A specific proteinase 3 activity footprint in α₁-antitrypsin deficiency

Abstract

α1-Antitrypsin (α1-AT) deficiency is a risk factor for emphysema due to tissue damage by serine proteases. Neutrophil elastase (NE) has long been considered the enzyme responsible. However, proteinase 3 (PR3) also produces the pathological features of chronic obstructive pulmonary disease (COPD), is present in the same granules in the neutrophil and is inhibited after NE. We developed a specific footprint assay for PR3 activity and assessed its relationship to an NE footprint in α1-AT deficiency.An ELISA was developed for the specific PR3 fibrinogen cleavage site Aα-Val541. Levels were measured in plasma from 239 PiZZ patients, 94 PiSZ patients, 53 nondeficient healthy smokers and 78 individuals with usual COPD. Subjects underwent extensive demographic characterisation including full lung function and lung computed tomography scanning.Aα-Val541 was greater than the NE footprint in all cohorts, consistent with differential activity. Values were highest in the PiZZ α1-AT-deficient patients and correlated with the NE marker Aα-Val360, but were ∼17 times higher than for the NE footprint, consistent with a greater potential contribution to lung damage. Aα-Val541 was related cross-sectionally to the severity of lung disease (forced expiratory volume in 1 s % pred: rs= −0.284; p<0.001) and was sensitive to augmentation therapy, falling from 287.2 to 48.6 nM (p<0.001).An in vivo plasma footprint of PR3 activity is present in greater quantities than an NE footprint in patients with α1-AT deficiency, is sensitive to augmentation therapy and represents a likely biomarker for dose-ranging studies.

The Course of AαVal541 as a Proteinase 3 Specific Neo-Epitope after Alpha-1-Antitrypsin Augmentation in Severe Deficient Patients

“…Here, we used an assay to detect AαVal541 levels as a marker of PR3 activity based on principles described in previous studies [ 11 ]. We applied this assay to monitor levels of AαVal541 in between two doses of AAT, which has not been previously reported.…”

“…We applied this assay to monitor levels of AαVal541 in between two doses of AAT, which has not been previously reported. Others showed that the PR3- and NE-specific fibrinopeptides were reduced after six months of weekly 60 mg/kg AAT augmentation [ 11 , 12 ]. Campos et al reported that the NE-specific fibrinopeptide AαVal360 showed a significantly stronger decrease in AATD patients who received 120 mg/kg weekly AAT infusion compared with those who received 60 mg/kg weekly AAT infusion [ 10 ].…”

“…More recent studies suggested that newly developed biomarkers or footprints of excess protease activity might be useful when performing dose-ranging studies of AAT augmentation therapy to evaluate its effect on the protease balance and to determine whether the current dose of 60 mg/kg is sufficient for an individual patient, as proposed by Campos et al [ 10 ]. Newby et al described a footprint of PR3 activity, the fibrinopeptide AαVal541, that is a neoepitope generated by PR3-mediated cleavage of fibrinogen [ 11 ]. They showed a reduction of plasma AαVal541 levels in AATD patients after six months of augmentation therapy to levels slightly higher than the AαVal541 they found in healthy controls.…”

Protease-Specific Biomarkers to Analyse Protease Inhibitors for Emphysema Associated with Alpha 1-Antitrypsin Deficiency. An Overview of Current Approaches

“…A PR3-specific fibrinogen epitope that is as promising and similar to Aα-Val360 is Aα-Val541. This fragment is also rather stable, can be measured in the blood of individuals, and is generated at the point of release of PR3 [ 62 ].…”

Relationship of CT densitometry to lung physiological parameters and health status in alpha-1 antitrypsin deficiency: initial report of a centralised database of the NIHR rare diseases translational research collaborative

“…The study was successful in enabling collaboration between academic centres by providing recruitment and in-depth phenotyping of a group of patients with AATD and a wide range of physiological impairment to an agreed standard. All subjects also had plasma taken and stored for the development and validation of assays to monitor serine proteinase footprints 16 and other disease-specific biomarkers. In addition, the network provides baseline cohort data for long-term follow-up and importantly a well-characterised patient database for collaboration with industry (one of the key aims for the NIHR programme).…”