Efficacy of STAT5 inhibition against LSCs from primary Lmo2^Tg T-ALL. (A) Relative levels of activated STAT5 (pSTAT5) in leukemic DN3a cells from primary Lmo2^Tg T-ALL treated with pimozide (1 μM), after in vitro stimulation of the IL-7 signaling pathway. Primary leukemias are indicated; unstimulated leukemic DN3a cells treated with vehicle were used as control. Mean ± SEM, 2-way ANOVA with Bonferroni correction test; ∗∗∗P < .001 as compared with vehicle. (B) Relative viability (%) of primary Lmo2^Tg leukemic cells treated with increasing concentration of pimozide for 48 hours (left). Viability was normalized to vehicle-treated cells with experiments performed in duplicates or triplicates. Mutations of growth factor-induced signaling pathways and median LC₅₀ of pimozide for each primary Lmo2^Tg T-ALL (right). (C) Experimental schematic for testing the efficacy of pimozide, VXL, and combination therapy in sublethally irradiated Cd45.1⁺ recipients injected with Lmo2^Tg primary leukemias. (D-E) Absolute numbers of leukemic DN3a cells in the thymus, bone marrow, and spleen (D), as well as in the peripheral blood (E) of recipients, 24 hours after the last administration of pimozide, VXL, and combination therapy. Primary leukemias are indicated on the right. Mean ± SEM, 2-way ANOVA with Tukey correction test; ∗∗∗P < .001 compared with vehicle; ^#P < .05 and ^##P < .01 compared with VXL. (F) Kaplan-Meier curves of sublethally irradiated recipients injected with Lmo2^Tg primary T-ALL, treated with pimozide, VXL, or combination therapy. Log-rank (Mantel-Cox) test; ∗P < .05, ∗∗P < .01, and ∗∗∗P < .001 compared with vehicle; ^##P < .01 and ^###P < .001 compared with VXL. The period of administration is indicated in light gray.