Bone marrow stromal cells as targets for gene therapy of hemophilia A
Bone marrow stromal cells as targets for gene therapy of hemophilia A
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DOI：
10.1089/hum.1998.9.3-353
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发表时间：
1998-02-10
影响因子：
4.2
通讯作者：
Vandendriessche, T
中科院分区：
文献类型：
--
作者：
Chuah, MKL;Brems, H;Vandendriessche, T
使用原代T细胞或骨髓干细胞/祖细胞来开发血友病A的体外基因治疗策略的尝试失败了，因为这些细胞类型不能表达凝血因子VIII(FVIII)。作为替代，我们评估了骨髓来源的基质细胞的潜力，这些细胞可以很容易地在体外获得和扩增。用含内含子的Moloney小鼠白血病病毒(MoMLV)逆转录病毒载体(MFG-FVIII Delta B)转导人和小鼠骨髓基质细胞。通过磷酸盐去除和离心，转导效率提高了10-15倍，这消除了对转导的BM基质细胞的选择性浓缩的需要，这导致在转导的人(每24小时180+/-4 ng FVIII/10(6)细胞)和小鼠(每24小时900+/-130 ng FVIII/10(6)细胞)BM基质细胞中FVIII高表达。带有长臂猿白血病病毒外壳的MFG-FVIII Delta B逆转录病毒载体(Galv-env)的假分型导致转导的人BM基质细胞的转导效率(100+/-20%)和FVIII表达水平(每24小时390+/-10 ng FVIII/10(6)个细胞)显著高于标准两性载体，这种转导效率的差异与Galv-env伪型病毒载体的较高滴度以及人BM基质细胞中Galv受体(GLVR-1)与两性受体(GLVR-2)的较高表达水平有关。这些发现证明了骨髓基质细胞在基因治疗方面的潜力，尤其是血友病A。
Attempts to develop an ex vivo gene therapy strategy for hemophilia A, using either primary T cells or bone marrow (BM) stem/progenitor cells have been unsuccessful, due to the inability of these cell types to express coagulation factor VIII (FVIII). As an alternative, we evaluated the potential of BM-derived stromal cells which can be readily obtained and expanded in vitro. Human and murine BM stromal cells were transduced with an intron-based Moloney murine leukemia virus (MoMLV) retroviral vector expressing a B-domain-deleted human factor VIII cDNA (designated as MFG-FVIII Delta B). Transduction efficiencies were increased 10- to 15-fold by phosphate depletion and centrifugation, which obviated the need for selective enrichment of the transduced BM stromal cells, This resulted in high FVIII expression levels in transduced human (180 +/- 4 ng FVIII/10(6) cells per 24 hr) and mouse (900 +/- 130 ng FVIII/10(6) cells per 24 hr) BM stromal cells, Pseudotyping of the MFG-FVIII Delta B retroviral vectors with the gibbon ape leukemia virus envelope (GALV-env) resulted in significantly higher transduction efficiencies (100 +/- 20%) and FVIII expression levels (390 +/- 10 ng FVIII/10(6) cells per 24 hr) in transduced human BM stromal cells than with standard amphotropic vectors, This difference in transduction efficiency correlated with the higher titer of the GALV-env pseudotyped viral vectors and with the higher GALV receptor (GLVR-1) versus amphotropic receptor (GLVR-2) mRNA expression levels in human BM stromal cells, These findings demonstrate the potential of BM stromal cells for gene therapy in general and hemophilia A in particular.