Site-specific Interaction of Doxorubicin in the Iron Responsive Element Rna: Implications in Cellular Iron Homeostasis and Non-iron Deficiency Anemia

Title of dissertation: SITE-SPECIFIC INTERACTION OF DOXORUBICIN IN THE IRON RESPONSIVE ELEMENT RNA: IMPLICATIONS IN CELLULAR IRON HOMEOSTASIS AND NON-IRON DEFICIENCY ANEMIA Luigi Jhon Alvarado, Doctor of Philosophy, 2014 Dissertation directed by: Professor T. Kwaku Dayie Department of Chemistry & Biochemistry A widely utilized chemotherapy drug, doxorubicin, has recently been shown to bind to a mammalian 5′ untranslated region Iron Responsive Element (IRE) RNA. In conjunction with the Iron Regulatory Protein (IRP), IRE RNA is involved in cellular iron homeostasis at the translational level. This tight RNA/protein complex prevents ribosomal assembly, hindering translation initiation of iron storage proteins, e.g. ferritin, under low cellular iron conditions. Conversely, iron overload is conducive to complex dissociation, allowing for up-regulation of the same proteins. However, this system is not entirely efficient. Some anemic patients receive adjuvant chelation therapies upon chronic blood transfusions to sequester excess labile iron. The use of doxorubicin to promote RNA/protein dissociation could potentially allow for downstream up-regulation of ferritin. In this work, I show how doxorubicin interacts with IRE RNA using multidimensional nuclear magnetic resonance, fluorescence spectroscopy, and electrophoretic mobility shift assays. All three approaches converge on the observation that the IRE/IRP complex formation is disrupted by doxorubicin. Obtaining further data on the RNA/protein/drug interactions may lead to unveiling a validated RNA target as a complementary treatment of iron overload disease, e.g. sickle cell anemia. SITE-SPECIFIC INTERACTION OF DOXORUBICIN IN THE IRON RESPONSIVE ELEMENT RNA: IMPLICATIONS IN CELLULAR IRON HOMEOSTASIS AND NON-IRON DEFICIENCY ANEMIA