CFH1b Artificial enzymes for selective antibiotic activation

Two prodrug forms of the fluoroquinolone antimicrobial ciprofloxacin were investigated and activity tests confirmed the masking of their antibacterial activity. Both were found suitable for further studies. Subsequently, a selection of synthetic catalysts was tested and found not to have antimicrobial activity in their own right at physiologically relevant concentrations. Upon addition of the prodrugs to the catalyst-containing growth medium, the antimicrobial activity of the fluoroquinolone was unmasked, confirming that the catalytic activation of the prodrug could be achieved in aqueous media and at physiological pH.
In the next step it was confirmed that the targeting siderophore group was required to allow the synthetic catalysts to enter bacterial cells. In addition, the antimicrobial activty of a series of siderophore-drug conjugates was assessed using E. coli and S. aureus strains under both iron-deficient and iron-sufficient conditions. Standard screening protocols are now established that allow time-dependent bacterial cell growth to be monitored and evaluated in an automated way and these will be used by chemists in the research group in the future. In addition, an E. coli mutant that is unable to transport iron siderophores into the cell was obtained to serve as a negative control. The insights obtained have allowed us to start a new collaboration with Prof. Eszter Boros at Stony Brook University in New York, who is now following the uptake of our siderophore conjugates into E. coli by means of radioactive Ga-67.
The results obtained from the antimicrobial activity screens and the radiolabelling revealed key structural requirements for the targeted delivery of cargo, such as catalysts or drugs, into the bacterial cell. Difficulties with the synthesis of the siderophore-catalyst conjugates could not be overcome for the planned intracellular catalytic tests to be completed within the timescale of the project. The studies are still ongoing and they are greatly facilitated by the protocols that were developed with CFH support.