The role of nitric oxide as a novel therapy to disrupt bacterial biofilms in patients with cystic fibrosis

Abstract

The lungs of most patients with cystic fibrosis (CF) eventually become chronically infected with the opportunistic pathogen Pseudomonas aeruginosa (PA). This infection is recognised as consisting of free-living bacteria (known as “planktonic bacteria”) and bacteria within aggregates known as “biofilms”. Bacteria within biofilms are more tolerant to antibiotics than their planktonic counterparts. Current CF treatment of infective exacerbations aims to eradicate or control PA infection using aggressive antibiotic regimes. Despite repeated courses of antibiotics used early after initial positive cultures, many patients become permanently infected. This chronic infection/colonisation results in progressive airways obstruction and worsening bronchiectasis. Patients colonised with PA are more unwell and die at a younger age. Work conducted by colleagues had established that low dose nitric oxide (NO) can disrupt pseudomonal biofilms in the laboratory and established a novel combination of assays to study biofilm obtained directly from CF sputa. This proof of concept study aimed to discover whether non-toxic levels of NO, administered to participants during an episode of acute infective exacerbation, could disrupt bacteria from biofilms and increase the effectiveness of antibiotic therapy. A randomised, participant and outcome-assessor blind, proof of concept study was carried out to compare treatment with nitric oxide gas during acute infection with placebo. Twelve participants with CF (aged 12 or above) were randomised to receive 5-7 days of either inhaled nitric oxide gas (10ppm) or placebo (air) in addition to standard intravenous (IV) antibiotics for a pulmonary exacerbation. The primary endpoint was the microbiological effect on proportion of bacteria in biofilms in sputum between baseline and follow-up. Secondary endpoints included laboratory and clinical parameters measured at all time-points including a number of safety measures. There was no difference between the groups in the primary outcome measure of proportion of bacteria in biofilm between baseline and follow-up. In the secondary outcomes, generalised estimating equation analysis showed a significant reduction in bacteria in biofilms in the NO group at the end of 7 days NO therapy (mean log difference between groups for measured number of >20 cell aggregates 3.49 (95%CI 0.32, 6.67; p=0.031) and for biofilm volume 4.47 (95%CI -.04, 8.98; p=0.052)). Safety measures were reassuring and other parameters (including lung function) demonstrated trends in the direction of treatment, but no other results reached significance. All results are impacted by the small number of participants and the wide variability between and within participants. There was no effect seen on bacterial biofilms at follow-up, however, the secondary microbiological outcome data shows preliminary evidence of benefit using low dose NO as adjunctive therapy during the period of treatment. Results show inhaled NO therapy is safe when administered to CF patients during a pulmonary exacerbation alongside IV antibiotic therapy. Further studies are needed to investigate whether inhaled NO, or new drugs in development (e.g. novel targeted NO donor therapies) may have any impact on PA related morbidity and mortality in CF.

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Identifiers

ORCID for Saul Faust: orcid.org/0000-0003-3410-7642

ORCID for Gary Connett: orcid.org/0000-0003-1310-3239

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