The MERTK/FLT3 inhibitor MRX-2843 overcomes resistance-conferring FLT3 mutations in acute myeloid leukemia
Katherine A. Minson, Catherine C. Smith, Deborah DeRyckere, Clara Libbrecht, Alisa B. Lee-Sherick, Madeline G. Huey, Elisabeth A. Lasater, Gregory D. Kirkpatrick, Michael A. Stashko, Weihe Zhang, Craig T. Jordan, Dmitri Kireev, Xiaodong Wang, Stephen V. Frye, H. Shelton Earp, Neil P. Shah, Douglas K. Graham
Katherine A. Minson, Catherine C. Smith, Deborah DeRyckere, Clara Libbrecht, Alisa B. Lee-Sherick, Madeline G. Huey, Elisabeth A. Lasater, Gregory D. Kirkpatrick, Michael A. Stashko, Weihe Zhang, Craig T. Jordan, Dmitri Kireev, Xiaodong Wang, Stephen V. Frye, H. Shelton Earp, Neil P. Shah, Douglas K. Graham
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Research Article Oncology

The MERTK/FLT3 inhibitor MRX-2843 overcomes resistance-conferring FLT3 mutations in acute myeloid leukemia

Abstract

FMS-like tyrosine kinase 3–targeted (FLT3-targeted) therapies have shown initial promise for the treatment of acute myeloid leukemia (AML) expressing FLT3-activating mutations; however, resistance emerges rapidly. Furthermore, limited options exist for the treatment of FLT3-independent AML, demonstrating the need for novel therapies that reduce toxicity and improve survival. MERTK receptor tyrosine kinase is overexpressed in 80% to 90% of AMLs and contributes to leukemogenesis. Here, we describe MRX-2843, a type 1 small-molecule tyrosine kinase inhibitor that abrogates activation of both MERTK and FLT3 and their downstream effectors. MRX-2843 treatment induces apoptosis and inhibits colony formation in AML cell lines and primary patient samples expressing MERTK and/or FLT3-ITD, with a wide therapeutic window compared with that of normal human cord blood cells. In murine orthotopic xenograft models, once-daily oral therapy prolonged survival 2- to 3-fold over that of vehicle-treated controls. Additionally, MRX-2843 retained activity against quizartinib-resistant FLT3-ITD–mutant proteins with clinically relevant alterations at the D835 or F691 loci and prolonged survival in xenograft models of quizartinib-resistant AML. Together, these observations validate MRX-2843 as a translational agent and support its clinical development for the treatment of AML.

Authors

Katherine A. Minson, Catherine C. Smith, Deborah DeRyckere, Clara Libbrecht, Alisa B. Lee-Sherick, Madeline G. Huey, Elisabeth A. Lasater, Gregory D. Kirkpatrick, Michael A. Stashko, Weihe Zhang, Craig T. Jordan, Dmitri Kireev, Xiaodong Wang, Stephen V. Frye, H. Shelton Earp, Neil P. Shah, Douglas K. Graham

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Figure 2

MRX-2843 inhibits MERTK activation and downstream signaling and has functional antitumor effects in MERTK+ FLT3-WT cell culture and animal models.

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MRX-2843 inhibits MERTK activation and downstream signaling and has func...
(A–D) Kasumi-1 and/or NOMO-1 AML cells were treated with the indicated concentrations of MRX-2843, vehicle (DMSO), or a nontargeting control TKI (Ctrl TKI). (A) After 1 hour, Kasumi-1 cells were treated with pervanadate phosphatase inhibitor for 10 minutes to stabilize p-MERTK, and cell lysates were prepared. MERTK protein was immunoprecipitated, and p-MERTK and total MERTK were detected by immunoblot analysis. (B) After 2 hours, lysates were prepared from Kasumi-1 cells without pervanadate treatment, and phosphorylated and total STAT6, AKT, and ERK1/2 proteins were detected by immunoblotting. Actin was used as a loading control. (C) After 48 hours, MTS reagent was added to cultures for an additional 2 hours, and absorbance was determined as an indicator of the number of viable cells. (D) After 72 hours, cells were stained with Yo-Pro-1 iodide and propidium iodide, and apoptotic and dead cells were detected by flow cytometry. ***P < 0.001, by 1-way ANOVA. (E) Kasumi-1 or NOMO-1 cells were cultured in soft agar overlaid with media containing the indicated concentrations of MRX-2843, vehicle (DMSO), or control TKI. Colonies were stained and counted after 14 to 21 days. Results are shown relative to vehicle-treated controls. Mean values and standard errors were derived from 3 independent experiments. **P < 0.01 and ***P < 0.001, by 1-way ANOVA. (F) Kaplan-Meier curve showing survival of NSGS mice with orthotopic NOMO-1 xenografts that were treated with MRX-2843 or vehicle (saline) beginning on day 21 after transplantation. Median survival was 51 days for MRX-2843–treated mice and 37 days for vehicle-treated control mice (n ≥10 per group). P < 0.001, by log-rank test. AML, acute myeloid leukemia; Ctrl TKI, control tyrosine kinase inhibitor; FLT3, FMS-like tyrosine kinase 3; NSGS, NOD-SCID-γ mice expressing Tg human cytokines; p, phosphorylated.