**5. Neutrophils as biomarkers of CF lung disease**

The mainstays of CF lung disease management are commenced early in infancy and presently include chest physiotherapy to remove mucus plugs from the airways and antibiotic therapy to control infections [12]. Other therapeutic approaches such as hypertonic saline, finalized to increase mucociliary clearance, should be corroborated by efficacy data [135]. Recombinant human DNAse (Dornase alpha) is a strong mucolytic which improves lung function [136] but is given to CF infants only on indication due to its cost [137]. The recent breakthrough in CF, represented by the use of CFTR‐correcting therapies, is a milestone in the clinical management of these patients. Ivacaftor (Kalydeco®, Vertex Pharmaceuticals, USA) is a CFTR potentiator given successfully to patients with class III gating mutations. This drug not only improves lung function and normalizes sweat chloride in children above 6 years of age [138], but its efficacy has also been proven in preschoolers [139].

At whatever age, the control of therapeutic efficacy of medications is granted by functional respiratory tests. However, more specific and sensitive assays are urgently needed to monitor the halt in the progression of lung disease, especially now that we entered the era of personalized medicine in CF [140]. Neutrophils, the main cell type involved in the onset and progression of CF lung disease, are clearly an interesting target in this context and are being evaluated for such a purpose. The best indication that neutrophils and their products are sensitive biomarkers of CF lung disease comes from the clinical data about NE. Sputum NE levels have been validated as the most predictive biomarker of lung decline and reduced survival [107, 141], being, however, of no utility in non‐expectorating young children. Being easy to isolate from the peripheral blood, circulating neutrophils are more at hand to being studied. Conese et al. [142] analysed blood neutrophils by microarray gene expression in 10 CF patients, homozygous for the F508del mutation, given a course of parenteral antibiotics for an acute exacerbation, before and after therapy. mRNAs of three genes were found downregulated in CF patients before therapy and returned to 'healthy' levels after therapy: phorbol‐12‐myristate‐13‐ acetate‐induced protein 1 (*PMAIP1*), hydrogen voltage‐gated channel 1 (*HVCN1*), and β‐arrestin 1 (*ARRB1*). Recently, we validated neutrophil *HVCN1* mRNA as a biomarker following the treatment of seven CF patients, homozygous or heterozygous for class III mutations, with ivacaftor, confirming that its expression levels are lower as compared with healthy controls before therapy, while they are increased after CF patients were treated for 6 months (Guerra et al., submitted). Overall, these data strongly indicate that *HVCN1* mRNA level is a neutrophil biomarker sensitive to therapy. In another study [77], ivacaftor treatment resulted in normalized ion homeostasis and corrected Rab27a activation as well as degranulation in blood neutrophils obtained from six CF patients with the genotype *F508del*/*G551D*. In line with these findings, extracellular Pseudomonas killing by CF neutrophils obtained from CF patients during treatment was significantly increased. Activated CD11b was investigated as a marker of neutrophil activation and whether it was downregulated by ivacaftor treatment in five patients with *F508del/G551D* and *G551D/N1303K* genotypes [143]. A cytofluorimetric assay showed that activated CD11b on PMNs was significantly higher at baseline in the CF patients compared to controls. However, after treatment, this marker was not significantly different from healthy controls, suggesting that ivacaftor treatment results in a decrease, towards normalization, of the activation status of blood neutrophils in vivo.
