Influence of highly effective modulator therapy on the sputum proteome in cystic fibrosis.

BACKGROUND: There have been dramatic clinical improvements in people with cystic fibrosis (PwCF) commenced on the cystic fibrosis conductance regulator (CFTR) modulator elexacaftor/tezacaftor/ivacaftor (ETI). Sputum proteomics is a powerful research technique capable of identifying important airway disease mechanisms. Using this technique, we evaluated how ETI changes the sputum proteome in PwCF. METHODS: Sputum samples from 21 CF subjects pre- and post- ETI, 6 CF controls ineligible for ETI, and 15 healthy controls were analysed by liquid chromatography mass spectrometry. RESULTS: Post-ETI, mean FEV1 % increased by 13.7 % (SD 7.9). Principal component and hierarchical clustering analysis revealed that the post-ETI proteome shifted to an intermediate state that was distinct from pre-ETI and healthy controls, even for those achieving normal lung function. Functional analysis showed incomplete resolution of neutrophilic inflammation. The CF control sputum proteome did not alter. At the protein-level many more proteins increased in abundance than decreased following ETI therapy (80 vs 30; adjusted p value <0.05), including many that have anti-inflammatory properties. Of those proteins that reduced in abundance many were pro-inflammatory neutrophil-derived proteins. Several important respiratory proteases were unchanged. CONCLUSIONS: Sputum proteomics can provide insights into CF lung disease mechanisms and how they are modified by therapeutic intervention, in this case ETI. This study identifies imbalances in pro- and anti- inflammatory proteins in sputum that partially resolve with ETI even in those achieving normal spirometry values. This post-ETI intermediate state could contribute to ongoing airway damage and therefore its relevance to clinical outcomes needs to be established.

The relationship between lung disease severity and the sputum proteome in cystic fibrosis.

BACKGROUND: Proteomics can reveal molecular pathways of disease and provide translational perspectives to inform clinical decision making. Although several studies have previously reported the cystic fibrosis airway proteome, the relationship with severity of lung disease has not been characterised. The objectives of this observational study were to investigate differences in the CF sputum proteome associated with disease severity and identify potential markers of disease with translational potential. METHODS: Sputum samples from healthy volunteers and cystic fibrosis subjects (some prescribed modulator therapies) were analysed using liquid-chromatography tandem mass spectrometry. Severity of lung disease was based on baseline spirometry (percentage predicted forced expiratory volume in 1 s, FEV1%). RESULTS: Multiple sputum proteins (108 increased; 202 decreased) were differentially expressed in CF (n = 38) and healthy volunteers (n = 32). Using principal component analysis and hierarchical clustering, differences in sputum proteome were observed associated with progressive lung function impairment. In CF subjects, baseline FEV1% correlated with 87 proteins (positive correlation n = 20, negative n = 67); most were either neutrophil derived, or opposed neutrophil-driven oxidant and protease activity. CONCLUSION: Predictable and quantifiable changes in the CF sputum proteome occurred associated with progressive lung function impairment, some of which might have value as markers of disease severity in CF sputum. Further work validating these markers in other patient cohorts and exploring their clinical utility is needed.

Quantitative proteomic analysis of bronchoalveolar lavage fluid in West Highland white terriers with canine idiopathic pulmonary fibrosis.

BACKGROUND: Canine idiopathic pulmonary fibrosis (CIPF) is a chronic, progressive, interstitial fibrosing lung disease, manifesting as cough, exercise intolerance and ultimately, dyspnea and respiratory failure. It mainly affects West Highland white terriers (WHWTs), lacks curable treatment and has a poor prognosis. Aspiration of gastroesophageal refluxate may play a role in the development of CIPF. In the first part of this study, we completed label-free quantitative proteomic analysis of bronchoalveolar lavage fluid (BALF) from CIPF and healthy WHWTs. In the second part, we evaluated potential protein markers of reflux aspiration from canine gastric juice and vomitus and whether these were present in BALF from the two groups. RESULTS: Across all BALF samples, 417 proteins were identified, and of these, 265 proteins were identified by two or more unique tryptic peptides. Using the 265 high confidence assignments, the quantitative proteome profiles were very similar in the two cohorts, but they could be readily resolved by principal component analysis on the basis of differential protein expression. Of the proteins that were differentially abundant in the two groups, several (including inflammatory and fibrotic markers) were elevated in CIPF, and a smaller, more diverse group of proteins were diminished in CIPF. No protein markers indicative of reflux aspiration were identified. CONCLUSIONS: Label-free proteomics allowed discrimination between CIPF and healthy WHWTs, consistent with fibrotic process but did not provide clear evidence for gastrointestinal aspiration. The measurement of proteins may provide a proteomics signature of CIPF that could be used to evaluate treatment options.