Abstract 3841: Therapeutic targeting of <i>FLT3</i> mutations in AML via menin-MLL1 and FLT3 inhibition
Authors: Margarita M. Dzama, Martha C. Taubert, Kerstin Kunz, Johanna Rausch, Chun-Wei Chen, Annalisa Mupo, Matthias Theobald, Thomas Kindler, Richard P. Koche, George S. Vassiliou, Scott A. Armstrong, Michael W. Kühn
Journal: Cancer Research (2019)

Abstract Acute myeloid leukemias (AML) that are driven by MLL1 (KMT2A)-fusion proteins (MLL-f) or NPM1 mutations (NPM1mut) are both associated with aberrant expression of HOX and MEIS1 transcription factors and commonly harbor mutations in the gene encoding the receptor tyrosine kinase FLT3. Inhibition of the menin-MLL1 interaction has been shown to be a therapeutic opportunity in MLL-f driven leukemias and we recently demonstrated that this interaction is a dependency in NPM1mut AML. MI503 a specific small molecule menin-MLL1 inhibitor reduces dramatically cell growth and reverses leukemogenic gene expression including MEIS1 and FLT3. To determine global transcriptional changes associated with menin inhibition we performed RNA sequencing upon MI503 treatment in NPM1mut OCI-AML3 and MLL-f driven MV411 cells. MEIS1 and its putative target gene FLT3 were found to be among the most significantly downregulated genes. MEIS1 and FLT3 were consistently downregulated in various human and murine leukemia cell lines driven by MLL-f or NPM1mut. Allele specific qPCR confirmed profound downregulation of the mutant FLT3 allele in the MLL-f driven MV411 and MOLM13 cells as well as murine Npm1mut/+Flt3ITD/+ cells that all harbor a FLT3-ITD mutation. FLT3 surface expression was also substantially reduced upon MI503 treatment as assessed by FACS. Next, we assessed combinatorial menin- and FLT3 tyrosine kinase inhibition using MI503 and the 2nd generation FLT3 inhibitor AC220. The two drugs worked in a synergistic way to promote growth inhibition and enhanced apoptosis compared to single drug treatment or vehicle alone in MOLM13 and MV411 cells. HL60 and NB4 AML cells lacking NPM1mut, MLL-f, or FLT3-ITD showed no response. Drug synergism was also observed in the murine NPM1mut/+FLT3ITD/+ AML cells when combining MI503 with ponatinib a tyrosine kinase inhibitor with activity against the FLT3-ITD F692L resistance mutation that has been described in these cells. Of interest, ectopic expression of Hoxa9-Meis1 resulted in upregulation of Flt3 and rescued the antiproliferative effect of combined menin- and FLT3-inhibition. Combined menin- and FLT3 inhibition reduced FLT3 phosphorylation more than AC220 or MI503 alone most likely reflecting the joint effect of AC220 mediated inhibition of FLT3 phosphorylation and transcriptional FLT3 suppression via MI503. Transcriptional profiling revealed substantial silencing of FLT3 downstream signature genes including MYC upon combinatorial treatment. In vivo treatment of leukemic MV411 xenografts with combined MI503 and AC220 resulted in significantly enhanced reduction of leukemia burden that was observed with single drug treatment compared to vehicle controls. Altogether, our data show that simultaneous menin- and FLT3 inhibition has a synergistic effect against leukemogenic FLT3 signaling and may represent a novel therapeutic concept for MLL-f and NPM1mut driven AML with concomitant FLT3-ITD. Citation Format: Margarita M. Dzama, Martha C. Taubert, Kerstin Kunz, Johanna Rausch, Chun-Wei Chen, Annalisa Mupo, Matthias Theobald, Thomas Kindler, Richard P. Koche, George S. Vassiliou, Scott A. Armstrong, Michael W. Kühn. Therapeutic targeting of FLT3 mutations in AML via menin-MLL1 and FLT3 inhibition [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3841.