Multiplex base editing of BCL11A regulatory elements to treat sickle cell disease

TL;DR

The paper introduces a new gene editing method to treat sickle cell disease by reactivating fetal hemoglobin without causing harmful DNA breaks.

Contribution

Multiplex base editing of BCL11A enhancers achieves higher fetal hemoglobin levels with minimal DNA damage compared to CRISPR-Cas9.

Findings

Base editing disrupts DNA motifs in BCL11A enhancers without generating double-strand breaks.

Combined editing of +58-kb and +55-kb enhancers boosts fetal hemoglobin induction in red blood cells.

Base editing is safe and durable in long-term repopulating hematopoietic stem cells.

Abstract

Sickle cell disease (SCD) is a genetic anemia caused by the production of an abnormal adult hemoglobin. Elevated levels of fetal hemoglobin (HbF) in adulthood reduce disease severity. A promising therapy involves the treatment of hematopoietic stem/progenitor cells (HSPCs) with CRISPR-Cas9 to downregulate the HbF repressor BCL11A via generation of double-strand breaks (DSBs) in the +58-kb enhancer. To improve safety and HbF induction, we use base editors to target both the +58-kb and +55-kb enhancers without generating DSBs. We dissect key DNA motifs recognized by transcriptional activators and identify critical nucleotides. Multiplex base editing efficiently disrupts these sites, reactivating HbF to levels exceeding those achieved with CRISPR-Cas9-induced editing, while minimizing DSBs and genomic rearrangements. Base editing is effective in long-term repopulating HSPCs and results in robust HbF reactivation in vivo. These findings demonstrate that multiplex base editing of BCL11A enhancers is a safe, efficient, and durable strategy to treat SCD.

•Base editing enables precise HbF tuning through the editing of critical DNA motifs•Base editing of +58/+55 enhancers reduces BCL11A mRNA without generation of DSBs•Combined editing of the +58-kb and +55-kb enhancers boosts HbF induction in RBCs•Base editing is safe, efficient, and durable in long-term repopulating HSCs

Base editing enables precise HbF tuning through the editing of critical DNA motifs

Base editing of +58/+55 enhancers reduces BCL11A mRNA without generation of DSBs

Combined editing of the +58-kb and +55-kb enhancers boosts HbF induction in RBCs

Base editing is safe, efficient, and durable in long-term repopulating HSCs

Fontana et al. demonstrate that multiplex base editing of the BCL11A +58-kb and +55-kb enhancers disrupts two key transcriptional activator motifs and leads to robust HbF induction with minimal generation of double-strand breaks and genomic rearrangements. This approach is safe, efficient, and durable in long-term repopulating hematopoietic stem cells.

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