The Development of Photo-activated Antimicrobial Dyes Against Opportunistic Infections

Jalab, Murhaf (2020) The Development of Photo-activated Antimicrobial Dyes Against Opportunistic Infections. Doctoral thesis, University of Central Lancashire.

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Microbial infections cause a major health threat and the growing incidence of invasive and opportunistic infections is usually associated with high rates of morbidity and mortality. Approximately 300,000 patients a year in England are affected by a healthcare-associated infection as a result of care within the NHS
and the NHS cost is estimated at approximately £1 billion a year. The treatment of infectious diseases is one of the most challenging problems in medicine due
to the emergence of microbial resistance, side effects and spectrum of activity.
Therefore, there is an obvious and urgent need to develop new and effective antimicrobial strategies. A possible alternative to traditional antimicrobial drugs is
photodynamic therapy (aPDT), which depends on the activation of a photosensitiser (PS) by a visible light source to produce reactive oxygen species
(ROS) and singlet oxygen, which can inactivate and kill microbial cells.
A library of novel photoactivatable compounds, based on acridine, flavin, acridine-isoalloxazine and anthraquinone dyes, has been characterised to
quantify singlet oxygen release following activation by blue light for 10 and 20 minutes. Candidate compounds were then screened, using the European Committee for Antimicrobial Susceptibility Testing (EUCAST) microbroth dilution method, for antimicrobial activity against a range of clinically important fungi, including Candida albicans (C. albicans), Saccharomyces cerevisiae (S. cerevisiae) and Aspergillus spp., and medically important bacteria, including Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus).
The chemical results demonstrated the ability of the studied compounds to generate singlet oxygen upon exposure to blue light. Determination of the
minimum inhibitory concentration (MIC) has identified a number of novel candidate compounds with activity against fungi and bacteria. These compounds
were further investigated to determine their mechanism of action using a hog1 and msn2/4 genomic deletion strain of S. cerevisiae. The findings suggest that the general stress HOG pathway (High-osmolarity glycerol) has a limited role in the cellular response to the compounds. However, msn2/4 deletion strains, which encode key transcription factors for the oxidative stress response, showed an increase in sensitivity suggesting these compounds are inhibiting microbial cell growth via oxidative stress. Further characterisation of the compounds indicates that these photo-activated compounds do not develop resistance in fungal and bacterial species following three repeated exposures. Additionally, a significant
effect against Candida albicans biofilm was shown for three tested PDT compounds. Finally, the tested compounds showed toxicity against HeLa cells in
the absence and presence of light. This suggests that, in their current form, they are not ideal compounds for clinical use.
The results from this project support ongoing work in this field which may help in the development of a new arsenal of antimicrobial drugs.

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