Non-invasive testing for Wilson disease: Revisiting the d-penicillamine challenge test☆
Article Outline
Wilson disease must be considered in patients with unexplained liver disease, in patients with liver disease and neurological or psychiatric symptoms or for first degree relatives of patients. The diagnosis of Wilson disease is most challenging in the pediatric population where most patients present with liver disease and Kayser-Fleischer rings are frequently absent. The spectrum of liver disease ranges from asymptomatic to cirrhosis and acute liver failure. The group with acute liver failure is the most easily recognized as having Wilson disease by virtue of the presence of non-immune hemolytic anemia and a low serum alkaline phosphatase level (yielding a serum alkaline phosphatase to bilirubin ratio <2). The remaining patients are first examined for signs of chronic liver disease and by slit lamp exam for corneal Kayser-Fleischer rings, tested for liver synthetic dysfunction and evidence of hepatic inflammation and for levels of serum ceruloplasmin and urinary copper excretion. If there remains suspicion of disease, a liver biopsy is recommended, the histology examined and the content of copper in the specimen quantitated. Prior to the more widespread use of transjugular liver biopsies, coagulopathy often precluded the performance of biopsy by percutaneous means. This led to a reliance on urine copper excretion and promotion of stimulation testing with the copper chelator d-penicillamine as a non-invasive means for diagnosing Wilson disease.
More recently our armamentarium for diagnostic testing for Wilson disease has expanded to include de-novo molecular testing for disease specific ATP7B mutations along with haplotype analysis for siblings of probands. However, this test is not universally available to all patients at this time, and the effects of the various mutations and polymorphisms of ATP7B on the protein function and disease expression as well as the impact of other genes that may regulate ATP7B expression or are involved in other aspects of hepatic copper metabolism are incompletely understood at this time.
Given the variance in disease phenotype and the difficulties often encountered in establishing the diagnosis of Wilson disease, we are still seeking an inexpensive and easily reproducible non-invasive test for disease diagnosis. Muller et al. [1] re-examined the diagnostic utility for Wilson disease of the penicillamine challenge test (PCT) for urine copper excretion. They compared a cohort of patients with Wilson disease (n
=38) and a control population of patients with active liver disease (n=60), but unfortunately did not include a group of heterozygous carriers. They evaluated various diagnostic parameters in their cohort, including basal copper excretion over 24h and the PCT. For the PCT, they utilized the same protocol previously used for patients with acute liver failure and severe liver injury, namely the administration of 500mg of d-penicillamine at the start of and midway through a 24-h collection of urine for copper content [2].
The first question is whether the PCT improved diagnosis compared with basal urine copper excretion. In their study, using a threshold for 24h urine basal copper excretion of 1.6μmol, 94% (15/16) symptomatic patients with Wilson disease, 69% (9/13) of asymptomatic siblings with Wilson disease, 100% (3/3) with fulminant hepatic failure but also 22% (13/60) of controls had positive results. We are told that using a more stringent cutoff of 0.6μmol/24h (see Ref. [3]) all of the 5 patients who failed to meet the 1.6μmol/24h had positive results suggesting further evaluation for Wilson disease would be warranted. We are not told what the false positive rate for this lower threshold for 24h urine copper excretion was for the controls with active liver disease. If the result for this control group was significantly increased from the 13/60 using the higher threshold of 1.6μmol/L, then using the lower threshold would increase diagnostic sensitivity at the cost of specificity for this test. Using the PCT, 9/38 patients with Wilson disease from all categories failed to reach the threshold value of >25μmol/24h, but only 4/60 (7%) of controls without Wilson disease reached this value. Therefore the PCT did not increase sensitivity but rather only the specificity of the test. However without results of testing for heterozygotes who are likely to have intermediate values of hepatic copper, we do not know if the false positive rate for this test would be increased or not.
Is the potential gain in diagnostic specificity above basal urine copper excretion for the PCT as shown by Muller et al. [1] worth the effort? In all four of the asymptomatic siblings with Wilson disease who had basal 24h urine Cu excretion <1.6μmol, serum ceruloplasmin values were <200μg/L, three of four had PCT>25μmol and none had Kayser-Fleischer rings. The combination of elevated urine copper excretion and low ceruloplasmin may itself be adequate for a diagnosis of Wilson disease with the backup of molecular diagnosis. This begs the question of whether we should just use the lower threshold of 24h basal urine copper excretion along with ceruloplasmin as initial diagnostic testing, and then proceed to molecular testing for those with positive results and save liver biopsy for those with indeterminate results on molecular testing. For siblings of probands, clearly proceeding directly to molecular testing would be reasonable – either by haplotype analysis or direct ATP7B mutation analysis – both now commercially but not universally available for patients. Even if molecular testing were universally available, a recent series showed that 43% of individuals with phenotypic Wilson disease lacked detectable mutations (ideally one on each allele) for ATP7B [4]. Whether the results of testing in these individuals are a failure of current technique or a lack of knowledge of the full spectrum of molecular changes that impact on ATP7B function remains to be determined.
Other important clinical points can be abstracted from this manuscript. Firstly, Kayser-Fleischer rings were present in 7/16 patients less than 10 years of age; all but one were symptomatic and the youngest was 7 year old. This reinforces the need to utilize ophthalmological examination for testing for Wilson disease in these very young patients. Secondly, four of six patients with fulminant hepatic failure were female, consistent with a prior series of patients with liver failure due to Wilson disease [5]. Lastly, the use of a proposed scoring system for Wilson disease showed all the patients with Wilson disease to have a score greater than 4 (>2 suggests further evaluation is necessary [6]). However we are not told of the results for the controls. A prior publication addresses the utility of this index in pediatric patients [7].
Therefore, while the PCT may have some utility in the pediatric population in symptomatic patients, lowering the threshold for basal urine copper excretion and combining this with measurement of serum ceruloplasmin may be equally useful or even better than the PCT. Those with a normal ceruloplasmin and who lack Kayser-Fleischer rings should undergo molecular testing or be evaluated by liver biopsy for hepatic histology and copper quantitation. The frequency of false positives for the PCT in 4/60 controls and false negative results in 9/38 Wilson disease patients as well as the lack of studies for PCT in unaffected heterozygous carriers for Wilson disease suggest that we still do not know the true sensitivity and specificity of this test for testing for Wilson disease in all pediatric patients.
We are still searching for the holy grail of a single, cost effective and highly specific non-invasive test for Wilson disease. Molecular diagnosis, even with the current limitations, will be the likely answer to this quest. However until molecular diagnosis for Wilson disease advances further, becomes less expensive and is universally available, we are well armed with respect to having adequate tools for diagnosing Wilson disease with our current and time proven tests. The real challenge is not in the use of a d-penicillamine challenge for urine testing for Wilson disease, but to consider the diagnosis in the first place.
References
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- Diagnosis and phenotypic classification of Wilson disease. Liver Int. 2003;23:139–142
- . Evaluation of the scoring system for the diagnosis of Wilson’s disease in children. Liver Int. 2005;25:680–681
☆ The authors declare that they do not have anything to disclose regarding funding or conflict of interest with respect to this manuscript.
PII: S0168-8278(07)00284-X
doi:10.1016/j.jhep.2007.05.002
© 2007 European Association for the Study of the Liver. Published by Elsevier Inc. All rights reserved.
