Suppression of NRF2-dependent cancer growth by a covalent allosteric molecular glue
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ABSTRACT:
PROVIDER: PRJNA1167342 | ENA |
REPOSITORIES: ENA
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Project description:The NRF2 transcription factor is constitutively active in cancer where it functions to maintain oxidative homeostasis and reprogram cellular metabolism. NRF2-active tumors exhibit NRF2-dependency and resistance to chemo/radiotherapy. Here we developed VVD-065, a first-in-class NRF2 inhibitor that acts via an unprecedented allosteric molecular glue mechanism. In the absence of stress or mutation, NRF2 is rapidly degraded by the KEAP1-CUL3 ubiquitin-ligase complex. VVD-065 specifically and covalently adducts C151 on KEAP1, which in turn promotes KEAP1-CUL3 complex formation, leading to dramatic enhancement of NRF2 degradation. Previously reported C151-directed compounds decrease KEAP1-CUL3 interactions and stabilize NRF2, thus establishing KEAP1_C151 as a tunable regulator of the KEAP1-CUL3 complex and NRF2 stability. VVD-065 inhibited NRF2-dependent tumor growth and sensitized cancers to chemo/radiotherapy, supporting an open Phase I clinical trial (NCT05954312).
2025-01-03 | GSE278482 | GEO
Project description:Discovery of a covalent molecular glue degrader that covalently interacts with UBE2D and engages in molecular glue interactions with NFKB1
2023-02-08 | PXD039924 | Pride
Project description:Small molecules that induce protein-protein interactions to exert proximity-driven pharmacology such as targeted protein degradation are a powerful class of therapeutics1-3. Molecular glues are of particular interest given their favorable size and chemical properties and represent the only clinically approved degrader drugs4-6. The discovery and development of molecular glues for novel targets, however, remains challenging. Covalent strategies could in principle facilitate molecular glue discovery by stabilizing the neo-protein interfaces. Here, we present structural and mechanistic studies that define a trans-labeling covalent molecular glue mechanism, which we term “template-assisted covalent modification”. We found that a novel series of BRD4 molecular glue degraders act by recruiting the CUL4DCAF16 ligase to the second bromodomain of BRD4 (BRD4BD2). BRD4BD2, in complex with DCAF16, serves as a structural template to facilitate covalent modification of DCAF16, which stabilizes the BRD4-degrader-DCAF16 ternary complex formation and facilitates BRD4 degradation. A 2.2 Å cryo-electron microscopy structure of the ternary complex demonstrates that DCAF16 and BRD4BD2 have pre-existing structural complementarity which optimally orients the reactive moiety of the degrader for DCAF16Cys58 covalent modification. Systematic mutagenesis of both DCAF16 and BRD4BD2 revealed that the loop conformation around BRD4His437, rather than specific side chains, is critical for BD2 selectivity. Supporting a general applicability, we find that a subset of compounds leads to a drug-induced GAK-BRD4 interaction stabilized by covalent modification of GAK. Together our work establishes “template-assisted covalent modification” as a mechanism for covalent molecular glues, which opens a new path to proximity driven pharmacology.
2024-08-09 | PXD047137 | Pride
Project description:Small molecules that induce protein-protein interactions to exert proximity-driven pharmacology such as targeted protein degradation are a powerful class of therapeutics1-3. Molecular glues are of particular interest given their favorable size and chemical properties and represent the only clinically approved degrader drugs4-6. The discovery and development of molecular glues for novel targets, however, remains challenging. Covalent strategies could in principle facilitate molecular glue discovery by stabilizing the neo-protein interfaces. Here, we present structural and mechanistic studies that define a trans-labeling covalent molecular glue mechanism, which we term “template-assisted covalent modification”. We found that a novel series of BRD4 molecular glue degraders act by recruiting the CUL4DCAF16 ligase to the second bromodomain of BRD4 (BRD4BD2). BRD4BD2, in complex with DCAF16, serves as a structural template to facilitate covalent modification of DCAF16, which stabilizes the BRD4-degrader-DCAF16 ternary complex formation and facilitates BRD4 degradation. A 2.2 Å cryo-electron microscopy structure of the ternary complex demonstrates that DCAF16 and BRD4BD2 have pre-existing structural complementarity which optimally orients the reactive moiety of the degrader for DCAF16Cys58 covalent modification. Systematic mutagenesis of both DCAF16 and BRD4BD2 revealed that the loop conformation around BRD4His437, rather than specific side chains, is critical for BD2 selectivity. Supporting a general applicability, we find that a subset of compounds leads to a drug-induced GAK-BRD4 interaction stabilized by covalent modification of GAK. Together our work establishes “template-assisted covalent modification” as a mechanism for covalent molecular glues, which opens a new path to proximity driven pharmacology.
2024-08-09 | PXD047138 | Pride
Project description:Small molecules that induce protein-protein interactions to exert proximity-driven pharmacology such as targeted protein degradation are a powerful class of therapeutics1-3. Molecular glues are of particular interest given their favorable size and chemical properties and represent the only clinically approved degrader drugs4-6. The discovery and development of molecular glues for novel targets, however, remains challenging. Covalent strategies could in principle facilitate molecular glue discovery by stabilizing the neo-protein interfaces. Here, we present structural and mechanistic studies that define a trans-labeling covalent molecular glue mechanism, which we term “template-assisted covalent modification”. We found that a novel series of BRD4 molecular glue degraders act by recruiting the CUL4DCAF16 ligase to the second bromodomain of BRD4 (BRD4BD2). BRD4BD2, in complex with DCAF16, serves as a structural template to facilitate covalent modification of DCAF16, which stabilizes the BRD4-degrader-DCAF16 ternary complex formation and facilitates BRD4 degradation. A 2.2 Å cryo-electron microscopy structure of the ternary complex demonstrates that DCAF16 and BRD4BD2 have pre-existing structural complementarity which optimally orients the reactive moiety of the degrader for DCAF16Cys58 covalent modification. Systematic mutagenesis of both DCAF16 and BRD4BD2 revealed that the loop conformation around BRD4His437, rather than specific side chains, is critical for BD2 selectivity. Supporting a general applicability, we find that a subset of compounds leads to a drug-induced GAK-BRD4 interaction stabilized by covalent modification of GAK. Together our work establishes “template-assisted covalent modification” as a mechanism for covalent molecular glues, which opens a new path to proximity driven pharmacology.
2024-08-09 | PXD047141 | Pride
Project description:The nuclear factor erythroid 2-related factor 2 (NRF2) transcription factor activates cytoprotective and metabolic gene expression in response to various electrophilic stressors. In disease, constitutive NRF2 activity promotes cancer progression while decreased NRF2 function contributes to neurodegenerative diseases. In contrast to the regulation of NRF2 protein stability in the cytoplasm, co-complexed proteins that govern NRF2 activity on chromatin are less clear. Using biotin proximity proteomics, we report networks for NRF2 and its family members NRF1, NRF3 and the NRF2 heterodimer MAFG. We found that the Parkinson’s disease zinc finger transcription factor ZNF746 (PARIS) physically associated with NRF2 and MAFG, resulting in suppression of NRF2-driven transcription. ZNF746 expression increased oxidative stress and apoptosis, phenotypes that were reversed by chemical and genetic hyperactivation of NRF2. This study presents a functionally annotated proximity network for NRF2 and suggests that ZNF746 overexpression in Parkinson’s disease directly inhibits NRF2-driven neuroprotection
2023-12-18 | PXD043182 | Pride
Project description:The nuclear factor erythroid 2-related factor 2 (NRF2) transcription factor activates cytoprotective and metabolic gene expression in response to various electrophilic stressors. In disease, constitutive NRF2 activity promotes cancer progression while decreased NRF2 function contributes to neurodegenerative diseases. In contrast to the regulation of NRF2 protein stability in the cytoplasm, co-complexed proteins that govern NRF2 activity on chromatin are less clear. Using biotin proximity proteomics, we report networks for NRF2 and its family members NRF1, NRF3 and the NRF2 heterodimer MAFG. We found that the Parkinson’s disease zinc finger transcription factor ZNF746 (PARIS) physically associated with NRF2 and MAFG, resulting in suppression of NRF2-driven transcription. ZNF746 expression increased oxidative stress and apoptosis, phenotypes that were reversed by chemical and genetic hyperactivation of NRF2. This study presents a functionally annotated proximity network for NRF2 and suggests that ZNF746 overexpression in Parkinson’s disease directly inhibits NRF2-driven neuroprotection.
2023-12-18 | PXD043248 | Pride
Project description:The activation of the transcription factor NF-E2-related factor 2 (Nrf2) maintains cellular homeostasis in response to oxidative stress by the regulation of multiple cytoprotective genes. Without stressors the activity of Nrf2 is inhibited by its interaction with the kelch-like ECH-associated protein 1 (Keap1). Here, we describe RA839, a small molecule that binds non-covalently to the Nrf2-interacting kelch domain of Keap1 with a Kd of approximately 6 µM, as demonstrated by X-ray co-crystallization and isothermal titration calorimetry. Whole-genome DNA arrays showed that at 10 µM RA839 significantly regulated 105 genes in bone marrow-derived macrophages. Canonical pathway mapping of these genes revealed an activation of pathways linked with Nrf2 signalling. These pathways were also activated after the activation of Nrf2 by the silencing of Keap1 expression. RA839 regulated only two genes in Nrf2 knockout macrophages. Similar to the activation of Nrf2 by either silencing of Keap1 expression or by the reactive compound CDDO-Me, RA839 prevented the induction of both inducible nitric oxide synthase expression and nitric oxide release in response to lipopolysaccharides in macrophages. In mice RA839 acutely induced Nrf2-target gene expression in liver. RA839 is a selective inhibitor of the Keap1/Nrf2 interaction and a useful tool compound to study the biology of Nrf2. Gene expression profile of bone marrow derived murine macrophages (BMDM) from Nrf2+/+ or Nrf2-/- mice treated with RA838, siKeap1-1 or siKeap1-2 were compared to untreated DMSO control or siControl. Four biological replicates were used for each sample group.
2015-10-31 | E-GEOD-71695 | biostudies-arrayexpress
Project description:Molecular glues are small molecules that exert their biologic or therapeutic activities by inducing gain-of-function interactions between pairs of proteins. In particular, molecular-glue degraders, which mediate interactions between target proteins and components of the ubiquitin proteasome system to cause targeted protein degradation, hold great promise as a unique modality for therapeutic targeting of proteins that are currently intractable. Here, we report a new molecular glue HQ461 discovered by high-throughput screening of small molecules that inhibited NRF2 activity. Using unbiased loss-of-function and gain-of-function genetic screening followed by biochemical reconstitution, we show that HQ461 acts by promoting interaction between CDK12 and DDB1-CUL4-RBX1 E3 ubiquitin ligase, leading to polyubiquitination and proteasomal degradation of CDK12’s interacting protein Cyclin K (CCNK). Degradation of CCNK mediated by HQ461 compromised CDK12 function, leading to reduced phosphorylation of CDK12 substrate, downregulation of DNA damage response genes, and cell death. Structure-activity relationship analysis of HQ461 revealed the importance of a 5-methylthiazol-2-amine pharmacophore and resulted in an HQ461 derivate with improved potency. Our studies reveal a new molecular glue that engages its target protein directly with DDB1 to bypass the requirement of a substrate-specific receptor, presenting a new strategy for targeted protein degradation.
2020-07-03 | GSE153700 | GEO
Project description:Molecular glues are small molecules that exert their biologic or therapeutic activities by inducing gain-of-function interactions between pairs of proteins. In particular, molecular-glue degraders, which mediate interactions between target proteins and components of the ubiquitin proteasome system to cause targeted protein degradation, hold great promise as a unique modality for therapeutic targeting of proteins that are currently intractable. Here, we report a new molecular glue HQ461 discovered by high-throughput screening of small molecules that inhibited NRF2 activity. Using unbiased loss-of-function and gain-of-function genetic screening followed by biochemical reconstitution, we show that HQ461 acts by promoting interaction between CDK12 and DDB1-CUL4-RBX1 E3 ubiquitin ligase, leading to polyubiquitination and proteasomal degradation of CDK12’s interacting protein Cyclin K (CCNK). Degradation of CCNK mediated by HQ461 compromised CDK12 function, leading to reduced phosphorylation of CDK12 substrate, downregulation of DNA damage response genes, and cell death. Structure-activity relationship analysis of HQ461 revealed the importance of a 5-methylthiazol-2-amine pharmacophore and resulted in an HQ461 derivate with improved potency. Our studies reveal a new molecular glue that engages its target protein directly with DDB1 to bypass the requirement of a substrate-specific receptor, presenting a new strategy for targeted protein degradation.
2020-07-03 | GSE153707 | GEO