Restoration of T and B cell generation in X-linked severe combined immunodeficiency mice through adenine base editing of hematopoietic stem cells.

Base editing of hematopoietic stem/progenitor cells (HSPCs) is an attractive strategy for treating immunohematologic diseases. However, the feasibility of using adenine base-edited HSPCs for treating X-linked severe combined immunodeficiency (SCID-X1), the influence of dose-response relationships on immune cell generation, and the potential risks have not been demonstrated in vivo. Here, a humanized SCID-X1 mouse model was established and 86.67% ± 2.52% (n = 3) of mouse hematopoietic stem cell (HSC) pathogenic mutations were corrected, with no sgRNA-dependent off-target effects detected. Analysis of peripheral blood over 16 weeks post-transplantation in mice with different immunodeficiency backgrounds revealed efficient immune cell generation following transplantation of different amounts of modified HSCs. Therefore, a large-scale infusion of gene-corrected HSCs within a safe range can achieve rapid, stable, and durable immune cells regeneration. Tissue section staining further demonstrated the restoration of immune organ tissue structures, with no tumor formation in multiple organs. Collectively, these data suggest that base-edited HSCs are a potential therapeutic approach for SCID-X1 and that a threshold infusion dose of gene-corrected cells is required for immune cell regeneration. This study lays a theoretical foundation for the clinical application of base-edited HSCs in treating SCID-X1.