Introduction Alpha-1 antitrypsin (AAT) deficiency (AATD) is associated with early onset emphysema. The aim of this study was to investigate whether AAT binding to plasma constituents could regulate their activation, and in AATD, exploit this binding event to better understand the condition and uncover novel biomarkers of therapeutic efficacy.
Methods To isolate AAT linker proteins, plasma samples were separated by size exclusion chromatography, followed by co-immunoprecipitation. AAT binding proteins were identified by mass spectrometry. Complement turnover and activation was determined by ELISA measurement of C3, C3a and C3d levels in plasma of healthy controls (n=15), AATD (n=51), non-AATD patients with obstructive airway disease (n=10) and AATD patients post AAT augmentation therapy (n=5).
Results Direct binding of complement C3 to AAT was identified in vivo and in vitro. Compared with healthy controls, a breakdown product of C3, C3d, was increased in AATD (0.04 µg/mL vs 1.96 µg/mL, p=0.0002), with a significant correlation between radiographic pulmonary emphysema and plasma levels of C3d (R2=0.37, p=0.001). In vivo, AAT augmentation therapy significantly reduced plasma levels of C3d in comparison to patients not receiving AAT therapy (0.15 µg/mL vs 2.18 µg/mL, respectively, p=0.001).
Discussion Results highlight the immune-modulatory impact of AAT on the complement system, involving an important potential role for complement activation in disease pathogenesis in AATD. The association between plasma C3d levels and pulmonary disease severity, that decrease in response to AAT augmentation therapy, supports the exploration of C3d as a candidate biomarker of therapeutic efficacy in AATD.
- alpha1 antitrypsin deficiency
- rare lung diseases
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EPR and NGM are joint senior authors.
Contributors EPR, NGM and MEOB conceived and planned the study design, designed experiments, performed quality assurance and interpreted the data. EPR, NGM and MEOB wrote the manuscript; MEOB, LF, OJM, NB and KM carried out experiments; PM and MH assisted with proteomics and LC-MS/MS analysis and data interpretation; MEOB, LF and MPM performed statistical analysis. OJM and TC contributed to patient accrual and performed clinical data collection and analyses.
Funding The US Alpha-1 Foundation (EPR), the Medical Research Charities Group/Health Research Board Ireland (EPR and NGM), the European Alpha-1-Antitrypsin Laurell’s Training Award, sponsored by Grifols Inc (MEOB), 3U Biomedical Research (DCU-NUI Maynooth-RCSI) (PM, NGM and EPR), Beaumont Hospital Foundation, Dublin, Ireland and the Programme for Research in Third Level Institutes (PRTLI) administered by the Higher Education Authority (NGM).
Competing interests None declared.
Patient consent for publication Not required.
Provenance and peer review Not commissioned; externally peer reviewed.
Data availability statement All data relevant to the study are included in the article or uploaded as supplementary information. All data relevant to this study are included in the manuscript or uploaded as supplementary information.
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