Figure 1.
CsH Is a Potent and Non-toxic Enhancer of Lentiviral Vector Transduction in Hematopoietic Stem Cells
(A) Human cord-blood (CB) cells were transduced with a VSV-g lentiviral (LV) at a multiplicity of infection (MOI) of 1 transducing unit (TU)/293T cell ± 8 μM CsA or CsH. Percentages of transduced cells and vector copy numbers/human genome (VCN/genome) were assessed at 5 or 14 days post-transduction, respectively (mean ± SEM; n = 20; one-way ANOVA with Bonferroni’s multiple comparison, ∗p ≤ 0.05, ∗∗p ≤ 0.01, ∗∗∗∗p ≤ 0.0001).
(B) Impact of CsA and CsH on apoptosis was assessed in hCB-CD34+ cells 48 hr post-transduction (mean ± SEM; n = 6; Dunn’s adjusted Kruskal-Wallis test; ns, not significant).
(C) VSV-g pseudotyped LV was titered in parallel in 293T cells or human CB-CD34+ cells ± 8 μM CsH (mean ± SEM; n = 4; one-way ANOVA with Bonferroni’s multiple comparison; ns, not significant, ∗p ≤ 0.05).
(D and E) Transduction efficiencies (MOI = 1) in human mobilized peripheral blood (mPB)-CD34+ cells (mean ± SEM, n = 4, Mann-Whitney test, ∗p ≤ 0.05) (D) or murine hematopoietic stem and progenitor cells (mHSPC) (mean ± SEM, n = 8, Wilcoxon signed rank test, ∗p = 0.0078) (E).
(F) Transduction efficiencies (MOI = 1) in the different subpopulations of human mPB-CD34+ cells (mean ± SEM, n = 4, Mann-Whitney test versus each DMSO, ∗p ≤ 0.05).
(G and H) The composition (G) and cell-cycle status (H) of human mPB- or CB-CD34+ cells, respectively, were evaluated 48 hr post-transduction.
(I) Impact of CsA and CsH on cell proliferation was assessed in hCB-CD34+ cells 48 hr post-transduction (mean ± SEM; n = 4; Dunn’s adjusted Kruskal-Wallis test, ∗p ≤ 0.05).
(J–L) hCB- (J and K) and mPB- (L) derived HSPC were transduced in the presence of different drug combinations with LV at an MOI of 1 or 10 (mean ± SEM; n = 7 for I and J, and n = 4 for K; one-way ANOVA with Bonferroni’s multiple comparison, ∗p ≤ 0.05, ∗∗p ≤ 0.01, ∗∗∗p ≤ 0.001, ∗∗∗∗p ≤ 0.0001).
(M) Transduction efficiencies ± 8 μM CsH (MOI = 10) in unstimulated hCB-CD34+ cells (unst. hHSPC) (mean ± SEM, n = 4, Mann-Whitney, ∗p ≤ 0.05).
(N) hCB-CD34+ cells were transduced with an integrase defective LV (IDLV) vector at MOI = 50 ± 8 μM CsH.