Publications by authors named "Anthony D Verderosa"

11 Publications

  • Page 1 of 1

Antivirulence DsbA inhibitors attenuate serovar Typhimurium fitness without detectable resistance.

FASEB Bioadv 2021 Apr 10;3(4):231-242. Epub 2021 Feb 10.

Institute of Health and Biomedical Innovation School of Biomedical Sciences Queensland University of Technology Herston QLD Australia.

Inhibition of the DiSulfide Bond (DSB) oxidative protein folding machinery, a major facilitator of virulence in Gram-negative bacteria, represents a promising antivirulence strategy. We previously developed small molecule inhibitors of DsbA from K-12 (EcDsbA) and showed that they attenuate virulence of Gram-negative pathogens by directly inhibiting multiple diverse DsbA homologues. Here we tested the evolutionary robustness of DsbA inhibitors as antivirulence antimicrobials against serovar Typhimurium under pathophysiological conditions in vitro. We show that phenylthiophene DsbA inhibitors slow . Typhimurium growth in minimal media, phenocopying . Typhimurium isogenic null mutants. Through passaging experiments, we found that DsbA inhibitor resistance was not induced under conditions that rapidly induced resistance to ciprofloxacin, an antibiotic commonly used to treat infections. Furthermore, no mutations were identified in the gene of inhibitor-treated . Typhimurium, and . Typhimurium virulence remained susceptible to DsbA inhibitors. Our work demonstrates that under in vitro pathophysiological conditions, DsbA inhibitors can have both antivirulence and antibiotic action. Importantly, our finding that DsbA inhibitors appear to be evolutionarily robust offers promise for their further development as next-generation antimicrobials against Gram-negative pathogens.
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http://dx.doi.org/10.1096/fba.2020-00100DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8019255PMC
April 2021

BcfH Is a Trimeric Thioredoxin-Like Bifunctional Enzyme with Both Thiol Oxidase and Disulfide Isomerase Activities.

Antioxid Redox Signal 2021 Apr 12. Epub 2021 Apr 12.

Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Bundoora, Australia.

Thioredoxin (TRX)-fold proteins are ubiquitous in nature. This redox scaffold has evolved to enable a variety of functions, including redox regulation, protein folding, and oxidative stress defense. In bacteria, the TRX-like disulfide bond (Dsb) family mediates the oxidative folding of multiple proteins required for fitness and pathogenic potential. Conventionally, Dsb proteins have specific redox functions with monomeric and dimeric Dsbs exclusively catalyzing thiol oxidation and disulfide isomerization, respectively. This contrasts with the eukaryotic disulfide forming machinery where the modular TRX protein disulfide isomerase (PDI) mediates thiol oxidation and disulfide reshuffling. In this study, we identified and structurally and biochemically characterized a novel Dsb-like protein from termed bovine colonization factor protein H (BcfH) and defined its role in virulence. In the conserved bovine colonization factor () fimbrial operon, the Dsb-like enzyme BcfH forms a trimeric structure, exceptionally uncommon among the large and evolutionary conserved TRX superfamily. This protein also displays very unusual catalytic redox centers, including an unwound α-helix holding the redox active site and a proline instead of the conserved -proline active site loop. Remarkably, BcfH displays both thiol oxidase and disulfide isomerase activities contributing to fimbrial biogenesis. Typically, oligomerization of bacterial Dsb proteins modulates their redox function, with monomeric and dimeric Dsbs mediating thiol oxidation and disulfide isomerization, respectively. This study demonstrates a further structural and functional malleability in the TRX-fold protein family. BcfH trimeric architecture and unconventional catalytic sites permit multiple redox functions emulating in bacteria the eukaryotic PDI dual oxidoreductase activity.
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http://dx.doi.org/10.1089/ars.2020.8218DOI Listing
April 2021

A high-throughput cell-based assay pipeline for the preclinical development of bacterial DsbA inhibitors as antivirulence therapeutics.

Sci Rep 2021 Jan 15;11(1):1569. Epub 2021 Jan 15.

Institute of Health and Biomedical Innovation and Centre for Immunology and Infection Control, School of Biomedical Sciences, Queensland University of Technology, Brisbane, QLD, 4059, Australia.

Antibiotics are failing fast, and the development pipeline remains alarmingly dry. New drug research and development is being urged by world health officials, with new antibacterials against multidrug-resistant Gram-negative pathogens as the highest priority. Antivirulence drugs, which inhibit bacterial pathogenicity factors, are a class of promising antibacterials, however, their development is stifled by lack of standardised preclinical testing akin to what guides antibiotic development. The lack of established target-specific microbiological assays amenable to high-throughput, often means that cell-based testing of virulence inhibitors is absent from the discovery (hit-to-lead) phase, only to be employed at later-stages of lead optimization. Here, we address this by establishing a pipeline of bacterial cell-based assays developed for the identification and early preclinical evaluation of DsbA inhibitors, previously identified by biophysical and biochemical assays. Inhibitors of DsbA block oxidative protein folding required for virulence factor folding in pathogens. Here we use existing Escherichia coli DsbA inhibitors and uropathogenic E. coli (UPEC) as a model pathogen, to demonstrate that the combination of a cell-based sulfotransferase assay and a motility assay (both DsbA reporter assays), modified for a higher throughput format, can provide a robust and target-specific platform for the identification and evaluation of DsbA inhibitors.
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http://dx.doi.org/10.1038/s41598-021-81007-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7810732PMC
January 2021

An Reconstructed Human Skin Equivalent Model to Study the Role of Skin Integration Around Percutaneous Devices Against Bacterial Infection.

Front Microbiol 2020 14;11:670. Epub 2020 May 14.

Infection and Immunity Research Program, Institute of Health and Biomedical Innovation, School of Biomedical Sciences, Faculty of Health, Queensland University of Technology, Brisbane, QLD, Australia.

Percutaneous devices are a key technology in clinical practice, used to connect internal organs to external medical devices. Examples include prosthesis, catheters and electrical drivelines. Percutaneous devices breach the skin's natural barrier and create an entry point for pathogens, making device infections a widespread problem. Modification of the percutaneous implant surface to increase skin integration with the aim to reduce subsequent infection is attracting a great deal of attention. While novel surfaces have been tested in various models used to study skin integration around percutaneous devices, no skin model has been reported, for the study of bacterial infection around percutaneous devices. Here, we report the establishment of an human skin equivalent model for driveline infections caused by , the most common cause of driveline-related infections. Three types of mock drivelines manufactured using melt electrowriting (smooth or porous un-seeded and porous pre-seeded with human fibroblasts) were implanted in human skin constructs and challenged with Our results show a high and stable load of in association with the skin surface and no signs of -induced tissue damage. Furthermore, our results demonstrate that bacterial migration along the driveline surface occurs in micro-gaps caused by insufficient skin integration between the driveline and the surrounding skin consistent with clinical reports from explanted patient drivelines. Thus, the human skin-driveline infection model presented here provides a clinically-relevant and versatile experimental platform for testing novel device surfaces and infection therapeutics.
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http://dx.doi.org/10.3389/fmicb.2020.00670DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7240036PMC
May 2020

Bacterial Biofilm Eradication Agents: A Current Review.

Front Chem 2019 28;7:824. Epub 2019 Nov 28.

School of Chemistry, Physics, and Mechanical Engineering, Queensland University of Technology, Brisbane, QLD, Australia.

Most free-living bacteria can attach to surfaces and aggregate to grow into multicellular communities encased in extracellular polymeric substances called biofilms. Biofilms are recalcitrant to antibiotic therapy and a major cause of persistent and recurrent infections by clinically important pathogens worldwide (e.g., , and ). Currently, most biofilm remediation strategies involve the development of biofilm-inhibition agents, aimed at preventing the early stages of biofilm formation, or biofilm-dispersal agents, aimed at disrupting the biofilm cell community. While both strategies offer some clinical promise, neither represents a direct treatment and eradication strategy for established biofilms. Consequently, the discovery and development of biofilm eradication agents as comprehensive, stand-alone biofilm treatment options has become a fundamental area of research. Here we review our current understanding of biofilm antibiotic tolerance mechanisms and provide an overview of biofilm remediation strategies, focusing primarily on the most promising biofilm eradication agents and approaches. Many of these offer exciting prospects for the future of biofilm therapeutics for a large number of infections that are currently refractory to conventional antibiotics.
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http://dx.doi.org/10.3389/fchem.2019.00824DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6893625PMC
November 2019

Nitroxide Functionalized Antibiotics Are Promising Eradication Agents against Staphylococcus aureus Biofilms.

Antimicrob Agents Chemother 2019 12 20;64(1). Epub 2019 Dec 20.

Institute of Health and Biomedical Innovation, Queensland University of Technology, Brisbane, Queensland, Australia

Treatment of biofilm-related infections represents an important medical challenge worldwide, as biofilms, even those involving drug-susceptible strains, are highly refractory to conventional antibiotic therapy. Nitroxides were recently shown to induce the dispersal of Gram-negative biofilms , but their action against Gram-positive bacterial biofilms remains unknown. Here, we demonstrate that the biofilm dispersal activity of nitroxides extends to , a clinically important Gram-positive pathogen. Coadministration of the nitroxide CTEMPO (4-carboxy-2,2,6,6-tetramethylpiperidin-1-yloxyl) with ciprofloxacin significantly improved the biofilm eradication activity of the antibiotic against Moreover, covalently linking the nitroxide to the antibiotic moiety further reduced the ciprofloxacin minimal biofilm eradication concentration. Microscopy analysis revealed that fluorescent nitroxide-antibiotic hybrids could penetrate biofilms and enter cells localized at the surface and base of the biofilm structure. No toxicity to human cells was observed for the nitroxide CTEMPO or the nitroxide-antibiotic hybrids. Taken together, our results show that nitroxides can mediate the dispersal of Gram-positive biofilms and that dual-acting biofilm eradication antibiotics may provide broad-spectrum therapies for the treatment of biofilm-related infections.
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http://dx.doi.org/10.1128/AAC.01685-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7187575PMC
December 2019

Eradicating uropathogenic biofilms with a ciprofloxacin-dinitroxide conjugate.

Medchemcomm 2019 May 25;10(5):699-711. Epub 2019 Feb 25.

Queensland University of Technology , School of Chemistry, Physics and Mechanical Engineering , 2 George St , Brisbane , Queensland 4001 , Australia . Email:

Urinary tract infections (UTIs) are amongst the most common and prevalent infectious diseases worldwide, with uropathogenic (UPEC) reported as the main causative pathogen. Fluoroquinolone antibiotics are commonly used to treat UTIs but for infections involving UPEC biofilms, which are commonly associated with catheter use and recurrent episodes, ciprofloxacin is often ineffective. Here we report the development of a ciprofloxacin-dinitroxide (CDN) conjugate with potent UPEC biofilm-eradication activity. CDN exhibited a 2-fold increase in potency over the parent antibiotic ciprofloxacin against UPEC biofilms. Moreover, CDN resulted in almost complete UPEC biofilm cell eradication (99.7%) at concentrations as low as 12.5 μM, and significantly potentiated ciprofloxacin's biofilm-eradication activity against UPEC upon co-administration. The biofilm-eradication activity of CDN highlights the potential of nitroxide functionalized antibiotics as a promising strategy for the treatment of biofilm-related UTIs.
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http://dx.doi.org/10.1039/c9md00062cDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6533797PMC
May 2019

Profluorescent Fluoroquinolone-Nitroxides for Investigating Antibiotic⁻Bacterial Interactions.

Antibiotics (Basel) 2019 Mar 4;8(1). Epub 2019 Mar 4.

Institute of Health and Biomedical Innovation, School of Biomedical Sciences, Queensland University of Technology, Brisbane, QLD 4006, Australia.

Fluorescent probes are widely used for imaging and measuring dynamic processes in living cells. Fluorescent antibiotics are valuable tools for examining antibiotic⁻bacterial interactions, antimicrobial resistance and elucidating antibiotic modes of action. Profluorescent nitroxides are 'switch on' fluorescent probes used to visualize and monitor intracellular free radical and redox processes in biological systems. Here, we have combined the inherent fluorescent and antimicrobial properties of the fluoroquinolone core structure with the fluorescence suppression capabilities of a nitroxide to produce the first example of a profluorescent fluoroquinolone-nitroxide probe. Fluoroquinolone-nitroxide (FN) exhibited significant suppression of fluorescence (>36-fold), which could be restored via radical trapping (fluoroquinolone-methoxyamine ) or reduction to the corresponding hydroxylamine . Importantly, FN was able to enter both Gram-positive and Gram-negative bacterial cells, emitted a measurable fluorescence signal upon cell entry (switch on), and retained antibacterial activity. In conclusion, profluorescent nitroxide antibiotics offer a new powerful tool for visualizing antibiotic⁻bacterial interactions and researching intracellular chemical processes.
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http://dx.doi.org/10.3390/antibiotics8010019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6466543PMC
March 2019

Moraxella catarrhalis NucM is an entry nuclease involved in extracellular DNA and RNA degradation, cell competence and biofilm scaffolding.

Sci Rep 2019 02 22;9(1):2579. Epub 2019 Feb 22.

Institute for Glycomics, Griffith University, Gold Coast, Queensland, 4215, Australia.

Moraxella catarrhalis is a host-adapted bacterial pathogen that causes otitis media and exacerbations of chronic obstructive pulmonary disease. This study characterises the conserved M. catarrhalis extracellular nuclease, a member of the ββα metal finger family of nucleases, that we have named NucM. NucM shares conserved sequence motifs from the ββα nuclease family, including the DRGH catalytic core and Mg co-ordination site, but otherwise shares little primary sequence identity with other family members, such as the Serratia Nuc and pneumococcal EndA nucleases. NucM is secreted from the cell and digests linear and circular nucleic acid. However, it appears that a proportion of NucM is also associated with the cell membrane and acts as an entry nuclease, facilitating transformation of M. catarrhalis cells. This is the first example of a ββα nuclease in a Gram negative bacteria that acts as an entry nuclease. In addition to its role in competence, NucM affects cell aggregation and biofilm formation by M. catarrhalis, with ΔnucM mutants having increased biofilm biomass. NucM is likely to increase the ability of cells to survive and persist in vivo, increasing the virulence of M. catarrhalis and potentially affecting the behaviour of other pathogens that co-colonise the otorhinolaryngological niche.
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http://dx.doi.org/10.1038/s41598-019-39374-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6384898PMC
February 2019

Ciprofloxacin-nitroxide hybrids with potential for biofilm control.

Eur J Med Chem 2017 Sep 28;138:590-601. Epub 2017 Jun 28.

ARC Centre of Excellence for Free Radical Chemistry and Biotechnology, Faculty of Science and Engineering, Queensland University of Technology, Queensland 4001, Australia. Electronic address:

As bacterial biofilms display extreme tolerance to conventional antibiotic treatments, it has become imperative to develop new antibacterial strategies with alternative mechanisms of action. Herein, we report the synthesis of a series of ciprofloxacin-nitroxide conjugates and their corresponding methoxyamine derivatives in high yield. This was achieved by linking various nitroxides or methoxyamines to the secondary amine of the piperazine ring of ciprofloxacin using amide bond coupling. Biological evaluation of the prepared compounds on preformed P. aeruginosa biofilms in flow cells revealed substantial dispersal with ciprofloxacin-nitroxide hybrid 25, and virtually complete killing and removal (94%) of established biofilms in the presence of ciprofloxacin-nitroxide hybrid 27. Compounds 25-28 were shown to be non-toxic in both human embryonic kidney 293 (HEK 293) cells and human muscle rhabdomyosarcoma (RD) cells at concentrations up to 40 μM. Significantly, these hybrids demonstrate the potential of antimicrobial-nitroxide agents to overcome the resistance of biofilms to antimicrobials via stimulation of biofilm dispersal or through direct cell killing.
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http://dx.doi.org/10.1016/j.ejmech.2017.06.058DOI Listing
September 2017

Synthesis and Evaluation of Ciprofloxacin-Nitroxide Conjugates as Anti-Biofilm Agents.

Molecules 2016 Jun 27;21(7). Epub 2016 Jun 27.

ARC Centre of Excellence for Free Radical Chemistry and Biotechnology, Faculty of Science and Engineering, Queensland University of Technology, Queensland 4001, Australia.

As bacterial biofilms are often refractory to conventional antimicrobials, the need for alternative and/or novel strategies for the treatment of biofilm related infections has become of paramount importance. Herein, we report the synthesis of novel hybrid molecules comprised of two different hindered nitroxides linked to the piperazinyl secondary amine of ciprofloxacin via a tertiary amine linker achieved utilising reductive amination. The corresponding methoxyamine derivatives were prepared alongside their radical-containing counterparts as controls. Subsequent biological evaluation of the hybrid compounds on preformed P. aeruginosa flow cell biofilms divulged significant dispersal and eradication abilities for ciprofloxacin-nitroxide hybrid compound 10 (up to 95% eradication of mature biofilms at 40 μM). Importantly, these hybrids represent the first dual-action antimicrobial-nitroxide agents, which harness the dispersal properties of the nitroxide moiety to circumvent the well-known resistance of biofilms to treatment with antimicrobial agents.
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http://dx.doi.org/10.3390/molecules21070841DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6273952PMC
June 2016