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Imitation of β-lactam binding enables broad-spectrum metallo-β-lactamase inhibitors

Abstract

Carbapenems are vital antibiotics, but their efficacy is increasingly compromised by metallo-β-lactamases (MBLs). Here we report the discovery and optimization of potent broad-spectrum MBL inhibitors. A high-throughput screen for NDM-1 inhibitors identified indole-2-carboxylates (InCs) as potential β-lactamase stable β-lactam mimics. Subsequent structure–activity relationship studies revealed InCs as a new class of potent MBL inhibitor, active against all MBL classes of major clinical relevance. Crystallographic studies revealed a binding mode of the InCs to MBLs that, in some regards, mimics that predicted for intact carbapenems, including with respect to maintenance of the Zn(II)-bound hydroxyl, and in other regards mimics binding observed in MBL–carbapenem product complexes. InCs restore carbapenem activity against multiple drug-resistant Gram-negative bacteria and have a low frequency of resistance. InCs also have a good in vivo safety profile, and when combined with meropenem show a strong in vivo efficacy in peritonitis and thigh mouse infection models.

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Fig. 1: InC binding to MBLs mimics that of intact β-lactam substrates and/or products.
Fig. 2: InCs inhibit B1 and B3 MBLs via an unprecedented MBL binding mode.
Fig. 3: Heat-map analysis comparing the potency of InCs for selected clinically important SBLs and MBLs with β-lactamase inhibitors.
Fig. 4: Strategies for Indole carboxylate synthesis.
Fig. 5: InC 58 potentiates meropenem activity in vitro against clinically relevant strains.
Fig. 6: InC 58 potentiates meropenem activity in vivo against clinically relevant strains.

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Acknowledgements

We thank the ELF Screening partners for efforts leading to the initial hits, the current and past ENABLE Portfolio Management Committee (PMC) members and EFPIA partners for guidance, and the Diamond Light Source synchrotron for access to the IO3, IO4 and IO4-1 beamlines. J.U.H. thanks N. Sipari from the Viikki Metabolomics Unit (Helsinki Institute of Life Science, University of Helsinki and Biocenter Finland) for her expertise with the liquid chromatography–mass spectroscopy analyses. L.E. thanks Y. Zhou for help in the laboratory, and H. Saif, R. Farzana, E. Portal, K. Sands, K. Thomson and B. Hassan for providing strains for the Enterobacterales collection. The ELF work and the ENABLE project that led to these results were supported by the Innovative Medicines Initiative Joint Undertaking (grant agreements no. 115489 and no. 115583), which have financial contributions from the European Union’s Seventh Framework Programme (FP7/2007–2013), and the EFPIA companies’ in-kind contributions. The ENABLE project is financially supported by contributions from Academic and SME partners. The Oxford work was also supported by Cancer Research UK (C9047/A24759), the Medical Research Council, the Biotechnology and Biological Research Council (BB/S50676X/1) and the Ineos Oxford Institute for Antimicrobial Research (C.J.S.). This research was funded in whole, or in part, by the Wellcome Trust (grant no. 106244/Z/14/Z and no. 099141/Z/12/Z). The Bristol work was also supported by the National Institute of Allergy and Infectious Diseases of the National Institutes of Health (NIH) grant R01AI100560 (J.S. and P.H.) (the content is the responsibility of the authors and does not necessarily represent official views of the NIH) and the BrisSynBio Biosuite (UK Biotechnology and Biological Sciences (BBSRC) and Engineering and Physical Sciences (EPSRC) Research Councils (BB/L01386X/1) and BBSRC ALERT14 initiative (BB/M012107/1).

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J.B., T.P., J.H., P.D., A.J.M.F., K.S., G.G.C., P.K., S.S., D.K., R.K.L., Juliane Schmidt (Adrian), D.L., A.G.B., J.R., L.R., A.M.R., S.D.S.P., A.D.P., M.M., M.P., A.K.B., Peter Brandt, J.Y.-K., E.B., M.G.P.P., Fredrik Björkling, P.S.J., E.S., A.M. and C.J.S. conducted the medicinal chemistry analysis and/or chemical synthesis. J.U.H., E.L., E.I.N., J.K. and S.G. led the safety and/or in vivo efficacy experiments. A.E., M.B. and Pawel Baranczewski led or conducted the physicochemical and ADME testing. L.E., M.-C.T., A.F.A., J.C.J.-C., E.W. and J.M.T. conducted the microbiological experiments under the guidance of M.B.A., M.G., R.C., Fernando Baquero, and T.R.W. K.C., M.E.K., G.W.L., M.S., A.M.R., I.H.N., P.A.L., S.P.M. and J.B. conducted the biochemical or biophysical testing. P.H., M.A.M., T.M.L., James Spencer and J.B. conducted the X-ray crystallography work and analysis. J.B. oversaw all the studies. J.B. and C.J.S. wrote the first draft of the manuscript with subsequent input from all the authors.

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Correspondence to Jürgen Brem or Christopher J. Schofield.

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Competing interests

A patent has been filed concerning the indole carboxylates as MBL inhibitors (WO2017093727A1)29. The inventors may benefit financially from the work. The work described in the manuscript was carried out prior to these employments, which are not relevant to the work. G.W.L. is an employee of Charles River Laboratories, M.S., J.R., L.R., S.P.M., P.S.J. and A.M. are employees of BioAscent Discovery Ltd, A.M.R. is an employee of AstraZeneca and E.B. is an employee of Evotec.

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Brem, J., Panduwawala, T., Hansen, J.U. et al. Imitation of β-lactam binding enables broad-spectrum metallo-β-lactamase inhibitors. Nat. Chem. 14, 15–24 (2022). https://doi.org/10.1038/s41557-021-00831-x

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