Discovery of Potent and Selective WDR5 Proteolysis Targeting Chimeras as Potential Therapeutics for Pancreatic Cancer

Yu, Xufen; Li, Dongxu; Kottur, Jithesh; Kim, Huen Suk; Herring, Laura E.; Yu, Yao; Xie, Ling; Hu, Xiaoping; Chen, Xian; Cai, Ling; Liu, Jing; Aggarwal, Aneel K.; Wang, Gang Greg; Jin, Jian

doi:10.1021/acs.jmedchem.3c01521

Cited by 15 publications

(5 citation statements)

References 58 publications

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“…The lack of WDR5 degradation is not due to a general inability of Cereblon to induce protein degradation RiPA conditions, since Cereblon efficiently decreased Aurora A levels. This is noteworthy because it recapitulates our experience and that of others with the WDR5 and Aurora A PROTACs: while Cereblon-based PROTACs were able to efficiently degrade Aurora A 10,11,33–35 , they were either nonfunctional 29 or much less potent 30,31 at degrading WDR5 compared to VHL-based PROTACs. A possible reason for the selective potency of both E3 ligases to degrade WDR5 by PROTACs could be their different ability to bind to WDR5 through protein-protein interactions and thus support ternary complex formation.…”

Section: Discussionsupporting

confidence: 87%

“…To test whether this rapamycin-induced dimerization assay (RiPA) could induce target degradation, we inserted the WD repeat-containing protein 5 (WDR5) and the von Hippel-Lindau tumour suppressor (VHL) into the FKBP12 and FRB-containing plasmid, respectively. We chose this target/E3 ligase pair because we 29 and others 30,31 have previously demonstrated robust degradation of WDR5 by PROTACs harnessing VHL. We started to optimize the RiPA system by transfecting different amounts of the WDR5 and VHL-encoding plasmids into HEK293 cells and incubating them with 0.1 µM rapamycin for 6 hours ( Figure 1C ).…”

Section: Resultsmentioning

confidence: 99%

“…We chose this target/E3 ligase pair because we 29 and others 30,31 have previously demonstrated robust degradation of WDR5 by PROTACs harnessing VHL. We started to optimize the RiPA system by transfecting different amounts of the WDR5 and VHL-encoding plasmids into HEK293 cells and incubating them with 0.1 µM rapamycin for 6 hours (Figure 1C).…”

Section: Rapamycin-induced Proximity Assay (Ripa) Induces Quantifiabl...mentioning

confidence: 99%

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Identification of Suitable Target/E3 Ligase Pairs for PROTAC Development using a Rapamycin-induced Proximity Assay (RiPA)

Adhikari,

Schneider,

Diebold

et al. 2024

Preprint

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The development of proteolysis targeting chimeras (PROTACs), which induce the degradation of target proteins by bringing them into proximity with cellular E3 ubiquitin ligases, has revolutionized drug development. While the human genome encodes more than 600 different E3 ligases, current PROTACs use only a handful of them, drastically limiting their full potential. Furthermore, many PROTAC development campaigns fail because the selected E3 ligase candidates are unable to induce degradation of the particular target of interest. As more and more ligands for novel E3 ligases are discovered, the chemical effort to identify the best E3 ligase for a given target is exploding. Therefore, a genetic system to identify degradation-causing E3 ligases and suitable target/E3 ligase pairs is urgently needed. Here we used the well-established dimerization of the FKBP12 protein and FRB domain by rapamycin to bring the target protein WDR5 into proximity with candidate E3 ligases. Strikingly, this rapamycin-induced proximity assay (RiPA) revealed that VHL, but not Cereblon, is able to induce WDR5 degradation - a finding previously made by PROTACs, demonstrating its predictive power. By optimizing the steric arrangement of all components and fusing the target protein with a minimal luciferase, RiPA can identify the ideal E3 for any target protein of interest in living cells, significantly reducing and focusing the chemical effort in the early stages of PROTAC development.

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Section: Discussionsupporting

confidence: 87%

Section: Resultsmentioning

confidence: 99%

Section: Rapamycin-induced Proximity Assay (Ripa) Induces Quantifiabl...mentioning

confidence: 99%

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Identification of Suitable Target/E3 Ligase Pairs for PROTAC Development using a Rapamycin-induced Proximity Assay (RiPA)

Adhikari,

Schneider,

Diebold

et al. 2024

Preprint

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“…Accordingly, this review focuses largely on this class of agents. PROTACs, or WDR5 degraders, are the most recent group of WDR5 inhibitors to emerge, all of which are built with a WIN site ligand as the WDR5-targeting entity [ 18 , 35 , 36 , 37 , 38 ]. By triggering WDR5 destruction, PROTACs should in theory supersede both WBM and WIN site inhibitors.…”

Section: Strategies To Target Wdr5mentioning

confidence: 99%

WD Repeat Domain 5 Inhibitors for Cancer Therapy: Not What You Think

Weissmiller,

Fesik,

Tansey

2024

JCM

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WDR5 is a conserved nuclear protein that scaffolds the assembly of epigenetic regulatory complexes and moonlights in functions ranging from recruiting MYC oncoproteins to chromatin to facilitating the integrity of mitosis. It is also a high-value target for anti-cancer therapies, with small molecule WDR5 inhibitors and degraders undergoing extensive preclinical assessment. WDR5 inhibitors were originally conceived as epigenetic modulators, proposed to inhibit cancer cells by reversing oncogenic patterns of histone H3 lysine 4 methylation—a notion that persists to this day. This premise, however, does not withstand contemporary inspection and establishes expectations for the mechanisms and utility of WDR5 inhibitors that can likely never be met. Here, we highlight salient misconceptions regarding WDR5 inhibitors as epigenetic modulators and provide a unified model for their action as a ribosome-directed anti-cancer therapy that helps focus understanding of when and how the tumor-inhibiting properties of these agents can best be understood and exploited.

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“…Although WDR5 PROTACs have been described ( Yu et al, 2021 ; Li et al, 2022 ; Yu et al, 2023 ), safety concerns over destroying a pan-essential protein such as WDR5 ( Siladi et al, 2022 ) means that most drug discovery efforts have focused on small-molecule inhibition of key binding sites on the protein. Some initiatives target a hydrophobic cleft on WDR5 known as the ‘WDR5-binding motif’ (WBM) site ( Macdonald et al, 2019 ; Chacón Simon et al, 2020 ) that contacts MYC ( Thomas et al, 2015 ).…”

Section: Introductionmentioning

confidence: 99%

Ribosome subunit attrition and activation of the p53–MDM4 axis dominate the response of MLL-rearranged cancer cells to WDR5 WIN site inhibition

Howard,

Wang,

Rose

et al. 2024

eLife

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The chromatin-associated protein WDR5 is a promising target for cancer drug discovery, with most efforts blocking an arginine-binding cavity on the protein called the “WIN” site that tethers WDR5 to chromatin. WIN site inhibitors (WINi) are active against multiple cancer cell types in vitro, the most notable of which are those derived from MLL-rearranged (MLLr) leukemias. Peptidomimetic WINi were originally proposed to inhibit MLLr cells via dysregulation of genes connected to hematopoetic stem cell expansion. Our discovery and interrogation of small molecule WIN site inhibitors, however, revealed that they act in MLLr cell lines to suppress ribosome protein gene (RPG) transcription, induce nucleolar stress, and activate p53. Because there is no precedent for an anti-cancer strategy that specifically targets RPG expression, we took an integrated multi-omics approach to further interrogate the mechanism of action of WINi in MLLr cancer cells. We show that WINi induce depletion of the stock of ribosomes, accompanied by a broad translational choke, induction of a DNA damage response, and changes in alternative mRNA splicing that inactivate the p53 antagonist MDM4. We also show that WINi are synergistic with agents including venetoclax and BET-bromodomain inhibitors. Together, these studies reinforce the concept that WINi are a novel type of ribosome-directed anti-cancer therapy and provide a resource to support their clinical implementation in MLLr leukemias and other malignancies.

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Discovery of Potent and Selective WDR5 Proteolysis Targeting Chimeras as Potential Therapeutics for Pancreatic Cancer

Cited by 15 publications

References 58 publications

Identification of Suitable Target/E3 Ligase Pairs for PROTAC Development using a Rapamycin-induced Proximity Assay (RiPA)

Identification of Suitable Target/E3 Ligase Pairs for PROTAC Development using a Rapamycin-induced Proximity Assay (RiPA)

WD Repeat Domain 5 Inhibitors for Cancer Therapy: Not What You Think

Ribosome subunit attrition and activation of the p53–MDM4 axis dominate the response of MLL-rearranged cancer cells to WDR5 WIN site inhibition

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