Phenotypic and Genetic Characterization of children with Wilson Disease from Northeast China

doi:10.21203/rs.3.rs-2858430/v1

Download PDF

Research Article

Phenotypic and Genetic Characterization of children with Wilson Disease from Northeast China

https://doi.org/10.21203/rs.3.rs-2858430/v1

This work is licensed under a CC BY 4.0 License

You are reading this latest preprint version

Background

Wilson's disease (WD) is an autosomal recessive genetic disease caused by ATP7B gene mutations and characterized by copper metabolism disorders.

Objective

This study aimed to highlight the phenotypic and genetic characteristics of children with WD in Northeast China.

Methods

We retrospectively analyzed clinical data and gene sequencing results of 65 children with WD from January 1, 2014 to December 31, 2022 in Shengjing Hospital of China Medical University.

Results

The mean age at the time of the diagnosis of WD was 6.3 ± 3.52 years (range 1.2–15 years). Fifty cases (50/65, 76.9%) were asymptomatic and only found abnormal liver function during physical examination. However, they had negative Kayser–Fleischer (KF) ring with significant difference (p < 0.05). Children with acute liver failure had significantly elevated 24hr urinary Cu excretion (p < 0.05). We detected a total of 46 gene mutations in ATP7B gene, including 7 novel mutations. The most frequent mutation was p.R778L with an allelic frequency of 38.7%. Phenotype–genotype correlation analysis suggested that p.R778L was significantly associated with lower levels of serum ceruloplasmin and higher zinc levels (p < 0.05). LOF (loss of function) mutation was significantly associated lower albumin (p < 0.05).

Conclusion

Most children with WD are asymptomatic, which makes early diagnosis of WD challenging, and it is necessary to perform genetic test. p.R778L is the most frequently mutation of ATP7B gene in China and may play an important role in lower levels of serum ceruloplasmin.

Wilson disease

ATP7B gene

children

China

Wilson disease (WD) is a rare autosomal recessive disorder caused by mutations in the ATP7B gene. It is characterized by decreased biliary copper excretion, leading to excessive copper accumulation in different organs, including the liver, brain and cornea^［¹^］. The prevalence of this disease worldwide is 1/2 600-1/30 000, and the carrier frequency is about 1/90^［^2-3^］.

WD can develop at any age, but it is common in 5-35 years old^［⁴^］.WD is increasingly diagnosed in children who are younger than 5 years old, and clinical findings may be nonspecific in children who are younger than 2 years old^［⁵^］. Most children showed dysfunction of liver ranging from incidental finding of increased serum transaminases in otherwise asymptomatic, acute hepatitis, hepatomegaly, to acute liver failure (ALF) or cirrhosis^［⁶^］. Diagnosis of WD is difficult in children because they are often asymptomatic. However genetic testing has become an increasingly important method of early, rapid, and accurate diagnosis, which is essential in diagnosis^［^5,6^］.

ATP7B gene was located on the short arm of chromosome 13^［⁷^］.To date, more than 700 mutations have been identified^［⁸^］. The most common mutation in patients from Europe is the point mutation of H1069Q (exon 14), and Asia is R778L (exon 8)^［⁹^］. More than 50% of the mutations are missense mutations and nonsense mutations, and the rest can be insertion/deletion and splicing site mutations^［^10,11^］. Some studies have shown that mutations affecting critical portions of the protein including copper-binding domains or the ATPase loop may lead to early onset of hepatic disease^［¹²^］. There are also studies indicating that p.H1069Q was more often associated to a neurologic presentation and a later onset of symptoms^［^13,14^］. Loss of function (LOF) of ATP7B gene variants have been associated with ALF and earlier age of disease onset^［^15,16^］. However, there are no confirmed genotype–phenotype correlations^［¹⁷^］.

The aim of our study is to summarize the clinical manifestations, laboratory and genetic characteristics of children with WD in Northeast China. We want to determine the spectrum and frequency of mutations in the ATP7B gene and genotype–phenotype correlation of pediatric WD patients.

Patients and groups

A total of 65 pediatric WD patients admitted to Shengjing Hospital of China Medical University between January 1, 2014 and December 31, 2022 were enrolled in this study. The diagnosis of WD was established according to the scoring system provided by the Eighth International Meeting on Wilson disease^［¹^］ and a score > 4 was considered reasonable for making a diagnosis of WD. The medical history, physical examination, laboratory examination and imaging findings were all collected as clinical data. All the patients and their relatives provided oral consent, and this study was approved by the Ethics Committee of Shengjing Hospital of China Medical University (2022PS107K).

All children with WD were classified into four groups according to the type of presentations as followings: Group 1 (n=50): Asymptomatic cases with only elevation of serum transaminases who had been diagnosed during routine physical examination; Group 2 (n=5): Patients with biochemical abnormalities and abnormal hepatic manifestations including jaundice, nausea, abdominal pain and ascites; Group 3 (n=4): patients with ALF were diagnosed in accordance with the criteria of the Pediatric Acute Liver Failure Study^[18-20]; Group 4 (n=6): patients with an impressive spectrum of neurological, behavioral or psychiatric disorders.

LOF mutations were defined as nonsense (substitution in coding base pair predicted to result in a premature stop codon), frameshift (insertion or deletion predicted to result in a shift in the reading frame), deletions (exon deletions), and splice site (substitutions predicted to involve a splice site), which produced a non-functional protein product^［²¹.

Mutational analysis of the ATP7B gene

Genomic DNA was extracted from peripheral blood samples. All of the cases were tested with ATP7B targeted gene panel sequencing or whole-exome sequencing. All test protocols, including DNA extraction, construction of gene library, high-throughput sequencing, data analysis, Sanger sequencing verification, and bioinformatics analysis, were carried out by Clinical genetics laboratory in Shengjing Hospital of China Medical University and commercial companies including My Genostic (Beijing) and Kingmed (The Guangzhou Genomics Institute, Shenyang).

Statistical Analysis

All numeric variables were expressed as mean ± standard deviation (SD). Discrete variables were presented using patient numbers or percentages (%). Comparison of different variables in various groups was done using analysis of variance (ANOVA) and Student's t tests. The Chi-square test was used to study discrete variables. Analyses were conducted using Statistical Product and Service Solutions (SPSS 22.0) software. A P value less than 0.05 was considered statistically significant.

Clinical features of pediatric WD patients

There were 65 children with WD, including 34 males and 31 females with mean age at the time of the diagnosis was 6.3 ± 3.52 years (range 1.2-15 years). Fifty cases (50/65, 76.9%) were only found abnormal liver function during health examination. Cases of children with symptoms included neuropsychiatric symptoms (6/65, 9.2%), ascites (3/65,4.6%), nausea (2/65, 3.1%), jaundice (2/65, 3.1%), lower limbs edema (2/65, 3.1%), and liver pain (1/65,1.5%). Five patients (5/65，7.7%) had a family history of WD. Fourteen patients (14/65，21.5%) showed positive Kayser–Fleischer (KF) ring under slit-lamp, and the youngest child with positive KF ring was 6 years. All asymptomatic patients showed negative KF ring, which was significantly different from the other three groups (p< 0.05). Brain MRI abnormalities, including abnormal signal intensity in the basal ganglia, thalamus, brainstem and corpus callosum, were seen in 6 patients (6/65，9.2%).

Among all the 65 patients, 64 (98.5%) had low serum ceruloplasmin levels (<20 mg/dl), 42 (64.6%) had urinary Cu excretion over 100μg/24hr. WD patients with ALF in Group 3 accompanied by increased total bilirubin and direct bilirubin, decreased albumin, increased white blood cell (WBC) and decreased hemoglobin (Hb). Group 3 also had significantly elevated 24hr urinary Cu excretion compared to the other three groups (p< 0.05). Group 3 and Group 4 had significantly lower blood platelet than other two groups (p< 0.05) (Table 1).

TABLE 1. Clinical and laboratory characteristics of 65 pediatric WD patients at diagnosis

Parameter	G1（n=50）	G2（n=5）	G3（n=4）	G4（n=6）	P value
Males	26（52.0）	3（60.0）	2（50.0）	3（50.0）	0.986
Age at diagnosis (y，mean+SD)	4.92.6	9.83.3	112.2	11.5+1.2	0.000
Hepatomegaly	14（28.0）	3（60.0）	1（25.0）	2（33.3）	0.519
Splenomegaly	1（2.0）	1（20.0）	3（75.0）	1（16.7）	0.000
Jaundice	0（0）	0（0）	2（50.0）	0（0）	0.000
Edema	0（0）	0（0）	3（75.0）	1（16.7）	0.000
Ascites	0（0）	0（0）	2（50.0）	0（0）	0.000
Hemolytic anemia	0（0）	0（0）	1（25.0）	0（0）	0.000
Neuropsychiatric deficit	0（0）	0（0）	0（0）	6（100.0）	0.000
Family history	5（10.0）	0（0）	0（0）	0（0）	0.654
KF ring (+)	0（0）	4（80.0）	4（100.0）	6（100.0）	0.000
Brain MRI	0（0）	0（0）	1（25.0）	6（100.0）	0.000
Ceruloplasmin (mg/dl)	5.04.5	6.75.3	6.85.9	4.22.4	0.691
Urinary copper (μg/24h)	124.493.2	262.3196.5	460.4428.8	296.06114.4	0.000
ALT (IU/L)	213.74123.5	146.8102.6	131.388.2	83.52.6108.6	0.048
AST (IU/L)	115.069.8	97.628.0	168.5131.0	67.543.1	0.166
TB (mg/dL)	7.42.6	22.5221.8	218.4277.0	14.77.8	0.000
DB (mg/dL)	2.31.0	14.317.9	183.7271.2	5.94.2	0.000
ALB (g/L)	43.12.5	34.46.9	26.12.5	37.77.1	0.000
WBC (mm3)	6.82.0	5.01.6	22.334.1	4.62.2	0.002
Hb (g/dL)	127.07.8	123.46.0	86.330.5	126.811.9	0.000
PLT (mm3)	327.183.2	206.0173.5	137.5132.2	144.3118.5	0.000
Zinc (mg/dl)	73.412.2	63.011.7	62.36.4	70.912.1	0.114

Abbreviations: WD, Wilson's disease; KF ring, Kayser–Fleischer ring; MRI, magnetic resonance imaging; ALT, alanine aminotransferase; AST, aspartate aminotransferase; TB, total bilirubin; DB, directly bilirubin; ALB, Albumin protein; WBC, white blood cell; Hb, hemoglobin; PLT, platelets; G1, Asymptomatic cases with only elevation of serum transaminases; G2, Patients with biochemical abnormalities and hepatic presentation; G3, WD patients with acute liver failure at diagnosis; G4, WD patients with neurological/psychiatric symptoms.

Gene mutation analysis

We identified 46 gene mutations in ATP7B (Table 2 and Figure 1), and seven of them are novel. The novel mutations included c.2572A>C, c.-362G>A, c.51+4A>C, c.1543+40G>A, c.3903+2T>C, c.2664-2665delCC, c.3524-3528delAAGGA.

The most frequent mutation was p.R778L with an allelic frequency of 38.7% followed by p.P992L (6.5%), p.R919G and p.A874V (4.9% each), p.V1216M(3.2%), p.N1270S and p. Q1372Ter (2.4% each).

Of all 46 identified mutations 34 were classified as mild mutations, including 31missense mutations, 2 synonymous and 1 in-frame deletion mutation. Twelve were LOF mutations, including 5 frameshift mutations, 3 splice site variants and 4 nonsense mutations.

The 46 mutations were distributed throughout all exons of ATP7B except for exons 5, 7, 9, and 21. All the exons harboring the highest percentage of mutations were exons 8, 11, 12, 13 and 18 (Figure 2). The total mutation detection rate on these 5 exons was 75%, suggesting that these exons could be important regions for detecting mutations in our study.

TABLE 2. Spectrum of gene mutations in the ATP7B gene of this study

Exon	Nucleotide mutation	Amino acid change	Mutation type	Pathogenicity	Doman	Allele frequency (%)
1	c.51+4A>C*	－	splice site	Uncertain	Before Cu1	0.81
2	c.122A>G	p. N41S	Missense	Likely Pathogenic	Before Cu1	0.81
2	c.-362G>A*	－	Nonsense	Uncertain	Cu1	0.81
2	c.525DupA*	p. V176Sfs*28	Frameshift	Pathogenic	Cu2	0.81
3	c.1488C>T	p. G496=	Synonymous	Uncertain	Cu2	0.81
4	c.1543+40G>A*	－	splice site	Uncertain	Cu5	0.81
6	c.1924G>T	p. D642Y	Missense	Pathogenic	bet Cu6/TM1	1.61
8	c.2128G>A	p. G710S	Missense	Pathogenic	TM2	1.61
8	c.2132G>A	p. G711E	Missense	Pathogenic	TM2	0.81
8	c.2145C>T	p. Y715=	Synonymous	Uncertain	TM4	0.81
8	c.2252C>A	p. A751D	Missense	Likely Pathogenic	TM4	0.81
8	c.2333G>T	p. R778L	Missense	Pathogenic	TM4	38.71
8	c.2337G>A*	p. W779X	Nonsense	Pathogenic	TM4	0.81
8	c.2293G>A	p. D765N	Missense	Pathogenic	TM4	0.81
10	c.2572A>C	p. T858P	Missense	Likely Pathogenic	Td	0.81
11	c.2621C>T	p. A874V	Missense	Pathogenic	bet Td/TM5	4.84
11	c.2633A>C	p. N878T	Missense	Pathogenic	bet Td/TM5	0.81
11	c.2662A>C	p. T888P	Missense	Pathogenic	bet Td/TM5	0.81
11	c.2664-2665delCC*	p. H889fs	Frameshift	Pathogenic	bet Td/TM5	0.81
11	c.2668G>A	p. V890M	Missense	Pathogenic	bet Td/TM5	0.81
12	c.2755C>G	p. R919G	Missense	Pathogenic	bet Td/TM5	4.84
12	c.2785A>G	p. I929V	Missense	Pathogenic	TM5	0.81
12	c.2790-2792delCAT	p. I930del	In-frame deletion	Likely Pathogenic	TM5	0.81
13	c.2924C>A	p. S975Y	Missense	Pathogenic	TM6	1.61
13	c.2930C>T	p. T977M	Missense	Pathogenic	TM6	0.81
13	c.2975C>T	p. P992L	Missense	Pathogenic	bet TM6/Ph	6.45
13	c.3029A>C	p. K1010T	Missense	Pathogenic	bet TM6/Ph	0.81
14	c.3140A>T	p. D1047V	Missense	Pathogenic	ATP loop	0.81
14	c.3168delT*	p. L1057fs	Frameshift	Pathogenic	ATP loop	0.81
15	c.3316G>A	p. V1106I	Missense	Pathogenic	ATP loop	1.61
16	c.3459G>T	p. Trp1153Cys	Missense	Pathogenic	ATP loop	0.81
16	c.3517G>A	p. E1173K	Missense	Pathogenic	ATP loop	1.61
16	c.3524-3528delAAGGA*	－	Frameshift	Pathogenic	ATP loop	0.81
16	c.3532A>G	p. T1178A	Missense	Pathogenic	ATP loop	0.81
17	c.3634A>G	p. M1212V	Missense	Likely Pathogenic	ATP bind	0.81
17	c.3646G>A	p. V1216M	Missense	Pathogenic	ATP bind	3.22
18	c.3797G>A	p. G1266E	Missense	Likely Pathogenic	ATP hinge	1.61
18	c.3809A>G	p. N1270S	Missense	Pathogenic	ATP hinge	2.42
18	c.3818C>T	p. P1273L	Missense	Pathogenic	ATP hinge	0.81
18	c.3851T>A	p. I1284N	Missense	Likely Pathogenic	ATP hinge	0.81
18	c.3903+2T>C*	－	splice site	Likely Pathogenic	bet ATP hinge/TM7	0.81
19	c.3955C>T*	p. R1319X	Nonsense	Pathogenic	bet ATP hinge/TM7	0.81
19	c.4006delA*	p. I1336fs	Frameshift	Pathogenic	TM7	1.61
20	c.4064G>T	p. G1355V	Missense	Likely Pathogenic	TM8	0.81
20	c.4112T>C	p. L1371P	Missense	Pathogenic	TM8	0.81
20	c.4114C>T*	p. Q1372X	Nonsense	Pathogenic	TM8	2.42

Novel mutations are highlighted in bold

*: LOF mutations were defined as nonsense, frameshift, deletions, and splice site.

Evaluation of the correlation of phenotype and genotype

We compared the two most common mutations c.2333G>T (p.R778L) and c.2975C>T (p.P992L) with other mutations of the age at the first onset of symptoms and clinical parameters. Among all the 65 patients, 41 (63.1%) had mutation of p.R778L, 8 were homozygous and 33 were heterozygous. We observed that patients carrying the p.R778L mutation had a lower serum ceruloplasmin levels and higher Zinc levels than patients with other mutations at both alleles (p<0.05) (Fig. 3a, Fig. 3b). Eight (12.3%) had mutation of p.P992L, which were all heterozygous, and there was no significant association between the types of mutation and phenotype.

Among all the 65 patients, 15 (23.1%) had LOF mutation on one allele chromosome. We observed that patients carrying LOF mutation had a lower level of albumin than patients without LOF mutation at both alleles (p<0.05) (Fig. 3c, Fig. 3b). A total of 4 WD patients with ALF were identified (Fig. 3d, Table 3). Three had LOF mutations on one allele chromosome and missense mutations on the other allele chromosome. One had missense mutation and synonymous mutation on each allele chromosome.

TABLE 3 Clinical and mutational data in four pediatric WD patients with ALF

Patient	ATP7B mutations	Mutation type	Sex	Age at diagnosis (y)	KF ring	Neuropsychiatric deficit	Hemolytic anemia	Outcome
1	c.2333G>T	missense	M	13	+	-	-	Survival
	c.3524-3528delAAGGA*	frameshift
2	c.2333G>T	missense	F	11	+	-	-	Death
	c.3955C>T*	nonsense
3	c.1924G>T	missense	M	8	+	-	-	Liver transplant
	c.3903+2T>C*	splice site
4	c.2128G>A	missense	F	12	+	-	+	Survival
	c.1488C>T	Synonymous

*: LOF mutations were defined as nonsense, frameshift, deletions, and splice site.

Most of the children (76.9%) in our study were asymptomatic, with only elevation of serum transaminases who had been diagnosed during routine physical examination. The onset age of diagnosis of asymptomatic children are 4.92.6 years, which was younger than the other groups with statistical significance. Asymptomatic children have no specific clinical manifestations, but manifestations of hepatomegaly and fatty liver are often found by liver ultrasound. Untreated, asymptomatic patients with evidence of organ damage typically progress to symptomatic WD^［⁵^］. Therefore, early diagnosis of WD is critical to achieve a better outcome. However, early diagnosis of asymptomatic children in WD can be challenging. Thus, it is necessarily to perform gene test for asymptomatic children with WD.

There have been researches that ALF is the leading cause of death in children with WD, if untreated, carries an almost 95% mortality^[4]. In our study, 4 pediatric WD patients with ALF were identified, with 2 improved after corresponding treatment, 1 improved after liver transplantation, and 1 died due to failure to receive liver transplantation in time. Expeditious diagnosis is critically important because these patients require urgent liver transplantation to survive ^[22].

There were 6 patients with an impressive spectrum of neurological, behavioral or psychiatric disorders. The onset age of children in neurological group is 11.5+1.2 years. Except involuntary movements, tremor, and dysarthria due to extrapyramidal involvement, it can be combined with cognitive difficulties, depressive disorder, sleep disorders and other psychiatric symptoms. Besides, 3 patients showed cirrhosis and splenomegaly by liver ultrasound. All patients had brain MRI abnormalities, including abnormal signal intensity in the basal ganglia, thalamus, brainstem and corpus callosum.

The KF ring, which is absent in asymptomatic children of our study, has been found in 4 children with hepatic manifestations. All patients with ALF as well as all patients with neurological symptoms in our study had a KF ring. In previous study, KF rings are present in 44%-62% of adult patients with mainly hepatic disease^[23-24]. However, KF rings are usually absent in children presenting with liver disease^[25]. This suggests that although the KF ring is a specific manifestation of WD, the positive rate is very low in the early stage, especially in asymptomatic children. KF ring is often associated with disease progression, especially with neurological and severe hepatic manifestations in WD. Therefore, KF ring cannot be used as a basis for early diagnosis.

Previous study suggested that 20% of pediatric WD patients had normal ceruloplasmin^[26]. However, only 1 (1.5%) patient in our study has normal level of ceruloplasmin (>0.20 g/L). It seems that in our study, it is very uncommon to have a normal ceruloplasmin level, suggesting that different races may have different levels of ceruloplasmin, which may be attributed to different genotype.

In our study, 92% of the children showed 24hr urinary Cu >40μg/24h, and 64.4% of the children showed 24hr urinary Cu >100μg/24h. In addition, children in Group 3 with ALF also had significantly elevated 24hr urinary Cu excretion compared to the other three groups with statistical significance (p<0.05). These results suggest that >40μg seems to be more sensitive to the abnormality of urinary Cu, especially in the early asymptomatic period. The abnormal high level of 24hr urinary Cu is often associated with ALF.

Previous studies mainly focus on southern China, while there are few reports on northern China. In our study, the high frequency are p.R778L, p.P992L, p.R919G and p.A874V, accounting for 54.8% of all mutations, which are different from previous studies in southern China, which top three mutations are p.R778L, p.P992L and p.T935M, accounting for 50%~60% of the all mutations^[27-29]. In addition, in southern China the major ATP7B variants focus on exons 8, 12, 13, and 16^[30,31], whereas the variants detected in our study are mostly in exons 8, 11, and 13, and the results are the same with the exons prevalent in Qingdao, also in northern China^[32]，suggesting that these regions might be the hotspot regions for variants in the northern China.

There are 7 novel mutations previously unreported, including c.2572A>C, c.-362G>A, c.51+4A>C, c.1543+40G>A, c.3903+2T>C, c.2664-2665delCC, c.3524-3528delAAGGA, that have not been included in the Exome Aggregation Consortium database and 1000 Genomes Project database so far. They are predicted to be pathogenic or likely pathogenic by bioinformatics software analysis.

The relationship between clinical phenotypes and genotypes is complex. Most studies considered the phenotypic variability in WD is possibly due to additional genetic, epigenetic, and even environmental factors, which play a role in the timing of disease onset and the clinical phenotype^[33-36]. First, we suggest the symptoms are mainly related to the age of onset because of progressive toxic accumulation of Cu in liver and other organ. In addition, there has been reports of predominance of female patients with ALF^[37-38] due to the effect of sex hormone, however our results are not consistent with prior findings, which may need further larger sample of study. Second, different clinical phenotype may be attributed to different genotype, but there is lack of large-scale trial results. In our study, LOF mutation was significantly associated lower albumin, which may be associated with ALF, which was similar to previous researches that LOF mutation may be associated with ALF^{[15, 16]}. LOF mutations include nonsense, frameshift, deletions, and splice site, which produced a non-functional protein product. It might be expected that nonsense and frameshift mutations will result in profoundly affected function of ATP7B due to the absence of a full-length gene product. Nonsense and frameshift mutations often result in the production of highly unstable mRNA, which disrupt any Cu transporting function^[15,39]. European studies suggested that patients with LOF mutations had lower serum ceruloplasmin oxidase activity and an earlier onset of WD than patients with missense mutations^{[15,16,21,38,39]}. Recent research suggests that the presence of at least 1 LOF variant is associated with a worse transplant-free survival over a long period of follow-up for patients on chelation^[40]. Furthermore, we observed that patients carrying the p.R778L mutation had a lower serum ceruloplasmin levels and higher Zinc levels than patients with other mutations at both alleles. Recent study showed that p.R778L was related to lower levels of ceruloplasmin as well^{[28, 31]}. In addition, clinical phenotype may depend on the age of diagnosis and early intervention. In recent years, due to the popularization and acceptance of gene technology by WD parents, more and more children have been diagnosed and treated early before clinical symptoms, which may delay the symptoms of liver diseases and nervous systems.

In conclusion, the diagnosis of WD can be challenging in children, particularly at the early stages of liver disease and in the case of neurological presentation. Therefore, clinical and laboratory characteristic and genetic testing support are essential.

Ethics approval and consent to participate:All the patients and their relatives provided oral consent, and this study was approved by the Ethics Committee of Shengjing Hospital of China Medical University (2022PS107K). This study was conducted in accordance with the Declaration of Helsinki.

Consent for publication:All participants agreed to their data being published anonymously in a journal article.

Availability of data and material: The datasets are available from the corresponding author on reasonable request.

Competing interests: All authors declare no conflict of interest.

Funding: This work has no funding.

Author contribution: Tianhe Zhang(ZTH) contributed to conceptualization, methodology and original draft preparation. Wenliang Song(SWL) contributed to visualization, software, data curation and editing. Zhiqin Mao(MZQ) contributed to conceptualization, validation, review and supervision, project administration, and funding acquisition. All authors have read and agreed to the published version of the manuscript(All authors read and approved the final Manuscript).

Acknowledgements: The author thank all the team members for corporation.

Scheinberg IH. Wilson's disease. J Rheumatol Suppl. 1981 Jan-Feb; 7:90-3.
Bandmann O, Weiss KH, Kaler SG. Wilson's disease and other neurological copper disorders. Lancet Neurol. 2015 Jan;14(1):103-13.
Lo C, Bandmann O. Epidemiology and introduction to the clinical presentation of Wilson disease. Handb Clin Neurol. 2017; 142:7-17.
European Association for Study of Liver. EASL Clinical Practice Guidelines: Wilson's disease. J Hepatol. 2012 Mar;56(3):671-85.
Schilsky ML, Roberts EA, Bronstein JM, Dhawan A, Hamilton JP, Rivard AM, Washington MK, Weiss KH, Zimbrean PC. A multidisciplinary approach to the diagnosis and management of Wilson disease: Executive summary of the 2022 practice guidance on Wilson disease from the American Association for the Study of Liver Diseases. Hepatology. 2022 Sep 24.
Socha P, Janczyk W, Dhawan A, Baumann U, D'Antiga L, Tanner S, Iorio R, Vajro P, Houwen R, Fischler B, Dezsofi A, Hadzic N, Hierro L, Jahnel J, McLin V, Nobili V, Smets F, Verkade HJ, Debray D. Wilson's Disease in Children: A Position Paper by the Hepatology Committee of the European Society for Paediatric Gastroenterology, Hepatology and Nutrition. J Pediatr Gastroenterol Nutr. 2018 Feb;66(2):334-344.
Bonne-Tamir B, Farrer LA, Frydman M, Kanaaneh H, Rao DC. Evidence for linkage between Wilson disease and esterase D in three kindreds: detection of linkage for an autosomal recessive disorder by the family study method. Genet Epidemiol 1986; 3:201–9.
Shribman S, Marjot T,et al.British Association for the Study of the Liver Rare Diseases Special Interest Group. Investigation and management of Wilson's disease: a practical guide from the British Association for the Study of the Liver. Lancet Gastroenterol Hepatol. 2022 Jun;7(6):560-575.
Ferenci P. Regional distribution of mutations of the ATP7B gene in patients with Wilson disease: impact on genetic testing. Hum Genet. 2006 Sep;120(2):151-9.
The Human Gene Mutation Database (HGMD®). Available at: http://www.hgmd.cf.ac.uk/ac/index.php. (Accessed: 3rd May 2018)
Stenson PD, et al. The Human Gene Mutation Database: towards a comprehensive repository of inherited mutation data for medical research, genetic diagnosis and next-generation sequencing studies. JHum Genet 2017; 136:665–677.
Thomas GR, Forbes JR, Roberts EA, Walshe JM, Cox DW. The Wilson disease gene: spectrum of mutations and their consequences. Nat Genet. 1995; 9:210–7.
Maier-Dobersberger T, Ferenci P, Polli P, Balac P, Dienes HP, Kaserer K, et al. Detection of the H1069Q mutation in Wilson disease by rapid polymerase chain reaction. Ann Intern Med 1997;127:21–26.
Firneisz G, Lakatos PL, Szalay F, Polli C, Glant TT, Ferenci P. Common mutations of ATP7B in Wilson disease patients from Hungary. Am J Med Genet 2002;108:23–28.
Merle U, et al. Truncating mutations in the Wilson disease gene A TP7B are associated with very low serum ceruloplasmin oxidase activity and an early onset of Wilson disease. JBMC Gastroenterol 2010; 10:8.
Okada T, et al. High prevalence of fulminant hepatic failure among patients with mutant alleles for truncation of ATP7B in Wilson’s disease. J Scandinavian Journal of Gastroenterology 2010;45: 1232–237.
Ferenci P, Roberts EA. Defining Wilson disease phenotypes: from the patient to the bench and back again. Gastroenterology. 2012; 142:692–6.
Núñez-Ramos R, Montoro S, Bellusci M, et al. Acute Liver Failure: Outcome and Value of Pediatric End-Stage Liver Disease Score in Pediatric Cases. J Pediatr Emerg Care 2018; 34: 409-412.
Squires RH Jr, Shneider BL, et al. Acute liver failure in children: the first 348 patients in the pediatric acute liver failure study group. J Pediatr 2006; 148: 652-658.
Ostapowicz G, Fontana RJ, et al, US. Acute Liver Failure Study Group. Results of a prospective study of acute liver failure at 17 tertiary care centers in the United States. Ann Intern Med 2002; 137: 947-954.
Panagiotakaki E, Tzetis M, Manolaki N et al. Genotypephenotype correlations for a wide spectrum of mutations in the Wilson disease gene (ATP7B). Am J Med Genet 2004: 131 (2): 168–173.
Roberts EA, Schilsky ML; American Association for Study of Liver Diseases (AASLD). Diagnosis and treatment of Wilson disease: an update. JHepatology 2008; 47:2089-111.
Medici V, Trevisan CP, D'Incà R, Barollo M, Zancan L, Fagiuoli S, et al. Diagnosis and management of Wilson's disease: results of a single center experience. J Clin Gastroenterol. 2006;40:936–41.
Merle U, Schaefer M, Ferenci P, Stremmel W. Clinical presentation, diagnosis and long-term outcome of Wilson's disease: a cohort study. Gut. 2007; 56:115-20.
Sánchez-Albisua I, Garde T, Hierro L, Camarena C, Frauca F, de la Vega A, et al. A high index of suspicion: the key to an early diagnosis of Wilson's disease in childhood. J Pediatr Gastroenterol Nutr. 1999; 28:186-90.
Dhawan A, Taylor RM, Cheeseman P, De Silva P, Katsiyiannakis L, Mieli Vergani G: Wilson’s disease in children: 37-year experience and revised King’s score for liver transplantation. Liver Transpl 2005, 11:441-448.
Dong Y, Ni W, Chen WJ, et al. Spectrum and classification of ATP7B variants in a large cohort of Chinese patients with Wilson′s disease guides genetic diagnosis[J]. Theranostics, 2016, 6(5): 638‑649.
Cheng N, Wang H, Wu W, et al. Spectrum of ATP7B mutations and genotype‑phenotype correlation in large‑scale Chinese patients with Wilson Disease[J]. Clin Genet, 2017, 92(1): 69‑79.
Li X, Lu Z, Lin Y, et al. Clinical features and mutational analysis in 114 young children with Wilson disease from South China[J]. Am J Med Genet A, 2019, 179(8): 1451‑1458.
Wu ZY, Wang N, Lin MT, Fang L, Murong SX, Yu L. Mutation analysis and the correlation between genotype and phenotype of Arg778Leu mutation in Chinese patients with Wilson disease. Arch Neurol. 2001;58(6):971–6
Li M, Ma J, Wang W, Yang X, Luo K. Mutation analysis of the ATP7B gene and genotype-phenotype correlation in Chinese patients with Wilson disease. BMC Gastroenterol. 2021 Sep 1;21(1):339.
Qiao L, Ge J, Li C, Liu Y, Hu C, Hu S, Li W, Li T. Pathogenic gene variation spectrum and carrier screening for Wilson's disease in Qingdao area. Mol Genet Genomic Med. 2021 Aug;9(8): e1741.
El-Youssef M. Wilson disease. Mayo Clin Proc 2003: 78: 1126–1136.
Ferenci P, Caca K, Loudianos G et al. Diagnosis and phenotypic classification of Wilson disease. Liver Int 2003: 23: 139–142.
Medici V, Weiss KH. Genetic and environmental modifiers of Wilson disease. Handb Clin Neurol 2017; 142:35-41.
Pfister E-D, Karch A, Adam R, et al. Predictive factors for survival inchildren receiving liver transplants for Wilson’s disease: a cohort studyusing European Liver Transplant Registry Data.Liver Transpl2018;24:1186 – 98
Ferenci P. Phenotype-genotype correlations in patients with Wilson’sdisease.Ann N Y Acad Sci2014;1315:1 – 5
Gromadzka G, Schmidt HH, Genschel J, Bochow B, Rodo M, Tarnacka B, Litwin T, Chabik G, Członkowska A. Frameshift and nonsense mutations in the gene for ATPase7B are associated with severe impairment of copper metabolism and with an early clinical manifestation of Wilson's disease. Clin Genet. 2005 Dec;68(6):524-32.
Nicastro E, Loudianos G, Zancan L, D'Antiga L, Maggiore G, Marcellini M, Barbera C, Marazzi MG, Francavilla R, Pastore M, Vajro P, D'Ambrosi M, Vegnente A, Ranucci G, Iorio R. Genotype-phenotype correlation in Italian children with Wilson's disease. J Hepatol. 2009 Mar;50(3):555-61.
Nayagam JS, Jeyaraj R, Foskett P, Dhawan A, Ala A, Joshi D, Bomford A, Thompson RJ. ATP7B Genotype and Chronic Liver Disease Treatment Outcomes in Wilson Disease: Worse Survival with Loss-of-Function Variants. Clin Gastroenterol Hepatol. 2022 Sep 9: S1542-3565(22)00837-0.

Download PDF

Reviewers invited by journal
17 May, 2023
Editor assigned by journal
26 Apr, 2023
First submitted to journal
25 Apr, 2023

You are reading this latest preprint version

Phenotypic and Genetic Characterization of children with Wilson Disease from Northeast China

Status:

Version 1

Abstract

Background

Objective

Methods

Results

Conclusion

Figures

Introduction

Material and methods

Results

Discussion

Declarations

References

Status:

Version 1