Cellectis S.A., 8 Rue de la Croix Jarry, Paris, France.
Université Paris Cité, Imagine Institute, Laboratory of Chromatin and Gene Regulation During Development, INSERM UMR 1163, Paris, France.
Nat Commun. 2024 Jun 11;15(1):4965. doi: 10.1038/s41467-024-49353-3.
Sickle cell disease is a devastating blood disorder that originates from a single point mutation in the HBB gene coding for hemoglobin. Here, we develop a GMP-compatible TALEN-mediated gene editing process enabling efficient HBB correction via a DNA repair template while minimizing risks associated with HBB inactivation. Comparing viral versus non-viral DNA repair template delivery in hematopoietic stem and progenitor cells in vitro, both strategies achieve comparable HBB correction and result in over 50% expression of normal adult hemoglobin in red blood cells without inducing β-thalassemic phenotype. In an immunodeficient female mouse model, transplanted cells edited with the non-viral strategy exhibit higher engraftment and gene correction levels compared to those edited with the viral strategy. Transcriptomic analysis reveals that non-viral DNA repair template delivery mitigates P53-mediated toxicity and preserves high levels of long-term hematopoietic stem cells. This work paves the way for TALEN-based autologous gene therapy for sickle cell disease.
镰状细胞病是一种严重的血液疾病，起源于血红蛋白 HBB 基因编码的单个点突变。在这里，我们开发了一种符合 GMP 标准的 TALEN 介导的基因编辑过程，通过 DNA 修复模板实现高效的 HBB 校正，同时最大限度地降低 HBB 失活相关的风险。在体外比较造血干细胞和祖细胞中病毒与非病毒 DNA 修复模板的传递，两种策略都能实现相当的 HBB 校正，并导致红细胞中正常成人血红蛋白的表达超过 50%，而不会诱导β-地中海贫血表型。在免疫缺陷的雌性小鼠模型中，与使用病毒策略编辑的细胞相比，使用非病毒策略编辑的移植细胞具有更高的植入和基因校正水平。转录组分析表明，非病毒 DNA 修复模板的传递减轻了 P53 介导的毒性，并保持了高水平的长期造血干细胞。这项工作为镰状细胞病的基于 TALEN 的自体基因治疗铺平了道路。
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