The gilteritinib/ruxolitinib combination effectively kills IgH-CRLF2-r ALL cells. Cells were treated with vehicle, gilteritinib (Gilt) (1 μM), ruxolitinib (Ruxo) (1 μM),^43,50,58,65 or both, and protein samples were obtained 2 hours after treatment. Patient samples were cultured in RPMI medium supplemented with 20% FBS. Western blots were performed using antibodies against indicated proteins. Three cell lines (A) and 4 primary samples (B), all of which harbor the IgH-CRLF2 rearrangements, were used. Mutation status of JAK2 and RAS is indicated below. *Pathogenic significance of this mutation is unclear (COSMIC ID: COSM303853). **NRAS G12S mutation was detected only in a minor fraction (VAF 0.07). (C) Cells were treated with gilteritinib (500 nM), ruxolitinib (500 nM), or both and cultured for 6 days. Viability of drug-treated relative to vehicle (DMSO)-treated cells is plotted. Each condition was assessed in triplicate, and data are represented as means ± SD. Cells were treated with ruxolitinib at 500 nM, which represented the IC50 when MUTZ5 cell viability was assessed after 6 days of culture with drug. (D) Proportions of apoptotic cells were assessed by Annexin V stain via FACS on days 3 and 6 after DMSO or drug treatment. Each condition was assessed in triplicate, and data are represented as means + SD. P values were calculated using one-way ANOVA with Dunnett’s posttest for multiple comparisons. *P < .05, **P < .0001.