A Self-Inactivating Retrovector Incorporating the IL-2 Promoter for Activation- Induced Transgene Expression in Genetically Engineered T-Cells - Genetic Engineering: Recent Developments in Applications - page 231

of SINIL-2pr-transduced Jurkat P116 T-cells showed level of EGFP fluorescence
(MnX = 1.86, Figure 4C) that was similar to background fluorescence of control
null cells (MnX = 1.12, Figure 4A). Co-stimulation of the SINIL-2 pr-modified
cells with 1 µ M ionomycin and 10 ng/ml PMA for ~6 hr resulted in a marked
increase in EGFP reporter expression (MnX = 8.28, Figure 4D) as compared to
stimulated null cells (MnX = 1.16, Figure 4B). Moreover, following drug stimula-
tion, there was a clear-cut segregation between the EGFP-expressing population
and the EGFP-negative population. Results obtained from three independent ex-
periments (Figure 4E) showed an average 3.4 ± 0.4 - fold increase in relative mean
EGFP expression in SINIL-2pr - transduced Jurkat P116 cells following ionomy-
cin and PMA treatment which was significantly higher than that in untreated
transduced cells (P = 0.029). This activation-induced expression was inhibited
when the Jurkat P116 cells were pretreated for 30 minutes with 300 nM CsA.
Figure 3. Activation-induced EGFP expression in Jurkat cells transduced with SINIL-2pr retroparticles. A-D.
Flow cytometry analysis of mean EGFP expression in Jurkat T-cells transduced with SINIL-2pr retroparticles
relative to Jurkat null cells. Expression was measured at ~48 hr post-activation with 1 µ M ionomycin and 10
ng/ml PMA relative to untreated cells. E. Cyclosporin A-sensitive induction of EGFP expression in Jurkat
cells transduced with SINIL-2pr. Co-stimulation of the SINIL-2pr-gene modified Jurkat T-cells with 1 µ M
ionomycin and 10 ng/ml PMA resulted in a 2.0 ± 0.1 - fold increase in relative mean EGFP expression measured
at ~48 hrs (P = 0.011). This activation-induced EGFP expression was abrogated when the cells were pretreated
for ~30 minutes with 300 nM CsA.