Potential novel antimicrobial therapies for burn wounds: peptides and cold plasma

Gabriëlle Sherella Dijksteel

    Research output: PhD ThesisPhD-Thesis - Research and graduation internal

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    Abstract

    More than 40 AMPs have been studied in clinical trials but only a few AMPs have been approved for clinical use against bacterial infections. Based on the literature review (Chapter 2), clinical trials using AMPs have been discontinued due to unexpected toxicity or unfavorable pharmacokinetics of peptides. However, the majority of AMPs were not approved due to lack of efficacy or superiority of AMPs over conventional antibiotics or treatments. To improve the performance of AMPs, several strategies are available such as chemical modifications, the preparation of delivery systems and specifically targeted AMPs (STAMPs), combination therapy and counter-ion selection during the final step of AMP synthesis. In addition to these strategies, an increasing number of AMPs might reach the clinic if clinical trial regulatory bodies could shift their focus from superiority trials to equivalence or non-inferiority trials (i.e. clinical trials whereby the performance of AMPs should be similar to that of antibiotics or standard treatments). One of the causes of unexpected and unfavorable performance of AMPs in trials might be the overestimated efficacy of AMPs due to inaccurate in vitro testing. To accurately and appropriately determine efficacy of antimicrobials, it is important to neutralize the remaining active antimicrobials before microbiological assessment of the number of surviving bacteria. Without effective neutralization, efficacy of antimicrobials could be overestimated due to ongoing bacterial killing during sample preparations. We studied the ability of sodium polyanethol sulfonate (SPS) to neutralize the bactericidal activity of several antimicrobials (Chapter 3). SPS effectively inactivated positively charge antimicrobials and therefore, we have used this neutralizing agent in all subsequent efficacy tests of the positively charged peptide SAAP-148. Efficacy of SAAP-148 against methicillin-resistant Staphylococcus aureus (MRSA) was determined using an excision wound model in rats. In Chapter 4, we have shown that SAAP-148 did not eradicate MRSA in this model. The potential factors that may have contributed to the limited bactericidal effect of SAAP-148 were investigated and we found that components in the wound micro-environment were most likely responsible for the limited bactericidal effect. Higher antimicrobial concentrations of SAAP-148 were required to eradicate bacteria effectively in the presence of ex vivo human skin, blood plasma, eschar and skin extract as compared to PBS. The impact of these biological environments on the bactericidal activity of SAAP-148 was dependent on the incubation time (Chapter 5). A short incubation time of 1 h in a biological environment did not affect the bactericidal activity of SAAP-148, whereas a 24 h incubation time considerably reduced its bactericidal efficacy. This was not caused by proteolytic degradation but presumably by protein binding, in particular binding of SAAP-148 to blood proteins resulting in a low bio-availability of the peptide to combat bacteria. Furthermore, SAAP-148 was toxic for human skin cells in vitro but not in ex vivo human skin, suggesting that like efficacy, the safety of AMPs should be assessed in environments that simulate the clinical situation. CP from the flexible surface DBD device did not eradicate Pseudomonas aeruginosa in excision wound models in rats (Chapter 6). This was most likely related to the micro-wound environment consisting of bacteria in biofilms and the presence of wound fluids, which can reduce the bactericidal efficacy of CP. Nonetheless, exposure to this CP for 2 min completely eradicated P. aeruginosa in vitro. Additionally, exposure to CP for 4-6 min did not induce mutations, apoptosis and DNA damage in human cells, or affect the wound healing process ex vivo. This suggests that CP from the flexible surface DBD device is safe. In Chapter 7 we discuss the results of the therapies investigated in this thesis and describe future possibilities for clinical application.
    Original languageEnglish
    QualificationPhD
    Awarding Institution
    • Vrije Universiteit Amsterdam
    Supervisors/Advisors
    • Middelkoop, E., Supervisor, -
    • Ulrich, M.W., Supervisor, -
    • Boekema, Bouke, Co-supervisor, -
    Award date24 Mar 2021
    Print ISBNs9789464231298
    Publication statusPublished - 24 Mar 2021

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