Ex vivo and in vivo sensitivity to checkpoint inhibitors and decitabine. (A) Activity of checkpoint/TP53 pathway inhibitors (n = 17) in human cell lines of AEL and NUP98-rearranged AML and in primary cells from the established mouse models of AEL, AML, and B/T-ALL. Cells were treated with increasing concentrations of the indicated compounds. Cell viability was determined after drug incubation by using CellTiter-Glo (Promega). Heatmap indicates area under the curve (AUC) values for each compound per leukemia sample. The positive control drugs (bortezomib and panobinostat) are in bold. (B) Ex vivo sensitivity to decitabine and talazoparib as single agents or in combination in AEL primary cells from #4491 (upper panel) and serial passages of #4497 (lower panel) cells. Data are reported as percent control (dimethyl sulfoxide) and represented as mean ± standard deviation of 3 replicates. The half-maximal inhibitory concentration (values in square brackets) was evaluated by nonlinear regression analysis using GraphPad Prism (GraphPad Software). In vivo studies: AEL mouse models #4497 (C) and #4491 (D) were randomized to receive 0.5 mg/kg intravenous decitabine once daily on treatment days 1 to 5 every 28-day cycle; 0.1 mg/kg oral talazoparib days 1 to 5 and days 14 to 19 once daily; 0.1 mg/kg oral talazoparib days 1 to 5 and days 14 to 19 once daily plus 0.5 mg/kg intravenous decitabine [Dec + Tal] once daily days 1 to 5; and a talazoparib vehicle control group. Tumor burden was monitored by spleen size reduction and percentage of GFP⁺RFP⁺ in bone marrow (BM) and spleen samples (SPL) from moribund mice (#4497) (panel C) or by bioluminescent imaging (#4491) (panel D). In panel C, some mice were found dead (n = 3 in Dec + Tal; n = 1 in decitabine; n = 2 in talazoparib; and n = 2 in vehicle). From those, spleen was weighted when possible, but BM and SPL samples were not available for analysis by flow cytometry. (E) Kaplan-Meier analysis of animal survival. Survival comparison was analyzed by log-rank test. Adj., adjusted; Anova, analysis of variance; Checkp. i., checkpoint inhibitors; GFP, green fluorescent protein; RFP, red fluorescent protein.