Friday, March 22, 2019

Development of a Selective CDK7 Covalent Inhibitor Reveals Predominant Cell-Cycle Phenotype


Calla M. Olson, Yanke Liang, Alan Leggett, Woojun D. Park, Lianbo Li, Caitlin E. Mills, Selma Z. Elsarrag, Scott B. Ficarro, Tinghu Zhang, Robert Düster, Matthias Geyer, Taebo Sim, Jarrod A. Marto, Peter K. Sorger, Ken D. Westover, Charles Y. Lin, Nicholas Kwiatkowski, Nathanael S. Gray,
Cell Chemical Biology2019
Cyclin-dependent kinase 7 (CDK7) regulates both cell cycle and transcription, but its precise role remains elusive. We previously described THZ1, a CDK7 inhibitor, which dramatically inhibits superenhancer-associated gene expression. However, potent CDK12/13 off-target activity obscured CDK7s contribution to this phenotype. Here, we describe the discovery of a highly selective covalent CDK7 inhibitor. YKL-5-124 causes arrest at the G 1/S transition and inhibition of E2F-driven gene expression; these effects are rescued by a CDK7 mutant unable to covalently engage YKL-5-124, demonstrating on-target specificity. Unlike THZ1, treatment with YKL-5-124 resulted in no change to RNA polymerase II C-terminal domain phosphorylation; however, inhibition could be reconstituted by combining YKL-5-124 and THZ531, a selective CDK12/13 inhibitor, revealing potential redundancies in CDK control of gene transcription. These findings highlight the importance of CDK7/12/13 polypharmacology for anti-cancer activity of THZ1 and posit that selective inhibition of CDK7 may be useful for treatment of cancers marked by E2F misregulation.

Tuesday, March 19, 2019

Dimethyl Fumarate Disrupts Human Innate Immune Signaling by Targeting the IRAK4–MyD88 Complex

Balyn W. ZaroEkaterina V. VinogradovaDaniel C. LazarMegan M. BlewettRadu M. SuciuJunichiro TakayaSean StuderJuan Carlos de la TorreJean-Laurent CasanovaBenjamin F. CravattJohn R. Teijaro


Dimethyl fumarate (DMF) is a prescribed treatment for multiple sclerosis and has also been used to treat psoriasis. The electrophilicity of DMF suggests that its immunosuppressive activity is related to the covalent modification of cysteine residues in the human proteome. Nonetheless, our understanding of the proteins modified by DMF in human immune cells and the functional consequences of these reactions remains incomplete. In this study, we report that DMF inhibits human plasmacytoid dendritic cell function through a mechanism of action that is independent of the major electrophile sensor NRF2. Using chemical proteomics, we instead identify cysteine 13 of the innate immune kinase IRAK4 as a principal cellular target of DMF. We show that DMF blocks IRAK4–MyD88 interactions and IRAK4-mediated cytokine production in a cysteine 13–dependent manner. Our studies thus identify a proteomic hotspot for DMF action that constitutes a druggable protein–protein interface crucial for initiating innate immune responses.

Azabicyclic Vinyl Sulfones for Residue-specific Dual Protein Labelling

Enrique Gil de Montes,  Ester Jiménez-Moreno,  Bruno Oliveira,  Claudio Daniel Navo,  Pedro M. S. D. Cal,  Gonzalo Jiménez-Osés,  Inmaculada Robina,  Antonio J Moreno-Vargas  and  Gonçalo J. L. Bernardes

Chemical Science, 2019
DOI: 10.1039/x0xx00000x

We have developed [2.2.1]azabicyclic vinyl sulfone reagents that simultaneously enable cysteine-selective protein modification and introduce a handle for further bioorthogonal ligation. The reaction is fast and selective for cysteine relative to other amino acids that have nucleophilic side-chains, and the formed products are stable in human plasma and are moderately resistant to retro Diels–Alder degradation reactions. A model biotinylated [2.2.1]azabicyclic vinyl sulfone reagent was shown to efficiently label two cysteine-tagged proteins, ubiquitin and C2Am, under mild conditions (1–5 equiv. of reagent in NaPi pH 7.0, room temperature, 30 min.). The resulting thioether-linked conjugates were stable and retained the native activity of the proteins. Finally, the dienophile present in the azabicyclic moiety on a functionalised C2Am protein could be fluorescently labelled through an inverse electron demand Diels–Alder reaction in cells to allow selective apoptosis imaging. The combined advantages of directness, site-specificity and easy preparation mean [2.2.1]azabicyclic vinyl sulfones can be used for residue-specific dual protein labelling/construction strategies with minimal perturbation of native function based simply on the attachment of an [2.2.1]azabicyclic moiety to cysteine.

Sunday, March 17, 2019

“Sleeping Beauty” Phenomenon: SuFEx-Enabled Discovery of Selective Covalent Inhibitors of Human Neutrophil Elastase

Qinheng Zheng and Jordan L. Woehl and Seiya Kitamura and Diogo Santos-Martins and Christopher J. Smedley and Gencheng Li and Stefano Forli and John E. Moses and Dennis W. Wolan and K. Barry Sharpless

ChemRxiv, 2019
DOI: 10.26434/chemrxiv.7842020.v1

Sulfur-Fluoride Exchange (SuFEx) has emerged as the new generation of click chemistry. We report here a SuFEx-enabled approach exploiting the “sleeping beauty” phenomenon of sulfur fluoride compounds in the context of the serendipitous discovery of selective covalent human neutrophil elastase (hNE) inhibitors. Evaluation of an ever-growing collection of SuFExable compounds toward various biological assays unexpectedly yielded a selective and covalent hNE inhibitor, benzene-1,2-disulfonyl fluoride. Derivatization of the initial hit led to a better agent, 2- triflyl benzenesulfonyl fluoride, itself made through a SuFEx trifluoromethylation process, with IC50 = 1.1 μM and ~200-fold selectivity over the homologous neutrophil serine protease, cathepsin G. The optimized probe only modified active hNE and not its denatured form, setting another example of the “sleeping beauty” phenomenon of sulfur fluoride capturing agents for the discovery of covalent medicines.


Thursday, February 28, 2019

The Alkyne Moiety as a Latent Electrophile in Irreversible Covalent Small Molecule Inhibitors of Cathepsin K

J. Am. Chem. Soc.2019141 (8), 3507–3514

Irreversible covalent inhibitors can have a beneficial pharmacokinetic/pharmacodynamics profile but are still often avoided due to the risk of indiscriminate covalent reactivity and the resulting adverse effects. To overcome this potential liability, we introduced an alkyne moiety as a latent electrophile into small molecule inhibitors of cathepsin K (CatK). Alkyne-based inhibitors do not show indiscriminate thiol reactivity but potently inhibit CatK protease activity by formation of an irreversible covalent bond with the catalytic cysteine residue, confirmed by crystal structure analysis. The rate of covalent bond formation (kinact) does not correlate with electrophilicity of the alkyne moiety, indicative of a proximity-driven reactivity. Inhibition of CatK-mediated bone resorption is validated in human osteoclasts. Together, this work illustrates the potential of alkynes as latent electrophiles in small molecule inhibitors, enabling the development of irreversible covalent inhibitors with an improved safety profile.

DUckCov: a Dynamic Undocking‐based Virtual Screening Protocol for Covalent Binders

Moira Rachman Andrea Scarpino Dávid Bajusz Gyula Palfy Istvan Vida Andras Perczel Xavier Barril György M Keseru

ChemMedChem, 2019. doi:10.1002/cmdc.201900078

Thanks to recent guidelines, the design of safe and effective covalent drugs has gained significant interest. Other than targeting non‐conserved nucleophilic residues, optimizing the non‐covalent binding framework is important to improve potency and selectivity of covalent binders towards the desired target. Strong efforts have been made in extending the computational toolkits to include a covalent mechanism of protein targeting, like in the development of covalent docking methods for binding mode prediction. To highlight the value of the non‐covalent complex in the covalent binding process, here we describe a new protocol utilizing tethered and constrained docking in combination with Dynamic Undocking (DUck) as a tool to privilege strong protein binders for the identification of novel covalent inhibitors. At the end of the protocol, dedicated covalent docking methods were used to rank and select the virtual hits based on the predicted binding mode. By validating the method on JAK3 and KRas, we demonstrate how this fast iterative protocol could be applied to explore a wide chemical space and identify potent targeted covalent inhibitors.

Tuesday, February 26, 2019

Allosteric Inhibition of Ubiquitin-like Modifications by a Class of Inhibitor of SUMO-Activating Enzyme

Yi-Jia Li, Li Du, Jianghai Wang, Ramir Vega, Terry D. Lee, Yunan Miao, Grace Aldana-Masangkay, Eric R. Samuels, Baozong Li, S. Xiaohu Ouyang, Sharon A. Colayco, Ekaterina V. Bobkova, Daniela B. Divlianska, Eduard Sergienko, Thomas D.Y. Chung, Marwan Fakih, Yuan Chen

Cell Chemical Biology, 2019
DOI: 10.1016/j.chembiol.2018.10.026

Ubiquitin-like (Ubl) post-translational modifications are potential targets for therapeutics. However, the only known mechanism for inhibiting a Ubl-activating enzyme is through targeting its ATP-binding site. Here we identify an allosteric inhibitory site in the small ubiquitin-like modifier (SUMO)-activating enzyme (E1). This site was unexpected because both it and analogous sites are deeply buried in all previously solved structures of E1s of ubiquitin-like modifiers (Ubl). The inhibitor not only suppresses SUMO E1 activity, but also enhances its degradation in vivo, presumably due to a conformational change induced by the compound. In addition, the lead compound increased the expression of miR-34b and reduced c-Myc levels in lymphoma and colorectal cancer cell lines and a colorectal cancer xenograft mouse model. Identification of this first-in-class inhibitor of SUMO E1 is a major advance in modulating Ubl modifications for therapeutic aims.

Development of a Selective CDK7 Covalent Inhibitor Reveals Predominant Cell-Cycle Phenotype

Calla M. Olson, Yanke Liang, Alan Leggett, Woojun D. Park, Lianbo Li, Caitlin E. Mills, Selma Z. Elsarrag, Scott B. Ficarro, Tinghu Zhan...