Factor VIII delivered by haematopoietic stem cell-derived B cells corrects the phenotype of haemophilia A mice.
Factor VIII delivered by haematopoietic stem cell-derived B cells corrects the phenotype of haemophilia A mice.
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DOI：
10.1160/th10-11-0725
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发表时间：
2011-04
影响因子：
6.7
通讯作者：
Hawley RG
中科院分区：
文献类型：
--
作者：
Ramezani A;Zweier-Renn LA;Hawley RG
临床应用造血干细胞（HSC）基因疗法治疗甲型血友病的主要障碍是骨髓移植相关风险以及逆转录病毒载体引起的插入突变的可能性。为了规避这些限制，我们采用了非清髓性预处理方案，并使用含有免疫球蛋白重链增强子-启动子的绝缘慢病毒载体将因子 VIII (FVIII) 蛋白定向合成到 B 谱系细胞。在研究期间（6 个月），慢病毒载体修饰的 HSC 移植导致所有移植小鼠循环中的 FVIII 达到治疗水平。脾细胞免疫染色显示大部分 FVIII 由 B220+ B 细胞和 CD138+ 浆细胞合成。随后用重组 FVIII 进行攻击最多仅引发轻微的抗 FVIII 抗体反应，表明诱导了免疫低反应。所有移植受者均表现出凝块形成并在剪尾后存活下来，表明其血友病表型得到纠正。治疗水平的 FVIII 可以通过骨髓移植转移至二级受体，证实基因转移至长期再生的 HSC 中。此外，通过 HSC 来源的脾 B 细胞的过继转移，也可以在二次受者中实现短期治疗性 FVIII 水平。我们的研究结果支持将 B 细胞定向蛋白质递送作为治疗 A 型血友病和其他可通过全身分布蛋白质纠正的疾病的潜在临床方法。
The main impediments to clinical application of hematopoietic stem cell (HSC) gene therapy for treatment of hemophilia A are the bone marrow transplant-related risks and the potential for insertional mutagenesis caused by retroviral vectors. To circumvent these limitations, we have adapted a nonmyeloablative conditioning regimen and directed factor VIII (FVIII) protein synthesis to B lineage cells using an insulated lentiviral vector containing an immunoglobulin heavy chain enhancer-promoter. Transplantation of lentiviral vector-modified HSCs resulted in therapeutic levels of FVIII in the circulation of all transplanted mice for the duration of the study (6 months). Immunostaining of spleen cells showed that the majority of FVIII was synthesized by B220+ B cells and CD138+ plasma cells. Subsequent challenge with recombinant FVIII elicited at most a minor anti-FVIII antibody response, demonstrating induction of immune hyporesponsiveness. All transplant recipients exhibited clot formation and survived tail clipping, indicating correction of their hemophilic phenotype. Therapeutic levels of FVIII could be transferred to secondary recipients by bone marrow transplantation, confirming gene transfer into long-term repopulating HSCs. Moreover, short-term therapeutic FVIII levels could also be achieved in secondary recipients by adoptive transfer of HSC-derived splenic B cells. Our findings support pursuit of B cell-directed protein delivery as a potential clinical approach to treat hemophilia A and other disorders correctable by systemically distributed proteins.