Combine iron Chelation therapy ameliorates decline of iron on iron loaded Zebrafish embryo

Abstract

Thalassemia is the most common genetically inherited blood disorder in the world arisen from defect in hemoglobin production, resulting in ineffective erythropoiesis and rapid destruction of RBC in the periphery that leads to severe anemia. While transfusion therapy corrects the anemia, it gives rise to secondary iron overload. Finally, recurrent blood exposure can induce alloimmunization to erythrocytes antigens, leading to difficulty in finding compatible blood. Current iron chelation therapy using Desferal® (DFO) is challenging due to its short vascular half-life, frequency of injections, toxicity and expense. To address these issues new iron chelating agents and improved iron chelation therapy we propose using combination of iron chelators. To test the utility of different chelators, in vivo studies performed on iron loaded zebrafish embryos (100 μM Fe3+; ferric ammonium citrate (FAC); 3dpf -6dpf). Iron chelation studies utilized either single or combinational treatment with Deferiprone (L1) or DFO for 7dpf-10dpf on zebrafish iron loaded model. The efficacy of treatment was assessed by total iron with ICP-MS, clinical chemistry analyzer spectrophotometry and with Perl's iron stain microscopy. Iron treatment alone resulted in a significant increase in total iron, histochemical iron staining, also resulted in an increase in stainable iron. Treatment with iron chelators either L1 or DFO alone demonstrated modestly decreased total iron and iron staining. Importantly, combination therapy (L1+ DFO), resulted in an additive effect in FAC-driven iron levels after 96 hours treatment. As expected, iron chelators (e.g., 100 μM DFO or L1) reduced the level of iron. These studies confirm that cellular iron utilization requires one chelator to have the properties to enter cells, chelate intracellular iron and subsequently that go to another chelator with higher affinity to iron binding. The development of improved combination iron chelation therapy using nigh and low molecular weight biocompatible iron chelators may provide better therapeutic value (less toxic and less frequent administration) in developing nations. These biocompatible iron chelators can improve shelf life of blood products to minimize oxidative damage due to iron-mediated oxidation. Consequent to this, iron mediated pathology to patients diminished resulting in less cost to already strained public health budgets in many poor countries.