Therapeutic adenine base editing of human hematopoietic stem cells

作者：Liao, Jiaoyang; Chen, Shuanghong^*; Hsiao, Shenlin; Jiang, Yanhong; Yang, Yang; Zhang, Yuanjin; Wang, Xin; Lai, Yongrong; Bauer, Daniel E.; Wu, Yuxuan^*

来源：Nature Communications, 2023, 14(1): 207.

DOI：10.1038/s41467-022-35508-7

• FETAL-HEMOGLOBIN
• HEREDITARY PERSISTENCE
• CAS9 CLEAVAGE
• GENOMIC DNA
• BCL11A
• PHENOTYPE
• ENHANCER

In beta-thalassemia, either gamma-globin induction to form fetal hemoglobin (alpha 2 gamma 2) or beta-globin repair to restore adult hemoglobin (alpha 2 beta 2) could be therapeutic. ABE8e, a recently evolved adenine base editor variant, can achieve efficient adenine conversion, yet its application in patient-derived hematopoietic stem cells needs further exploration. Here, we purified ABE8e for ribonucleoprotein electroporation of beta-thalassemia patient CD34(+) hematopoietic stem and progenitor cells to introduce nucleotide substitutions that upregulate gamma-globin expression in the BCL11A enhancer or in the HBG promoter. We observed highly efficient on-target adenine base edits at these two regulatory regions, resulting in robust gamma-globin induction. Moreover, we developed ABE8e-SpRY, a near-PAMless ABE variant, and successfully applied ABE8e-SpRY RNP to directly correct HbE and IVS II-654 mutations in patient-derived CD34(+) HSPCs. Finally, durable therapeutic editing was produced in self-renewing repopulating human HSCs as assayed in primary and secondary recipients. Together, these results support the potential of ABE-mediated base editing in HSCs to treat inherited monogenic blood disorders. @@@ Here, Liao and colleagues apply adenine base editor ABE8e and its PAM-less variant ABE8e-SpRY to beta-thalassemia patient hematopoietic stem cells in the form of ribonucleoprotein complexes, resulting in efficient long-term editing and beta-thalassemia alleviation.

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