Prevalence and Clinical Characteristics of Fabry Disease in Chinese Patients With Hypertrophic Cardiomyopathy.

BACKGROUND: The prevalence of Fabry disease (FD) in Chinese patients with hypertrophic cardiomyopathy (HCM) is unclear. We aimed to evaluate the prevalence, clinical characteristics, and outcomes of FD in Chinese patients with HCM. METHODS: Of 217 patients with HCM, FD probands were screened by next-generation sequencing at Fuwai Hospital. Medical data from α-galactosidase A activity, electrocardiography, echocardiography, coronary angiography, cardiac magnetic resonance, pathological examination, and follow up was analyzed. RESULTS: Two FD probands were observed (0.93% of patients with HCM), both of which were diagnosed with symptomatic obstructive HCM at 49 years of age. One proband had a GLA mutation (c.887T>C [p.M296T]) with a late-onset cardiac variant, which was characterized by dual ventricular hypertrophy and conduction disease with a permanent pacemaker. The other patient had a GLA mutation (c.758T>C [p.I253T]) with a classic phenotype and dual ventricular hypertrophy, atrioventricular block, renal failure, and recurrent cerebral infarction. Both probands had late gadolinium enhancement mainly in the basal segment of the inferolateral wall. Follow up revealed no exertional symptoms or outflow obstruction after surgical septal myectomy in the two probands, and stable renal function was observed after 6 months of migalastat therapy in the later one. A family study revealed six female carriers and three sudden cardiac deaths. CONCLUSIONS: FD is not uncommon in Chinese patients with HCM. Multiple organic involvement, dual ventricular hypertrophy, and conduction disease provide clinical clues for suspected FD, and early genetic screening is necessary. Surgical septal myectomy and migalastat improve the long-term prognosis of patients with FD.

Truncated Epithelial Sodium Channel ß Subunit Responsible for Liddle Syndrome in a Chinese Family.

BACKGROUND/AIMS: Liddle syndrome (LS) is a rare autosomal dominant disease caused by mutations in genes coding for epithelial sodium channel (ENaC) subunits. The aim of this study was to identify the mutation responsible for the LS in an extended Chinese family. METHODS: DNA samples from the proband with early-onset, treatment-resistant hypertension, and hypokalemia and 19 additional relatives were all sequenced for mutations in exon 13 of the ß-ENaC and γ-ENaC genes, using amplification by polymerase chain reaction and direct DNA sequencing. RESULTS: Genetic testing of exon 13 of SCNN1B revealed duplication of guanine into a string of 3 guanines located at codon 602. This frameshift mutation is predicted to generate a premature stop codon at position 607, resulting in truncated ß-ENaC lacking the remaining 34 amino acids, including the crucial PY motif. Among a total of 9 participants with the identical mutation, different phenotypes were identified. Tailored treatment with amiloride was safe and effective in alleviating disease symptoms in LS. No mutation of SCNN1G was identified in any of the examined participants. CONCLUSIONS: We report here a family affected by LS harboring a frameshift mutation (c.1806dupG) with a premature stop codon deleting the PY motif of ß-ENaC. Our study demonstrates that the earlier LS patients are diagnosed by genetic testing and treated with tailored medication, the greater the likelihood of preventing or minimizing complications in the vasculature and target organs.

A novel phenotype with splicing mutation identified in a Chinese family with desminopathy.

BACKGROUND: Desminopathy, a hereditary myofibrillar myopathy, mainly results from the desmin gene (DES) mutations. Desminopathy involves various phenotypes, mainly including different cardiomyopathies, skeletal myopathy, and arrhythmia. Combined with genotype, it helps us precisely diagnose and treat for desminopathy. METHODS: Sanger sequencing was used to characterize DES variation, and then a minigene assay was used to verify the effect of splice-site mutation on pre-mRNA splicing. Phenotypes were analyzed based on clinical characteristics associated with desminopathy. RESULTS: A splicing mutation (c.735+1G>T) in DES was detected in the proband. A minigene assay revealed skipping of the whole exon 3 and transcription of abnormal pre-mRNA lacking 32 codons. Another affected family member who carried the identical mutation, was identified with a novel phenotype of desminopathy, non-compaction of ventricular myocardium. There were 2 different phenotypes varied in cardiomyopathy and skeletal myopathy among the 2 patients, but no significant correlation between genotype and phenotype was identified. CONCLUSIONS: We reported a novel phenotype with a splicing mutation in DES, enlarging the spectrum of phenotype in desminopathy. Molecular studies of desminopathy should promote our understanding of its pathogenesis and provide a precise molecular diagnosis of this disorder, facilitating clinical prevention and treatment at an early stage.

Liddle syndrome misdiagnosed as primary aldosteronism resulting from a novel frameshift mutation of SCNN1B.

Liddle syndrome (LS), a monogenetic autosomal dominant disorder, is mainly characterized by early-onset hypertension and hypokalemia. Clinically, misdiagnosis or missing diagnosis is common, since clinical phenotypes of LS are variable and nonspecific. We report a family with misdiagnosis of primary aldosteronism (PA), but identify as LS with a pathogenic frameshift mutation of the epithelial sodium channel (ENaC) ß subunit. DNA samples were collected from a 32-year-old proband and 31 other relatives in the same family. A designed panel including 41 genes associated with monogenic hypertension was screened using next-generation sequencing. The best candidate disease-causing variants were verified by Sanger sequencing. Genetic analysis of the proband revealed a novel frameshift mutation c.1838delC (p.Pro613Glnfs*675) in exon 13 of SCNN1B. This heterozygous mutation involved the deletion of a cytosine from a string of three consecutive cytosines located at codons 612 to 613 and resulted in deletion of the crucial PY motif and elongation of the ß-ENaC protein. The identical mutation was also found in 12 affected family members. Amiloride was effective in alleviating LS for patients. There were no SCNN1A or SCNN1G mutations in this family. Our study emphasizes the importance of considering LS in the differential diagnosis of early-onset hypertension. The identification of a novel frameshift mutation of SCNN1B enriches the genetic spectrum of LS and has allowed treatment of this affected family to prevent severe complications.

Genetic screening of SCNN1B and SCNN1G genes in early-onset hypertensive patients helps to identify Liddle syndrome.

BACKGROUND: Liddle syndrome is an autosomal dominant form of monogenic hypertension. Phenotypic variability makes it difficult to identify patients with Liddle syndrome, resulting in misdiagnosis and severe complications at early age. OBJECTIVES: To identify mutation in SCNN1B and SCNN1G genes in an adolescent with suspicious Liddle syndrome and his family members and to explore the screening target subjects of Liddle syndrome. METHODS: Genetic analysis of the C-terminus of SCNN1B and SCNN1G genes was conducted in an adolescent, with treatment-resistant hypertension and hypokalemia, who was suspected of having Liddle syndrome, and his family members. A Medline research of the reported cases with Liddle syndrome was also performed. RESULTS: A recurrent SCNN1B mutation, c.1853C>A (p.P618H), was detected in the 19-year-old male patient, and family screening identified five additional members who were heterozygous for the mutation. The diagnosis of Liddle syndrome was made in all affected individuals. Despite the phenotypic variability, a systematic review of 54 reported index cases revealed the early-onset hypertension, aged no more than 30 years, as a common feature. CONCLUSIONS: Genetic screening for Liddle syndrome should be considered in hypertensive subjects with early penetrance, maybe no more than 30 years, after exclusion of common secondary causes of hypertension.

A novel PRKAG2 mutation in a Chinese family with cardiac hypertrophy and ventricular pre-excitation.

PRKAG2 syndrome is a rare autosomal dominant inherited disorder that is characterized by cardiac hypertrophy, ventricular pre-excitation and conduction system abnormalities. There is little knowledge in cardiovascular magnetic resonance (CMR) characteristics of PRKAG2 cardiomyopathy. This study investigated the genetic defect in a three-generation Chinese family with cardiac hypertrophy and ventricular pre-excitation using whole-exome sequencing. A novel missense mutation, c.1006 G > T (p.V336L), was identified in PRKAG2. This mutation had not been identified in the ExAC database, and the prediction result of MutationTaster indicated a deleterious effect. Furthermore, it cosegregated with the disease in the present family and was absent in unrelated 300 healthy controls. cDNA analysis did not detect any splicing defects, although the variant occurred in the first base of exon 9. CMR evaluation in five affected members showed diffuse hypertrophy in a concentric pattern, with markedly increased left ventricular mass above age and gender limits (median 151.3 g/m2, range 108.4-233.4 g/m2). Two patients in progressive stage and one patient with sudden cardiac death exhibited extensive subendocardial late gadolinium enhancement. In conclusion, molecular screening for PRKAG2 mutations should be considered in patients who exhibit cardiac hypertrophy coexisting with ventricular pre-excitation. CMR offers promising advantages for evaluation of PRKAG2 cardiomyopathy.

Restrictive Cardiomyopathy Resulting from a Troponin I Type 3 Mutation in a Chinese Family.

Objective To identify the pathogenic variant responsible for restrictive cardiomyopathy (RCM) in a Chinese family.Methods Next generation sequencing was used for detecting the mutation and Results verified by sequencing. We used restriction enzyme digestion to test the mutation in the family members and 200 unrelated normal subjects without any cardiac inherited diseases when the mutation was identified.Results Five individuals died from cardiac diseases, two of whom suffered from sudden cardiac death. Two individuals have suffered from chronic cardiac disorders. Mutation analysis revealed a novel missense mutation in exon 7 of troponin I type 3 (TNNI3), resulting in substitution of serine (S) with proline (P) at amino acid position 150, which cosegregated with the disease in the family, which is predicted to be probably damaging using PolyPhen-2. The mutation was not detected in the 200 unrelated subjects we tested.Conclusion Using next generation sequencing, which has very recently been shown to be successful in identifying novel causative mutations of rare Mendelian disorders, we found a novel mutation of TNNI3 in a Chinese family with RCM.

Whole-exome sequencing identifies a missense mutation in hnRNPA1 in a family with flail arm ALS.

OBJECTIVE: To identify the disease-causing gene of a family with upper limb predominant, slowly progressive amyotrophic lateral sclerosis (ALS), which was diagnosed as flail arm syndrome (FAS). METHODS: After causation of 24 known ALS genes was excluded by targeted next-generation sequencing, whole-exome sequencing was applied in the FAS family. Cellular localization of mutant hnRNPA1 was examined in transfected HeLa cells. An additional 251 Chinese patients with ALS (including 7 sporadic FAS) underwent mutation screening of hnRNPA1. RESULTS: We detected a novel missense mutation in hnRNPA1, c.862/1018C>T (p.P288S/P340S), which cosegregated with disease in the FAS family. The residue is highly conserved across species and exists in the encoded PY nuclear localization signal of hnRNPA1 protein. Mutant hnRNPA1 showed altered intracellular localization, resulting in formation of cytoplasmic inclusions that colocalized with stress granules in transfected cells. Further mutation screening of hnRNPA1 in additional patients with FAS and typical ALS detected 2 rare variants with unknown significance. These variants lie in the prion-like domain of hnRNPA1 long isoform, which was detected exclusively in the CNS. CONCLUSIONS: Our results suggest that hnRNPA1 is the causative gene in the family with flail arm ALS. This further expanded the disease phenotype of hnRNPA1 mutations.

Identification of a novel loss-of-function C9orf72 splice site mutation in a patient with amyotrophic lateral sclerosis.

Abnormal expansion of a hexanucleotide GGGGCC repeat in the C9orf72 gene is the most common cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia in Caucasians. However, the underlying pathologic mechanisms remain controversial, and both loss-of-function and gain-of-function models have been proposed. To gain further insight into these mechanisms, we performed mutation analysis of C9orf72 in 276 Han Chinese patients with ALS. We identified GGGGCC expansions in 2 cases of sporadic ALS with 38 and 63 repeats, as well as a novel splice site mutation (c.601-2A>G) in a third case. These genetic alterations were not detected in 332 control patients without neurological disease. Intriguingly, functional analysis revealed that the splice site mutation disrupted the reading frame, creating a premature stop codon (p.I201fsX235). Decreased levels of the mutant messenger RNA were detected in patient cells, suggesting that it may undergo nonsense-mediated messenger RNA decay. Taken together, these results demonstrate that C9orf72 mutation is infrequently associated with ALS in Han Chinese patients and suggest that a splice site mutation in C9orf72 may lead to loss of function due to haploinsufficiency of the resulting protein.

A Novel Splicing Mutation Identified in a Chinese Family with X-linked Alport Syndrome Using Targeted Next-Generation Sequencing.

AIMS: Alport syndrome (AS) is a genetically heterogeneous disorder, characterized by hematuria, progressive renal failure, sensorineural hearing loss, and ocular abnormalities caused by mutations in the COL4A3, COL4A4, and COL4A5 genes. The aim of this study was to identify underlying mutations in individuals from a Chinese family with X-linked AS. METHODS: We performed targeted next-generation sequencing (NGS) to identify mutations associated with AS. The results were processed and visualized using an Integrated Genomics Viewer software. The most likely disease-causing variants were identified and confirmed by Sanger sequencing of reverse transcription-polymerase chain reaction products. RESULTS: Visual inspection using Integrative Genomics Viewer software found that COL4A5 exon 10 was not covered by the disease panel, while coverage of exons 4, 17, 20, 21, 37, and 45 was incomplete. Sanger sequencing of these regions identified a novel splice-site mutation in intron 9 (c.547-3C>A) of the COL4A5 gene. Subsequent cDNA analysis revealed that c.547-3C>A led to skipping of exon 10, which resulted in an in-frame deletion of 21 amino acids from the α5 chain of type IV collagen. CONCLUSION: We determined the molecular basis of AS in a Chinese family by targeted NGS and cDNA analysis. This is the first report of the novel c.547-3C>A splicing mutation in the collagen domain of COL4A5 gene.

Novel Phenotype-Genotype Correlations of Restrictive Cardiomyopathy With Myosin-Binding Protein C (MYBPC3) Gene Mutations Tested by Next-Generation Sequencing.

BACKGROUND: MYBPC3 dysfunctions have been proven to induce dilated cardiomyopathy, hypertrophic cardiomyopathy, and/or left ventricular noncompaction; however, the genotype-phenotype correlation between MYBPC3 and restrictive cardiomyopathy (RCM) has not been established. The newly developed next-generation sequencing method is capable of broad genomic DNA sequencing with high throughput and can help explore novel correlations between genetic variants and cardiomyopathies. METHODS AND RESULTS: A proband from a multigenerational family with 3 live patients and 1 unrelated patient with clinical diagnoses of RCM underwent a next-generation sequencing workflow based on a custom AmpliSeq panel, including 64 candidate pathogenic genes for cardiomyopathies, on the Ion Personal Genome Machine high-throughput sequencing benchtop instrument. The selected panel contained a total of 64 genes that were reportedly associated with inherited cardiomyopathies. All patients fulfilled strict criteria for RCM with clinical characteristics, echocardiography, and/or cardiac magnetic resonance findings. The multigenerational family with 3 adult RCM patients carried an identical nonsense MYBPC3 mutation, and the unrelated patient carried a missense mutation in the MYBPC3 gene. All of these results were confirmed by the Sanger sequencing method. CONCLUSIONS: This study demonstrated that MYBPC3 gene mutations, revealed by next-generation sequencing, were associated with familial and sporadic RCM patients. It is suggested that the next-generation sequencing platform with a selected panel provides a highly efficient approach for molecular diagnosis of hereditary and idiopathic RCM and helps build new genotype-phenotype correlations.

A novel frameshift mutation of epithelial sodium channel ß-subunit leads to Liddle syndrome in an isolated case.

OBJECTIVE: Liddle syndrome, an autosomal dominant form of monogenic hypertension, is attributed to mutations in the genes encoding ß and γ subunits (SCNN1B and SCNN1G) of the epithelial sodium channel (ENaC). The aim of this study was to search for pathogenic mutations of SCNN1B and SCNN1G in an adolescent under the impression of Liddle syndrome and no family history of hypertension. DESIGN AND PATIENTS: We screened the C-terminus of SCNN1B and SCNN1G in an adolescent with poorly controlled hypertension who was clinically diagnosed as having Liddle syndrome. We also screened for the mutation in his parents, 100 hypertensive patients and 100 controls. RESULTS: Genetic analysis of SCNN1B revealed a frameshift mutation induced by insertion of an additional cytosine into a string of six located between codons 617 and 618, which is predicted to introduce a new termination codon at position 621 and produce a protein truncated by 20 amino acids. This frameshift mutation was not detected in the patient's parents, the 100 hypertensive patients or the 100 controls, indicating that this is a de novo mutation and not a common genetic polymorphism. There was no mutation of SCNN1G in any of the individuals examined. CONCLUSION: Based on direct DNA sequencing, we identified a novel frameshift mutation in the ßENaC gene in an isolated case of Liddle syndrome. Confirmation of the diagnosis and effective tailored treatment in the patient were achieved, implying that genetic testing is a useful tool to diagnose Liddle syndrome.

[Mutation in the SLC37A4 gene of glycogen storage disease type Ib in 15 families of the mainland of China].

OBJECTIVE: Glycogen storage disease type Ib (GSDIb, MIM: 232220) is an autosomal recessive inborn error of metabolism caused by deficiency of the glucose-6-phosphate translocase. The clinical manifestations include symptoms and signs of both the typical GSDIa, including hepatomegaly, fasting hypoglycemia, lactic acidemia and hyperlipidemia, and the dysfunction of neutrophils of recurrent infection and neutropenia. More than 84 mutations have been identified since the discovery of the SLC37A4 gene as the disease causing gene. Up to date, 5 mutations in 4 Chinese patients were reported from Hong Kang and Taiwan. In order to see the spectrum of the SLC37A4 gene mutations and the correlation between genotype and phenotype in patients with GSDIb of the mainland of China, the authors investigated 17 GSDIb patients from 15 families in this study. METHOD: Data of 17 patients from 12 provinces, 11 male and 6 female, aged 6 months to 35 years, were collected from the genetic clinics of Peking Union Medical College Hospital from Oct. 2006 to Mar. 2009. All of them were Han Chinese in ethnicity. Consanguineous status was confirmed in 2 unrelated patients. All patients were presented with hepatomegaly, fasting hypoglycemia, lactic acidemia, hyperlipidemia and neutropenia with variable frequency of infections. The full coding exons, their relevant exon-intron boundaries, and the 5'- and 3'-flanking regions of the SLC37A4 gene were amplified and directly sequenced. RT-PCR was performed to verify the effect of the 2 novel splicing mutations. RESULT: A total of 11 mutations were identified in 15 families. Four mutations, p.Gly149Glu, p.Pro191Leu, p.Arg415X and c.1042_1043 del CT, were previously reported, and seven mutations, p. Leu23Arg, p.Gly115Arg, p.Gly281Val, p.Arg415Gly, c.784 + 1G > A, c.870 + 5G > A and c.1014_1120del107, were novel. The frequent mutations are p.Pro191Leu, p.Gly149Glu and c.870 + 5G > A, accounting for 37%, 15% and 11% of mutant alleles respectively. RT-PCR analysis of novel mutation c.784 + 1G > A confirmed the splicing of exon 5 of 159 bp, causing inframe deletion. While mutation c.870 + 5G > A was proved to cause exon 6, 86 bp, deletion causing frame-shift. Among 15 families, 12 genotypes were identified, including 3 with homozygous mutation and 9 with compound heterozygous mutations. Homozygous p.Pro191Leu mutation was the only genotype detected in more than 1 family and was found in 4 unrelated families, including 1 patient from consanguineous marriage. CONCLUSION: A total of 11 SLC37A4 gene mutations were identified in 15 families of the mainland of China. The frequent mutations are p.Pro191Leu, p.Gly149Glu and c.870 + 5G > A. The number of Chinese SLC37A4 gene mutations was extended from 5 to 14.

[Sequence analysis of a novel human leukocyte antigen allele B*5827].

OBJECTIVE: To investigate the molecular basis for a novel human leukocyte antigen (HLA) allele B*5827. METHODS: DNA from the proband was analyzed by polymerase chain reaction-sequence specific oligonucleotide (PCR-SSO) typing. The amplified product was sequenced bidirectionally. RESULTS: Abnormal HLA-B locus was observed and its nucleotide sequence was different from the known HLA-B allele sequences, with highest homology to HLA-B*5820 allele. It differs from HLA-B*5820 by 8 nucleotide substitutions in exon 3, i.e., nt 290 (G > C), nt 346 (T > A), nt 390 (A > C), nt 404 (G > C), nt 413 (C > G), nt 471 (A > G), nt 486 (A > G) and nt 487 (C > A), resulting in an amino acid change from ser > arg at nt 97, phe >tyr at nt 115, ser > arg at nt 130, thr > ala at nt 157 and thr > glu at nt 162. Nucleotide differences of nt 404 (G > C) and nt 413( C > G) did not change amino acid. CONCLUSION: The sequences of the novel allele have been submitted to GenBank (access No.GU071234). A novel HLA class I allele B*5827 has been officially assigned by the WHO HLA Nomenclature Committee in Jan. 2010.

[Mutation analysis of phenylalanine hydroxylase gene in 55 patients with phenylketonuria from Hebei province].

OBJECTIVE: To identify the pathogenic mutations of phenylalanine hydroxylase gene (PAH) in patients with phenylketonuria (PKU) from Hebei Province. METHODS: Genomic DNA was extracted from 55 unrelated PKU patients from September 2007 to July 2009. All PAH exons and exon-intron junctions were amplified by polymerase chain reaction (PCR) and sequenced. Multiplex ligation-dependent probe amplifications (MLPA) was performed to detect the deletions or duplications of PAH. Gap-PCR was used to determine the breakpoints of large deletions. RESULTS: Among them, 108 mutant alleles (98.2%) were found. All PAH exons with the exceptions of exons 9 and 13 were affected. A total of 41 different mutations were detected, including missense (n = 24), nonsense (n = 7), splicing (n = 7), small deletion (n = 1) and large deletion (n = 2). Among them, 4 missense mutations (p.Pro147Leu, p.Gly289Arg, p.Phe392Ser, p.Ile421Thr) and 2 large deletions (-4163_-406del and -1932_+3402del) were novel. The most common mutations were p.Arg243Gln (12.7%), c.611A > G (11.8%) and c.1197A > T (9.1%). CONCLUSION: The mutations of PKU patients with from Hebei Province are scattered throughout the PAH gene. Most of them are of single nucleotide substitutions, but large deletions are not rare.

[Diagnosis of glycogen storage disease type IIIA by detecting glycogen debranching enzyme activity, glycogen content and structure in muscle].

OBJECTIVE: Glycogen storage disease type III (GSD III) is an autosomal recessive disease caused by glycogen debranching enzyme (GDE) gene (AGL gene) mutation resulting in hepatomegaly, hypoglycemia, short stature and hyperlipidemia. GSD IIIA, involves both liver and muscle, and accounts for up to 80% of GSD III. The definitive diagnosis depends on either mutation analysis or liver and muscle glycogen debranching enzyme activity tests. This study aimed to establish enzymologic diagnostic method for GSD IIIA firstly in China by detecting muscular GDE activity, glycogen content and structure and to determine the normal range of muscular GDE activity, glycogen content and structure in Chinese children. METHOD: Muscle samples were collected from normal controls (male 15, female 20; 12-78 years old), molecularly confirmed GSD III A patients (male 8, female 4, 2-27 years old) and other myopathy patients (male 9, 2-19 years old). Glycogen in the muscle homogenate was degraded into glucose by amyloglucosidase and phosphorylase respectively. The glycogen content and structure were identified by glucose yield determination. The debranching enzyme activity was determined using limit dextrin as substrate. Independent samples Kruskal-Wallis H test, Nemenyi-Wilcoxson-Wilcox test, and Chi-square test were used for statistical analyses by SPSS 11.5. RESULT: (1) GSD III A patients' glycogen content were higher, but G1P/G ratio and GDE activity were lower than those of the other two groups (P < 0.01). In all of the three parameters, there were no significant difference between normal controls and other myopathy patients. (2) The range of normal values: glycogen content 0.31%-0.43%, G1P/G ratio 22.37%- 26.43%, GDE activity 0.234-0.284 micromol/(g. min). (3) Enzymologic diagnostic method had a power similar to that of gene analysis in diagnosis of GSD-IIIA patients. The sensitivity and specificity of enzymologic diagnostic method and mutation detection were 91.7% and 100% respectively. CONCLUSION: Enzymologic diagnostic method of GSD IIIA was firstly established in China. The range of normal values was determined. This method could be used in diagnosing suspected GSD IIIA patients in the clinic.

[Identification of a novel TP53 germline mutation in one child with adrenocortical carcinoma].

OBJECTIVE: To detect the germline TP53 gene mutation in a child with pediatric adrenocortical carcinoma (ADCC) in order to provide genetic diagnosis and counseling. METHODS: Genomic DNA was extracted from peripheral blood from a girl with ADCC and her parents. All TP53 exons and their flanking intronic sequences were PCR-amplified and subjected to automatic DNA sequencing. RESULTS: Direct sequencing of PCR products revealed a heterozygous G insertion between nucleotide 522 and 523 (c.522-523insG) in TP53 exon 5. This novel mutation is predicted to result in a frame shift at codon 175, producing a new reading frame ending in a stop at position 6 (p.R175AfsX6). The same heterozygous mutation was also found in her father, but not in her mother. CONCLUSION: A novel germline mutation in the TP53 gene has been identified in one case with pediatric ADCC.