ABSTRACT
The rising prevalence of antibiotic resistance underscores the urgent need for novel antimicrobial agents. Antimicrobial peptides (AMPs) are potentially effective therapeutics that disrupt bacterial membranes regardless of resistance to traditional antibiotics. We have developed engineered cationic AMPs (eCAPs) with broad activity against multidrug-resistant (MDR) bacteria, but stability remains an important concern. Therefore, we sought to enhance the clinical utility of eCAP WLBU2 in biological matrices relevant to respiratory infection. A designed substitution of d-Val for l-Val resulted in increased resistance to protease enzymatic degradation. We observed multiple gains of functions such as higher activity against bacteria in biofilm mode of growth, significantly lower toxicity to erythrocytes and white blood cells compared to WLBU2, with increased safety in mice. Direct airway delivery revealed a therapeutic index of >140 for the selected enantiomer compared to that of <35 for WLBU2. The data warrant clinical exploration by aerosolized delivery to mitigate MDR-related respiratory infection.
Subject(s)
Anti-Bacterial Agents , Antimicrobial Cationic Peptides , Bacteria , Respiratory Tract Infections , Animals , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/therapeutic use , Antimicrobial Cationic Peptides/pharmacology , Antimicrobial Cationic Peptides/therapeutic use , Bacteria/drug effects , Bacteria/metabolism , Drug Resistance, Multiple, Bacterial , Mice , Microbial Sensitivity Tests , Protein Engineering , Therapeutic IndexABSTRACT
OBJECTIVES: Pseudomonas aeruginosa is a common cause of pneumonia in patients with cystic fibrosis with the property to generate multidrug resistance against clinically used antibiotics. Antimicrobial peptides (AMPs) are a diverse group of effector molecules of the innate immune system that protect the host against pathogens. However, the lack of activity in common biological matrices has hampered efforts towards clinical development. In this study, we evaluated the therapeutic potential of the engineered AMP WLBU2 via direct airway delivery in a murine model of P. aeruginosa infection. METHODS: The human AMPs LL37 and WLBU2 were compared for (i) antibiofilm activity using P. aeruginosa on polarized human bronchial epithelial cells, and (ii) efficacy in P. aeruginosa pneumonia in mice using intratracheal instillation of bacteria and AMPs. RESULTS: WLBU2 (16 µM) prevents biofilm formation by up to 3-log compared with 1-log reduction by LL37. With a single dose of 1 µg (0.05 mg/kg) delivered intratracheally, the initial effect of LL37 was moderate and transitory, as bacterial load and inflammatory cytokines increased at 24 h with observed signs of disease such as lethargy and hypothermia, consistent with moribund state requiring euthanasia. In sharp contrast, WLBU2 reduced bacterial burden (by 2 logs) and bacteria-induced inflammation (leucocytic infiltrates, cytokine and chemokine gene expression) at 6 h and 24 h post-exposure, with no observed signs of disease or host toxicity. CONCLUSION: These promising results now establish a much lower minimum therapeutic dose of WLBU2 (a net gain of 80-fold) compared with the previously reported 4 mg/kg systemic minimum therapeutic dose, with significant implications for clinical development.
