There is a global requirement to identify novel treatments to combat multidrug-resistant (MDR) Gram-negative bacteria. Incidence of infections caused by these pathogens is rising and effective treatment is problematic resulting in increased morbidity and mortality . Few new treatment options are in the drug ‘pipeline’ so the use of combination treatments with two or more antibiotics is a realistic option. Combinations consisting of multiple antibiotics are employed clinically and have been extensively studied. In contrast, combinations of approved drugs (that are not in the first instance antimicrobial) with antibiotics that could act as ‘resistance breakers’ is under-explored, particularly with in vivo studies . The aim of this research is to employ an invertebrate infection model, Galleria mellonella larvae, to screen the efficacy of a range of ‘repurposed’ approved drugs with existing antibiotics against MDR strains of Pseudomonas aeruginosa, Klebsiella pneumoniae and Escherichia coli. The ‘repurposed’ drugs that will be employed have been selected on the basis of a screen of published literature showing evidence of enhanced activity in combinations in vitro. This type of in vivo efficacy screen is only possible because of the use of an invertebrate infection model . Screening for efficacious combinations in mammalian models would not be possible due to ethical constraints and costs. Any efficacious combinations identified by this study would be the subject of further funding applications to assess possible clinical utility.
Hypotheses to be investigated – Combination therapies consisting of ‘repurposed’ approved drugs with existing antibiotics can act as ‘resistance breakers’ and effectively treat real infections caused by multi-drug resistant Gram-negative bacteria.
Aims of the proposed research project – The aim of this study is to identify combination treatments of a range of ‘repurposed’ drugs with existing antibiotics that show enhanced efficacy versus real infections in G. mellonella larvae. After conducting a literature search on ‘repurposed’ drugs with potential activity versus MDR Gram-negative pathogens, a number of compounds were identified that possessed inhibitory activity, mainly in vitro, and merit further investigation in vivo against real infections and in combination with existing antibiotics. The selected compounds include:
1. Nucleoside and nucleobase analogue drugs (NNADs), including 5-fluorouracil, 5-flucytosine, theophylline, zidovudine and caffeine .
2. Zn2+-dependent inhibitors of bacterial metallo-β-lactamases including metal chelating drugs such as captopril, thiorphan, tiopronin and dimercaprol .
3. Bismuth containing drugs that inhibit metallo-β-lactamases such as bismuth subcitrate and ranitidine bismuth citrate .
4. Non-steroidal anti-inflammatory drugs (NSAID) such as celecoxib  and meloxicam .
5. Immunomodulatory drugs such as glatiramer acetate used for treatment of multiple sclerosis 
6. Thioredoxin inhibitors such as auranofin (used for rheumatoid arthritis), ebselen (anti-inflammatory) and PX-12 .
These compounds will be tested in combination with a range of existing antibiotics that are adversely affected by resistance mechanisms in clinical practice. Infections will be induced by characterized multidrug resistant strains (in addition to clinical isolates provided by Dr Ben Parcell, Consultant Microbiologist, Ninewells Hospital, NHS Tayside) of Pseudomonas aeruginosa, Klebsiella pneumoniae and Escherichia coli. Dr Parcell has collected MDR isolates from 23 patients that have the added advantage of associated healthcare informatics data including antibiotic regimens and doses used to treat the infections associated with these strains. This data will allow the comparison of real antibiotic efficacy in patients with that observed in G. mellonella larvae. This comparison will provide additional important insight into the suitability and relevance of employing the G. mellonella infection model for developing novel antibiotic therapies for use in patients.
Furthermore, any combination(s) identified to have enhanced efficacy will be investigated to determine their inhibitory mode of action compared to the constituent monotherapies.
Informal enquiries can be made to Dr Peter Coote via e-mail or telephone.
Email: [email protected]
Tel: (44) (0)1334 463406
 Centers for Disease Control and Prevention (2013) Antibiotic resistance threats in the United States, 2013. Atlanta: CDC, 2013. http://www.cdc.gov/drugresistance/threat-report-2013/pdf/ar-threats- 2013-508.pdf. Accessed 11 November 2015.  Ejim L, et al. (2011) Combinations of antibiotics and non-antibiotic drugs enhance antimicrobial efficacy. Nature Chem Biol 7: 348-350.  Desbois A, Coote P (2012) Utility of greater wax moth larva (Galleria mellonella) for evaluating the toxicity and efficacy of new antimicrobial agents. Adv Appl Microbiol 2012:78: 25-53.  Yssel AEJ et al. (2017) Repurposing of nucleoside- and nucleobase-derivative drugs as antibiotics and biofilm inhibitors. J Antimic Chemother 72: 2156-2170.  Rotondo CM, Wright GD (2017) Inhibitors of metallo-β-lactamases. Curr Opin Microbiol 39:96-105.  Wang R, et al. (2018) Bismuth antimicrobial drugs serve as broad-spectrum metallo-β-lactamase inhibitors. Nat Comm 9: 439-450.  Annamanedi M et al. (2017) Celecoxib enhances the efficacy of low-dose antibiotic treatment against polymicrobial sepsis in mice and clinical isolates of ESKAPE pathogens. Front Microbiol 8: 805.  She P et al. (2018) Meloxicam inhibits bofilm formation and enhances antimicrobial agents efficacy by Pseudomonas aeruginosa. Microbiol Open 7:e545.  Christiansen SH et al. (2017) The immunomodulatory drug glatiramer acetate is also an effective antimicrobial agent that kills Gram-negative bacteria. Scientif Rep 7: 15653.  May HC et al. (2018) Repurposing auranofin, ebselen and PX-12 as antimicrobial agents targeting the thioredoxin system. Front Microbiol 9: 336.