Synergistic targeting of KMT2A-rearranged AML with combined LSD1 and menin inhibitors Free

KMT2A-rearranged (KMT2Ar) acute myeloid leukemia (AML) is a high-risk subtype of AML associated with poor prognosis and frequent relapse. Standard therapies often fail to achieve durable remission, underscoring the urgent need for novel treatments. Menin inhibitors have emerged as promising therapeutics in relapsed/refractory (R/R) KMT2Ar AML with revumenib approval in November 2024, marking it the first-in-class menin inhibitor approved to date. However, the response duration in the AUGMENT-101 trial averaged just over 6 months. Here, we hypothesized that combining menin inhibitors with other epigenetic-modifying drugs could lead to synergistic combinations that would spark a more durable response by promoting stable differentiation and disrupting leukemic transcriptional networks, thereby improving therapeutic outcomes in KMT2Ar AML.To test our hypothesis, we conducted a high-throughput screen (HTS) consisting of 932 epigenetic modulators to identify compounds that synergize with menin inhibitors in KMT2Ar AML cell lines. Our epigenetic screen validated previously published menin inhibitor synergy with known targets such as BRD4 and KAT6A. Notably, all seven lysine specific demethylase (LSD1) inhibitors tested, including iadademstat (ORY-1001), enhanced cell death when combined with menin inhibition. In MV4-11 (MLL-AF4) and MOLM-13 (MLL-AF9) cells, combination therapy reduced viability with a synergistic IC50 compared to single-agent IC50s for revumenib and iadademstat alone.RNA-seq and ATAC-seq were conducted to assess transcriptomic alterations and changes in chromatin accessibility in MOLM-13 and MV4-11 cells following treatment with each drug individually and in combination. Chromatin profiling by CUT&RUN assessed co-occupancy of LSD1 and menin-MLL components. Co-immunoprecipitation and mass spectrometry defined LSD1's interactome, including its association with PSIP1 (LEDGF), a component of the MLL interactome. Functional effects on apoptosis and differentiation suggested that the combination induced greater apoptosis and more pronounced differentiation than either agent alone. RNA-seq analysis of AML cells revealed that the combination therapy induced higher gene expression changes compared to each drug alone, particularly in differentiation-associated genes such as CD11B and CD86. This was accompanied by increased chromatin accessibility at relevant loci.To evaluate in vivo efficacy and toxicity of the combination, we first optimized a dosing regimen and then treated NSG mice engrafted with a patient-derived xenograft (PDX) harboring the MLL-AF6 fusion (CPDM-0984X) randomized to treatment with vehicle, revumenib (40mg/kg twice daily for 5 out of 7 days), iadademstat (0.02 mg/kg M/W/F), or the combination (all 6 mice/group). Notably, after 3 weeks of treatment, the combination therapy dramatically reduced human CD45+ leukemic burden, outperforming monotherapies. Importantly, despite an initial decline in platelets in all treatment groups at week one, combination treatment was well-tolerated with no evidence of hematologic toxicity. Additionally, mice body weight did not change by >20% except in animals that succumbed to disease. Preliminary survival analysis indicates a survival benefit with the combination therapy. Five out of 6 animals treated with the combination remain alive at 4 weeks, while only 2/6 iadamestat treated mice and no revumenib or vehicle mice remain alive.In conclusion, these findings provide strong preclinical evidence that dual inhibition of LSD1 and menin is a rational and potent strategy to provide more durable treatment responses in KMT2Ar AML. The demonstrated synergy disrupts core leukemic transcriptional programs, potentially driving irreversible differentiation, and significantly reduces leukemic burden both in vitro and in vivo. Mechanistically, LSD1 shares genomic binding sites with KMT2A, suggesting a cooperation with KMT2A fusions to maintain leukemia stemness and suppress differentiation. Combining LSD1 inhibition shows promise for a dual-targeted epigenetic strategy for translation into early-phase clinical trials, particularly for patients with R/R KMT2A-rearranged AML. Given the depth of response and low toxicity observed in vivo in our PDX, this combination and dosing strategy may be important for limiting the toxicities of menin inhibition, such as differentiation syndrome or the thrombocytopenia associated with LSD1 inhibition, when planning future clinical trials.