Targeting IgG Autoantibodies for Improved Cytotoxicity of Bactericidal Permeability Increasing Protein in Cystic Fibrosis

Abstract

In people with cystic fibrosis (PWCF), inflammation with concurrent infection occurs from a young age and significantly influences lung disease progression. Studies indicate that neutrophils are important effector cells in the pathogenesis of CF and in the development of anti-neutrophil cytoplasmic autoantibodies (ANCA). ANCA specific for bactericidal permeability increasing protein (BPI-ANCA) are detected in people with CF, and correlate with infection with Pseudomonas aeruginosa. The aim of this study was to determine the signaling mechanism leading to increased BPI release by CF neutrophils, while identifying IgG class BPI-ANCA in CF airways samples as the cause for impaired antimicrobial activity of BPI against P. aeruginosa. Plasma and/or bronchoalveolar lavage fluid (BAL) was collected from PWCF (n = 40), CF receiving ivacaftor therapy (n = 10), non-CF patient cohorts (n = 7) and healthy controls (n = 38). Plasma and BAL BPI and BPI-ANCA were measured by ELISA and GTP-bound Rac2 detected using an in vitro assay. The antibacterial effect of all treatments tested was determined by colony forming units enumeration. Levels of BPI are significantly increased in plasma (p = 0.007) and BALF (p < 0.0001) of PWCF. The signaling mechanism leading to increased degranulation and exocytosis of BPI by CF neutrophils (p = 0.02) involved enhancement of Rac2 GTPloading (p = 0.03). The full-length BPI protein was detectable in all CF BAL samples and patients displayed ANCA with BPI specificity. IgG class autoantibodies were purified from CF BAL complexed to BPI (n=5), with IgG autoantibody cross-linking of antigen preventing BPI induced P. aeruginosa killing (p < 0.0001). Results indicate that the immune-mediated diminished antimicrobial defense, attributed to anti-BPI-IgG, necessitates the formation of a drug/immune complex intermediate that can maintain cytotoxic effects of BPI towards Gram-negative pathogens, with the potential to transform the current treatment of CF airways disease.

C3d Elicits Neutrophil Degranulation and Decreases Endothelial Cell Migration, with Implications for Patients with Alpha-1 Antitrypsin Deficiency

“…Our own studies have previously shown AATD as a risk factor for developing neutrophilic ulcerative panniculitis [ 50 ], increased anti-lactoferrin autoantibodies [ 14 ], and increased titers of anti-CCP in MZ-AATD patients with rheumatoid arthritis (RA) [ 51 ]. In patients with cystic fibrosis (CF), increased anti-BPI autoantibodies have been detected [ 52 , 53 ], but although data of the current study demonstrate that BPI levels are significantly increased in the plasma of AATD individuals, no link between AATD and anti-BPI antibodies has been established [ 54 ].…”

Altered Degranulation and pH of Neutrophil Phagosomes Impacts Antimicrobial Efficiency in Cystic Fibrosis

“…In the current study we observed increased primary granule degranulation and MPO accumulation in the CF phagosome, yet decreased bacterial killing, supporting the concept of unsuitable conditions in the CF phagosome for optimal MPO peroxidase activity. The exact mechanism leading to increased primary granule release by CF cells most likely involves increased Rac2 activation (59). Moreover, impaired GTP-Rab27a activation in CF blood neutrophils has been shown to decrease secondary and tertiary granule degranulation to the outside of the cell in response to soluble stimuli (20).…”

Clinical significance of BPI-ANCA in patients with cystic fibrosis: a single center prospective study

“…These ANCA antibodies with BPI specificity (BPI-ANCA) have been identified in several inflammatory disorders and infectious diseases, such as CF, chronic obstructive lung disease, rheumatoid arthritis, vasculitis, and inflammatory bowel disease 3 , 4 . This BPI-ANCA complex prevents BPI-induced P. aeruginosa killing 4 , 5 . In a recent meta-analysis, the prevalence of BPI-ANCA in patients with CF varied from 17.9 to 83% with a pooled prevalence of nearly 50% 6 .…”