Development and evaluation of an unlabeled probe high-resolution melting assay for detection of <i>ATP7B</i>
 mutations in Wilson's disease

Xu, Anjian; Zhang, Bei; Zhang, Wei; Ou, Xiaojuan; Huang, Jian

doi:10.1002/jcla.22064

Cited by 6 publications

(3 citation statements)

References 24 publications

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“…In populations in which a small number of pathological changes predominate, the prioritization of these changes, likely founder events, is a cost-effective strategy. For this, HRM (high resolution melting) techniques, customized SNP (single nucleotide polymorphism) microarray, or RFLPs (restriction-fragment-length-polymorphism) test can be used [ 72 , 73 , 74 , 75 ].…”

Section: A Mendelian Disease Caused By Mutations In Atp7bmentioning

confidence: 99%

Wilson’s Disease: Facing the Challenge of Diagnosing a Rare Disease

et al. 2021

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Wilson disease (WD) is a rare disorder caused by mutations in ATP7B, which leads to the defective biliary excretion of copper. The subsequent gradual accumulation of copper in different organs produces an extremely variable clinical picture, which comprises hepatic, neurological psychiatric, ophthalmological, and other disturbances. WD has a specific treatment, so that early diagnosis is crucial to avoid disease progression and its devastating consequences. The clinical diagnosis is based on the Leipzig score, which considers clinical, histological, biochemical, and genetic data. However, even patients with an initial WD diagnosis based on a high Leipzig score may harbor other conditions that mimic the WD’s phenotype (Wilson-like). Many patients are diagnosed using current available methods, but others remain in an uncertain area because of bordering ceruloplasmin levels, inconclusive genetic findings and unclear phenotypes. Currently, the available biomarkers for WD are ceruloplasmin and copper in the liver or in 24 h urine, but they are not solid enough. Therefore, the characterization of biomarkers that allow us to anticipate the evolution of the disease and the monitoring of new drugs is essential to improve its diagnosis and prognosis.

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Section: A Mendelian Disease Caused By Mutations In Atp7bmentioning

confidence: 99%

Wilson’s Disease: Facing the Challenge of Diagnosing a Rare Disease

et al. 2021

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“…Different ethnic groups have different frequencies of specific variants of the ATP7B [ 23 , 24 ]. In populations where a small number of pathogenic ATP7B variants predominate, it is possible to design relatively simple, inexpensive, and low-time-consuming genetic screening tests based on high-resolution melting (HRM), customized microarrays, or restriction fragment length polymorphism (RFLP) tests [ 25 , 26 , 27 , 28 ]. For example, in the Czech Republic, as well as in Poland, the screening for the five or six (respectively) most frequent pathogenic variants in ATP7B allows the confirmation of diagnosis in 70–80% of patients [ 29 , 30 ].…”

Section: Resultsmentioning

confidence: 99%

Wilson’s Disease—Genetic Puzzles with Diagnostic Implications

2023

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(1) Introduction: Wilson’s disease (WND) is an autosomal recessive disorder of copper metabolism. The WND gene is ATP7B, located on chromosome 13. WND is characterized by high clinical variability, which causes diagnostic difficulties. (2) Methods: The PubMed, Science Direct, and Wiley Online Library medical databases were reviewed using the following phrases: “Wilson’s disease”, “ATP7B genotype”, “genotype-phenotype”, “epigenetics”, “genetic modifiers”, and their combinations. Publications presenting the results of experimental and clinical studies, as well as review papers, were selected, which concerned: (i) the diversity of genetic strategies and tests used in WND diagnosis; (ii) the difficulties of genetic diagnosis, including uncertainty as to the pathogenicity of variants; (iii) genetic counseling; (iv) phenotypic effects of ATP7B variants in patients with WND and in heterozygous carriers (HzcWND); (v) genetic and epigenetics factors modifying the clinical picture of the disease. (3) Results and conclusions: The genetic diagnosis of WND is carried out using a variety of strategies and tests. Due to the large number of known variants in the ATP7B gene (>900), the usefulness of genetic tests in routine diagnostics is still relatively small and even analyses performed using the most advanced technologies, including next-generation sequencing, require additional tests, including biochemical evidence of abnormal copper metabolism, to confirm the diagnosis of WND. Pseudodominant inheritance, the presence of three various pathogenic variants in the same patient, genotypes indicating the possibility of segmental uniparental disomy, have been reported. Genotype–phenotype relationships in WND are complex. The ATP7B genotype, to some extent, determines the clinical picture of the disease, but other genetic and epigenetic modifiers are also relevant.

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“…Therefore, the p.P992L variant was not included in our WD microarray. However, based on the high‐resolution melting curve (HRM), we have developed a detection method for p.P992L and applied for a patent in order to make up for the shortcomings of this site 17 . With the identification of novel recurrent ATP7B mutation and specifically, hotspot mutations in other countries such as p.H1069Q in the western countries, more probes can be included in the microarray, and this method can be used for the detection of more ATP7B mutations in patients worldwide.…”

Section: Discussionmentioning

confidence: 99%

Laboratory and clinical evaluation of a microarray for the detection of ATP7B mutations in Wilson disease in China

Jia

Zhang

et al. 2022

Clinical Laboratory Analysis

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Background and objective Wilson disease (WD) is an autosomal recessive copper metabolic disorder caused by mutations in ATP7B. Sanger sequencing is currently used for ATP7B variant identification. However, the ATP7B gene contains 21 exons, which makes sequencing of the entire gene both complex and time‐consuming. Therefore, a simpler assay is urgently needed. Methods We performed a laboratory and clinical evaluation of an oligonucleotide microarray for the detection of 24 ATP7B recurrent mutations (except p.P992L) in Chinese patients with WD. Results The accuracy of the microarray was evaluated by screening for ATP7B mutations in 126 patients including 106 suspected WD samples and 20 patients with other liver diseases as negative control. Results were confirmed by Sanger sequencing. We established a reliable microarray system for the rapid detection of the 24 ATP7B mutations, with a sensitivity of 30 ng/test genomic DNA and specificity of 100% for all loci; the coefficient of variation in repeatability tests was <10%. Clinical evaluation showed an overall concordance between the microarray detection and sequencing of 100%, and 81.13% (86/106) of suspected WD cases showed ATP7B mutations by microarray detection. Microarray and Sanger sequencing identified p.R778L (50.94%), p.A874V (17.92%), p.P992L (11.32%), p.V1106I (11.32%), and p.I1148T (6.60%) as the most common mutations in WD patients. Conclusions Our microarray system is customizable and easily used for high‐throughput detection of certain recurrent ATP7B mutations, providing a simpler method suitable for WD genetic diagnosis in China.

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Development and evaluation of an unlabeled probe high-resolution melting assay for detection of ATP7B mutations in Wilson's disease

Abstract: The newly developed assay to rapidly genotype the ATP7B mutations is convenient, accurate, and reproducible, and represents a favorable alternative to Sanger sequencing in the identification of specific ATP7B mutations.

Cited by 6 publications

References 24 publications

Wilson’s Disease: Facing the Challenge of Diagnosing a Rare Disease

Wilson’s Disease: Facing the Challenge of Diagnosing a Rare Disease

Wilson’s Disease—Genetic Puzzles with Diagnostic Implications

Laboratory and clinical evaluation of a microarray for the detection of ATP7B mutations in Wilson disease in China

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