Truncating mutations in the Wilson disease gene ATP7B are associated with very low serum ceruloplasmin oxidase activity and an early onset of Wilson disease

Abstract

Background: Mutations in the gene ATP7B cause Wilson disease, a copper storage disorder with a high phenotypic and genetic heterogeneity. We aimed to evaluate whether 'severe' protein-truncating ATP7B mutations (SMs) are associated with low serum ceruloplasmin oxidase activities and an early age of onset when compared to missense mutations (MMs).

Methods: The clinical phenotype of 59 genetically confirmed WD patients was analyzed retrospectively. Serum ceruloplasmin was measured by its oxidase activity with o-dianisidine dihydrochloride as substrate and immunologically.

Results: Thirty-nine patients had two MMs, 15 had the genotype SM/MM, and 5 patients had two SMs on their ATP7B alleles. Enzymatic and immunologic serum ceruloplasmin levels differed significantly between the three groups (P < 0.001 and P < 0.01, respectively). The lowest levels were measured in patients with two SMs (0.0 U/L; IQR, 0.0-0.0 U/L and 0.02 g/L; IQR, 0.01-0.02 g/L, respectively) and the highest in patients with two MMs (17.8 U/L; IQR, 5.8-35.1 U/L and 0.11 g/L; IQR,0.10-0.17 g/L, respectively). The age of onset was also significantly different between the three patient groups (P < 0.05), with SM/SM patients showing the earliest onset (13 years; IQR, 9-13 years) and patients with two MMs showing the latest onset (22 years; IQR, 14-27 years). By ROC curve analysis a ceruloplasmin oxidase level <or= 5 U/L can predict the presence of at least one SM with a sensitivity of 80% and a specificity of 79.5%.

Conclusions: In our German study cohort truncating ATP7B mutations were associated with lower ceruloplasmin serum oxidase levels and an earlier age of onset when compared to MMs. Measurement of serum ceruloplasmin oxidase might help to predict presence of truncating ATP7B mutations and might facilitate the mutation analysis.

Figure 1

Serum ceruloplasmin oxidase activity and immunoreactive serum ceruloplasmin in Wilson disease patients grouped according to their type of ATP7B mutation. Analysis of Wilson disease (WD) patients possessing either (1) two 'severe' mutations (SM/SM); (2) one 'severe' mutation and one missense mutation (SM/MM), or (3) two missense mutations (MM/MM). Distribution of data presented as box and whisker plots: 25^th percentile, median, 75^th percentile, maximum, outliers (circles), and extreme values (x). Both assays resulted in no indeterminate results or missing data. a, p-value for the comparison SM/SM with SM/MM patients; b, p-value for the comparison SM/SM with MM/MM patients. (A) Oxidase activity is significantly lower in SM/SM patients (0.0 U/L; IQR, 0.0 to 0.0) compared to SM/MM patients (3.1 U/L; IQR, 0.0 - 5.9) and MM/MM patients (17.8 U/L; IQR, 5.8 - 35.1). (B) Immunoreactive serum ceruloplasmin is significantly lower in SM/SM patients (0.02 g/L; IQR, 0.01 - 0.02) compared to SM/MM patients (0.07, g/L; IQR 0.06 - 0.10) and MM/MM patients (0.11 g/L; IQR, 0.10 - 0.17).

Figure 2

Age of disease onset in Wilson disease patients grouped according to their type of ATP7B mutation. Analysis of Wilson disease (WD) patients possessing either (1) two 'severe' mutations (SM/SM); (2) one 'severe' mutation and one missense mutation (SM/MM), or (3) two missense mutations (MM/MM). Distribution of data presented as box and whisker plots: 25^th percentile, median, 75^th percentile, maximum, outliers (circles), and extreme values (x). Both assays resulted in no indeterminate results or missing data. a, difference for the comparison with SM/MM patients; b, difference for the comparison with MM/MM patients. Age of onset is significantly earlier in SM/SM patients (13 years; IQR, 9 - 13) compared to SM/MM patients (16 years; IQR, 14 - 18) and MM/MM patients (22 years; IQR, 14 - 27).

Figure 3

ROC (receiver operating characteristic) curves for the prediction of presence of 'severe' ATP7B mutations by serum ceruloplasmin oxidase activity and immunoreactive serum ceruloplasmin. For serum ceruloplasmin oxidase activity (bold line) the area under the curve is 0.838 (95% CI, 0.724 - 0.953) and for immunoreactive ceruloplasmin (light line) the area under the curve is 0.785 (95% CI, 0.658 - 0.911).