Autoantibodies Against Lysosome Associated Membrane Protein-2 (LAMP-2) in Pediatric Chronic Primary Systemic Vasculitis

Abstract

Background: Anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) is a small vessel vasculitis in adults and children that commonly affects the kidneys. Although the frequent antigenic, and presumed pathogenic, targets of ANCA in AAV are proteinase-3 (PR3) and myeloperoxidase (MPO), ANCA against lysosome associated membrane protein-2 (LAMP-2), a lesser known ANCA antigen that is expressed on the glomerular endothelium, are present in some adults with AAV-associated renal disease. LAMP-2-ANCA has not been assessed in children with chronic systemic vasculitis, and, if present, would be a potentially valuable biomarker given that treatment decisions for these pediatric patients at diagnosis are largely informed by kidney function.

Methods: A custom ELISA, using commercially available reagents, was designed to detect autoantibodies to human LAMP-2 in serum. Sera obtained from 51 pediatric patients at the time of diagnosis of chronic primary systemic vasculitis (predominantly AAV) were screened. LAMP-2-ANCA titers were evaluated for correlation with clinical metrics of disease activity (pediatric vasculitis activity score [pVAS], C-reactive protein [CRP] concentration, and erythrocyte sedimentation rate [ESR]), MPO- and PR3-ANCA titers, and renal function (glomerular filtration rate [GFR], renal-specific pVAS, and serum creatinine concentration).

Results: LAMP-2-ANCA (>1,000 ng/ml) were detected in 35% (n = 18) of pediatric systemic vasculitis patients, of which, 10 (20% of all patients) were found to have high positive titers (>1,500 ng/ml). Undetectable or negative titres (<500 ng/ml) were identified in 12% (n = 6) of patients, those with titers between 500 and 1,000 ng/ml were considered low with unknown clinical relevance (53%, n = 27). Although LAMP-2-ANCA titers did not significantly differ between patients with AAV versus ANCA-negative vasculitis, only AAV patients had high concentrations (>1,500 ng/ml) of LAMP-2-ANCA. LAMP-2-ANCA titers did not correlate with measures of disease activity (pVAS, CRP, or ESR) at the time of diagnosis. In contrast, for patients with 12-month post diagnosis follow-up, a negative correlation was observed between the change in GFR (from diagnosis to 12-month follow-up) and LAMP-2-ANCA titer at diagnosis.

Conclusions: Moderate to high LAMP-2-ANCA titers were detected in 35% (18/51) of children with chronic systemic vasculitis affecting small-to-medium vessels. Although the highest concentrations of LAMP-2-ANCA in this population were observed in individuals positive for classic ANCA (MPO- or PR3-ANCA), similar to previous reports on adult patients, LAMP-2-ANCA titers do not correlate with classic ANCA titers or with overall disease activity at diagnosis. Renal disease is a common manifestation in systemic small-medium vessel vasculitis (both in adults and children, though more severe in children) and our preliminary data suggest LAMP-2-ANCA at diagnosis may be a risk factor for more severe renal disease.

Keywords: ANCA-associated vasculitis; LAMP-2; anti-neutrophil cytoplasmic antibody; lysosome-associated membrane protein-2; pediatric; systemic vasculitis.

Figure 1

Concentration of LAMP-2-ANCA in pediatric chronic small-to-medium vessel vasculitis patients. (A) LAMP-2-ANCA concentration (y-axis; ng/mL) in serum of individuals with early-onset vasculitis that are known to be positive (n = 5) and negative (n = 1) for LAMP-2-ANCA (squares) (14), children with vasculitis (n = 51, circles), and children with systemic autoinflammatory disease (n = 5, triangles). (B) LAMP-2-ANCA concentration (y-axis; ng/mL) in pediatric patients grouped (x-axis) based on positivity for MPO-ANCA (n = 19), PR3-ANCA (n = 23), or neither MPO- or PR3-ANCA (ANCA-, n = 7), and (C, D) LAMP-2-ANCA concentration (y-axis; ng/mL) plotted against (C) MPO-ANCA (x-axis; U/L) (n =19) and (D) PR3-ANCA (x-axis; U/L) (n = 23). Bars show median. Horizontal line divided low (<1,000 ng/mL) and moderate-high positive LAMP-2-ANCA (>1,000 ng/mL). Open symbols on the x-axis denote samples below the lower limit of detection of the assay (n = 4 patients with vasculitis, and n = 1 patient with autoinflammatory disease).

Figure 2

Comparison of LAMP-2-ANCA titer with standard clinical measures of disease activity. Concentration of LAMP-2-ANCA (y-axis; ng/mL) in pediatric vasculitis patients plotted against (A) C-reactive protein (CRP) concentration (x-axis; mg/L) (n = 51), (B) erythrocyte sedimentation rate (ESR) (x-axis; mm/h) (n = 44), and (C) pediatric vasculitis activity score (pVAS) (x-axis) (n = 51) at the time of diagnosis, and (D) blood samples taken prior to (naïve, n = 14), or after (treated, n = 37), induction of immune suppressive therapy. Bars show median. Horizontal line divided low (<1,000 ng/mL) and moderate-high positive LAMP-2-ANCA (>1000 ng/mL). Open symbols on the x-axis denote samples below the lower limit of detection of the assay (n = 4).

Figure 3

Comparison of LAMP-2-ANCA titer with renal metrics. Concentration of LAMP-2-ANCA (y-axis: ng/ml) in pediatric vasculitis patients: (A) with the absence (n = 15) and presence (n = 36) of proteinuria (x-axis) at the time of diagnosis. (B) plotted against change in GFR from the time of diagnosis to 12-month follow-up (x-axis, ml/min/1.73m²) (n = 25, p = 0.0314). (C) with no renal involvement (renal PVAS < 4; n = 5), and either renal improvement (increase in GFR at 12-month > 10 ml/min/1.73m², n = 8) or worsening (decrease in GFR at 12-month > 10 ml/min/1.73m², n = 12) from diagnosis to 12-month follow-up. (D) plotted against serum creatinine concentration (x-axis; µmol/L) (n = 26, p = 0.2149). Bars show median.