Activity of the De Novo Engineered Antimicrobial Peptide WLBU2 against Pseudomonas aeruginosa in Human Serum and Whole Blood: Implications for Systemic Applications

Abstract

Cationic amphipathic peptides have been extensively investigated as a potential source of new antimicrobials that can complement current antibiotic regimens in the face of emerging drug-resistant bacteria. However, the suppression of antimicrobial activity under certain biologically relevant conditions (e.g., serum and physiological salt concentrations) has hampered efforts to develop safe and effective antimicrobial peptides for clinical use. We have analyzed the activity and selectivity of the human peptide LL37 and the de novo engineered antimicrobial peptide WLBU2 in several biologically relevant conditions. The host-derived synthetic peptide LL37 displayed high activity against Pseudomonas aeruginosa but demonstrated staphylococcusspecific sensitivity to NaCl concentrations varying from 50 to 300 mM. Moreover, LL37 potency was variably suppressed in the presence of 1 to 6 mM Mg 2؉ and Ca 2؉ ions. In contrast, WLBU2 maintained its activity in NaCl and physiologic serum concentrations of Mg 2؉ and Ca 2؉ . WLBU2 is able to kill P. aeruginosa (10 6 CFU/ml) in human serum, with a minimum bactericidal concentration of <9 M. Conversely, LL37 is inactive in the presence of human serum. Bacterial killing kinetic assays in serum revealed that WLBU2 achieved complete bacterial killing in 20 min. Consistent with these results was the ability of WLBU2 (15 to 20 M) to eradicate bacteria from ex vivo samples of whole blood. The selectivity of WLBU2 was further demonstrated by its ability to specifically eliminate P. aeruginosa in coculture with human monocytes or skin fibroblasts without detectable adverse effects to the host cells. Finally, WLBU2 displayed potent efficacy against P. aeruginosa in an intraperitoneal infection model using female Swiss Webster mice. These results establish a potential application of WLBU2 in the treatment of bacterial sepsis.

Antimicrobial Polymers: Mimicking Amino Acid Functionali ty, Sequence Control and Three-dimensional Structure of Host-defen se Peptides

“…It was found for antimicrobial peptides that a high ion strength has a negative influence on the antimicrobial activity, 47 which was ascribed to a masking of cationic and anionic charges by small ions. 48 A similar effect was observed for methacrylate based AMP mimics. 46a Furthermore, bivalent cations such as Ca 2+ or Mg 2+ were found to have a higher inhibiting potential, which was ascribed to the bridging of membrane components, increasing the membrane stability.…”

Cathelicidin LL-37 Increases Lung Epithelial Cell Stiffness, Decreases Transepithelial Permeability, and Prevents Epithelial Invasion by Pseudomonas aeruginosa

“…1A, 1B). LL-37 increased A549 cell stiffness ∼30% more compared with the effects of truncated LL-37 moieties, such as RK-30 and KR-20, and ∼85% higher compared with the longer peptide WLBU2, which in several experimental conditions showed stronger bactericidal activity than LL-37 (32,33) (Fig. 1C).…”

Covalent immobilization of antimicrobial peptides (AMPs) onto biomaterial surfaces

“…These are distinct from those in nature, with simpler but rationally engineered composition, obtained by varying the amino acid content and sequence and overall peptide length to achieve enhanced activity and very low cytotoxic properties [57]. For example, Mietzner and co-workers [58][59][60][61] have developed a series of de novo peptides based on structure-function properties observed in natural AMPs. They engineered the peptide composition to achieve enhanced potency, selectivity and stability.…”