Disulfiram and Copper Ions Kill Mycobacterium tuberculosis in a Synergistic Manner

Dalecki, Alex G.; Haeili, Mehri; Shah, Silvi; Speer, Alexander; Niederweis, Michael; Kutsch, Olaf; Wolschendorf, Frank

doi:10.1128/aac.00692-15

Cited by 83 publications

(70 citation statements)

References 68 publications

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“…To exclude the possibility of medium-specific effects and to determine the full extent of the synergy between Cu and 8HQ, we repeated the experiment in Hartmans-de Bont minimal medium (HdB SM ), which permits the growth of M. tuberculosis in the absence of potentially interfering albumin supplements. Albumin is known to bind Cu and other metal ions and may therefore reduce the sensitivity of the assay (35,45). As expected, the Cu dependency was much more pronounced, yielding an MIC of ϳ0.16 M for M. tuberculosis H37Rv (Fig.…”

Section: Resultsmentioning

confidence: 52%

“…The activity is therefore more likely related to the intrinsic properties of the associated metal ion (e.g., antibacterial activity), with the ligand being responsible for the disguise and delivery of the proper metal ion to a sensitive target site. The entire complex could then resemble a Trojan horse, as previously proposed by us in the context of the Cu-active drug disulfiram (45). In such a scenario, the complex would be expected to act on targets within the cell.…”

Section: Discussionmentioning

confidence: 89%

“…As a class, Cudependent inhibitors are known to act through a wide range of mechanisms. They can act as ionophores, overloading cells with Cu ions (e.g., 8HQ against Cryptococcus neoformans [28]); as liberators, growing the pool of bioavailable (and toxic) Cu without an overall increase (e.g., disulfiram against M. tuberculosis [45]); and even as traditional antibiotics, disrupting specific enzymatic targets (e.g., glyoxal-bis[N(4)-methylthiosemicarbazonato]copper(II) complex against Neisseria gonorrhoeae [55]). To gain insight into the mechanism of mycobacterial inhibition, we utilized PhenGreen FL (PGFL), a fluorescent metal sensor, to determine net changes in the amounts of cell-associated labile metal ions upon treatment with Cu or 8HQ alone or cotreatment with Cu ions and 8HQ.…”

Section: Resultsmentioning

confidence: 99%

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8-Hydroxyquinolines Are Boosting Agents of Copper-Related Toxicity in Mycobacterium tuberculosis

Shah

Dalecki

Malalasekera

et al. 2016

Antimicrob Agents Chemother

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c Copper (Cu) ions are likely the most important immunological metal-related toxin utilized in controlling bacterial infections. Impairment of bacterial Cu resistance reduces viability within the host. Thus, pharmacological enhancement of Cu-mediated antibacterial toxicity may lead to novel strategies in drug discovery and development. Screening for Cu toxicity-enhancing antibacterial molecules identified 8-hydroxyquinoline (8HQ) to be a potent Cu-dependent bactericidal inhibitor of Mycobacterium tuberculosis. The MIC of 8HQ in the presence of Cu was 0.16 M for replicating and nonreplicating M. tuberculosis cells. We found 8HQ's activity to be dependent on the presence of extracellular Cu and to be related to an increase in cell-associated labile Cu ions. Both findings are consistent with 8HQ acting as a Cu ionophore. Accordingly, we identified the 1:1 complex of 8HQ and Cu to be its active form, with Zn, Fe, or Mn neither enhancing nor reducing its Cu-specific action. This is remarkable, considering that the respective metal complexes have nearly identical structures and geometries. Finally, we found 8HQ to kill M. tuberculosis selectively within infected primary macrophages. Given the stark Cu-dependent nature of 8HQ activity, this is the first piece of evidence that Cu ions within macrophages may bestow antibacterial properties to a Cu-dependent inhibitor of M. tuberculosis. In conclusion, our findings highlight the metal-binding ability of the 8-hydroxyquinoline scaffold to be a potential focus for future medicinal chemistry and highlight the potential of innate immunity-inspired screening platforms to reveal molecules with novel modes of action against M. tuberculosis.

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Section: Resultsmentioning

confidence: 52%

Section: Discussionmentioning

confidence: 89%

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

8-Hydroxyquinolines Are Boosting Agents of Copper-Related Toxicity in Mycobacterium tuberculosis

Shah

Dalecki

Malalasekera

et al. 2016

Antimicrob Agents Chemother

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“…Furthermore, auxotrophic Mtb strains have already been used in well-established drug evaluation studies for Mtb. [19][20][21] For our experiments, we utilized a gfp-expressing strain of Mtb mc 2 6206 to infect THP-1 cells, a monocyte cell line widely used in Mtb research. 22,23 The ability of this Mtb strain to express GFP allows for the visual control of macrophage phagocytosis efficacy and direct quantification of infection levels.…”

Section: Resultsmentioning

confidence: 99%

A Macrophage Infection Model to Predict Drug Efficacy AgainstMycobacterium Tuberculosis

Schaaf

Hayley

Speer

et al. 2016

ASSAY and Drug Development Technologies

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In the last 40 years, only a single new antituberculosis drug was FDA approved. New tools that improve the drug development process will be essential to accelerate the development of next-generation antituberculosis drugs. The drug development process seems to be hampered by the inefficient transition of initially promising hits to candidate compounds that are effective in vivo. In this study, we introduce an inexpensive, rapid, and BSL-2 compatible infection model using macrophage-passaged Mycobacterium tuberculosis (Mtb) that forms densely packed Mtb/macrophage aggregate structures suitable for drug efficacy testing. Susceptibility to antituberculosis drugs determined with this Mtb/macrophage aggregate model differed from commonly used in vitro broth-grown single-cell Mtb cultures. Importantly, altered drug susceptibility correlated well with the reported ability of the respective drugs to generate high tissue and cerebrospinal fluid concentrations relative to their serum concentrations, which seems to be the best predictors of in vivo efficacy. Production of these Mtb/macrophage aggregates could be easily scaled up to support throughput efforts. Overall, its simplicity and scalability should make this Mtb/macrophage aggregate model a valuable addition to the currently available Mtb drug discovery tools.

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“…These compounds are highly inhibitory in the presence of copper, yet impotent in its absence. Example inhibitors have been described for Gram negatives 17 , Gram positives 18 , and mycobacteria 19 , as well as eukaryotic targets including pathogenic fungi 20 and cancer cells 21–23 . The broad range of potential targetable pathogens, coupled with the wide array of novel mechanisms of action, has generated interest in exploiting copper’s antibiotic properties 24, 25 .…”

Section: Introductionmentioning

confidence: 99%

Combinatorial phenotypic screen uncovers unrecognized family of extended thiourea inhibitors with copper-dependent anti-staphylococcal activity

et al. 2016

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The continuous rise of multi-drug resistant pathogenic bacteria has become a significant challenge for the health care system. In particular, novel drugs to treat infections of methicillin-resistant Staphylococcus aureus strains (MRSA) are needed, but traditional drug discovery campaigns have largely failed to deliver clinically suitable antibiotics. More than simply new drugs, new drug discovery approaches are needed to combat bacterial resistance. The recently described phenomenon of copper-dependent inhibitors has galvanized research exploring the use of metal-coordinating molecules to harness copper’s natural antibacterial properties for therapeutic purposes. Here, we describe the results of the first concerted screening effort to identify copper-dependent inhibitors of Staphylococcus aureus. A standard library of 10,000 compounds was assayed for anti-staphylococcal activity, with hits defined as those compounds with a strict copper-dependent inhibitory activity. A total of 53 copper-dependent hit molecules were uncovered, similar to the copper independent hit rate of a traditionally executed campaign conducted in parallel on the same library. Most prominent was a hit family with an extended thiourea core structure, termed the NNSN motif. This motif resulted in copper-dependent and copper-specific S. aureus inhibition, while simultaneously being well tolerated by eukaryotic cells. Importantly, we could demonstrate that copper binding by the NNSN motif is highly unusual and likely responsible for the promising biological qualities of these compounds. A subsequent chemoinformatic meta-analysis of the ChEMBL chemical database confirmed the NNSNs as an unrecognized staphylococcal inhibitor, despite the family’s presence in many chemical screening libraries. Thus, our copper-biased screen has proven able to discover inhibitors within previously screened libraries, offering a mechanism to reinvigorate exhausted molecular collections.

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Disulfiram and Copper Ions Kill Mycobacterium tuberculosis in a Synergistic Manner

Cited by 83 publications

References 68 publications

8-Hydroxyquinolines Are Boosting Agents of Copper-Related Toxicity in Mycobacterium tuberculosis

8-Hydroxyquinolines Are Boosting Agents of Copper-Related Toxicity in Mycobacterium tuberculosis

A Macrophage Infection Model to Predict Drug Efficacy AgainstMycobacterium Tuberculosis

Combinatorial phenotypic screen uncovers unrecognized family of extended thiourea inhibitors with copper-dependent anti-staphylococcal activity

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