Resensitizing carbapenem- and colistin-resistant bacteria to antibiotics using auranofin

Global emergence of Gram-negative bacteria carrying the plasmid-borne resistance genes, blaMBL and mcr, raises a significant challenge to the treatment of life-threatening infections by the antibiotics, carbapenem and colistin (COL). Here, we identify an antirheumatic drug, auranofin (AUR) as a dual inhibitor of metallo-β-lactamases (MBLs) and mobilized colistin resistance (MCRs), two resistance enzymes that have distinct structures and substrates. We demonstrate that AUR irreversibly abrogates both enzyme activity via the displacement of Zn(II) cofactors from their active sites. We further show that AUR synergizes with antibiotics on killing a broad spectrum of carbapenem and/or COL resistant bacterial strains, and slows down the development of β-lactam and COL resistance. Combination of AUR and COL rescues all mice infected by Escherichia coli co-expressing MCR-1 and New Delhi metallo-β-lactamase 5 (NDM-5). Our findings provide potential therapeutic strategy to combine AUR with antibiotics for combating superbugs co-producing MBLs and MCRs.


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MS data was processed using FlexAnalysis (version 1.2, Bruker Daltonics).The diffraction data were reduced with XDS54. The Phaser from CCP4 suite (7.0.078) and Phenix (version 1.15-3459) were used for data refinement and finalization. Cycles of refinement with the anomalous data and with careful manual rebuilding were done by using Refmac 5.8.0135 and Coot version 6, respectively. TLS refinement was used in the later stages of data processing. The final models were analyzed with MolProbity version 4.4. Structural alignment was done over C! residues using DaliLite version 5. All of the structural illustrations were generated using the software PyMOL1.8.0.0. Image J (1.52a) was used to quantify the signals of each band for cellular thermal shift analysis. All other analyses were carried out using GraphPad Prism software 8.0 (GraphPad Software, Inc., La Jolla, CA) All data are available in the main manuscript or in the Supplementary Information, except that the structure factors for Au-NDM-1, Zn-NDM-1, Au-MCR-1-S, Zn-MCR-1-S, and apo-MCR-1-S are deposited at Protein Data Bank with accessing codes of 6LHE, 5ZGE, 6LI6, 6LI4 and 6LI5, respectively.

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All studies must disclose on these points even when the disclosure is negative. The sample size and n/group was chosen based on similar experiments conducted by previous studies (refs 31, 34 and 43) using the same or similar mouse strains. The sample size of n= 4 each group was chosen for CFU counting in animal tissue and n= 6 for animal survival studies. For in vitro biochemical studies, three biological repeats, followed by repeating the experiments twice for confirmation were widely accepted and used in published papers.
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All the experiments were subjected to three biological replicates unless specified. All attempts at replication were successful.
All the animals were numbered and allocated into groups using a simple randomization of excel-generated random numbers. To avoid biases, we also assured that different treatments were performed on the same day. All the animals were randomized to cages for each experiment and had free access to food and water.
Mice were blindly allocated to each group. The sample classification were replaced by simple marks during data collection and analysis. The investigators were not blinded to the allocation during experiments and outcome assessment. Data collection and analysis were performed by the same people.