Dana-Farber/Boston Children's Cancer and Blood Disorders Center, Harvard Medical School, Boston, MA, USA.
Gene Therapy Consulting, Padua, Italy.
Nat Commun. 2024 Nov 25;15(1):10192. doi: 10.1038/s41467-024-54515-4.
Transplantation of engineered hematopoietic stem/progenitor cells (HSPCs) showed curative potential in patients affected by neurometabolic diseases treated in early stage. Favoring the engraftment and maturation of the engineered HSPCs in the central nervous system (CNS) could allow enhancing further the therapeutic potential of this approach. Here we unveil that HSPCs haplo-insufficient at the Cx3cr1 (Cx3cr1) locus are favored in central nervous system (CNS) engraftment and generation of microglia-like progeny cells (MLCs) as compared to wild type (Cx3cr1) HSPCs upon transplantation in mice. Based on this evidence, we have developed a CRISPR-based targeted gene addition strategy at the human CX3CR1 locus resulting in an enhanced ability of the edited human HSPCs to generate mature MLCs upon transplantation in immunodeficient mice, and in lineage specific, regulated and robust transgene expression. This approach, which benefits from the modulation of pathways involved in microglia maturation and migration in haplo-insufficient cells, may broaden the application of HSPC gene therapy to a larger spectrum of neurometabolic and neurodegenerative diseases.
移植经过工程改造的造血干/祖细胞（HSPCs）在早期接受治疗的神经代谢疾病患者中显示出潜在的治愈效果。有利于工程化 HSPCs 在中枢神经系统（CNS）中的植入和成熟，可以进一步增强这种方法的治疗潜力。在这里，我们揭示了与野生型（Cx3cr1）HSPCs 相比，在 Cx3cr1（Cx3cr1）基因座处单倍体不足的 HSPCs 在移植到小鼠后更有利于中枢神经系统（CNS）植入和生成类似于小神经胶质细胞的祖细胞（MLCs）。基于这一证据，我们在人类 CX3CR1 基因座开发了一种基于 CRISPR 的靶向基因添加策略，从而使编辑后的人类 HSPCs 在移植到免疫缺陷小鼠后产生成熟的 MLC 的能力增强，并实现谱系特异性、调控和稳健的转基因表达。这种方法受益于对单倍体不足细胞中参与小神经胶质细胞成熟和迁移的途径的调节，可能会将 HSPC 基因治疗的应用扩展到更广泛的神经代谢和神经退行性疾病谱。
