In vivo HSC transduction studies in rhesus macaques with HDAd5/35++GFP, HDAd5/3+GFP, HDAd5/35++GFP/cxcr4, and HDAd5/3+GFP/cxcr4. (A) Schematic of the experiment showing the injection time of the mobilization drugs (G-CSF and AMD3100) and cytokine prophylaxis drugs (dexamethasone, anakinra, and tocilizumab), as well as the injection time of the HDAd vector at a dose of 0.4 × 10¹² vp/kg. The time point of HDAd injection is taken as “day 0, 0 hours.” HDAd5/35++GFP– and HDAd5/3+GFP–injected animals were monitored for 7 days. HDAd5/35++GFP/cxcr4– and HDAd5/3+GFP/cxcr4–injected animals were observed for 8 weeks to capture late effects of cxcr4 expression. Bone marrow aspirates (BMAs) were performed at days 3, 7, and (for HDAd5/35++GFP/cxcr4 and HDAd5/3+GFP/cxcr4) at weeks 4 and 8. Blood samples were taken at 0 hours (before HDAd injection) and 2, 6, 9, and 12 hours on day 0; on days 1, 2, 3, and 7; and then weekly. (B) HSC mobilization. Shown are numbers of primitive (CD34⁺/CD45RA^–/CD90⁺) HSCs in peripheral blood. HDAd vectors were IV injected at day 0, 0 hours. Notably, there was no correlation between the weight of the animals and the efficacy of mobilization. The weights were 9.70 kg (HDAd5/35++GFP), 6.64 kg (HDAd5/3+GFP), 7.13 kg (HDAd5/35++GFP/cxcr4), and 7.19 kg (HDAd5/3+GFP/cxcr4). (C) Vector clearance from blood. HDAd vector genome copies in serum samples were measured by using quantitative polymerase chain reaction.