Using human ESCs as a genetic model for PNH and other blood diseases

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DESCRIPTION (provided by applicant): Paroxysmal Nocturnal Hemoglobinuria (PNH) is a clonal disorder of hematopoietic stem cells (HSCs) acquiring mutations in the PIG-A gene. Clonal PIG-A mutations are found in nearly all PNH patents tested resulting in the lack of all GPI-anchored proteins (GPI-APs) in affected HSCs and all the derived hematopoietic progeny. Despite evidences that the biochemical and molecular mechanisms for PNH have been brilliantly elucidated, mechanisms of PIG-A mutant clonal dominance in PNH patients and the close relationship of PNH to other marrow failure diseases such as aplastic anemia (AA) and myelodysplasia syndrome (MDS) are still unknown. Monitoring the PIG-A mutations in patients is impossible at early stage before the onset of clinical features. Creating a PIG-A mutation in human HSCs from healthy volunteers is not feasible currently and existing Pig-a null mice lacking GPI-APs in. blood cells have not replicated the PNH symptoms seen in patients. The lone term objective of this research is understanding effects of GPI-APs in bone marrow failure syndrome and hematopoesis. In response to PA-05-013, a human ESC-based [NIH-approved, WA01(H1)], prospective experimental system is proposed here to investigate effects of the PIG-A/GPI-AP deficiency in human hematopoietic cells. The NIH-approved human ESC that contain an induced PIG-A mutation and are GPI-APs deficient will be used, and effects of PIG-A/GPI-AP deficiency using human ESC-initiated hematopoiesis systems recently developed will be examined. The successful completion of this project may also provide a novel genetic model to investigate normal and abnormal human hematopoiesis and HSCs. Improved understanding of PIG-A/GPI-AP deficiency in human hematopoiesis and HSCs will in turn help us to improve and develop new treatments for PNH and other related blood diseases such as AA & MDS.