Effects of single and combined menin-MLL and FLT3 inhibition on primary NPM1^mut FLT3^ITD AML patient samples and on survival of in vivo–treated MLL-r FLT3-ITD leukemic xenograft mice. (A) The human stromal cell coculture assay, performed to maintain and treat patients’ primary AML blasts. (B) Summary of characteristics of patients providing the samples used in panels C-E. (C-D) Number of viable cells inde novoAML samples treated in coculture for 7 days with DMSO, MI-503 (2 µM), quizartinib (6 nM), or combinatorial MI-503 and quizartinib treatment. (C) Five independent samples of de novo NPM1^mutFLT3^ITD AML. (D) Two independent samples of de novo AML, WT for NPM1, FLT3, and MLL. Depicted are 4′,6-diamidino-2-phenylindole [DAPI]⁻, human CD45⁺ cell numbers as assessed by flow cytometry. (E) Effect of MI-503 (2.5 µM), quizartinib (3 nM), and combinatorial treatment (2.5 µM and 3 nM) on total and blast-like CFUs in primary patient sample cells. (F) Experimental setup for the treatment of MV411-derived leukemic xenograft mice (left); percentage of human CD45⁺ cells in the bone marrow of leukemic mice (right) after treatment with drug vehicles, MI-503 (50 mg/kg; twice daily IP), quizartinib (10 mg/kg; PO; once daily), or combined MI-503 and quizartinib. (G) Kaplan-Meier survival analysis of MV411-derived leukemic xenograft mice treated with drug vehicles, MI-503 (50 mg/kg; twice daily IP), quizartinib (10 mg/kg; PO; once daily), or combinatorial MI-503 and quizartinib (n = 5 mice/group). The treatment period is displayed in blue. The log-rank (Mantel-Cox) test was used to calculate the P-values.