Analysis of NGAL Metabolism Identifies a Therapy for Iron Overload

NGAL 代谢分析确定了铁过载的治疗方法

基本信息

批准号：
8095786
负责人：
JONATHAN M. BARASCH
金额：
$ 24.58万
依托单位：
COLUMBIA UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-09-01 至 2013-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8095786
关键词：
Acute Renal Failure with Renal Papillary Necrosis Aerobic Affinity Agranulocytosis Animals Bacteria Binding Biological Availability Biological Markers Biology Blood Circulation Blood Transfusion Bypass Carrier Proteins Catechols Cell Lineage Cells Chelating Agents Chemicals Chronic Kidney Failure Clinical Medicine Color Complex Data Deferoxamine Development Disease Distal Endocrine Endocytosis Engineering Enterobactin Evaluation Excretory function Family Ferritin Gelatinase A Gene Deletion Generations Genetic Growth Harvest Hepatitis Hereditary Disease Histocompatibility Testing Hour Human In Transferrin Inherited Interruption Iron Iron Chelation Iron Overload Kidney Knock-out LDL-Receptor Related Protein 2 Learning Left Length Ligands Liver Lung Lysosomes Measures Mediating Metabolism Modeling Molecular Mus Nature Nephrons Neutropenia New Agents Nutrient Organ Organism Oxidation-Reduction Oxidative Stress Oxygen Paper Pathway interactions Patients Peritoneum Phenotype Physiological Process Proteins Reaction Reactive Oxygen Species Role Series Serum Siderophores Skin Solutions Stimulus Structure Sum Surface Plasmon Resonance Syndrome System Techniques Testing Therapeutic Toxic effect Transferrin Transport Process Urine Wild Type Mouse X-Ray Crystallography base chelation cofactor extracellular genotoxicity hearing impairment in vivo member microorganism mutant non-genetic novel novel therapeutics prevent receptor receptor recycling small molecule trafficking urinary

项目摘要

DESCRIPTION (provided by applicant): The transport of iron poses a significant problem because free ferric iron is insoluble (< 10-18 M) in aerobic solutions at physiologic pH, while upon solubilization by some chelators, a reactive form of iron is created that can produce toxic oxygen species. Specialized mechanisms are consequently required to traffic iron and these specialized mechanisms are found in proteins which utilize conserved motifs to directly bind iron (transferrin and ferritin) or utilize embedded cofactors. While extracellular iron transport is largely mediated by transferrin, mice carrying deletions of these genes displayed surprisingly limited phenotypes (Barasch, Developmental Cell, 2009). We found that a member of the lipocalin superfamily called Ngal acted as a high affinity iron carrier (Barasch, Molecular Cell, 2002) when binding a family of novel cofactors called the catechols or else the related bacterial siderophores constructed from catechol. In the presence of iron, formation of the Ngal:siderophore:FeIII complex occurred at subnanomolar affinity (Barasch, Nature Chemical Biology, 2010) forming a bright red protein, which was stable for many days in solution and stable in vivo for transport of its tightly bound iron. Ngal is expressed in vivo, but a number of "damage" stimuli raise its concentration by orders of magnitude. Thereafter, Ngal traffics in the serum and is thought to be captured by the kidney receptor megalin, where Ngal clears the siderophore:Fe complex. While we have learned a great deal about the metabolism of the urinary form of Ngal (it is expressed from the distal nephron and is excreted in the urine as a full length protein), we know much less about this clearance system and the role of the megalin receptor, which is the only confirmed receptor for Ngal. To study this process in depth we will examine a conditional mutant of megalin, and for studies in wild type mice we are creating a series of Ngal mutants, some of which bypass the proximal tubule where megalin is located, resulting in their presence in the urine. During these studies we realized that the mutants could still bind to siderophore:FeIII at high affinity (they were red colored proteins), and that they could definitely excrete iron, we speculate in a redox inactive manner. Indeed, rather than donate iron to micro-organisms, which is a major concern for small molecule chelators, the Ngal:siderophore:Fe complexes sequester iron from bacteria. In sum, in this proposal, we test the hypothesis that megalin is the key recycling receptor for Ngal and as a result of this idea, we propose that when the megalin-Ngal complex is inhibited, Ngal can carry tightly bound iron in the urine, hence serving as a safe, novel therapeutic for the common syndromes of iron overload diseases. PUBLIC HEALTH RELEVANCE: Iron overload diseases are common occurrences in clinical medicine, and their therapies have proved toxic to many cell lineages as well as inductive of bacterial growth. Iron overload is a common sequel of blood transfusions, but it is well known in hepatitis, chronic kidney disease as well as in common hereditary diseases such as hemachromatosis. We have discovered a novel iron trafficking pathway based on the protein Ngal, which is massively expressed in the human in different types of tissue damage. Our studies in Ngal metabolism have led to a proof of concept that Ngal can be used as a safe, therapeutic iron chelator.

描述（由申请人提供）：铁的运输构成了一个重要的问题，因为在生理pH下的有氧溶液中游离铁不溶于（<10-18 m），而在某些螯合剂的溶解后，产生了一种可产生有毒氧的铁的反应性铁。因此，需要专门的机制才能交通铁，这些专业机制是在蛋白质中发现的，这些机制利用保守的基序直接结合铁（转铁蛋白和铁蛋白）或利用嵌入式辅助因子。虽然细胞外铁的转运大部分是由转铁蛋白介导的，但携带这些基因缺失的小鼠表现出了令人惊讶的有限表型（Barasch，发育细胞，2009年）。我们发现，当结合一种称为Catechols的新型辅助因子家族时，Lipocalin超家族的成员被称为NGAL（Barasch，Molecular Cell，2002），或者是由儿茶酚构成的相关细菌Siderophores。在存在铁的存在下，ngal的形成：辅助载体：FeIII复合物发生在亚摩尔亲和力（Barasch，Nature Chemical Biology，2010年），形成了鲜红色的蛋白，在溶液中稳定稳定，在溶液中稳定，体内用于其紧密结合的铁。 Ngal在体内表示，但许多“损害”刺激通过数量级提高了其集中度。此后，Ngal流量在血清中，被认为是由肾脏受体Megalin捕获的，Ngal清除了铁载体：FE Complex。尽管我们对Ngal的尿形式的代谢有了很多了解（它是从远端的肾单位表达的，并在尿液中排泄为全长蛋白），但我们对这种清关系统的了解和大甲蛋白受体的作用少得多，这是Ngal的唯一确认的受体。为了深入研究这一过程，我们将检查一个有条件的Megalin突变体，对于野生型小鼠的研究，我们正在创建一系列的Ngal突变体，其中一些绕过Megalin所在的近端小管，导致它们在尿液中的存在。在这些研究中，我们意识到突变体仍然可以与铁载体结合：高亲和力的FEIII（它们是红色蛋白质），并且它们肯定可以排泄铁，我们以氧化还原的方式推测。实际上，NGAL：铁载体：Fe Compleffers从细菌中持续了铁，而不是将铁捐赠给微生物，这是小分子螯合剂的主要关注点。总而言之，在这一提议中，我们检验了梅加林是Ngal的关键回收受体的假设，并且由于这个想法，我们建议，当Megalin-ngal复合体受到抑制时，Ngal可以在尿液中携带紧密结合的铁，因此可以作为一种安全的，可作为一种安全的，可作为常见的铁超综合症的新型治疗方法。公共卫生相关性：铁超负荷疾病是临床医学中常见的发生，其疗法已被证明对许多细胞谱系有毒以及细菌生长的诱导。铁超负荷是输血的常见续集，但在肝炎，慢性肾脏疾病以及常见的遗传性疾病（如血调理病）中众所周知。我们已经发现了一种基于蛋白质NGAL的新型铁运输途径，该途径在人类中以不同类型的组织损伤表达。我们在NGAL代谢方面的研究导致了概念证明NGAL可以用作安全的治疗性铁螯合剂。