MCC950 directly targets the NLRP3 ATP-hydrolysis motif for inflammasome inhibition

Abstract

Inhibition of the NLRP3 inflammasome is a promising strategy for the development of new treatments for inflammatory diseases. MCC950 is a potent and specific small-molecule inhibitor of the NLRP3 pathway, but its molecular target is not defined. Here, we show that MCC950 directly interacts with the Walker B motif within the NLRP3 NACHT domain, thereby blocking ATP hydrolysis and inhibiting NLRP3 activation and inflammasome formation.

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Fig. 1: MCC950 directly interacts with NLRP3.
Fig. 2: MCC950 interacts with the NACHT domain of NLRP3.
Fig. 3: MCC950 binds non-covalently to NLRP3, proximal to the Walker B motif and blocks NLRP3 ATPase activity.

Data availability

The data that support the findings of this study are available from the corresponding author upon reasonable request.

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Acknowledgements

This work was supported by the National Health and Medical Research Council of Australia (Fellowship 1138466 and Program Grant no. 1071659 to M.P.J.; Fellowship no. 1141131 to K.S.; Project Grant no. 1086786 to A.A.B.R. and K.S.), the Australian Research Council (Fellowship no. FT130100361 to K.S.), the Institute for Molecular Bioscience (Research Advancement Award to J.H.) and The University of Queensland (Postdoctoral Fellowships to R.C.C. and D.B.; Research Scholarship to J.H.). We thank D. Edwards for chemical purification and analytical support, M. Cooper (University of Queensland) for providing MCC950 and K. Stacey (University of Queensland) for providing ASC-deficient mice.

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Contributions

R.C.C. designed and performed most experiments. J.R.H. conceived and synthesized the photoaffinity probe and performed labeling experiments. C.J.D. and M.P.J. designed and performed SPR analysis. A.Z. designed and cloned the NLRP3 expression plasmids and mutants. D.B. expressed and purified recombinant NLRP3 and assisted with experimental design. N.L.M synthesized the photoaffinity probe. J.L.C. and J.A.F. assisted with the expression and purification of recombinant NEK7, A.A.B.R. formulated MCC950, conceived the photoaffinity probe and expressed and purified recombinant NEK7. A.A.B.R. and K.S. designed experiments and supervised the study. R.C.C and K.S wrote the manuscript, with assistance from J.R.H. and A.A.B.R. and input from all authors.

Corresponding authors

Correspondence to Rebecca C. Coll or Avril A. B. Robertson or Kate Schroder.

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Competing interests

R.C.C., A.A.B.R. and K.S. are co-inventors on patent applications for NLRP3 inhibitors (WO2018215818, WO2017140778 and WO2016131098), which are licensed to Inflazome Ltd, a company headquartered in Dublin, Ireland. Inflazome is developing drugs that target the NLRP3 inflammasome to address unmet clinical needs in inflammatory disease.

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Supplementary Table 1, Supplementary Figures 1–15

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Synthetic Procedures

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Coll, R.C., Hill, J.R., Day, C.J. et al. MCC950 directly targets the NLRP3 ATP-hydrolysis motif for inflammasome inhibition. Nat Chem Biol 15, 556–559 (2019). https://doi.org/10.1038/s41589-019-0277-7

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