Optimizing Exogenous Surfactant as a Pulmonary Delivery Vehicle for Chicken Cathelicidin-2

Brandon Baer,Shehrazade Jekhmane,Markus Weingarth,Cory Yamashita,Ruud Veldhuizen,Håvard Jenssen,Natalia Molchanova,Annelise E. Barron,Jennifer S. Lin,Edwin J. A. Veldhuizen

doi:10.1038/s41598-020-66448-1

Abstract

The rising incidence of antibiotic-resistant lung infections has instigated a much-needed search for new therapeutic strategies. One proposed strategy is the use of exogenous surfactants to deliver antimicrobial peptides (AMPs), like CATH-2, to infected regions of the lung. CATH-2 can kill bacteria through a diverse range of antibacterial pathways and exogenous surfactant can improve pulmonary drug distribution. Unfortunately, mixing AMPs with commercially available exogenous surfactants has been shown to negatively impact their antimicrobial function. It was hypothesized that the phosphatidylglycerol component of surfactant was inhibiting AMP function and that an exogenous surfactant, with a reduced phosphatidylglycerol composition would increase peptide mediated killing at a distal site. To better understand how surfactant lipids interacted with CATH-2 and affected its function, isothermal titration calorimetry and solid-state nuclear magnetic resonance spectroscopy as well as bacterial killing curves against Pseudomonas aeruginosa were utilized. Additionally, the wet bridge transfer system was used to evaluate surfactant spreading and peptide transport. Phosphatidylglycerol was the only surfactant lipid to significantly inhibit CATH-2 function, showing a stronger electrostatic interaction with the peptide than other lipids. Although diluting the phosphatidylglycerol content in an existing surfactant, through the addition of other lipids, significantly improved peptide function and distal killing, it also reduced surfactant spreading. A synthetic phosphatidylglycerol-free surfactant however, was shown to further improve CATH-2 delivery and function at a remote site. Based on these in vitro experiments synthetic phosphatidylglycerol-free surfactants seem optimal for delivering AMPs to the lung.

Highlights

The increasing incidence of antimicrobial resistance in bacterial pneumonia has instigated a much-needed search for new therapeutic approaches for these types of infections[1,2]
To investigate how exogenous surfactant and its lipid components affect antimicrobial peptide function, bacterial killing curves were performed with CATH-2 combined with 10 mg/mL bovine lipid extract surfactant (BLES) or 1–2 mg/mL of individual surfactant lipids (Fig. 1)
To explore if the PG effect was specific to CATH-2, killing curves for CRAMP, PMAP-23 and LL-37 were performed in the presence of BLES or individual surfactant lipids

Summary

Introduction

The increasing incidence of antimicrobial resistance in bacterial pneumonia has instigated a much-needed search for new therapeutic approaches for these types of infections[1,2]. The primary benefit of utilizing CATH-2, and other cathelicidins, for therapeutic purposes is their diverse range of pathways to kill bacteria since their positive charge allows them to interact directly with both the negatively charged lipids of the bacterial cell wall as well as intracellular targets such as DNA or RNA9–12. This multi-target approach to killing bacteria has been demonstrated to be effective against a wide spectrum of antibiotic-resistant organisms[5,6,7]. It is hypothesized that the PG component of surfactant inhibits CATH-2 function and that an exogenous surfactant, with a reduced PG composition would increase peptide mediated killing at a distal site

Methods

Results

Conclusion