Combating Bacterial Biofilm Infections on Medical Implants by Using Substrate Mediated Enzyme Prodrug Therapy

Student: Signe Maria Nielsen, Raoul Walter and Rikke Christiansen

Supervisors: Rikke Louise Meyer and Alexander Zelikin

Biofilm-related implant infections are a serious and increasing health issue, often resulting in significant clinical implications for the patient, such as additional surgery and lengthy systemic antimicrobial treatment. Biofilm infections on medical implants are caused by bacteria entering the site of the implant, either during surgery or post-operatively. The bacteria then settle on the surface of the implant where they will adhere to it and form a biofilm. A biofilm is a community of bacteria encased in a thick slime, which is produced by the bacteria themselves. Bacteria in a biofilm are protected from antimicrobial agents and host immune cells and are therefore difficult to eradicate with conventional antimicrobial therapy. Effective treatments are lacking, and these infections may therefore result in removal of the implant or life-long suppressive treatment.

To overcome these problems, new strategies for disrupting biofilm growth using local drug delivery are being developed. Local drug delivery has the potential to increase the antimicrobial load achievable directly at the site of infection as well as reducing potential side effects of systemic antimicrobial treatment. Substrate Mediated Enzyme Prodrug Therapy (SMEPT) is a novel approach that allows for localized drug synthesis directly on the surface of the medical implant. Before the implant is inserted into the body, it can be coated with polymer films containing an immobilized enzyme.  In SMEPT, an externally administered, non-toxic, non-therapeutic prodrug is converted to its corresponding active drug when encountering the immobilized enzyme in the coating. This makes it possible to control the administration, to alter the treatment dose and to choose between multiple drugs.

This project aims to provide proof-of-concept for the ability of SMEPT to combat in vitro grown biofilms of medically relevant microorganisms.

The ability of SMEPT to kill bacterial cells in in vitro grown biofilms is investigated in biofilms grown on surfaces coated with multi-layered polyelectrolyte thin films containing the enzyme β-glucuronidase. Upon administration of a non-therapeutic glucuronide prodrug of moxifloxacin, β-glucuronidase converts the prodrug to active moxifloxacin. The killing-effect of the converted prodrug of moxifloxacin on clinical isolates of Staphylococcus aureus and Staphylococcus epidermidis will be investigated in vitro by evaluating the reduction in biomass on statically grown biofilms and by assessing the ratio of living and dead cells in biofilms grown under continuous flow, differentially stained to distinguish living and dead bacteria and visualized by confocal laser scanning microscopy.

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