In vitro, 19mz/IL-12–modified T cells, compared with 19mz⁺ T cells, exhibit enhanced cytotoxicity, resistance to Treg-mediated inhibition, a proinflammatory cytokine profile and enhanced CD25 expression. (A) 19mz/IL-12 and 19mz transduced T-cell populations were composed of equivalent CD4 and CD8 T cells, as detected by flow cytometry with the former exhibiting enhanced but not statistically significant increase in expression of memory T-cell markers (B). (C) 19mz/IL-12⁺ T cells have significantly higher expression of perforin and granzyme B compared with 19mz⁺ T cells. (D) As determined by standard ⁵¹Cr release assays 19mz/IL-12⁺ T cells have significantly increased ability to lyse EL4(hCD19) tumor cells compared with 19mz⁺ T cells. (E) 19mz/IL-12⁺ T cells, but not 19mz⁺ T cells retain capacity to lyse EL4(hCD19) tumor after preincubation with Tregs. (F) 19mz/IL-12⁺ T cells secrete increased levels of IFNγ, GM-CSF but fail to secrete IL-2 compared with 19mz⁺ T cells after coculture with EL4(hCD19) tumor cells at an E:T of 1:10 for 24 hours. (G) 19mz/IL-12⁺ T cells (thick lines) express higher levels of CD25 compared with 19mz⁺ T cells (thin lines), as detected by flow cytometry. Results are representative of at least 2 independent experiments.