The sensitivity of LV'VFas⁺ T cells to AP1903 correlates with the level of transgene expression. (A) Analysis of ΔLNGFR expression in LV'VFas⁺ T cells by flow cytometry either prior to rest (thin line), rested for 12 days in the presence of autologous γ-irradiated feeder cells (dashed line), or after subsequent in vitro reactivation using anti-CD3 and anti-CD28 mAbs and culture for 14 days (thick line). The LV'VFas⁺ T cells were stained with anti-CD3 and anti-LNGFR mAbs, and expresssion of ΔLNGFR was evaluated by gating on CD3⁺ T cells. Unmodified T cells (dotted line) were stained in an identical fashion. (B) Aliquots of the T-cell cultures either prior to rest, rested, or after reactivation, were exposed to 10 nM of AP1903 (▪) or to medium alone (□), and viability was assessed by trypan blue exclusion after 24 hours. (C-D) Residual LV'V and LV'VFas⁺ T cells display reduced levels of ΔLNGFR expression. Analysis of PBMCs for the in vivo persistence of LV'V⁺ and LV'VFas⁺ T cells after simultaneous transfer to macaque no. 2. Dosing of AP1903 (0.2 mg/kg) was begun 1 day after the T-cell infusion and was then given every other day for a total of 5 doses. PBMCs collected from macaque no. 2 1 day (C) or 14 days (D) after the T-cell infusion were stained with both anti-LNGFR and anti-CD3 mAbs, and evaluated by flow cytometry. The cells are gated on CD3⁺ T cells. The mean fluorescence intensities (MFIs) of the ΔLNGFR expression are indicated in the upper right of each panel.