Abstract 1227: Novel combination therapies to overcome non-genetic/adaptive menin inhibitor resistance in AML with MLL1r or mtNPM1
Authors: Fiskus Warren C. , Mill Christopher P. , Piel Jessica , Hentemann Murphy , Birdwell Christine E. , Das Kaberi , Davis John A. , Hou Hanxi , Kadia Tapan M. , Daver Naval , Loghavi Sanam , Cuglievan Branko , DiNArdo Courtney D. , Bhalla Kapil N. 
Published: March 2024

In AML with MLL1 rearrangement or MLL1 fusion proteins (FP), the N-terminus of MLL1 becomes fused to over 80 partner proteins resulting in an aberrant leukemic transcriptional program including dysregulated expression of HOXA9, MEIS1, PBX3, FLT3, MEF2C and CDK6. On the other hand, in mutant NPM1 AML cells, wild type MLL1 is the main regulator of HOXA9, MEIS1 and FLT3, promoting self-renewal of myeloid progenitor cells. Menin inhibitors have been developed that disrupt the binding of Menin to MLL1 leading to reduced activity of HOXA9 and MEIS1 and repression of MLL1 or MLL1-FP target genes. Clinical trials have demonstrated that Menin inhibitors (MI) are well-tolerated and clinical remissions have been observed in patients with relapsed/refractory AML harboring MLLr. Unfortunately, most patients either fail to respond or eventually relapse. MI-resistance in AML with MLL1r or mtNPM1 is either due to recently described mutations in the MI-binding domain of Menin or is non-genetic/adaptive, due to dysregulated epigenome/transcriptome/proteome. In the present studies, through repeated shocks with LD90 dosing of MI, followed by drug washout and recovery, we have generated MLL1r (MV4-11-MITR) and mtNPM1 (OCI-AML3-MITR) AML cell lines that are tolerant/resistant (TR) to MI with LD50 values greater than 1 μM. Whole exome sequencing confirmed that these cells were lacking in new driver mutations. We determined that resistance in the MITR cells was not due to presence of hotspot mutations in Menin. ChIP-Seq analysis of H3K27Ac showed that the activities of super enhancers and core transcriptional regulatory circuitry was altered in both MITR cell types. Non-genetic/adaptive resistance to MI also concordantly reduced genome wide ATAC-Seq and RNA-Seq peaks in the OCI-AML3-MITR cells with significant depletion of MEIS1, IGF2BP2, BCL2, and PBX3, with concomitant induction of leukemia stem cell associated CLEC12A and CD244 expressions. A domain-specific CRISPR screen in MV4-11-MITR cells identified several druggable co-dependencies (e.g. EP300 and MOZ) and co-enrichments (SMARCA4, CREBBP and BRD4) with MI, suggesting them as potential mechanisms of non-genetic/adaptive MI resistance. Consistent with these findings, co-treatment with SMARCA2/SMARCA4 (chromatin remodeler ATPases) inhibitor FHD-286 and the BETi OTX015 or the MI SNDX-50469 exerted synergistic lethality in MV4-11-MITR and OCI-AML3-MITR cells as well as in PD MITR AML cells with MLL1r or mtNPM1. Co-treatment with CBP/p300 inhibitor GNE049 or GNE781 was also synergistic with MI in these cells. In vivo treatment with FHD-286 and OTX015 or SNDX-5613 significantly reduced the AML burden in mice bearing OCI-AML3-MITR xenografts. These findings underscore preclinical activity of epigenetically-targeted agent-based combinations and highlight their promise in overcoming MI resistance in AML with MLL1r or mtNPM1.

Citation Format: Warren C. Fiskus, Christopher P. Mill, Jessica Piel, Murphy Hentemann, Christine E. Birdwell, Kaberi Das, John A. Davis, Hanxi Hou, Tapan M. Kadia, Naval Daver, Sanam Loghavi, Branko Cuglievan, Courtney D. DiNArdo, Kapil N. Bhalla. Novel combination therapies to overcome non-genetic/adaptive menin inhibitor resistance in AML with MLL1r or mtNPM1 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1227.