Novel Conjugates of Butyrate and δ-Aminolevulinate Increase γ-Globin Gene Expression and Fetal Hemoglobin Synthesis in Erythroid Progenitors

Abstract Pharmacologic induction of fetal hemoglobin (HbF) expression reduces the clinical severity of sickle cell disease (SCD) and improves long-term survival. Research efforts to develop effective and safe therapies have focused on a wide variety of agents however, Hydroxyurea is the only FDA-approved drug proven to mediate HbF induction in about 50% of adult SCD patients. Our group previously reported potent HbF-induction by sodium butyrate through p38 MAPK activation and CREB1 binding to the Gγ-globin cyclic AMP response element (Sangerman et al., Blood 2006). However, oral administration of butyrate derivatives to SCD patients was ineffective due to rapid metabolic inactivation by the liver. Therefore to address this critical barrier of limited treatment options for SCD, we investigated the prodrugs 1-(butyryloxy)ethyl 5-amino-4-oxopentanoate (AN233) and 1-(butyryloxy)butyl 5-amino-4-oxopentanoate (AN908), esters of δ-aminolevulinate (ALA) and butyric acid as HbF inducers in erythroid cells. Oral administration of AN233 and AN908 to BALB/c mice increased total hemoglobin levels without causing tissue toxicity (Rephaeli et al., Eur J Pharm Sci 2016). To test the ability of prodrugs to induce HbF, both K562 and KU812 cells and the immortalized human umbilical cord blood-derived erythroid progenitor 1 (HUDEP1) model were used. Treatment of K562 cells with 0.125-0.50 mM concentrations of AN233 produced cell toxicity at the 0.50 mM concentration, however >90% viability was observed by Trypan blue exclusion for 0.25 mM AN233 without evidence of caspace-3 cleavage. Using this concentration of AN233, a 1.5-fold and 6-fold (p Collectively, these findings support the oral active prodrugs AN233 and AN908 as HbF inducers in erythroid cells. Furthermore, the combination of AN908 and hydroxyurea produced an additive effect without increasing cell toxicity. Studies are underway to determine the EC50 of both prodrugs in normal and sickle primary erythroid progenitors and mechanisms of γ-globin regulation through epigenetic modifications. Disclosures No relevant conflicts of interest to declare.