSL). A significant number of patients experience severe neurotoxicity with these therapeutic approaches, and is a limiting factor in the management of patients with intracerebral cancer. Erythropoietin (EPO) is an important hormone for the development of fetal brain and homeostasis of the adult brain, and both EPO and EPO receptor (EPO-R) are expressed on astrocytes and neurons in adult brain. Exogenous EPO has demonstrated a protective effect on neuronal cells cultures after injuries such as nitric oxide or glutamate exposure, as well as in animal models of ischemia, inflammation, seizures, and subarachnoid hemorrhage. In clincial trials, EPO has been safely and efficiently used to prevent neurological damage after acute stroke. These findings suggest that EPO may be neuroprotective against chemotherapy and radiotherapy-induced neurotoxicity. In this proposal, the potential therapeutic role of erythropoietin in reducing therapy-induced neurotoxicity will be tested in molecular and cellular studies in vitro, and in in vivo animal models. We will assess the effect of EPO on tumor cell growth and chemotherapy toxicity in vitro, and determine the protection provided by EPO in an in vitro neuronal cell model. We hypothesize that EPO has no growth and survival activity on lymphomatous cells, but is neuroprotective against chemotherapy and radiotherapy toxicity in cultured neuronal cells. We will search for EPO-induced alteration of the blood-brain barrier permeability in normal rats and rats with intracerebral tumors. We hypothesize that EPO does not prevent the entry of chemotherapy in the brain. Finally, we will determine if there is a neuroprotective effect of EPO in rats exposed to neurotoxic doses of chemotherapy and radiotherapy. We hypothesize that EPO chemoprotection can improve the management of treatment-related CNS toxicities.
Progress report available:
 |
Dr. Christina Warnecke
University Erlangen-Nürnberg
Germany
Molecular mechanisms underlying the hypoxic induction of erythropoietin by HIF-2a
|
The hormone erythropoietin (EPO), which is predominantly produced in the kidneys, is the primary regulator of red blood cell production. Hypoxia, i.e. insufficient oxygen supply as en-countered at high altitudes or after blood loss, leads to a compensatory increase of EPO production due to an increase in EPO gene expression, which is mediated by hypoxia-inducible transcription factors. In disease states that impair kidney function such as diabetes, chronic renal disease or heart failure the production of erythropoietin in the kidney fails to meet the needs and patients suffer from anemia, which deteriorates their general condition and often aggravates the primary disease. Treatment with human recombinant EPO is highly effective and so far the therapy of choice for erythropoietin-deficiency anaemias. An alternative and very attractive approach, because of the high costs of recombinant EPO, could be the stimulation of endogenous EPO production.
In 2001 a family of transcription factors inducible by hypoxia (Hypoxia-inducible factor=HIF) was identified and their regulation has since then been actively investigated. Interestingly, although HIF was primarily identified as a regulator of EPO expression, it turned out that it also induces many other genes that serve to protect against oxygen deficiency. Although this underscores the importance of HIF, this lack of specificity makes it more difficult to use it as a molecular target to stimulate EPO generation. Only recently, we and others demonstrated that one specific isoform of HIF, called HIF-2a, is responsible for the regulation of erythropoietin, but not that of most other HIF-dependent genes. By studying the molecular basis of this specificity in EPO-producing cells lines we attempt to increase our understanding of the nor-mal regulation of red cell production, as well as its impairment in different diseases and hope to identify novel targets for medical intervention.
Progress report available:
|