Erythropoiesis involves the gradual progression of a hematopoietic stem cell into a mature red blood cell. During the terminal differentiation step late stage erythroblasts extrude their nucleus to produce reticulocytes in a process termed erythroid enucleation. Further maturation results in the formation of biconcave shaped erythrocytes that are found in the peripheral blood. Recent work has found that enucleation takes around 10 minutes and is speculated to involve multiple pathways. Although similarities with cytokinesis and apoptosis have been reported, the molecular mechanisms employed remain largely unknown.
In order to address the cellular program governing enucleation, a medium-throughput functional drug screen was performed in vitro using a 326 compound library consisting of structurally diverse, medicinally active and cell permeable drugs. Here, compounds that inhibited enucleation were selected as lead compounds for further validation. Initial results led to 37 hit compounds, which were validated in a second screen that also incorporated a compound concentration gradient to examine does-response relationships. Importantly, this approach validates previous work, demonstrating for example a role for p38 MAPK (1) and HDAC (2) in enucleation and has identified several novel candidates including CDKs and topoisomerases. Future studies aim to functionally assess candidate hits in vitro using shRNA knockdown and in vivo using various mouse models.