Development of B-lineage Predominant Lentiviral Vectors for Use in Genetic Therapies for B Cell Disorders
Development of B-lineage Predominant Lentiviral Vectors for Use in Genetic Therapies for B Cell Disorders
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DOI：
10.1038/mt.2010.259
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发表时间：
2011-03-01
影响因子：
12.4
通讯作者：
Rawlings, David J.
中科院分区：
文献类型：
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作者：
Sather, Blythe D.;Ryu, Byoung Y.;Rawlings, David J.
在原代B淋巴细胞中持续、靶向、高水平的转基因表达可能有助于B细胞疾病的基因治疗。我们开发了几种候选的B系优势自失活慢病毒载体(LV)，其中包括：免疫球蛋白β(Igβ)(B29)启动子与免疫球蛋白Mu增强子(E Mu B29)相结合的启动子；以及内源性BtK启动子(E Mu Btkp或Btkp)。LV驱动的增强型绿色荧光蛋白(EGFP)报告蛋白在人或小鼠造血干细胞(HSC)来源的细胞系和原代细胞中的表达被评估。在小鼠原代细胞中，与所有其他谱系相比，EmuB29和EmuBtkp LV介导的未成熟和成熟B细胞的高水平表达。随着B细胞的成熟，其表达增加，并维持在外周血细胞亚群中。T细胞和髓样细胞的表达百分率和表达强度均较低。同样，E-Mu-B29和E-Mu-Btkp LV在人原代B细胞中都表现出高水平的活性。与EmuB29相比，Btkp和EmuBtkp LV在髓系细胞中也表现出适度的活性，这与内源性Bruton‘s酪氨酸激酶(BTK)的表达谱一致。值得注意的是，在所有的表达模型中，EmuB29和EmuBtkp的活性都优于另一种B系靶向载体，该载体含有EmuS.CD19增强子/启动子。综上所述，E u B29和E u Btkp LV构成了B系细胞基因表达的高效传递平台。
Sustained, targeted, high-level transgene expression in primary B lymphocytes may be useful for gene therapy in B cell disorders. We developed several candidate B-lineage predominant self-inactivating lentiviral vectors (LV) containing alternative enhancer/promoter elements including: the immunoglobulin beta (Ig beta) (B29) promoter combined with the immunoglobulin mu enhancer (E mu B29); and the endogenous BTK promoter with or without E mu (E mu Btkp or Btkp). LV-driven enhanced green fluorescent protein (eGFP) reporter expression was evaluated in cell lines and primary cells derived from human or murine hematopoietic stem cells (HSC). In murine primary cells, E mu B29 and E mu Btkp LV-mediated high-level expression in immature and mature B cells compared with all other lineages. Expression increased with B cell maturation and was maintained in peripheral subsets. Expression in T and myeloid cells was much lower in percentage and intensity. Similarly, both E mu B29 and E mu Btkp LV exhibited high-level activity in human primary B cells. In contrast to E mu B29, Btkp and E mu Btkp LV also exhibited modest activity in myeloid cells, consistent with the expression profile of endogenous Bruton's tyrosine kinase (Btk). Notably, E mu B29 and E mu Btkp activity was superior in all expression models to an alternative, B-lineage targeted vector containing the E mu S. CD19 enhancer/promoter. In summary, E mu B29 and E mu Btkp LV comprise efficient delivery platforms for gene expression in B-lineage cells.