Disruption of the BCL11A Erythroid Enhancer Reactivates Fetal Hemoglobin in Erythroid Cells of Patients with β-Thalassemia Major

Abstract

In the present report, we carried out clinical-scale editing in adult mobilized CD34+ hematopoietic stem and progenitor cells (HSPCs) using zinc-finger nuclease-mediated disruption of BCL11a to upregulate the expression of γ-globin (fetal hemoglobin). In these cells, disruption of the erythroid-specific enhancer of the BCL11A gene increased endogenous γ-globin expression to levels that reached or exceeded those observed following knockout of the BCL11A coding region without negatively affecting survival or in vivo long-term proliferation of edited HSPCs and other lineages. In addition, BCL11A enhancer modification in mobilized CD34+ cells from patients with β-thalassemia major resulted in a readily detectable γ-globin increase with a preferential increase in G-gamma, leading to an improved phenotype and, likely, a survival advantage for maturing erythroid cells after editing. Furthermore, we documented that both normal and β-thalassemia HSPCs not only can be efficiently expanded ex vivo after editing but can also be successfully edited post-expansion, resulting in enhanced early in vivo engraftment compared with unexpanded cells. Overall, this work highlights a novel and effective treatment strategy for correcting the β-thalassemia phenotype by genome editing.

Lentiviral and genome-editing strategies for the treatment of β-hemoglobinopathies

“…In vitro, studies targeting the BCL11A erythroid enhancer using ZFNs or CRISPR/Cas9 have reported a high genome editing rate (up to ;90%) and high g-globin levels (accounting for up to ;40% of total b-like globin chains) in erythroid cells derived from genetically modified adult HSPCs from healthy donors and patients with b-thalassemia or SCD. 74,79,82 Elevated HbF levels were associated with an amelioration of the b-thalassemic and SCD cell phenotypes. 79 Transplantation studies have shown that after ZFN or CRISPR/ Cas9 transfection, BCL11A-edited HSPCs can stably engraft in immunodeficient mice, with an NHEJ frequency of #95%.…”

Genome editing of HBG1 and HBG2 to induce fetal hemoglobin

“…5,[7][8][9][10][11][12] Second, fetal hemoglobin (HbF, a2g2) can be induced in adult red blood cells (RBCs) by using nonhomologous end-joining (NHEJ) mediated mutations to disrupt noncoding DNA regulatory elements that repress transcription of the genes encoding g-globin (HBG1 and HBG2) postnatally. 4,[13][14][15][16][17][18][19] Numerous clinical studies show that elevated HbF production is associated with reduced morbidity and mortality in SCD and b-thalassemia. 20,21 In extreme cases, a rare, benign genetic condition called hereditary persistence of HbF (HPFH) induces high-level pancellular HbF expression in RBCs and eliminates the pathologies of coinherited b-hemoglobinopathies.…”

Gene therapies for transfusion dependent β‐thalassemia: Current status and critical criteria for success

“…BCL11a, the TF that controls the switch from HbF to HbA and functions as a repressor of HbF, provides an excellent target for gene‐therapy approaches for hemoglobinopathies 40,41 . By suppressing BCL11A , it is postulated that HbF production can be re‐triggered in erythroid progenitors of thalassemia patients to a sufficient degree to ameliorate anemia and avoid transfusions.…”