flt3 inhibitor  (Janssen)

Journal: bioRxiv

Article Title: Sphingosine-1-phosphate receptor modulators resensitize FLT3-ITD acute myeloid leukemia cells with NRAS mutations to FLT3 inhibitors

doi: 10.1101/2025.11.21.689510

Figure Lengend Snippet: A. M14(R)701 cells were treated with the FLT3 inhibitors gilteritinib and quizartinib at increasing concentrations alone and in combination with the S1PR modulator FTY720 (2.5 μM) for 48 hours. Cytotoxicity was measured with the WST1 assay. B. M14(R)701 cells seeded at 10,000 cells/well in 96-well plates were treated for 48 hours with gilteritinib or quizartinib and/or FTY720 as single drugs and in combinations at the concentrations shown, in triplicate. Cytotoxicity was measured with the WST-1 assay, and drug combination effects by the Chou-Talalay method. Synergy was defined by combination index values < 1.0. C. M14(R)701 were treated with gilteritinib (10 nM), quizartinib (1 nM), FTY720 (2.5 μM) or KRP-203 (5 μM) as single drugs and in the combinations shown, in triplicate experiments. Apoptosis was analyzed by Annexin V staining, measured by flow cytometry. *p=0.01; **p=0.008; ****p<.0001. D. NRG mice injected intravenously with M14(R)701-luc cells were treated with gilteritinib 7.5 mg/kg and/or FTY720 10 mg/kg, or vehicle control, starting on Day 1; serial non-invasive luciferin imaging of mice is shown. E. Changes in photon intensity, measured by bioluminescence imaging, over time, with p =0.015, comparing gilteritinib and FTY720 combination versus gilteritinib alone by 2-way ANOVA with Sidak’s multiple comparison test. F. Survival curves, with p =0.0035 comparing gilteritinib and FTY720 combination versus gilteritinib alone by Kaplan–Meier analysis.

Article Snippet: FLT3 inhibitors included lestaurtinib (CEP-701; HY-50867, MedChem Express, Monmouth Junction, NJ) and gilteritinib (ASP 2215; S7754) (Type I), and quizartinib (AC220, S1526) (Type II) (Selleck Chemicals, Houston, TX).

Techniques: WST-1 Assay, Staining, Flow Cytometry, Injection, Control, Imaging, Comparison

Journal: bioRxiv

Article Title: Sphingosine-1-phosphate receptor modulators resensitize FLT3-ITD acute myeloid leukemia cells with NRAS mutations to FLT3 inhibitors

doi: 10.1101/2025.11.21.689510

Figure Lengend Snippet: A. M14(R)701 cells were treated with the FLT3 inhibitors gilteritinib or quizartinib at increasing concentrations as single drugs and in combination with the PP2A-activating drug (PAD) DT-061 (12 μM) for 48 hours. Cytotoxicity was measured with the WST1 assay. B. M14(R)701 cells seeded at 10,000 cells/well in 96-well plates were treated for 48 hours with gilteritinib or quizartinib and/or DT-061 as single drugs and in combinations at diverse concentrations, in triplicate. Cytotoxicity was measured with the WST-1 assay, and drug combination effects were analyzed by the Chou-Talalay method. C. MOLM-14 and M14(R)701 cells were treated with the FLT3 inhibitors gilteritinib (10 nM) or quizartinib (1 nM) and/or the PAD DT-061 (12 μM) or DMSO control. Apoptosis was measured by Annexin V staining. D. PP2A phosphatase activity was measured in MOLM-14 and M14(R)701 cells treated with the PAD DT-061 (12 μM), the PAD and S1PR1 modulator FTY720 (2.5 μM) or the S1PR1 modulator KRP-203 (5 μM) and in M14(R)701 cells treated with the FLT3 inhibitors gilteritinib (10 nM) or quizartinib (1 nM) with or without DT-061, FTY720 or KRP-203 at the concentrations above. ****p<0.0001. Results are mean of two independent experiments.

Article Snippet: FLT3 inhibitors included lestaurtinib (CEP-701; HY-50867, MedChem Express, Monmouth Junction, NJ) and gilteritinib (ASP 2215; S7754) (Type I), and quizartinib (AC220, S1526) (Type II) (Selleck Chemicals, Houston, TX).

Techniques: WST-1 Assay, Control, Staining, Activity Assay

Journal: bioRxiv

Article Title: Sphingosine-1-phosphate receptor modulators resensitize FLT3-ITD acute myeloid leukemia cells with NRAS mutations to FLT3 inhibitors

doi: 10.1101/2025.11.21.689510

Figure Lengend Snippet: A. Blasts from Patient 1, with FLT3-ITD and an NRAS G13D mutation treated with 1nM quizartinib and/or 2.5µM FTY720, or DMSO control, were harvested for immunoblotting at serial time points and SPHK1, p-AKT/AKT, p-p70 S6K/p70 S6K and p-BAD/BAD protein expression was measured. B . Graphic representation of data in A is shown. SPHK1, p-AKT, p-p70 S6K and p-BAD were downregulated by combination treatment, but not by single-drug treatments. C. Blasts from Patient 1 were incubated for 48 hours with 1nM quizartinib and/or 2.5µM FTY720, or DMSO control, and cytotoxicity was measured using the WST-1 assay. Combination treatment produced markedly greater cytotoxicity, compared to single drugs. D. Blasts from Patient 2, with FLT3-ITD and an NRAS G13V mutation, were treated with 1nM quizartinib and/or 2.5µM FTY720, or DMSO control, for 6 hours, and cells were harvested for immunoblotting. E. Graphic representation of the data in D is shown. SPHK1, p-AKT/AKT, p-p70 S6K/p70 S6K and p-BAD/BAD were downregulated by combination treatment, but not by single-drug treatments.

Article Snippet: FLT3 inhibitors included lestaurtinib (CEP-701; HY-50867, MedChem Express, Monmouth Junction, NJ) and gilteritinib (ASP 2215; S7754) (Type I), and quizartinib (AC220, S1526) (Type II) (Selleck Chemicals, Houston, TX).

Techniques: Mutagenesis, Control, Western Blot, Expressing, Incubation, WST-1 Assay, Produced

Journal: bioRxiv

Article Title: Sphingosine-1-phosphate receptor modulators resensitize FLT3-ITD acute myeloid leukemia cells with NRAS mutations to FLT3 inhibitors

doi: 10.1101/2025.11.21.689510

Figure Lengend Snippet:

Article Snippet: FLT3 inhibitors included lestaurtinib (CEP-701; HY-50867, MedChem Express, Monmouth Junction, NJ) and gilteritinib (ASP 2215; S7754) (Type I), and quizartinib (AC220, S1526) (Type II) (Selleck Chemicals, Houston, TX).

Techniques:

Journal: Clinical Cancer Research

Article Title: Synergistic Activity of Combined FLT3-ITD and MDM2 Inhibition with Quizartinib and Milademetan in FLT3 -ITD Mutant/ TP53 Wild-type Acute Myeloid Leukemias

doi: 10.1158/1078-0432.CCR-24-2764

Figure Lengend Snippet: Quizartinib plus milademetan exerts highly synergistic proapoptotic activity through suppression of FLT3 downstream signaling and induction of apoptotic signaling and PUMA in FLT3 -ITD mutant/ TP53 WT AML cell lines. A, MOLM-13, MOLM-14, and MV-4-11 cell lines (all of these cell lines bear FLT3 -ITD mutations and p53 WT) and ( B ) OCI/AML3 ( FLT3 WT/ TP53 WT) and Tohoku Hospital Pediatrics 1 (THP-1; FLT3 WT/ TP53 mutation) cell lines were treated with indicated concentrations of milademetan and/or quizartinib for 48 hours. Apoptosis induction was measured by FCM with annexin V staining and CI. All experiments were repeated ≥3 times, and data are presented as mean ± SD. CIs were calculated using CalcuSyn software (version 2.0, PREMIER Biosoft). C, MOLM-13, MOLM-14, and MV-4-11 cell lines were treated with milademetan (30 nmol/L) or quizartinib (2 nmol/L) or Q/M combination for 24 hours. Phospho (p)-FLT3 and its downstream signaling proteins and Bcl-2 family proteins were measured by immunoblotting. Bim, Bcl-2–interacting mediator; CI, confidence interval; GAPDH, glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: The small-molecule inhibitor of FLT3 quizartinib, human-specific MDM2 antagonist milademetan (DS-3032b), and murine-specific MDM2i (DS-5272) were provided by Daiichi Sankyo Co., Ltd.. Molecular structures are shown in Supplementary Figs. S1 and S2.

Techniques: Activity Assay, Mutagenesis, Staining, Software, Western Blot

Journal: Clinical Cancer Research

Article Title: Synergistic Activity of Combined FLT3-ITD and MDM2 Inhibition with Quizartinib and Milademetan in FLT3 -ITD Mutant/ TP53 Wild-type Acute Myeloid Leukemias

doi: 10.1158/1078-0432.CCR-24-2764

Figure Lengend Snippet: Combination of Q/M-induced proapoptotic activity is p53-dependent, and the combination treatment impairs clonogenicity in FLT3 mutant/ TP53 WT leukemia blasts. A, MOLM-13-p53-Vec or MOLM-13-p53-KD cells were treated with increasing concentrations of single-agent milademetan or quizartinib or Q/M combination for 48 hours. Apoptosis was measured by FCM with annexin V staining. CIs were calculated using CalcuSyn software (version 2.0, PREMIER Biosoft). B, MOLM-13-p53-Vec or MOLM-13-p53-KD cells were treated with MOLM-13 and milademetan (60 nmol/L) or quizartinib (3 nmol/L) or the combination for 24 hours. Correlated signaling pathway proteins were measured with immunoblotting. C, Primary AML samples (three cases; harboring FLT3 mutant/ TP53 WT) and two normal BM samples from normal donors were treated with indicated concentrations of quizartinib and/or milademetan for 2 weeks in methylcellulose medium. CFU-GM colonies were counted, and the percentage of colony forming against the control group was calculated. The data were obtained from triplicated wells, and error bars are presented as mean ± SD. Asterisks indicate the level of statistical significance. *, P < 0.05; **, P < 0.01; ***, P < 0.001 as determined using a two-tailed unpaired t test. BAX, Bcl-2–associated X; CFU-GM, colony-forming unit granulocyte–macrophage; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; NS, no statistical significance.

Article Snippet: The small-molecule inhibitor of FLT3 quizartinib, human-specific MDM2 antagonist milademetan (DS-3032b), and murine-specific MDM2i (DS-5272) were provided by Daiichi Sankyo Co., Ltd.. Molecular structures are shown in Supplementary Figs. S1 and S2.

Techniques: Activity Assay, Mutagenesis, Staining, Software, Western Blot, Control, Two Tailed Test

Journal: Clinical Cancer Research

Article Title: Synergistic Activity of Combined FLT3-ITD and MDM2 Inhibition with Quizartinib and Milademetan in FLT3 -ITD Mutant/ TP53 Wild-type Acute Myeloid Leukemias

doi: 10.1158/1078-0432.CCR-24-2764

Figure Lengend Snippet: Co-targeting FLT3 and MDM2 synergistically induces apoptosis in quizartinib-sensitive and -resistant FLT3 - ITD and TKD double-mutant AML cells and in venetoclax-resistant cells. A, Murine FLT3 -ITD/ TP53 WT cells Ba/F3- FLT3 -ITD, Ba/F3- FLT3 -ITD + D835Y, and Ba/F3- FLT3 -ITD + F691L were treated with increasing concentrations of quizartinib and/or DS-5272 (murine MDM2i) for 48 hours. Apoptosis induction was measured by FCM with annexin V staining. CIs were calculated using CalcuSyn software (version 2.0, PREMIER Biosoft). B, MOLM-13 parental and venetoclax-resistant (MOLM-13-VEN-Resis) cells were treated with venetoclax (Bcl-2i) for 48 hours. Viable cells were measured using the CellTiter-Glo luminescent cell viability assay (Promega). C, MOLM-13-VEN-Resis cells were treated with quizartinib and/or milademetan for 48 hours. Apoptosis was determined by FCM with Annexin V staining. D, MOLM-13-VEN-Resis cells were treated with indicated concentrations of milademetan and/or quizartinib for 24 hours. Correlated proteins were determined using immunoblotting. Asterisks indicate the level of statistical significance. **, P < 0.01; ***, P < 0.001. BAX, Bcl-2–associated X; Bim, Bcl-2–interacting mediator; CI, confidence interval; GAPDH, glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: The small-molecule inhibitor of FLT3 quizartinib, human-specific MDM2 antagonist milademetan (DS-3032b), and murine-specific MDM2i (DS-5272) were provided by Daiichi Sankyo Co., Ltd.. Molecular structures are shown in Supplementary Figs. S1 and S2.

Techniques: Mutagenesis, Staining, Software, Cell Viability Assay, Western Blot

Journal: Clinical Cancer Research

Article Title: Synergistic Activity of Combined FLT3-ITD and MDM2 Inhibition with Quizartinib and Milademetan in FLT3 -ITD Mutant/ TP53 Wild-type Acute Myeloid Leukemias

doi: 10.1158/1078-0432.CCR-24-2764

Figure Lengend Snippet: Quizartinib plus milademetan significantly extended survival in a PDX model of treatment-resistant AML ( FLT3 -ITD positive + D835 mutant/ TP53 WT). Prevalence of xenografted cells in ( A ) PB and ( B ) BM was measured using FCM by gating with the mCD45 – /hCD45 + population. C, Spleen mass was determined using a digital balance after dissection from three mice per group. D, The survival curve for xenografted mice was analyzed using the Kaplan–Meier method, and log-rank statistics were used to assess survival differences between the groups. Asterisks indicate the level of statistical significance. *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001. NS, no statistical significance.

Article Snippet: The small-molecule inhibitor of FLT3 quizartinib, human-specific MDM2 antagonist milademetan (DS-3032b), and murine-specific MDM2i (DS-5272) were provided by Daiichi Sankyo Co., Ltd.. Molecular structures are shown in Supplementary Figs. S1 and S2.

Techniques: Mutagenesis, Dissection

Journal: Journal of advanced research

Article Title: Tubular epithelial cell-derived Flt3L is required for type 1 conventional dendritic cell (cDC1) activation and expansion in promoting the recovery in acute kidney injury.

doi: 10.1016/j.jare.2025.02.036

Figure Lengend Snippet: Fig. 3. Effect of Flt3 inhibitor gilteritinib on kidney damage in WT mice with AKI. (a) Experimental design. As illustrated, WT mice were pre-treated with PBS or 5 % DMSO or 10 mg/kg gilteritinib for fourteen consecutive days prior to Sham surgery or IR surgery and administrated daily after surgery. (b) The phosphorylation expression levels of Flt3 and Erk in kidneys on day 7 after Sham or IR surgery. n = 5 per group. (c-d) BUN and CREA levels. n = 3–5 per group. (e) Representative macroscopic images of kidney atrophy. n = 5 per group. (f) Representative images of kidney sections from PBS-, DMSO-, and gilteritinib-treated mice on day 7 after AKI, stained with H&E, PAS, LTL and UMOD. Semi- quantitative morphometry of tubular injury is presented as PAS scores under HPF (40X). Quantitative assessment of LTL- and UMOD-positive cells under HPF (40X) is indicated by IOD per area. n = 5 per group. Scale bars: 200 lm. Data are shown as mean ± SD and analyzed using one-way or two-way ANOVA.

Article Snippet: WT mice received i.p. injections of either the FLT3 inhibitor gilteritinib (10 mg/kg in 5 % DMSO [31], sourced from Selleck, China), a control solution of 5 % DMSO, or PBS as control.

Techniques: Phospho-proteomics, Expressing, Staining