Abstract

Rhinoviruses (RVs) are the pathogens most often responsible for the common cold, and are a frequent cause of exacerbations in asthma, chronic obstructive pulmonary disease and cystic fibrosis. Here we report the discovery of IMP-1088, a picomolar dual inhibitor of the human N-myristoyltransferases NMT1 and NMT2, and use it to demonstrate that pharmacological inhibition of host-cell N-myristoylation rapidly and completely prevents rhinoviral replication without inducing cytotoxicity. The identification of cooperative binding between weak-binding fragments led to rapid inhibitor optimization through fragment reconstruction, structure-guided fragment linking and conformational control over linker geometry. We show that inhibition of the co-translational myristoylation of a specific virus-encoded protein (VP0) by IMP-1088 potently blocks a key step in viral capsid assembly, to deliver a low nanomolar antiviral activity against multiple RV strains, poliovirus and foot and-mouth disease virus, and protection of cells against virus-induced killing, highlighting the potential of host myristoylation as a drug target in picornaviral infections.

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References

  1. 1.

    Ritchie, A. I. et al. Pathogenesis of viral infection in exacerbations of airway disease. Ann. Am. Thorac. Soc. 12, S115–132 (2015).

  2. 2.

    Kieninger, E. et al. High rhinovirus burden in lower airways of children with cystic fibrosis. Chest 143, 782–790 (2013).

  3. 3.

    Flight, W. G. et al. Incidence and clinical impact of respiratory viruses in adults with cystic fibrosis. Thorax 69, 247–253 (2014).

  4. 4.

    Thibaut, H. J., De Palma, A. M. & Neyts, J. Combating enterovirus replication: state-of-the-art on antiviral research. Biochem. Pharmacol. 83, 185–192 (2012).

  5. 5.

    Jiang, P., Liu, Y., Ma, H. C., Paul, A. V. & Wimmer, E. Picornavirus morphogenesis. Microbiol. Mol. Biol. Rev. 78, 418–437 (2014).

  6. 6.

    Liu, Y. et al. Atomic structure of a rhinovirus C, a virus species linked to severe childhood asthma. Proc. Natl Acad. Sci. USA 113, 8997–9002 (2016).

  7. 7.

    Wright, M. H., Heal, W. P., Mann, D. J. & Tate, E. W. Protein myristoylation in health and disease. J. Chem. Biol. 3, 19–35 (2010).

  8. 8.

    Marc, D., Masson, G., Girard, M. & van der Werf, S. Lack of myristoylation of poliovirus capsid polypeptide VP0 prevents the formation of virions or results in the assembly of noninfectious virus particles. J. Virol. 64, 4099–4107 (1990).

  9. 9.

    Marc, D., Drugeon, G., Haenni, A. L., Girard, M. & van der Werf, S. Role of myristoylation of poliovirus capsid protein VP4 as determined by site-directed mutagenesis of its N-terminal sequence. EMBO J. 8, 2661–2668 (1989).

  10. 10.

    Marc, D., Girard, M. & van der Werf, S. A Gly1 to Ala substitution in poliovirus capsid protein VP0 blocks its myristoylation and prevents viral assembly. J. Gen. Virol. 72, 1151–1157 (1991).

  11. 11.

    Moscufo, N., Simons, J. & Chow, M. Myristoylation is important at multiple stages in poliovirus assembly. J. Virol. 65, 2372–2380 (1991).

  12. 12.

    Ritzefeld, M., Wright, M. H. & Tate, E. W. New developments in probing and targeting protein acylation in malaria, leishmaniasis and African sleeping sickness. Parasitology 145, 157–174 (2018).

  13. 13.

    Thinon, E. et al. Global profiling of co- and post-translationally N-myristoylated proteomes in human cells. Nat. Commun. 5, 4919 (2014).

  14. 14.

    Bell, A. S. et al. Selective inhibitors of protozoan protein N-myristoyltransferases as starting points for tropical disease medicinal chemistry programs. PLoS Negl. Trop. Dis. 6, e1625 (2012).

  15. 15.

    Brannigan, J. A. et al. N-myristoyltransferase from Leishmania donovani: structural and functional characterisation of a potential drug target for visceral leishmaniasis. J. Mol. Biol. 396, 985–999 (2010).

  16. 16.

    Goncalves, V. et al. Discovery of Plasmodium vivax N-myristoyltransferase inhibitors: screening, synthesis, and structural characterization of their binding mode. J. Med. Chem. 55, 3578–3582 (2012).

  17. 17.

    Schärfer, C. et al. Torsion angle preferences in druglike chemical space: a comprehensive guide. J. Med. Chem. 56, 2016–2028 (2013).

  18. 18.

    Goncalves, V. et al. A fluorescence-based assay for N-myristoyltransferase activity. Anal. Biochem. 421, 342–344 (2012).

  19. 19.

    Wright, M. H. et al. Validation of N-myristoyltransferase as an antimalarial drug target using an integrated chemical biology approach. Nat. Chem. 6, 112–121 (2014).

  20. 20.

    Thinon, E., Morales-Sanfrutos, J., Mann, D. J. & Tate, E. W. N-myristoyltransferase inhibition induces ER-stress, cell cycle arrest, and apoptosis in cancer cells. ACS Chem. Biol. 11, 2165–2176 (2016).

  21. 21.

    Broncel, M. et al. Multifunctional reagents for quantitative proteome-wide analysis of protein modification in human cells and dynamic profiling of protein lipidation during vertebrate development. Angew. Chem. Int. Ed. 54, 5948–5951 (2015).

  22. 22.

    Broncel, M. et al. Myristoylation profiling in human cells and zebrafish. Data Brief. 4, 379–383 (2015).

  23. 23.

    Tyanova, S., Temu, T. & Cox, J. The MaxQuant computational platform for mass spectrometry-based shotgun proteomics. Nat. Protocols 11, 2301–2319 (2016).

  24. 24.

    Reddel, H. K. et al. A summary of the new GINA strategy: a roadmap to asthma control. Eur. Respir. J. 46, 622–639 (2015).

  25. 25.

    Lee, W. M., Monroe, S. S. & Rueckert, R. R. Role of maturation cleavage in infectivity of picornaviruses: activation of an infectosome. J. Virol. 67, 2110–2122 (1993).

  26. 26.

    Koh, C. et al. Oral prenylation inhibition with lonafarnib in chronic hepatitis D infection: a proof-of-concept randomised, double-blind, placebo-controlled phase 2A trial. Lancet Infect. Dis. 15, 1167–1174 (2015).

  27. 27.

    Vizcaino, J. A. et al. ProteomeXchange provides globally coordinated proteomics data submission and dissemination. Nat. Biotechnol. 32, 223–226 (2014).

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Acknowledgements

The authors gratefully acknowledge financial support from the Medical Research Council (MRC) of the UK (grant G0900278 and a confidence-in-concept award to Imperial College, MC-PC-14100), the EPSRC (grant EP/F500416/1), Cancer Research UK (grant C29637/A20183), MRC Asthma UK Centre in Allergic Mechanisms of Asthma (grant G1000758) and Imperial Innovations. A.M. is supported by an MRF/Asthma UK Research Grant (MRFAUK-2015-311). S.L.J. is supported by a Chair from Asthma UK (CH11SJ) and is a National Institute of Health Research (NIHR) Senior Investigator. The Pirbright Institute receives strategic funding from the Biotechnology and Biological Research Council of the UK. We thank Diamond Light Source for access to beamlines I04 and I04-1 (proposal nos mx12579, mx7864 and mx9948). We thank E. Cota (Imperial College London) for advice on structure determination, and S. Roberts for expert crystal handling. The authors thank K.-K. Conzelmann (Max von Pettenkofer-Institut) for permission to use the BSR-T7/5 cell line.

Author information

Author notes

    • Robin J. Leatherbarrow

    Present address: Liverpool John Moores University, Liverpool, UK

  1. These authors contributed equally: Aurélie Mousnier, Andrew S. Bell.

Affiliations

  1. National Heart & Lung Institute, Imperial College London, London, UK

    • Aurélie Mousnier
    • , Dawid P. Swieboda
    • , Anabel Guedán
    • , Sebastian L. Johnston
    •  & Roberto Solari
  2. Centre for Experimental Medicine, Queen’s University Belfast, Belfast, UK

    • Aurélie Mousnier
  3. Department of Chemistry, Imperial College London, London, UK

    • Andrew S. Bell
    • , Julia Morales-Sanfrutos
    • , Inmaculada Pérez-Dorado
    • , Markus Ritzefeld
    • , Jennie A. Hutton
    • , Robin J. Leatherbarrow
    •  & Edward W. Tate
  4. Department of Life Sciences, Imperial College London, London, UK

    • Inmaculada Pérez-Dorado
  5. Structural Biology Laboratory, Department of Chemistry, University of York, York, UK

    • James A. Brannigan
    •  & Anthony J. Wilkinson
  6. The Pirbright Institute, Woking, UK

    • Joseph Newman
    • , Amin S. Asfor
    •  & Tobias J. Tuthill
  7. Kinetic Discovery Limited, Dundee, UK

    • Sean W. Robinson
    •  & Iva Hopkins-Navratilova
  8. School of Life Sciences, University of Dundee, Dundee, UK

    • Iva Hopkins-Navratilova

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Contributions

A.M. and D.P.S. designed and executed the RV infection experiments, with assistance from A.G. and R.S.; A.S.B. designed and synthesized the NMT inhibitors, with E.W.T., J.A.H. and R.J.L.; J.N., A.S.A. and T.J.T. undertook the PV and FMDV infections, and the sucrose gradient fractionation. J.M.-S. undertook chemical proteomics experiments and data analysis, with E.W.T.; I.P.-D., J.A.B. and A.J.W. undertook protein purification, X-ray crystallography and structure refinement. M.R. undertook cell viability and tagging experiments. S.W.R. and I.H.N. undertook SPR experiments and data analysis. S.L.J. provided advice on virus experiments, and access to essential facilities. E.W.T. and R.S. conceived the project and, with A.M. and A.B., directed the project. All of the authors contributed to the writing of the manuscript.

Competing interests

A.B., E.W.T., R.J.L., J.A.H. and J.A.B. are inventors on a patent application that describes NMT inhibitors that include IMP-1031 and IMP-1088 (Bell, A.S.; Tate, E.W.; Leatherbarrow, R.J.; Hutton, J.A.; Brannigan, J.A., Compounds and their use as inhibitors of N-myristoyl transferase, Patent Cooperation Treaty International Application (2017) WO 2017001812).

Corresponding authors

Correspondence to Roberto Solari or Edward W. Tate.

Supplementary information

  1. Supplementary Information

    Supplementary Figures 1–15, Table 1, Methods, NMR Spectra and References

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DOI

https://doi.org/10.1038/s41557-018-0039-2