Generation, expansion and phenotipic properties of T-bodies.

<p>(A) Set up of transduction protocol and growth kinetics. <i>Left panel</i>, PBMC were activated with OKT3 at time 0 and transduced concomitantly (Td0), or after 48-h (Td2) or 72-h (Td3) hours. Flow cytometry analysis of c-myc tag expression was performed 72 hours post infection. <i>Central panel</i> reports the expression of CD27, CD28 and CD62L in the three different conditions of activation/infection reported in the <i>left panel</i>. <i>Right panel,</i> Expansion of T-body-hPSMA/fluc and T-body-hPSMA/eGFP transduced after 48 h activation in response to weekly restimulation with PC3-PIP cells. Untransduced PBMC were used as control. (B) Phenotype and CAR expression in transduced T cells upon stimulation. <i>Left panel</i>, Expression of surface markers in T-body-anti-hPSMA/fluc at different time points (3, 11 and 18 days) post transduction, as assessed by flow cytometry. Data are representative of 3 independent experiments. Overlapping results were obtained in T-body-anti-hPSMA/eGFP. <i>Right panel,</i> percentage of c-myc⁺ cells in LV CAR hPSMA/fluc and LV CAR hPSMA/eGFP-transduced T cell populations at different time points post transduction. Figure shows mean +/− SD of at least three independent experiments. (C) Kinetics of CD4 and CD8 subsets in transduced T cell populations. Expression of CD4 and CD8 markers in c-myc⁺-gated T-body-hPSMA/eGFP <i>(left panel)</i> and T-body-hPSMA/fluc (<i>right panel)</i> populations at different time points post transduction, as assessed by flow cytometry. Data are from a representative experiment out of three that produced similar results.