Abstract

PAD4 has been strongly implicated in the pathogenesis of autoimmune, cardiovascular and oncological diseases through clinical genetics and gene disruption in mice. New selective PAD4 inhibitors binding a calcium-deficient form of the PAD4 enzyme have validated the critical enzymatic role of human and mouse PAD4 in both histone citrullination and neutrophil extracellular trap formation for, to our knowledge, the first time. The therapeutic potential of PAD4 inhibitors can now be explored.

  • Compound C25H26F3N5O3

    (3-Aminopiperidin-1-yl)(1-methyl-2-(1-methyl-1H-indol-2-yl)-1H-benzo[d]imidazol-5-yl)methanone trifluoroacetic acid salt

  • Compound C53H53N7O8

    (R)-2-((6-((3-(5-(3-Aminopiperidine-1-carbonyl)-2-(1-ethyl-1H-indol-2-yl)-1H-benzo[d]imidazol-1-yl)propyl)amino)-6-oxohexyl)carbamoyl)-6-(6-hydroxy-3-oxo-3H-xanthen-9-yl)benzoic acid

  • Compound C53H53N7O8

    (R)-5-((6-((3-(5-(3-Aminopiperidine-1-carbonyl)-2-(1-ethyl-1H-indol-2-yl)-1H-benzo[d]imidazol-1-yl)propyl)amino)-6-oxohexyl)carbamoyl)-2-(6-hydroxy-3-oxo-3H-xanthen-9-yl)benzoic acid

  • Compound C24H29ClN6O2

    (R)-(3-Aminopiperidin-1-yl)(2-(1-ethyl-1H-pyrrolo[2,3-b]pyridin-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-5-yl)methanone hydrochloride

  • Compound C27H32ClN5O3

    ((3S,4R)-3-Amino-4-hydroxypiperidin-1-yl)(2-(1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-5-yl)methanone hydrochloride

  • Compound C26H31ClN6O3

    ((3S,4R)-3-Amino-4-hydroxypiperidin-1-yl)(2-(1-(cyclopropylmethyl)-1H-pyrrolo[2,3-b]pyridin-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazol-5-yl)methanone hydrochloride

  • Compound C24H28ClN5O

    (3-Aminopiperidin-1-yl)(2-(1-ethyl-1H-indol-2-yl)-1-methyl-1H-benzo[d]imidazol-6-yl)methanone hydrochloride

  • Compound C24H28ClN5O

    (R)-(3-Aminopiperidin-1-yl)(2-(1-ethyl-1H-indol-2-yl)-1-methyl-1H-benzo[d]imidazol-5-yl)methanone hydrochloride

  • Compound C18H26N2O5

    trans-Benzyl 3-((tert-butoxycarbonyl)amino)-4-hydroxypiperidine-1-carboxylate

  • Compound C25H30N2O6

    cis-Benzyl 4-(benzoyloxy)-3-((tert-butoxycarbonyl)amino)piperidine-1-carboxylate

  • Compound C18H26N2O5

    (3S,4R)-Benzyl 3-((tert-butoxycarbonyl)amino)-4-hydroxypiperidine-1-carboxylate

  • Compound C10H20N2O3

    tert-Butyl ((3S,4R)-4-hydroxypiperidin-3-yl)carbamate

  • Compound C13H13NO

    1-(Cyclopropylmethyl)-1H-indole-2-carbaldehyde

  • Compound C10H12N2O5

    Methyl 3-methoxy-4-(methylamino)-5-nitrobenzoate

  • Compound C23H23N3O3

    Methyl 2-(1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylate

  • Compound C22H21N3O3

    2-(1-(Cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylic acid

  • Compound C32H39N5O5

    tert-Butyl ((3S,4R)-1-(2-(1-(cyclopropylmethyl)-1H-indol-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-4-hydroxypiperidin-3-yl)carbamate

  • Compound C13H10N2O2S

    1-(Phenylsulfonyl)-1H-pyrrolo[2,3-b]pyridine

  • Compound C14H10N2O3S

    1-(Phenylsulfonyl)-1H-pyrrolo[2,3-b]pyridine-2-carbaldehyde

  • Compound C10H10N2O

    1-Ethyl-1H-pyrrolo[2,3-b]pyridine-2-carbaldehyde

  • Compound C9H10N2O5

    3-Methoxy-4-(methylamino)-5-nitrobenzoic acid

  • Compound C19H28N4O6

    (R)-tert-Butyl (1-(3-methoxy-4-(methylamino)-5-nitrobenzoyl)piperidin-3-yl)carbamate

  • Compound C29H36N6O4

    (R)-tert-Butyl (1-(2-(1-ethyl-1H-pyrrolo[2,3-b]pyridin-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)piperidin-3-yl)carbamate

  • Compound C12H12N2O

    1-(Cyclopropylmethyl)-1H-pyrrolo[2,3-b]pyridine-2-carbaldehyde

  • Compound C22H22N4O3

    Methyl 2-(1-(cyclopropylmethyl)-1H-pyrrolo[2,3-b]pyridin-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylate

  • Compound C21H20N4O3

    2-(1-(Cyclopropylmethyl)-1H-pyrrolo[2,3-b]pyridin-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carboxylic acid

  • Compound C31H38N6O5

    tert-Butyl ((3S,4R)-1-(2-(1-(cyclopropylmethyl)-1H-pyrrolo[2,3-b]pyridin-2-yl)-7-methoxy-1-methyl-1H-benzo[d]imidazole-5-carbonyl)-4-hydroxypiperidin-3-yl)carbamate

  • Compound C11H11NO

    1-Ethyl-1H-indole-2-carbaldehyde

  • Compound C17H22ClN3O5

    (R)-tert-Butyl (1-(4-chloro-3-nitrobenzoyl)piperidin-3-yl)carbamate

  • Compound C20H31N5O5

    (R)-tert-Butyl (1-(4-((3-aminopropyl)amino)-3-nitrobenzoyl)piperidin-3-yl)carbamate

  • Compound C35H41N5O7

    1,1-Dimethylethyl {(3R)-1-[(4-{[3-({[(9H-fluoren-9-ylmethyl)oxy]carbonyl}amino)propyl]amino}-3-nitrophenyl)carbonyl]-3-piperidinyl}carbamate

  • Compound C46H50N6O5

    1,1-Dimethylethyl [(3R)-1-({2-(1-ethyl-1H-indol-2-yl)-1-[3-({[(9H-fluoren-9-ylmethyl)oxy]carbonyl}amino)propyl]-1H-benzimidazol-5-yl}carbonyl)-3-piperidinyl]carbamate

  • Compound C31H40N6O3

    (R)-tert-Butyl (1-(1-(3-aminopropyl)-2-(1-ethyl-1H-indol-2-yl)-1H-benzo[d]imidazole-5-carbonyl)piperidin-3-yl)carbamate

  • Compound C58H61N7O10

    (R)-2-((6-((3-(5-(3-((tert-Butoxycarbonyl)amino)piperidine-1-carbonyl)-2-(1-ethyl-1H-indol-2-yl)-1H-benzo[d]imidazol-1-yl)propyl)amino)-6-oxohexyl)carbamoyl)-6-(6-hydroxy-3-oxo-3H-xanthen-9-yl)benzoic acid

  • Compound C58H61N7O10

    (R)-2-((6-((3-(5-(3-((tert-Butoxycarbonyl)amino)piperidine-1-carbonyl)-2-(1-ethyl-1H-indol-2-yl)-1H-benzo[d]imidazol-1-yl)propyl)amino)-6-oxohexyl)carbamoyl)-2-(6-hydroxy-3-oxo-3H-xanthen-9-yl)benzoic acid

  • Compound C18H26N4O5

    1,1-Dimethylethyl ((3R)-1-{[4-(methylamino)-3-nitrophenyl]carbonyl}-3-piperidinyl)carbamate

  • Compound C29H35N5O3

    1,1-Dimethylethyl ((3R)-1-{[2-(1-ethyl-1H-indol-2-yl)-1-methyl-1H-benzimidazol-5-yl]carbonyl}-3-piperidinyl)carbamate

  • Compound C34H43N5O5

    (R)-tert-Butyl 2-(5-(3-((tert-butoxycarbonyl)amino)piperidine-1-carbonyl)-2-(1-ethyl-1H-indol-2-yl)-1H-benzo[d]imidazol-1-yl)acetate

  • Compound C25H27N5O3

    (R)-2-(5-(3-Aminopiperidine-1-carbonyl)-2-(1-ethyl-1H-indol-2-yl)-1H-benzo[d]imidazol-1-yl)acetic acid

  • Compound C20H19N3O2

    Methyl 2-(1-ethyl-1H-indol-2-yl)-1-methyl-1H-benzimidazole-6-carboxylate

  • Compound C19H17N3O2

    2-(1-Ethyl-1H-indol-2-yl)-1-methyl-1H-benzimidazole-6-carboxylic acid

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Acknowledgements

This work was supported in part by US National Institutes of Health grants GM079357 (to P.R.T.) and R01 HL102101 (to D.D.W.). We thank the following colleagues for their medicinal and synthetic chemistry contributions to this manuscript: D. Amans, H. Diallo, C. Douault, N. Garton, K. Jones, J. Renaux, T. Shipley, A. Walker, B. Watson and C. Wellaway. We also gratefully acknowledge S. Ready for diligently preparing and quantifying S. aureus for neutrophil studies. For the chemoproteomics data, we would like to thank M. Boesche and Cellzome's tissue culture and target validation teams for technical expertise and M. Bantscheff for stimulating discussion. We thank M. Neu and P. Homes for assistance for structural studies and B. Nolte for advice on crystallography.

Author information

Author notes

    • Robert J Sheppard
    • , Daniel J Slade
    • , Paul R Thompson
    •  & David M Wilson

    Present addresses: AstraZeneca, Oncology iMed, Cambridge Science Park, Cambridge, UK (R.J.S. and D.M.W.); Virginia Polytechnic Institute and State University, Blacksburg, Virginia, USA (D.J.S.); University of Massachussetts Medical School, Worcester, Massachusetts, USA (P.R.T.).

Affiliations

  1. EpiNova DPU, Immuno-Inflammation Therapy Area, GlaxoSmithKline,Medicines Research Centre, Stevenage, Hertfordshire, UK.

    • Huw D Lewis
    • , John Liddle
    • , Stephen J Atkinson
    • , Michael D Barker
    • , Matthew Campbell
    • , Rob P Davis
    • , Claire Maller
    • , Chris Patten
    • , Robert J Sheppard
    • , Rab K Prinjha
    •  & David M Wilson
  2. Molecular Discovery Research, GlaxoSmithKline, Medicines Research Centre, Stevenage, Hertfordshire, UK.

    • Jim E Coote
    • , Benjamin D Bax
    • , Ryan P Bingham
    • , Yu Hua Chen
    • , Chun-wa Chung
    • , Peter D Craggs
    • , Kelly Locke
    • , Oxana Polyakova
    • , Martin Rüdiger
    • , Pamela Thomas
    •  & Jim Thorpe
  3. Department of Chemistry, Scripps Florida, The Scripps Research Institute, Jupiter, Florida, USA.

    • Kevin L Bicker
    • , Daniel J Slade
    •  & Paul R Thompson
  4. Cellzome GmbH, a GSK company, Heidelberg, Germany.

    • Dirk Eberhard
    • , Gerard Joberty
    •  & Gerard Drewes
  5. ELT Boston, Platform Technology and Science, GlaxoSmithKline, Waltham, Massachusetts, USA.

    • Kenneth E Lind
    • , Cecil E Rise
    •  & Gang Yao
  6. Program in Cellular and Molecular Medicine, Boston Children's Hospital, Boston, Massachusetts, USA.

    • Kimberly Martinod
    •  & Denisa D Wagner
  7. Immunology Graduate Program, Division of Medical Sciences, Harvard Medical School, Boston, Massachusetts, USA.

    • Kimberly Martinod
  8. Division of Hematology/Oncology, Boston Children's Hospital, Boston, Massachusetts, USA.

    • Denisa D Wagner
  9. Department of Pediatrics, Harvard Medical School, Boston, Massachusetts, USA.

    • Denisa D Wagner

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Contributions

H.D.L., J.L. and J.E.C. led the project, designed the research, interpreted data and drafted the manuscript with input from all authors. S.J.A., M.D.B., M.C., R.J.S. and D.M.W. synthesized or designed key compounds. K.E.L., C.E.R. and G.Y. performed and analyzed the screening, which identified the PAD4 inhibitors. K.L.B., R.P.B., P.D.C., K.L. and D.J.S. designed the biochemical assays and deduced the mechanism of action of compounds. D.E., G.J. and G.D. generated chemoproteomic data. Y.H.C., R.P.D., D.E., C.M., K.M., C.P. and M.R. performed cellular assays. For structures, O.P. purified protein; J.T. crystallized protein, soaked crystals and collected data; B.D.B. refined structures; and P.T. performed sequence and structural analysis. C.-w.C. D.D.W., P.R.T., R.K.P. and D.M.W. guided aspects of this work. C.M., S.J.A., C.-w.C. and H.D.L. also contributed invaluably to the revision and formatting of the final manuscript.

Competing interests

P.R.T. is a co-founder and consultant to Padlock Therapeutics. The majority of the other authors are pharmaceutical industry employees and shareholders.

Corresponding author

Correspondence to Huw D Lewis.

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    Supplementary Text and Figures

    Supplementary Results, Supplementary Figures 1–19 and Supplementary Tables 1–6.

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DOI

https://doi.org/10.1038/nchembio.1735

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