Structure-Based Development of Novel Spiro-Piperidine ASH1L Inhibitors.

The absent, small, or homeotic-like 1 (ASH1L) protein is a histone lysine methyltransferase that plays a crucial role in various cancers, including leukemia. Despite representing an attractive therapeutic target, only one class of ASH1L inhibitors was identified to date. Herein, we report development of advanced ASH1L inhibitors targeting the catalytic SET domain, which were designed to access previously unexplored binding pocket on ASH1L.

Combination of menin and kinase inhibitors as an effective treatment for leukemia with NUP98 translocations.

Chromosomal translocations of the nucleoporin 98 (NUP98) gene are found in acute myeloid leukemia (AML) patients leading to very poor outcomes. The oncogenic activity of NUP98 fusion proteins is dependent on the interaction between Mixed Lineage Leukemia 1 and menin. NUP98-rearranged (NUP98-r) leukemia cells also rely on specific kinases, including CDK6 and/or FLT3, suggesting that simultaneous targeting of these kinases and menin could overcome limited sensitivity to single agents.

Small-molecule inhibitors targeting Polycomb repressive complex 1 RING domain.

Polycomb repressive complex 1 (PRC1) is an essential chromatin-modifying complex that monoubiquitinates histone H2A and is involved in maintaining the repressed chromatin state. Emerging evidence suggests PRC1 activity in various cancers, rationalizing the need for small-molecule inhibitors with well-defined mechanisms of action. Here, we describe the development of compounds that directly bind to RING1B-BMI1, the heterodimeric complex constituting the E3 ligase activity of PRC1.

Discovery of first-in-class inhibitors of ASH1L histone methyltransferase with anti-leukemic activity.

ASH1L histone methyltransferase plays a crucial role in the pathogenesis of different diseases, including acute leukemia. While ASH1L represents an attractive drug target, developing ASH1L inhibitors is challenging, as the catalytic SET domain adapts an inactive conformation with autoinhibitory loop blocking the access to the active site. Here, by applying fragment-based screening followed by medicinal chemistry and a structure-based design, we developed first-in-class small molecule inhibitors of the ASH1L SET domain.

EWS-FLI1 and Menin Converge to Regulate ATF4 Activity in Ewing Sarcoma.

Ewing sarcomas are driven by EWS-ETS fusions, most commonly EWS-FLI1, which promotes widespread metabolic reprogramming, including activation of serine biosynthesis. We previously reported that serine biosynthesis is also activated in Ewing sarcoma by the scaffolding protein menin through as yet undefined mechanisms. Here, we investigated whether EWS-FLI1 and/or menin orchestrate serine biosynthesis via modulation of ATF4, a stress-response gene that acts as a master transcriptional regulator of serine biosynthesis in other tumors.

Covalent and noncovalent constraints yield a figure eight-like conformation of a peptide inhibiting the menin-MLL interaction.

The interaction between menin and mixed lineage leukemia (MLL) was identified as an interesting target for treating some cancers including acute leukemia. On the basis of the known crystal structure of the MBM1-menin complex (MBM - menin binding motif), several cyclic peptides were designed. Elaboration of the effective cyclization strategy using a metathesis reaction allowed for a successfully large number of derivatives to be obtained.

Covalent inhibition of NSD1 histone methyltransferase.

The nuclear receptor-binding SET domain (NSD) family of histone methyltransferases is associated with various malignancies, including aggressive acute leukemia with NUP98-NSD1 translocation. While NSD proteins represent attractive drug targets, their catalytic SET domains exist in autoinhibited conformation, presenting notable challenges for inhibitor development. Here, we employed a fragment-based screening strategy followed by chemical optimization, which resulted in the development of the first-in-class irreversible small-molecule inhibitors of the nuclear receptor-binding SET domain protein 1 (NSD1) SET domain.

Menin inhibitor MI-3454 induces remission in MLL1-rearranged and NPM1-mutated models of leukemia.

The protein-protein interaction between menin and mixed lineage leukemia 1 (MLL1) plays a critical role in acute leukemias with translocations of the MLL1 gene or with mutations in the nucleophosmin 1 (NPM1) gene. As a step toward clinical translation of menin-MLL1 inhibitors, we report development of MI-3454, a highly potent and orally bioavailable inhibitor of the menin-MLL1 interaction. MI-3454 profoundly inhibited proliferation and induced differentiation in acute leukemia cells and primary patient samples with MLL1 translocations or NPM1 mutations.

Complexity of Blocking Bivalent Protein-Protein Interactions: Development of a Highly Potent Inhibitor of the Menin-Mixed-Lineage Leukemia Interaction.

The protein-protein interaction between menin and mixed-lineage leukemia 1 (MLL1) plays an important role in development of acute leukemia with translocations of the MLL1 gene and in solid tumors. Here, we report the development of a new generation of menin-MLL1 inhibitors identified by structure-based optimization of the thienopyrimidine class of compounds. This work resulted in compound 28 (MI-1481), which showed very potent inhibition of the menin-MLL1 interaction (IC = 3.

Theoretical models of inhibitory activity for inhibitors of protein-protein interactions: targeting menin-mixed lineage leukemia with small molecules.

Development and binding affinity predictions of inhibitors targeting protein-protein interactions (PPI) still represent a major challenge in drug discovery efforts. This work reports application of a predictive non-empirical model of inhibitory activity for PPI inhibitors, exemplified here for small molecules targeting the menin-mixed lineage leukemia (MLL) interaction. Systematic analysis of menin-inhibitor complexes was performed, revealing the physical nature of these interactions.

Property Focused Structure-Based Optimization of Small Molecule Inhibitors of the Protein-Protein Interaction between Menin and Mixed Lineage Leukemia (MLL).

Development of potent small molecule inhibitors of protein-protein interactions with optimized druglike properties represents a challenging task in lead optimization process. Here, we report synthesis and structure-based optimization of new thienopyrimidine class of compounds, which block the protein-protein interaction between menin and MLL fusion proteins that plays an important role in acute leukemias with MLL translocations. We performed simultaneous optimization of both activity and druglike properties through systematic exploration of substituents introduced to the indole ring of lead compound 1 (MI-136) to identify compounds suitable for in vivo studies in mice.

Rational Design of Orthogonal Multipolar Interactions with Fluorine in Protein-Ligand Complexes.

Multipolar interactions involving fluorine and the protein backbone have been frequently observed in protein-ligand complexes. Such fluorine-backbone interactions may substantially contribute to the high affinity of small molecule inhibitors. Here we found that introduction of trifluoromethyl groups into two different sites in the thienopyrimidine class of menin-MLL inhibitors considerably improved their inhibitory activity.

Pharmacologic inhibition of the Menin-MLL interaction blocks progression of MLL leukemia in vivo.

Chromosomal translocations affecting mixed lineage leukemia gene (MLL) result in acute leukemias resistant to therapy. The leukemogenic activity of MLL fusion proteins is dependent on their interaction with menin, providing basis for therapeutic intervention. Here we report the development of highly potent and orally bioavailable small-molecule inhibitors of the menin-MLL interaction, MI-463 and MI-503, and show their profound effects in MLL leukemia cells and substantial survival benefit in mouse models of MLL leukemia.

The same site on the integrase-binding domain of lens epithelium-derived growth factor is a therapeutic target for MLL leukemia and HIV.

Lens epithelium-derived growth factor (LEDGF) is a chromatin-associated protein implicated in leukemia and HIV type 1 infection. LEDGF associates with mixed-lineage leukemia (MLL) fusion proteins and menin and is required for leukemic transformation. To better understand the molecular mechanism underlying the LEDGF integrase-binding domain (IBD) interaction with MLL fusion proteins in leukemia, we determined the solution structure of the MLL-IBD complex.

High-affinity small-molecule inhibitors of the menin-mixed lineage leukemia (MLL) interaction closely mimic a natural protein-protein interaction.

The protein-protein interaction (PPI) between menin and mixed lineage leukemia (MLL) plays a critical role in acute leukemias, and inhibition of this interaction represents a new potential therapeutic strategy for MLL leukemias. We report development of a novel class of small-molecule inhibitors of the menin-MLL interaction, the hydroxy- and aminomethylpiperidine compounds, which originated from HTS of ∼288000 small molecules. We determined menin-inhibitor co-crystal structures and found that these compounds closely mimic all key interactions of MLL with menin.

Structural insights into inhibition of the bivalent menin-MLL interaction by small molecules in leukemia.

Menin functions as a critical oncogenic cofactor of mixed lineage leukemia (MLL) fusion proteins in the development of acute leukemias, and inhibition of the menin interaction with MLL fusion proteins represents a very promising strategy to reverse their oncogenic activity. MLL interacts with menin in a bivalent mode involving 2 N-terminal fragments of MLL. In the present study, we reveal the first high-resolution crystal structure of human menin in complex with a small-molecule inhibitor of the menin-MLL interaction, MI-2.

Menin-MLL inhibitors reverse oncogenic activity of MLL fusion proteins in leukemia.

Translocations involving the mixed lineage leukemia (MLL) gene result in human acute leukemias with very poor prognosis. The leukemogenic activity of MLL fusion proteins is critically dependent on their direct interaction with menin, a product of the multiple endocrine neoplasia (MEN1) gene. Here we present what are to our knowledge the first small-molecule inhibitors of the menin-MLL fusion protein interaction that specifically bind menin with nanomolar affinities.

Collagen fragmentation promotes oxidative stress and elevates matrix metalloproteinase-1 in fibroblasts in aged human skin.

Aged human skin is fragile because of fragmentation and loss of type I collagen fibrils, which confer strength and resiliency. We report here that dermal fibroblasts express increased levels of collagen-degrading matrix metalloproteinases-1 (MMP-1) in aged (>80 years old) compared with young (21 to 30 years old) human skin in vivo. Transcription factor AP-1 and alpha2beta1 integrin, which are key regulators of MMP-1 expression, are also elevated in fibroblasts in aged human skin in vivo.