Division of Medical Genetics, Department of Medicine, University of Washington, Seattle, WA.
Magenta Therapeutics, Cambridge, MA.
Blood Adv. 2021 Mar 9;5(5):1239-1249. doi: 10.1182/bloodadvances.2020003714.
We have developed an in vivo hemopoietic stem cell (HSC) gene therapy approach without the need for myelosuppressive conditioning and autologous HSC transplantation. It involves HSC mobilization and IV injection of a helper-dependent adenovirus HDAd5/35++ vector system. The current mobilization regimen consists of granulocyte colony-stimulating factor (G-CSF) injections over a 4-day period, followed by the administration of plerixafor/AMD3100. We tested a simpler, 2-hour, G-CSF-free mobilization regimen using truncated GRO-β (MGTA-145; a CXCR2 agonist) and plerixafor in the context of in vivo HSC transduction in mice. The MGTA-145+plerixafor combination resulted in robust mobilization of HSCs. Importantly, compared with G-CSF+plerixafor, MGTA-145+plerixafor led to significantly less leukocytosis and no elevation of serum interleukin-6 levels and was thus likely to be less toxic. With both mobilization regimens, after in vivo selection with O6-benzylguanine (O6BG)/BCNU, stable GFP marking was achieved in >90% of peripheral blood mononuclear cells. Genome-wide analysis showed random, multiclonal vector integration. In vivo HSC transduction after mobilization with MGTA-145+plerixafor in a mouse model for thalassemia resulted in >95% human γ-globin+ erythrocytes at a level of 36% of mouse β-globin. Phenotypic analyses showed a complete correction of thalassemia. The γ-globin marking percentage and level were maintained in secondary recipients, further demonstrating that MGTA145+plerixafor mobilizes long-term repopulating HSCs. Our study indicates that brief exposure to MGTA-145+plerixafor may be advantageous as a mobilization regimen for in vivo HSC gene therapy applications across diseases, including thalassemia and sickle cell disease.
我们开发了一种无需骨髓抑制预处理和自体造血干细胞（HSC）移植的体内 HSC 基因治疗方法。它涉及 HSC 动员和 IV 注射辅助依赖性腺病毒 HDAd5/35++载体系统。目前的动员方案包括为期 4 天的粒细胞集落刺激因子（G-CSF）注射，然后给予plerixafor/AMD3100。我们在小鼠体内 HSC 转导的背景下，测试了一种更简单的、2 小时的、无 G-CSF 的动员方案，使用截断的 GRO-β（MGTA-145；CXCR2 激动剂）和 plerixafor。MGTA-145+plerixafor 联合使用导致 HSC 大量动员。重要的是，与 G-CSF+plerixafor 相比，MGTA-145+plerixafor 导致白细胞计数显著减少，血清白细胞介素-6 水平没有升高，因此可能毒性更小。使用这两种动员方案，在体内用 O6-苯甲基鸟嘌呤（O6BG）/BCNU 选择后，外周血单核细胞中稳定的 GFP 标记达到>90%。全基因组分析显示随机的、多克隆载体整合。MGTA-145+plerixafor 动员后在β地中海贫血小鼠模型中进行体内 HSC 转导，导致>95%的人类γ-球蛋白+红细胞，水平为 36%的小鼠β-球蛋白。表型分析显示β地中海贫血完全纠正。γ-球蛋白标记百分比和水平在二级受者中得以维持，进一步表明 MGTA145+plerixafor 动员长期重编程 HSC。我们的研究表明，短暂接触 MGTA-145+plerixafor 可能是一种有利的动员方案，适用于包括β地中海贫血和镰状细胞病在内的各种疾病的体内 HSC 基因治疗应用。
