Horn P A, Morris J C, Bukovsky A A, Andrews R G, Naldini L, Kurre P, Kiem H-P
Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, WA, USA.
Gene Ther. 2002 Nov;9(21):1464-71. doi: 10.1038/sj.gt.3301820.
Efficient transduction of hematopoietic stem cells is a prerequisite for successful hematopoietic stem cell gene therapy. Oncoretroviral vectors are the most widely used vectors for hematopoietic gene therapy studies. However, these vectors require cell division, and thus efficient transduction of quiescent stem cells has been difficult to achieve. Lentiviral vectors can transduce non-dividing cells and therefore may be more efficient in transducing quiescent hematopoietic stem cells. We have used a competitive repopulation assay in the baboon to compare transduction of hematopoietic repopulating cells by lentiviral and oncoretroviral vectors. Baboon CD34-enriched marrow cells were transduced in the presence or absence of multiple hematopoietic growth factors using a short, 2-day, transduction protocol. Here, we show that efficient lentiviral transduction of hematopoietic repopulating cells was only achieved when cells were transduced in the presence of multiple growth factors. Using these conditions, up to 8.6% of hematopoietic repopulating cells were genetically modified by the lentiviral vector more than 1 year after transplant. Interestingly, the number of lentivirally marked cells increased over time in three of four animals. In conclusion, these results suggest that lentiviral vectors are able to tranduce multilineage hematopoietic stem cells, and thus, may provide an alternative vector system for clinical stem cell gene therapy applications.
造血干细胞的高效转导是成功进行造血干细胞基因治疗的前提条件。γ-逆转录病毒载体是造血基因治疗研究中使用最广泛的载体。然而，这些载体需要细胞分裂，因此难以实现对静止干细胞的高效转导。慢病毒载体可以转导非分裂细胞，因此在转导静止造血干细胞方面可能更有效。我们在狒狒中使用竞争性再增殖试验来比较慢病毒载体和γ-逆转录病毒载体对造血再增殖细胞的转导情况。使用为期2天的短时间转导方案，在有或没有多种造血生长因子的情况下，对富集狒狒CD34的骨髓细胞进行转导。在此，我们表明只有在存在多种生长因子的情况下转导细胞时，才能实现造血再增殖细胞的高效慢病毒转导。在这些条件下，移植1年多后，高达8.6%的造血再增殖细胞被慢病毒载体进行了基因改造。有趣的是，在四只动物中的三只中，慢病毒标记的细胞数量随时间增加。总之，这些结果表明慢病毒载体能够转导多谱系造血干细胞，因此，可能为临床干细胞基因治疗应用提供一种替代载体系统。
