Modeling of sickle cell anemia utilizing disease-specific induced pluripotent stem cells.

摘要

Sickle cell anemia, caused by a point mutation that affects the HBB gene, is one of the most common human genetic disorders world-wide and has a high morbidity and mortality. A single FDA approved drug, hydroxyurea, is available for its ability to induce fetal hemoglobin expression, a major modulator of disease severity. Not every patient responds to treatment and additional HbF-inducing drugs are needed. In this thesis, I outline an induced pluripotent stem cell-based approach to the study of sickle cell disease (SCD). In the lab, we are currently building a library of SCD-induced pluripotent stem cell (iPSC) lines from a cohort of SCD patients with different genetic backgrounds and fetal hemoglobin levels. Utilizing a directed-differentiation approach, iPSC can give rise to hematopoietic progenitors that are similar to megakaryocyte-erythroid progenitors and can be further specified to become cells of either lineage. I examined the hypothesis that an iPSC-based system would be capable of producing fully functional erythroid cells and also recapitulate the variation in fetal hemoglobin levels seen in SCD patients. Directed-differentiation of iPSCs produced erythroid-lineage cells that were responsive to oxygen levels and erythropoietin, and were capable of further maturation and increased hemoglobin production. A humanized mouse model demonstrated the ability of these cells to localize to the bone marrow, contribute to the peripheral blood, and survive in vivo for over two weeks. The maturation capability of SCD-specific iPSC-derived erythroid lineage cells was correlated with hemoglobin expression and compared to control cells. Characterization of in vitro and in vivo differences between control and SCD-specific iPSC-derived erythroid-lineage cells demonstrated variation amongst individuals, similar to the variation seen in patients. Both of these patient-specific iPSC-based in vitro and in vivo models allow for the examination of the effect of genetic variability on fetal hemoglobin expression and also for the modeling of patient-specific responses to drug treatment. This information will facilitate better clinical treatment of the disease.