Identifying a nasal gene expression signature associated with hyperinflation and treatment response in severe COPD

Abstract

Hyperinflation contributes to dyspnea intensity in COPD. Little is known about the molecular mechanisms underlying hyperinflation and how inhaled corticosteroids (ICS) affect this important aspect of COPD pathophysiology. To investigate the effect of ICS/long-acting β2-agonist (LABA) treatment on both lung function measures of hyperinflation, and the nasal epithelial gene-expression profile in severe COPD. 117 patients were screened and 60 COPD patients entered a 1-month run-in period on low-dose ICS/LABA budesonide/formoterol (BUD/F) 200/6 one inhalation b.i.d. Patients were then randomly assigned to 3-month treatment with either a high dose BDP/F 100/6 two inhalations b.i.d. (n = 31) or BUD/F 200/6 two inhalations b.i.d. (n = 29). Lung function measurements and nasal epithelial gene-expression were assessed before and after 3-month treatment and validated in independent datasets. After 3-month ICS/LABA treatment, residual volume (RV)/total lung capacity (TLC)% predicted was reduced compared to baseline (p < 0.05). We identified a nasal gene-expression signature at screening that associated with higher RV/TLC% predicted values. This signature, decreased by ICS/LABA treatment was enriched for genes associated with increased p53 mediated apoptosis was replicated in bronchial biopsies of COPD patients. Finally, this signature was increased in COPD patients compared to controls in nasal, bronchial and small airways brushings. Short-term ICS/LABA treatment improves RV/TLC% predicted in severe COPD. Furthermore, it decreases the expression of genes involved in the signal transduction by the p53 class mediator, which is a replicable COPD gene expression signature in the upper and lower airways.Trial registration: ClinicalTrials.gov registration number NCT01351792 (registration date May 11, 2011), ClinicalTrials.gov registration number NCT00848406 (registration date February 20, 2009), ClinicalTrials.gov registration number NCT00158847 (registration date September 12, 2005).

Conflict of interest statement

IMB reports fees paid to the University from GlaxoSmithKline. GHK reports institutional grants from the Lung Foundation of the Netherlands, TEVA, GlaxoSmithKline, Ubbo Emmius Foundation, and TETRI Foundation. HAMK reports institutional grants from Novartis, Boehringer Ingelheim, and GlaxoSmithKline; speaking fees from Boehringer Ingelheim; consultant fees from AstraZeneca, Chiesi, GlaxoSmithKline, and Novartis; board membership with Chiesi, GlaxoSmithKline, and Novartis; and institutional patient fees from Fluidda. WT reports fees paid to the University from Pfizer, GlaxoSmithKline, Chiesi, Roche Diagnostics/Ventana, Biotest, Merck Sharp Dohme, Novartis, Lilly Oncology and a grant from the Dutch Asthma Fund. KS has a patent 9,677,138 issued for the Boston University, Annual royalties from UpToDate for co-authoring an UpToDate chaptor on Gene Expression, honoraria from Acdemic Research Coalition LLC for serving as expert faculty member and speaker at annual Respiratory Disease Young Investigators’ Forum, Grant from Lungevity Foundation. MEL reports stock from Metera Pharmaceuticals. DSP reports consultancy fees or research grants paid to the University from AstraZeneca, Chiesi, Genentec, GlaxoSmithKline, Roche. MvdB reports research grants paid to the University from Chiesi, GlaxoSmithKline and TEVA. AF, KI, EvdW, CAB, SV, HRP, WgB, PW, FvdE, KM, AS and IHH have no competing interests.

Figures

Figure 1

Schematic diagram of FAIR study outline.

Figure 2

Influence of ICS/LABA (BDP/F and BUD/F treatment arms combined) on measures of hyperinflation and lung function. Lung function measurements were taken before study medication at baseline, 1 month and 3 months. (A) FEV1% predicted, (B) RV% predicted (C) TLC% predicted (D) RV/TLC% predicted and E) FEF 25–75% predicted. Linear mixed effects model comparing each time point. *p < 0.05 FEV1, forced expiratory volume in one second; RV, residual volume; TLC, total lung capacity; RV/TLC, residual volume/total lung capacity; FEF 25–75% predicted, the average forced expiratory flow during the mid (25–75%) portion of the FVC. All data are expressed as mean ± SEM.

Figure 3

Nasal gene-expression associated with RV/TLC% predicted. Heatmap of genes associated with RV/TLC% predicted at baseline in all screened patients with severe COPD (FDR 

Figure 4

The relationship between the hyperinflation gene expression signature and presence of COPD in the upper, large and small airway brushings. (A) Enrichment of genes increased with RV/TLC% predicted in the upper airway among genes increased with RV/TLC% in bronchial biopsies (p < 0.001). The color bar indicates the genes ranked according to their association with RV/TLC% predicted in bronchial biopsies (blue representing a negative association with RV/TLC% predicted and red indicating a positive association). Enrichment of genes increased with RV/TLC% in the nose among genes increased in COPD (B) nasal epithelium, (C) large airway brushings and (D) small airway brushings, compared to healthy controls (GSEAp < 0.001). The color bar indicates the genes ranked according to their expression in COPD epithelial cells compared to healthy/non-COPD controls (blue representing genes decreased in COPD epithelium compared to healthy/non-COPD controls, while red indicates an increase in epithelial gene-expression). (E) Heatmap of rank metric score of the hyperinflation signature GSEA in comparison to the upper, large and small airway brushing from COPD and non-COPD controls. The enrichment of genes in the hyperinflation signature among genes increased in COPD in the upper, large and small airway brushing is represented with the green scale and illustrated in a (F) Venn diagram.

Figure 5

Gene set enrichment analysis (GSEA) comparison of genes associated with RV/TLC% predicted in bronchial biopsies (GLUCOLD). (A) Enrichment of genes increased with RV/TLC% predicted in the nose among genes down-regulated in nasal epithelium following increased dosage with ICS/LABA (p < 0.001) or treatment with (B) ICS/LABA (fluticasone/salmeterol) (p < 0.001) and (C) solely ICS (fluticasone) (p < 0.001) for 6 months compared to placebo in bronchial biopsies. The color bar indicates the genes ranked according to their change in expression with ICS (blue representing a treatment-induced decrease and red an increase in gene-expression). The vertical bars in all plots indicate the genes increased with RV/TLC% predicted with the location of the bar indicating the occurrence of that gene within the ranked gene list and the height of the bars indicating the running GSEA enrichment score (black bars = core enriched genes, grey bars = core non-enriched genes). (D) Heatmap of the change of expression of genes associated with RV/TLC% predicted core enriched to be decreased in bronchial biopsies from patients with COPD after 6 months treatment with ICS/LABA (fluticasone/salmeterol) or solely ICS (fluticasone). (E) Heatmap of rank metric score of the RV/TLC signature GSEA comparison to increased dosage with ICS/LABA (p < 0.001) in nasal epithelium, and treatment with ICS/LABA (fluticasone/salmeterol) or solely ICS (fluticasone) compared to placebo in bronchial biopsies. The enrichment of genes in the RVTLC signature among genes decreased within the three datasets is represented in the green scale and illustrated in a (F) Venn diagram.