Epigenetic control of leukemic gene expression by menin and wildtype MLL1 complex members.

• Applicant: Professor Dr. Michael Kühn
• Subject Area: Hematology, Oncology
• Term: from 2017 to 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 385973529

In 2016 we described that the interaction between the histone methyltransferase MLL1 (MLL, KMT2A) and the adaptor protein menin (MEN1) is a dependency and provides a potential opportunity for the treatment of NPM1mut (NPM1mut) leukemia. We could also demonstrate that menin-MLL1 interaction inhibitors, which were originally developed to target the oncogenic MLL-fusion complexes in MLL1-rearranged (MLL-r) leukemias, reversed leukemogenic gene expression (Kühn et al. Cancer Discovery 2016). Since the start of the Emmy Noether project, more specific novel menin-MLL1 inhibitors with outstanding drug-like properties have been developed and various groups independently confirmed our initial results. The topic became even more clinically relevant as encouraging reports from first clinical trials provided evidence for clinical activity of these compounds in relapsed / refractory AML. During the first funding period of the Emmy Noether Program, we have now provided evidence that menin-MLL1 inhibitors also target activating FLT3 mutations transcriptionally via MEIS1 in NPM1mut and MLL-r leukemias. Our data show that combined menin-MLL1 and FLT3 inhibition is a synergistic therapeutic opportunity in these leukemia subtypes with concurrent FLT3 mutations (Dzama et al. Blood 2020). These data are clinically relevant as activating FLT3 mutations are common AML drivers, particularly in the NPM1mut subset and this drug combination is already available for clinical testing. Other experiments were aimed at the question of how the menin-MLL1 complex specifically modifies chromatin to control leukemogenic gene expression. Based on the hypothesis that the menin-MLL1 complex drives leukemogenesis by recruiting interacting proteins to specific chromatin loci, we performed a high-definition CRISPR-domain scan of MEN1 and MLL1. Interrogating the coding sequence of MEN1 and MLL1 every eight nucleotides on average, we identified specific sequences encoding binding sites for interacting proteins as novel dependencies. Direct genetic deletion of these menin-MLL1 interacting proteins confirmed 3 of them as being required for NPM1mut AML. These 3 included two histone modifiers of which 1 is MOF, an H4K16 acetyltransferase, known to co-localize with MLL1 at its target loci. Characterization of the 3 interacting proteins in NPM1mut AML models is ongoing. This includes the assessment of chromatin binding and potential co-localization with menin and MLL1, the definition of transcriptional and binding targets, functional studies upon genetic deletion, and in vivo assessment of pharmacological inhibition in PDX models. Also, we developed a menin-MLL1-i resistant model of NPM1mut AML that we are characterizing. These ongoing studies within the 2nd and (extended) 3rd funding period will further delineate the molecular mechanisms of how the menin-MLL1 complex drives leukemogenesis and may define novel therapeutic targets for NPM1mut leukemia.

• DFG Programme: Independent Junior Research Groups