Superior efficacy of combination STI treatment. Mice engrafted with (A) CRLF2/JAK-mutant Ph-like ALL were treated with vehicle, ruxolitinib, gedatolisib, or both ruxolitinib and gedatolisib for 3 or 4 weeks. Human ALL in murine spleens after treatment completion was measured by quantitative flow cytometry as in Figure 1. Significantly greater inhibition of ALL proliferation was observed with combined ruxolitinib and gedatolisib treatment (orange asterisks), as measured by 1-way ANOVA with the Tukey posttest for multiple comparisons with α = 0.05. (B) ABL1-rearranged Ph-like ALL PDX models were treated with vehicle, dasatinib, gedatolisib, or both dasatinib and gedatolisib for 3 weeks. Enhanced antileukemia efficacy was also observed in these models with combined dasatinib and gedatolisib treatment (orange asterisks) vs dasatinib alone or gedatolisib alone. (C) Combination inhibitor treatment markedly reduced splenomegaly in ALL-engrafted PDX mice treated as in panels A or B. (D) Immunoblotting of total and phosphorylated signal transduction proteins from murine splenic lysates (obtained after 3 or 4 weeks of treatment) demonstrate greatest inhibition of target phosphoproteins with combination inhibitor treatment of PDX models. Total protein loss is observed with inhibitor treatment in some models. MUTZ5 (a CRLF2/JAK2-mutant ALL cell line) and K562 (a BCR-ABL1–rearranged chronic myeloid leukemia cell line) lysates were used as positive signaling controls, and β-actin immunoblotting was used as a protein loading control. *P < .05, **P < .01, ***P < .001, ****P < .0001. D, dasatinib; DG, dasatinib + gedatolisib combination; G, gedatolisib; R, ruxolitinib; RG, ruxolitinib + gedatolisib combination; V, vehicle.