L.D. Lozeau, T.E. Alexander, Z. Lipsky, T.A. Camesano
Keywords: antimicrobial peptides, mechanisms, surface tethering
Summary:Healthcare associated infections cost US hospitals over $45 billion dollars each year in direct medical costs, which does not account for the indirect costs and unmeasurable costs: patient pain, suffering, and quality of life. Catheter-associated urinary tract infections (CAUTIs) are the number one cause of these infections affecting 450,000 patients per year, significantly increasing patient length of stay and contribute to antibiotic resistance. At AMProtection, we believe in the proactive prevention of device-associated infection such as CAUTI. Thus, we have developed a broadly active antimicrobial coating for urinary catheters that will prevent and fight infection, combat resistance and reduce gold-standard antibiotic use. Our patent pending technology for surface-tethered antimicrobial peptides creates a strong competitive advantage for our potential customers, urinary catheter manufacturers. While current treatments may be toxic and are ineffective in the long-term, antimicrobial peptides (AMPs) such as Chrysophsin-1 (CHY1) are promising alternatives due to their broad-spectrum antimicrobial activity and unique mechanisms that make resistance less likely. Clinical development of AMPs has been hindered by their short half-life, potential toxicity and high cost. Many studies have explored methods for tethering AMPs onto surfaces to localize AMPs and retain activity while minimizing toxicity in a more cost-efficient manner, but the effects of AMP density and tether length on activity remain unclear. To examine this effect we covalently tethered cysteine-modified Chrysophsin-1 (C-CHY1) via the different molecular weight polyethylene glycol (PEG) molecules (MW866, MW2000 and MW7500). Quartz crystal microbalance with dissipation was used to characterize thickness and grafting density of the tethered C-CHY1. We also used QCM-D to determine the mechanism of Chrysophsin-1 against model mammalian membranes. Antimicrobial activity was determined against Staphylococcus aureus and Escherichia coli using minimum inhibitory activity assays. C-CHY1 activity was found to be independent of bacteria type at 23oC, while different tether lengths lead to different action mechanisms. The highest activity was achieved with MW7500, which allowed a native pore formation mechanism, followed by MW866, which forced a non-native ion-displacement mechanism. With the middle tether length MW2000, C-CHY1 demonstrated lower activity due to its inability to adopt either mechanism effectively. Increasing peptide density had an inverse effect on activity for Gram-positive and Gram-negative bacteria. Bacterial incubation temperature also had a significant effect on activity. These results will help develop a more clinically relevant design of catheter-tethered AMPs to prevent CAUTIs.