Fig. 6. Dual-gene transfer, selection, and expression in vivo.

Three different cohorts of mice (n = 7–9 animals per cohort) were transplanted with cotransduced (M+G), MIG-transduced, or MAG-transduced LSK cells. (A) Total MGMT⁺ and GFP⁺ percentages in murine PBLs at 5 and 16 weeks after transplant. At 5 weeks post-transplant animals were mock treated or selected with 3 rounds of 30 mg/kg BG and 10 mg/kg BCNU, with 3 weeks between each round of selection. Values at 5 weeks are the expression percentages prior to the first treatment. Values depicted at 16 weeks are expression percentages in mock treated (UT; n = 2 animals per transduction cohort) or thrice-treated (T; n = 5–7 animals per transduction cohort) animals. (B–E) Recipient animals in A were sacrificed at 17 weeks post-transplant for BMC analysis. (B) Total MGMT⁺ and GFP⁺ expression percentages in total BMCs. (C) Total GFP⁺ expression percentages in lymphoid (B220⁺) and granulocytic (Gr-1⁺) BMCs. (D) Relative MFI of MGMT or GFP expression (measured separately) in BMCs. (E) The average number of vector insertions per cell, calculated from individual CFU. Values were determined using primers for either MGMT or GFP on individual CFU (n = 8–15 CFU per animal). Mean MGMT and GFP values were calculated from all of the CFU analyzed in each animal. Mean MGMT and GFP values were totaled (cotransduced CFU), or averaged (MIG and MAG transduced CFU) to get the total number of vector insertion per cell. For individual animal data, please see Supplementary Fig. S2. Data in A are means ± s.e.m. *, P<0.05 compared to 5 wks UT; **, P<0.05 compared to 5 or 16 wks UT; #, P<0.05 for MAG compared to either M+G or MIG; ##, P>0.05 for MAG compared to either M+G or MIG; ns, not significant (P>0.05). Data in B–E are means ± s.e.m. *, P<0.05 (T compared to UT); #, P<0.05 for MAG compared to either M+G or MIG; ##, P>0.05 for MAG compared to either M+G or MIG; ns, not significant (P>0.05).