Sputum and blood transcriptomics characterisation of the inhaled PDE4 inhibitor CHF6001 on top of triple therapy in patients with chronic bronchitis

Abstract

Background: Although phosphodiesterase-4 (PDE4) inhibitors have been shown to reduce COPD exacerbation rate, their biological mechanism of action is not completely elucidated at the molecular level. We aimed to characterise the whole genome gene expression profile of the inhaled PDE4-inhibitor CHF6001 on top of triple therapy in sputum cells and whole blood of patients with COPD and chronic bronchitis.

Methods: Whole genome gene expression analysis was carried out by microarray in 54 patients before and after 32 days treatment with CHF6001 800 and 1600 μg and placebo twice daily (BID) in a randomised crossover study.

Results: CHF6001 had a strong effect in sputum, with 1471 and 2598 significantly differentially-expressed probe-sets relative to placebo (p-adjusted for False Discovery Rate < 0.05) with 800 and 1600 μg BID, respectively. Functional enrichment analysis showed significant modulation of key inflammatory pathways involved in cytokine activity, pathogen-associated-pattern-recognition activity, oxidative stress and vitamin D with associated inhibition of downstream inflammatory effectors. A large number of pro-inflammatory genes coding for cytokines and matrix-metalloproteinases were significantly differentially expressed for both doses; the majority (> 87%) were downregulated, including macrophage inflammatory protein-1-alpha and 1-beta, interleukin-27-beta, interleukin-12-beta, interleukin-32, tumour necrosis factor-alpha-induced-protein-8, ligand-superfamily-member-15, and matrix-metalloproteinases-7,12 and 14. The effect in blood was not significant.

Conclusions: Inhaled PDE4 inhibition by CHF6001 on top of triple therapy in patients with COPD and chronic bronchitis significantly modulated key inflammatory targets and pathways in the lung but not in blood. Mechanistically these findings support a targeted effect in the lung while minimising unwanted systemic class-effects.

Trial registration: ClinicalTrial.gov, EudraCT, 2015-005550-35. Registered 15 July 2016.

Keywords: Biomarkers; Chronic obstructive pulmonary disease; Gene expression; Inflammation; Phosphodiesterase 4 inhibitors.

Conflict of interest statement

MG, MB, SV, GL, AE, MAN and SP are employees of Chiesi, the sponsor of this study.

GLJ and EKA are employees of the Almac group; Almac received funding from Chiesi as part of a consultation fee for the work reported in this manuscript.

BC and BL have nothing to disclose.

OK’s institution received fees from Chiesi for conducting this study as a participating site. He reports personal fees as speaker or Advisory Board member from AstraZeneca, Boehringer Ingelheim, GlaxoSmithKline, Sanofi and Novartis.

KMB declares that no personal payments were received from any pharmaceutical entity in the past 5 yrs. He is a full time employee of insaf Respiratory Research Institute. The institution has received compensation for services on advisory boards or consulting for Ablynx, Almirall, AstraZeneca, Berlin Chemie, Boehringer, Chiesi, Cytos, Mundipharma, Novartis, Pohl Boskamp, Zentiva. The institution has received compensation for speaker activities in scientific meetings supported by Almirall, AstraZeneca, Berlin Chemie, Boehringer, Cytos, ERT, GSK, Novartis, Pfizer, Pohl Boskamp, Takeda. The institution has received compensation for design and performance of clinical trials from Almirall, Altana/Nycomed, AstraZeneca, Boehringer, Cytos, GSK, Infinity, Medapharma, MSD, Mundipharma, Novartis, Parexel, Pearl Therapeutics, Pfizer, Revotar, Teva, Sterna, and Zentiva.

HW reports personal fees from Chiesi during the conduct of the study. Outside the submitted work, Dr. Watz reports personal fees from Bayer, personal fees from GSK, personal fees from Boehringer Ingelheim, personal fees from Novartis, personal fees from AstraZeneca, personal fees from BerlinChemie, and personal fees from Roche.

DS received personal fees from Chiesi during the conduct of this study. Outside the submitted work, he reports grants and personal fees from AstraZeneca, Boehringer Ingelheim, Chiesi, GlaxoSmithKline, Glenmark, Menarini, Mundipharma, Novartis, Pfizer, Pulmatrix, Therevance, and Verona, and personal fees from Cipla, Genentech and Peptinnovate.

Figures

Fig. 1

Differentially expressed probe sets for CHF6001 800 μg and 1600 μg BID relative to placebo in (A) blood and (B) sputum. Volcano plot depicting all detected probe sets and coloured by fold change (FC) and adjusted p-values: orange, |FC| > 1.3; red, pFDR< 0.05; green, |FC| > 1.3 and pFDR< 0.05. Abbreviation: BID, twice daily

Fig. 2

Log2 fold change with CHF6001 1600 μg BID vs. log2 fold change with CHF6001 800 μg BID. Equality line in red, fitted loess curve in blue. Only differentially expressed probe sets (pFDR

Fig. 3

Network of pro-inflammatory cytokines and matrix metalloproteinases differentially expressed after treatment with CHF6001 A) 800 μg and B) 1600 μg BID relative to placebo. Each node represents all the proteins produced by a single, protein-coding gene locus, edges represent proteins that jointly contribute to a shared function, and the information inside the circle describes protein structure. Edges: a red line indicates the presence of fusion evidence; a green line, neighbourhood evidence; a blue line, co-occurrence evidence; a magenta line, experimental evidence; a yellow line, text mining evidence; a light blue line, database evidence; a black line, co-expression evidence; a purple line, protein homology evidence. A green halo around the nodes: downregulation, a red halo: upregulation. Unmarked nodes: pFDRp < 0.05 and |FC| > 1.3

Fig. 4

Cytokines and inflammation response from WikiPathways. Downregulated genes are coloured in different shades of green

Fig. 5

Key pathophysiological pathways significantly modulated by CHF6001 in sputum with associated inhibition of downstream inflammatory effectors. Created with Biorender.com