Development of safety vector for the gene therapy of ribosomal protein S19 deficient Diamond-Blackfan anemia

我们开发了新的慢病毒载体，用于基因治疗核糖体蛋白S19（RPS19）缺陷型Diamond-Blackfan贫血（DBA）。由于该载体需要较强的转基因表达活性，我们在LTR（long term repeat）中构建了慢病毒背生和逆转录病毒U3区之间的杂交载体。利用该载体在DBA患者造血干细胞中观察到较高的转导效率。, 2003)。为了分析DBA红细胞期细胞异常分化的机制，我们与Niklas Dahl合作建立了rps19敲除小鼠。在这种小鼠中，我们无法检测到与人类疾病相关的典型贫血表型。提示小鼠RPS19的功能与人RPS19 （Mol.Cell.Biol）不同。, 2004)。为了进一步分析人RPS19的功能，我们开发了含有sirna表达盒的慢病毒载体。当我们在人脐带血CD34+细胞中转导siRNA基因对抗RPS19时，在转导的细胞中，红系青春期细胞的生长和分化受到干扰（blood, 2005）。结合这些结果，我们揭示了RPS19的功能，并利用siRNA载体建立了DBA的干细胞模型。利用这一模型，我们希望分析DBA的整个机制，并开发新的治疗方法。

We have developed new lentiviral vector for the gene therapy of ribosomal protein S19(RPS19) deficient Diamond-Blackfan anemia(DBA). Since the strong transgene expression activity was required for this vector, we developed hybrid vector between lentiviral backborn and retrovirus U3 region in the LTR(long term repeat). Using this vector high transduction efficiency was observed in the DBA patient hematopoietic stem cells (Mol.Ther., 2003).In order to analyze the mechanisms for abnormal differentiation of erythroid leneage cells in DBA, RPS19-knock out mice were generated in the corraboration study with Niklas Dahl. In this mice we could not detect the typical phenotype of anemia correlated to human disease. It is suggested that the function of mouse RPS19 is different from human RPS19 (Mol.Cell.Biol., 2004).For further analyze of human RPS19 function, we developed lentiviral vector containing siRNA-expression cassette against RPS19. When we transduced siRNA gene against RPS19 in human cord blood CD34+ cells, the growth and differentiation of erythroid leneage cells were disturbed in the transduced cells (Blood, 2005).Together with these results, we have revealed the function of RPS19 and developed the stem cell model for DBA using siRNA vector. By using this model we would like to analyze the whole mechanisms of DBA, and develop the new therapy.