Selective inhibitors of JAK1 targeting an isoform-restricted allosteric cysteine

Kavanagh, Madeline E.; Horning, Benjamin D.; Khattri, Roli; Roy, Nilotpal; Lu, Justine P; Whitby, Landon R.; Ye, Elva; Brannon, Jaclyn C; Parker, Albert E.; Chick, Joel M.; Eissler, Christie L.; Wong, Ashley P; Rodriguez, Joe L; Rodiles, Socorro; Masuda, Kim; Teijaro, John R.; Simón, Gabriel; Patricelli, Matthew P.; Cravatt, Benjamin F.

doi:10.1038/s41589-022-01098-0

Cited by 47 publications

(46 citation statements)

References 50 publications

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“…3c and Extended Data Fig. 3d, e ), and we have recently described covalent ligands targeting this cysteine that allosterically inhibit JAK1-dependent signaling in human cells 28 .…”

Section: Resultsmentioning

confidence: 96%

“…Note that C810 and C817 on the same tryptic peptide, so their ligandability cannot be distinguished by cysteine-directed MS-ABPP. However, other studies have verified that C817 is the liganded cysteine in JAK1 28 . e, Scatter plots for the JAK1_C817 region showing the correlation between base editing-induced cell line dropout data acquired herein and gene disruption-induced cell line dropout data in the Cancer Dependency Map.…”

Section: Extended Data Figmentioning

confidence: 95%

“…Note that C810 and C817 on the same tryptic peptide, so their ligandability cannot be distinguished by cysteine-directed MS-ABPP. However, other studies have verified that C817 is the liganded cysteine in JAK1 28 The conserved catalytic residues are highlighted in blue and the EERY sequence is highlighted in red. j, TOE1 homology model based on the crystal structure of PARN in complex with RNA (PDB: 2A1R).…”

Section: Data Availabilitymentioning

confidence: 99%

“…The copyright holder for this preprint this version posted November Note that C810 and C817 on the same tryptic peptide, so their ligandability cannot be distinguished by cysteine-directed MS-ABPP. However, other studies have verified that C817 is the liganded cysteine in JAK1 28 (which was not certified by peer review) is the author/funder. All rights reserved.…”

Section: Data Availabilitymentioning

confidence: 99%

“…Advantages of this approach include the deployment of covalent chemistry 19,[21][22][23] , which can address shallow and dynamic pockets on protein surfaces that are less amenable to binding reversibly to small molecules, as well as provide a selectivity filter in the form of targeting isotype-restricted nucleophilic residues (e.g., cysteines) within paralogous proteins, as have been demonstrated by several recent cancer therapeutics (e.g., osimertinib for EGFR_C797 24,25 , sotorasib for KRAS_G12C 26,27 ). Additionally, by evaluating small molecules directly in native biological systems, chemical proteomics methods such as ABPP can identify cryptic ligandable pockets regulating aspects of protein function that are difficult to discern with purified proteins or protein domains 17,28,29 .…”

Section: Introductionmentioning

confidence: 99%

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Assigning functionality to cysteines by base editing of cancer dependency genes

Remsberg

Won

et al. 2022

Preprint

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Chemical probes are lacking for most human proteins. Covalent chemistry represents an attractive strategy for expanding the ligandability of the proteome, and chemical proteomics has revealed numerous electrophile-reactive cysteines on diverse proteins. Determining which of these covalent binding events impact protein function, however, remains challenging. Here, we describe a base-editing strategy to infer the functionality of cysteines by quantifying the impact of their missense mutation on cell proliferation. We show that the resulting atlas, which covers >13,800 cysteines on >1,750 cancer dependency proteins, correctly predicts the essentiality of cysteines targeted by cancer therapeutics and, when integrated with chemical proteomic data, identifies essential, ligandable cysteines on >110 cancer dependency proteins. We further demonstrate how measurements of reactivity in native versus denatured proteomes can discriminate essential cysteines amenable to chemical modification from those buried in protein structures, providing a valuable resource to prioritize the pursuit of small-molecule probes with high function-perturbing potential.

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“…3c and Extended Data Fig. 3d, e ), and we have recently described covalent ligands targeting this cysteine that allosterically inhibit JAK1-dependent signaling in human cells 28 .…”

Section: Resultsmentioning

confidence: 96%

Section: Extended Data Figmentioning

confidence: 95%

“…Note that C810 and C817 on the same tryptic peptide, so their ligandability cannot be distinguished by cysteine-directed MS-ABPP. However, other studies have verified that C817 is the liganded cysteine in JAK1 28 The conserved catalytic residues are highlighted in blue and the EERY sequence is highlighted in red. j, TOE1 homology model based on the crystal structure of PARN in complex with RNA (PDB: 2A1R).…”

Section: Data Availabilitymentioning

confidence: 99%

Section: Data Availabilitymentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Assigning functionality to cysteines by base editing of cancer dependency genes

Remsberg

Won

et al. 2022

Preprint

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Chemical Proteomic Discovery of Isotype‐Selective Covalent Inhibitors of the RNA Methyltransferase NSUN2

Tao,

Felber,

Zou

et al. 2023

Angewandte Chemie

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Abstract5‐Methylcytosine (m5C) is an RNA modification prevalent on tRNAs, where it can protect tRNAs from endonucleolytic cleavage to maintain protein synthesis. The NSUN family (NSUN1‐7 in humans) of RNA methyltransferases are capable of installing the methyl group onto the C5 position of cytosines in RNA. NSUNs are implicated in a wide range of (patho)physiological processes, but selective and cell‐active inhibitors of these enzymes are lacking. Here, we use cysteine‐directed activity‐based protein profiling (ABPP) to discover azetidine acrylamides that act as stereoselective covalent inhibitors of human NSUN2. Despite targeting a conserved catalytic cysteine in the NSUN family, the NSUN2 inhibitors show negligible cross‐reactivity with other human NSUNs and exhibit good proteome‐wide selectivity. We verify that the azetidine acrylamides inhibit the catalytic activity of recombinant NSUN2, but not NSUN6, and demonstrate that these compounds stereoselectively disrupt NSUN2‐tRNA interactions in cancer cells, leading to a global reduction in tRNA m5C content. Our findings thus highlight the potential to create isotype‐selective and cell‐active inhibitors of NSUN2 with covalent chemistry targeting a conserved catalytic cysteine.

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