Prevalence of LasB elastase variants from Pseudomonas aeruginosa respiratory isolates and their inhibition by novel LasB inhibitor ANT3273

Abstract

Introduction: P. aeruginosa (PA) elastase (LasB) is a key virulence factor associated with increased severity in chronic respiratory diseases and 30-day mortality in ICU patients. ANT3273 is a potent and specific inhibitor of LasB being developed as an inhaled treatment for PA respiratory infections.Objective: To identify the variants of LasB commonly encountered in PA respiratory isolates and determine their inhibition by ANT3273.Methods: The lasB genes of PA isolates from three clinical isolate collections, two from Cystic Fibrosis patients (Purpan Hospital, Toulouse, n=255; Seattle Children’s Hospital, n=248) and one from Bronchiectasis patients (University of Liverpool, n=189), were analysed. Culture supernatants from strains representing different lasB variants were tested for elastase activity and inhibition by ANT3273.Results: The full-length lasB gene was amplified from all Toulouse CF isolates. Three common lasB variants accounted for 83-95 % across all collections, designated LasB-1 (PA01 Wt sequence), LasB-2 (S241G) and LasB-3 (Q102R, S241G, D244N, K282N, R471S). These variants were also the most prevalent in the Pseudomonas.com genome database, representing 4955 PA strains from diverse environmental, veterinary and human sources. Culture supernatants representing seven variants, including the three most common, all exhibited active elastase and were inhibited by ANT3273 with similar IC50 values (0.16 – 0.25 µM).Conclusions: These results indicate that lasB is highly conserved among PA isolates and that all common LasB variants are inhibitable by ANT3273. Our findings support LasB as a valid target for ANT3273-based therapy.