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Aspergillomarasmine A overcomes metallo-β-lactamase antibiotic resistance

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Abstract

The emergence and spread of carbapenem-resistant Gram-negative pathogens is a global public health problem. The acquisition of metallo-β-lactamases (MBLs) such as NDM-1 is a principle contributor to the emergence of carbapenem-resistant Gram-negative pathogens that threatens the use of penicillin, cephalosporin and carbapenem antibiotics to treat infections. To date, a clinical inhibitor of MBLs that could reverse resistance and re-sensitize resistant Gram-negative pathogens to carbapenems has not been found. Here we have identified a fungal natural product, aspergillomarasmine A (AMA), that is a rapid and potent inhibitor of the NDM-1 enzyme and another clinically relevant MBL, VIM-2. AMA also fully restored the activity of meropenem against Enterobacteriaceae, Acinetobacter spp. and Pseudomonas spp. possessing either VIM or NDM-type alleles. In mice infected with NDM-1-expressing Klebsiella pneumoniae, AMA efficiently restored meropenem activity, demonstrating that a combination of AMA and a carbapenem antibiotic has therapeutic potential to address the clinical challenge of MBL-positive carbapenem-resistant Gram-negative pathogens.

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Figure 1: AMA inactivates MBLs.
Figure 2: AMA potentiates the activity of meropenem against carbapenem-resistant Gram-negative pathogens.
Figure 3: AMA rescues meropenem activity in vivo.

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Acknowledgements

We thank M. Mulvey (Public Health Agency of Canada) and R. Melano (Public Health Ontario) for clinical strains. We thank L. Rossi for her work in constructing the screening strain. AMA inhibition activity on clinical strains by T.R.W. was funded by the UK Medical Research Council (G1100135). This research was funded by a Canadian Institutes of Health Research Grant (MT-13536), Natural Sciences and Engineering Research Council Grant (237480), and by a Canada Research Chair in Infectious Disease Pathogenesis (to B.K.C.) and Antibiotic Biochemistry (to G.D.W.).

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Contributions

A.M.K., G.D.W., S.A.R.-Y., B.K.C., T.R.W. and N.C.S. designed experiments; G.D.P. and A.M.K. designed and engineered the E. coli strains and screened extracts; A.M.K. synthesized nitrocefin; A.M.K. cloned constructs; A.M.K. and D.T.K. purified enzymes; A.M.K. performed enzyme kinetics; W.W. and A.M.K. fermented WAC-138 and purified AMA; W.W. elucidated AMA structure; A.M.K. performed FIC experiments; D.T.K. performed ICP-MS; S.A.R.-Y. and B.K.C. designed the animal studies; S.A.R.-Y. and A.M.K. performed animal experiments; T.R.W. performed the clinical isolate screen; and A.M.K. and G.D.W. principally wrote the manuscript with input from all.

Corresponding author

Correspondence to Gerard D. Wright.

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The authors declare no competing financial interests.

Extended data figures and tables

Extended Data Figure 1 1H NMR spectrum of AMA in D2O.

Extended Data Figure 2 13C NMR spectrum of AMA in D2O.

Extended Data Figure 3 1H-1H COSY NMR spectrum of AMA in D2O.

Extended Data Figure 4 1H-13C HSQC NMR spectrum of AMA in D2O.

Extended Data Figure 5 1H-13C HMBC NMR spectrum of AMA in D2O.

Extended Data Figure 6 IC50 inhibition profiles for select SBLs and ACE.

a, b, Experiments were done as in Fig. 1b for ACE and CTX-M-15 (black circles), KPC-2 (white circles), and TEM-1 (black squares). Error bars denote standard deviation of at least two replicates.

Extended Data Figure 7 Effects of meropenem dosage on spleen burden.

CD-1 mice were infected with K. pneumoniae N11-2218 by i.p. injection. Mice were treated with either PBS (n = 6) or various doses of meropenem (n = 3 per group) by s.c. injection. Mice were euthanized 48 h after infection, and the bacterial load in the spleen was determined by selective plating. Data are the means with standard error.

Extended Data Table 1 1H and 13C NMR Data of aspergillomarasmine in D2O
Extended Data Table 2 Per cent residual activity following metalloenzyme incubation with AMA
Extended Data Table 3 FIC indices against select clinical isolates of CRE

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King, A., Reid-Yu, S., Wang, W. et al. Aspergillomarasmine A overcomes metallo-β-lactamase antibiotic resistance. Nature 510, 503–506 (2014). https://doi.org/10.1038/nature13445

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