Inherited beta-globinopathies, including sickle cell disease (SCD) and ?-thalassemia (HLS13-04), are chronic blood disorders resulting from defects in hemoglobin (Hb) structure or synthesis. These diseases affect an estimated 70,000 people in the U.S. alone and much larger numbers in Africa, Europe, India, the Middle East and the Caribbean. The complications of SCD, by far the most common beta-globin disorder, include hemolytic anemia, painful ischemic crises, bacterial infections, stroke, and chronic lung and kidney disease. The major therapies for SCD and ?-thalassemia include lifelong blood transfusions, treatment with hydroxyurea-a cytotoxic agent that modestly induces fetal hemoglobin (HbF, ?2?2)-or high risk transplantation of hematopoietic cells. Elevation of HbF is considered the most promising approach to treat SCD or to effect a remission or cure. A benign condition, hereditary persistence of fetal hemoglobinemia (HPFH), dramatically ameliorates or even prevents the symptoms of ?-thalassemia and SCD. Significantly, in one form of HPFH, naturally-occurring mutations in the gene promoters of the fetal ?-globin chains A? and G? induce HbF to 10- 30% of the level of ?-globin in every red blood cell, sufficient to inhibit SCD.It has long been suggested that a pharmacological agent that replicates pancellular HbF induction could transform current therapy. The goal of this Phase I SBIR proposal is to identify small molecule ligands to a transcription factor, an orphan nuclear receptor (NR), that has a central role in silencing embryonic and fetal ?-globin expression in the adult, by binding to the promoters of the fetal ?-globin genes. To date, no valid ligands have been described for this target. Assay development and structural analysis also show that the NR target is likely to function similarly to other NRs, such as the steroid, retinoid, and thyroid receptors, by allosteri modulation of peptide affinity in response to ligand. We developed a cell-free high throughput screening (HTS) assay for this receptor that measures interaction with a short peptide derived from a transcriptional cofactor, a putative corepressor for this orphan NR. Our objective is to identify small molecules that disrupt corepressor peptide interaction with the orphan NR and to evaluate whether these negative modulators can derepress fetal ?-globin expression.
In Aim 1 we will screen diverse compound libraries in the biochemical HTS assay to identify small molecule ligands that disrupt the interaction between the orphan NR and corepressor peptide. We will then determine in Aim 2 whether, in addition to regulating binding of corepressors, the ligands identified in Aim 1 also regulate corepressor interactions via the target receptor in cell culture. We also propose to increase ligand affinity (IC50 < 0.5 M) by purchase and synthesis of analogs to initial hits. Finally, in Aim 3, we will test the ability of the identified ligands to ativate ?-globin expression in hematopoietic cells cultured in vitro. This pharmacological evaluation will enable more intensive drug discovery with SBIR Phase II support leading to testing of lead compounds in animal models of the ?-globin disorders.
About 70,000 Americans suffer from sickle cell disease and ?-thalassemia. Annual medical costs exceed $1 billion. Elevation of fetal hemoglobin (HbF) is considered a highly promising approach to treat SCD or the ?-thalassemias since the effects of the defective or absent ?-globin chain can be overcome by induction of fetal ?-chain production. Safe and effective therapies that elevate HbF will be rapidly accepted. The goal of this Phase I project is to identify small molecule drug-like compounds to a unique transcription factor that regulates the levels of HbF. Our long-term goal is development of a small molecule oral medication to treat these very serious inherited ?-globin disorders.
