Enhanced γ-globin reactivation and sickle cell correction through a repressor-to-activator motif switch in the HBG1/2 promoters.

Authors

Anne Chalumeau, Panagiotis Antoniou, Maria Bou Dames, Pierre Martinucci, Elena Retana, Pragya Gupta, Mike Firth, Muralidhar Reddivari, Aathira Sarath Chandran, Jonathan S Yen, Martin Peterka, Marcello Maresca, Mégane Brusson, Annarita Miccio

Published in

Molecular therapy. Nucleic acids. Volume 37. Issue 3. Pages 102974. Sep 08, 2026. Epub Jun 06, 2026.

Abstract

Sickle cell disease (SCD) is caused by the production of an abnormal adult hemoglobin that generates sickle-shaped red blood cells (RBCs). Transplantation of autologous genetically corrected hematopoietic stem/progenitor cells (HSPCs) represents a promising therapy. Persistent fetal hemoglobin expression improves SCD. Here, we engineered the fetal HBG1/2 promoters by replacing the BCL11A repressor-binding site (BS) with a TAL1:GATA1 motif recognized by transcriptional activators. We exploited the prime editing nuclease (PEn) that efficiently installed the TAL1:GATA1 motif in K562 cells, outperforming the original PE. Non-homologous end joining (NHEJ) and/or alternative-end joining (alt-EJ) pathway inhibition enhanced precise editing. However, this strategy was poorly efficient in patients' HSPCs. Alternatively, we used CRISPR-Cas9 nuclease to either disrupt the BCL11A BS via NHEJ and/or alt-EJ or to replace it with the TAL1:GATA1 motif via homology-directed repair (HDR) using a donor ssODN template. NHEJ and alt-EJ inhibition improved product purity, reducing insertions and deletions (indels), and achieving superior precise editing efficiency compared to PEn in K562 and HSPCs. HDR-edited HSPCs preserved clonogenic capacity and differentiated into RBCs showing elevated HBG expression and correction of the sickling phenotype. These results indicate that replacing the BCL11A BS with a TAL1:GATA1 motif is a potent strategy for reactivating HBG1/2 to treat SCD.

PMID:
42405279
Bibliographic data and abstract were imported from PubMed on 06 Jul 2026.

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