Figure 2. In vivo HSPC transduction with HDAd-long-LCR containing the 32.4 kb transposon and HDAd-short-LCR containing an 11.8 kb transposon.

(A) Treatment regimen: hCD46tg mice were mobilized and IV injected with either HDAd-short-LCR + HDAd-SB or HDAd-long-LCR +HDAd-SB (2 times each 4 × 10¹⁰ viral particles (vp) of a 1:1 mixture of both viruses). Five weeks later, O⁶BG/BCNU treatment was started. With each cycle, the BCNU concentration was increased from 5 mg/kg, to 7.5 mg/kg, and to 10 mg/kg. The O⁶BG concentration was 30 mg/kg in all 4 treatments. Mice were followed until week 20, when animals were sacrificed for analysis. Bone marrow Lin^– cells were used for transplantation into secondary recipients. Secondary recipients were then followed for 16 weeks. (B) Percentage of human γ-globin–positive cells in peripheral red blood cells (RBCs) measured by flow cytometry. Each symbol is an individual animal. In mice that were mock transduced, fewer than 0.1% of cells were γ-globin–positive. The arrows indicate O⁶BG/BCNU treatment. (C) Levels of γ-globin protein chain measured by HPLC in RBCs at week 20 after in vivo HSPC transduction. Shown are the percentages of human γ-globin to mouse α-globin protein chains. (D) Levels of γ-globin mRNA measured by quantitative reverse transcription (qRT-PCR) in total blood at week 20 after in vivo HSPC transduction. Shown are the percentages of human γ-globin mRNA to mouse α-globin mRNA. (E) VCN per cell in bone marrow MNCs, harvested at week 20 after in vivo HSPC transduction. The difference between the 2 groups is not significant. Statistical analyses were performed using 2-way ANOVA.