Wongborisuth C.Innachai P.Saisawang C.Tubsuwan A.Jearawiriyapaisarn N.Kaewprommal P.Piriyapongsa J.Chiangjong W.Anurathapan U.Songdej D.Tangprasittipap A.Hongeng S.Mahidol University2025-07-222025-07-222025-12-01Scientific Reports Vol.15 No.1 (2025)https://repository.li.mahidol.ac.th/handle/123456789/111309CRISPR/Cas9 genome editing has emerged as a promising treatment for genetic diseases like β-thalassemia. Editing γ-globin promoters to disrupt ZBTB7A/LRF or BCL11A binding sites has shown potential for reactivating fetal hemoglobin and treating sickle cell disease. However, its application to β⁰-thalassemia/HbE disease remains unclear. This study utilized CRISPR/Cas9 to disrupt these sites in mobilized CD34 + hematopoietic stem /progenitor cells from healthy donors and β⁰-thalassemia/HbE patients. The editing efficiency for the BCL11A site (75–92%) was higher than for the ZBTB7A/LRF site (57–60%). Both disruptions similarly increased fetal hemoglobin production in healthy donors (BCL11A 26.2 ± 1.4%, ZBTB7A/LRF 27.9 ± 1.5%) and β⁰-thalassemia/HbE cells (BCL11A 62.7 ± 0.9%, ZBTB7A/LRF 64.0 ± 1.6%). Off-target effects were absent in BCL11A-edited cells but observed at low frequencies in ZBTB7A/LRF-edited cells. Neither disruption significantly affected erythroid differentiation. These findings highlight the comparable contributions of ZBTB7A/LRF and BCL11A binding sites to γ-globin reactivation. CRISPR/Cas9 editing of either site may offer a potential therapeutic strategy for β⁰-thalassemia/HbE disease.MultidisciplinaryArticleSCOPUS10.1038/s41598-025-10791-82-s2.0-10501059897620452322