Induction of Fetal Hemoglobin by Introducing Natural Hereditary Persistence of Fetal Hemoglobin Mutations in the γ-Globin Gene Promoters for Genome Editing Therapies for β-Thalassemia.

Reactivation of γ-globin expression is a promising therapeutic approach for β-hemoglobinopathies. Here, we propose a novel Cas9/AAV6-mediated genome editing strategy for the treatment of β-thalassemia: Natural HPFH mutations -113A > G, -114C > T, -117G>A, -175T > C, -195C > G, and -198T > C were introduced by homologous recombination following disruption of BCL11A binding sites in promoters. Precise on-target editing and significantly increased γ-globin expression during erythroid differentiation were observed in both HUDEP-2 cells and primary HSPCs from β-thalassemia major patients. Moreover, edited HSPCs maintained the capacity for long-term hematopoietic reconstitution in B-NDG hTHPO mice. This study provides evidence of the effectiveness of introducing naturally occurring HPFH mutations as a genetic therapy for β-thalassemia.