High frequency of the TARDBP p.Ala382Thr mutation in Sardinian patients with amyotrophic lateral sclerosis

Recently, rare mutations in the TARDBP gene have been identified in familial and sporadic amyotrophic lateral sclerosis (ALS) patients. The purpose of this study was to characterize the genetic variability of the TARDBP gene in a cohort of Sardinian ALS patients. The coding region of the gene was analyzed in 97 unrelated patients previously tested negative for superoxide dismutase (SOD1) mutations. The p.Ala382Thr (c.1144G>A) mutation was found in 30 patients (30.9%). The mutation was predominant in familial ALS patients (FALS) as it was represented in 24 of 30 FALS cases (80%) (p < 0.0003). Six cases were apparently sporadic (9% of sporadic ALS patients). No further mutation of TARDBP was found in our cohort of ALS patients. Patients carrying the mutation showed spinal site of onset in 24 cases (80%), an average age at onset of 54.7 ± 11.1 years, not significantly different from patients not harboring TARDBP mutations (56.7 ± 9.6) and a female:male gender ratio of 1:1.1. The haplotype analysis carried out using eight microsatellite markers flanking the gene showed a founder effect for this mutation. Finally, we estimated the age-specific penetrance of the TARDBP p.Ala382Thr mutation in an additional sample of 47 carriers (20 affected and 27 unaffected). The average penetrance to 70 years was 60% (95% confidence interval 41-79%). A trend toward a higher penetrance in males was observed. Even in the presence of a causal mutation, most of the ALS clinical heterogeneity, however, draws upon from a multifactorial context.     

RB患者及家庭成员Rb基因突变和患病风险的基因诊断及遗传咨询

目的建立Ｒｂ基因突变的基因诊断方法，正确估计视网膜母细胞瘤（ＲＢ）患者的预后及其家庭成员的患病风险。方法综合利用Ｓｏｕｔｈｅｒｎｂｌｏｔ杂交、ＳＳＣＰ分析、直接ＤＮＡ序列测定等多种分子生物学技术作染色体单体型分析及Ｒｂ基因点突变的直接检测。结果７９个有Ｒｂ基因突变的ＲＢ家系中２５例先证者仅查出体细胞起源的Ｒｂ基因突变，其余５４例存在生殖细胞起源的Ｒｂ基因突变，其中３６例突变是新产生的，１５例突变由亲代遗传而来，此外，尚有３例Ｒｂ基因突变嵌合体。结论直接检测Ｒｂ基因点突变可不依赖于患者家庭成员ＲＢ发病情况的遗传背景资料诊断患者是否属于遗传型，正确估计患者的预后；并能在肿瘤发生前甚至产前检出Ｒｂ基因突变携带者，正确估计患病风险     

Mild phenotypes in a series of patients with Opitz GBBB syndrome with MID1 mutations

Opitz syndrome (OS; MIM 145410 and MIM 300000) is a congenital midline malformation syndrome characterized by hypertelorism, hypospadias, cleft lip/palate, laryngotracheoesophageal (LTE) abnormalities, imperforate anus, developmental delay, and cardiac defects. The X-linked form (XLOS) is caused by mutations in the MID1 gene, which encodes a microtubule-associated RBCC protein. In this study, phenotypic manifestations of patients with and without MID1 mutations were compared to determine genotype-phenotype correlations. We detected 10 novel mutations, 5 in familial cases, 2 in sporadic cases, and 3 in families for whom it was not clear if they were familial or sporadic. The genotype and phenotype was compared for these 10 families, clinically diagnosed OS patients found not to have MID1 mutations, and 4 families in whom we have previously reported MID1 mutations. This combined data set includes clinical and mutation data on 70 patients. The XLOS patients with MID1 mutations were less severely affected than patients with MID1 mutations reported in previous studies, particularly in functionally significant neurologic, LTE, anal, and cardiac abnormalities. Minor anomalies were more prevalent in patients with MID1 mutations compared to those without mutations in this study. Female MID1 mutation carriers had milder phenotypes compared to male MID1 mutation carriers, with the most common manifestation being hypertelorism in both sexes. Most of the anomalies found in the patients of the present study do not correlate with the MID1 mutation type, with the possible exception of LTE malformations. This study demonstrates the wide spectrum of severity and manifestations of OS. It also shows that XLOS patients with MID1 mutations may be less severely affected than indicated in prior reports.     

CACNA1A R1347Q: a frequent recurrent mutation in hemiplegic migraine

Of the 18 missense mutations in the CACNA1A gene, which are associated with familial hemiplegic migraine type 1 (FHM1), only mutations S218L, R583Q and T666M were identified in more than two independent families. Including the four novel families presented here, of which two represent de novo cases, the R1347Q mutation has now been identified in six families. A genotype-phenotype comparison of R1347Q mutation carriers revealed a wide clinical spectrum ranging from (trauma triggered) hemiplegic migraine with and without ataxia, loss of consciousness and epilepsy. R1347Q is the third most frequent mutation in hemiplegic migraine patients and should therefore be screened with priority for confirmation of clinical diagnosis. This study clearly demonstrates that the availability of multiple families better reflects the full clinical spectrum associated with FHM1 mutations.     

Multiple lipomas linked to an RB1 gene mutation in a large pedigree with low penetrance retinoblastoma

Hereditary predisposition to lipomas is observed in familial multiple lipomatosis (OMIM 151900) and benign cervical lipomatosis (OMIM 151800) and can also be associated with mutations in the MEN1 and PTEN genes (OMIM 131100 and 153480, respectively). In addition, a recent report indicates that a few patients with hereditary retinoblastoma also have lipomas. Here we report on an extended family segregating a splice site mutation in the RB1 gene. Almost all adult carriers of this mutation had multiple lipomas while penetrance for retinoblastoma was incomplete. In an unrelated pedigree, which was reported previously, the identical mutation was only associated with low-penetrance retinoblastoma but not lipomas. Our data indicate that lipoma predisposition in hereditary retinoblastoma is not associated with specific RB1 gene mutations but is influenced by modifying factors linked to this gene.     

Wolfram syndrome in French population: characterization of novel mutations and polymorphisms in the WFS1 gene

Wolfram syndrome (WS), a rare autosomal recessive neurodegenerative disorder, results in most cases from mutations in the WFS1 gene. In this study, a total of 19 patients with Wolfram syndrome and 36 relatives from 17 families were screened for mutations in the WFS1 gene. WFS1 mutations were identified on both alleles in 16 of 19 patients and on 1 allele of 3 patients, showing that WFS1 is the major gene involved in WS in the french population. We identified 25 different mutations, twelve of which were novel. We found 6 frameshift mutations, 6 nonsense mutations, 6 missense mutations, 6 in-frame deletions, and one new homozygous mutation in the splice donor site of exon 7 (c.861+1G>A) resulting in a frameshift. Most patients were compound heterozygotes. No common founder mutation or mutational hot spot were found in the WFS1 gene. Although most mutations occurred in exon 8, in some cases molecular screening requires analysis of all exons, including the non-coding exon 1. We also identified 3 new polymorphisms. Furthermore, genotype-phenotype correlation suggests that the presence of inactivating mutations on both alleles may be associated with an early onset of diabetes mellitus.     

Molecular screening of SHH, ZIC2, SIX3, and TGIF genes in patients with features of holoprosencephaly spectrum: Mutation review and genotype-phenotype correlations

Holoprosencephaly (HPE; 1 out of 16,000 live births; 1 out of 250 conceptuses) is a complex brain malformation resulting from incomplete cleavage of the prosencephalon, affecting both the forebrain and the face. Clinical expressivity is variable, ranging from a single cerebral ventricle and cyclopia to clinically unaffected carriers in familial dominant autosomic HPE. The disease is genetically heterogeneous, but additional environmental agents also contribute to the etiology of HPE. In our cohort of 200 patients, 34 heterozygous mutations were identified, 24 of them being novel ones: 13 out of 17 in the Sonic hedgehog gene (SHH); 4 out of 7 in ZIC2; and 7 out of 8 in SIX3. The two mutations identified in TGIF have already been reported. Novel phenotypes associated with a mutation have been described, such as abnormalities of the pituitary gland and corpus callosum, colobomatous microphthalmia, choanal aperture stenosis, and isolated cleft lip. This study confirms the great genetic heterogeneity of the disease, the important phenotypic variability in HPE families, and the difficulty to establish genotype-phenotype correlations.     

Molecular and phenotypic characteristics of metachromatic leukodystrophy patients from Poland

The occurrence and genotype-phenotype correlations of the eight most common mutations in the arylsulfatase A (ARSA) gene were studied in 43 unrelated Polish patients suffering from different types of metachromatic leukodystrophy (MLD). Screening for mutations p.R84Q, p.S96F, c.459+1G>A, p.I179S, p.A212V, c.1204+1G>A, p.P426L, and c.1401-1411del allowed the identification of 53.5% of the mutant alleles. In the whole investigated group of patients, mutations c.459+1G>A and p.P426L were the most frequent, 19 and 17%, respectively. The prevalence of the third most frequent mutation, i.e. p.I179S (13%), seems to be higher than that in other populations. The incidence of c.1204+1G>A was 5%, which is higher than reported earlier (2%). It seems that p.I179S and c.1204+1G>A are more prevalent in MLD patients from Poland than from other countries. In the group examined by us, mutations p.R84Q, p.S96F, p.A212V, and c.1401-1411del were not detected; thus, 46.5% of MLD alleles remained unidentified. This indicates that other, novel or already described, but rare, mutations exist in Polish population. In late infantile homozygotes for c.459+1G>A and one homozygote for c.1204+1G>A, first clinical symptom was motor deterioration. In adult homozygotes for p.P426L, the disease onset manifested as gait disturbances, followed by choreoathetotic movements, difficulties in swallowing, dysarthria, tremor, and nystagmus. In the carriers of the p.I179S mutation, the hallmark of the clinical picture was psychotic disturbances.     

Enlarged parietal foramina caused by mutations in the homeobox genes ALX4 and MSX2: from genotype to phenotype

Heterozygous mutations of the homeobox genes ALX4 and MSX2 cause skull defects termed enlarged parietal foramina (PFM) and cranium bifidum (CB); a single MSX2 mutation has been documented in a unique craniosynostosis (CRS) family. However, the relative mutational contribution of these genes to PFM/CB and CRS is not known and information on genotype-phenotype correlations is incomplete. We analysed ALX4 and MSX2 in 11 new unrelated cases or families with PFM/CB, 181 cases of CRS, and a single family segregating a submicroscopic deletion of 11p11.2, including ALX4. We explored the correlations between skull defect size and age, gene, and mutation type, and reviewed additional phenotypic manifestations. Four PFM cases had mutations in either ALX4 or MSX2; including previous families, we have identified six ALX4 and six MSX2 mutations, accounting for 11/13 familial, but only 1/6 sporadic cases. The deletion family confirms the delineation of a mental retardation locus to within 1.1 Mb region of 11p11.2. Overall, no significant size difference was found between ALX4- and MSX2-related skull defects, but the ALX4 mutation p.R218Q tends to result in persistent CB and is associated with anatomical abnormalities of the posterior fossa. We conclude that PFM caused by mutations in ALX4 and MSX2 have a similar prevalence and are usually clinically indistinguishable. Mutation screening has a high pickup rate in PFM, especially in familial cases, but is not indicated in CRS.     

GJB2 mutations in patients with non-syndromic hearing loss from Northeastern Hungary

Mutations in the GJB2 gene encoding the gap-junction protein connexin 26 have been identified in many patients with childhood hearing impairment (HI). One single mutation, c.35delG, accounts for the majority of mutations in Caucasian patients with HI. In the present study we screened 500 healthy control individuals and a group of patients with HI from Northeastern Hungary for GJB2 mutations. The patients' group consisted of 102 familial from 28 families and 92 non-familial cases. The most common mutation in the Hungarian population is the c.35delG, followed by the c.71G>A (p.W24X) mutation. 34.3% of the patients in the familial group were homozygous, and 17.6% heterozygous for 35delG. In the non-familial group the respective values were 37% and 18% (allele frequency: 46.2%). In the general population an allele frequency of 2.4% was determined. Several patients were identified with additional, already described or new GJB2 mutations, mostly in heterozygous state. The mutation c.380G>A (p.R127H) was formerly found only in heterozygous state and its disease relation was controversial. We demonstrated the presence of this mutation in a family with three homozygous patients and 4 heterozygous unaffected family members, a clear indication of recessively inherited HI. Furthermore, we provided evidence for the pathogenic role of two new mutations, c.51C>A (p.S17Y) and c.177G>T (p.G59V), detected in the present study. In the latter case the pattern of inheritance might be dominant. Our results confirm the importance of GJB2 mutations in the Hungarian population displaying mutation frequencies that are comparable with those in the Mediterranean area.     

Correlation between RB1germline mutations and second primary malignancies in hereditary retinoblastoma patients treated with external beam radiotherapy

Retinoblastoma (Rb) results from biallelic inactivation of the RB1 gene. Hereditary Rb patients i. e germline carriers of a RB1 mutation also have a risk of developing subsequent malignant neoplasms (SMN) such as osteosarcomas. This SMN risk is maximized by external beam radiotherapy treatments (EBRT), which is why these treatments are now avoided. Nevertheless, EBRT is still a matter of great concern, as EBRT-treated patients are in their adulthood and SMNs remain the major cause of death for patients.To decipher the relationship between RB1 genotype and SMN development in EBRT treated patients, we conducted a retrospective study in a cohort of 160 irradiated hereditary Rbs with fully resolved RB1 mutational status. Median follow-up was 22 years [1-51] and median age of patients was 27 years old [7-53]. Among these 160 Rb patients, 120 did not develop any SMN (75%) and 40 developed SMNs (25%). The age at which EBRT is given (i.e. before or after the age of 12 months) was not correlated to SMN development (p?=?0.6). We didn't find any difference in RB1 mutation type between patients with or without SMN, neither could we detect any linkage between mutation type and SMN location, SMN type and age at diagnosis. Interestingly, among 13 carriers of a RB1 low penetrance mutation, 3 of them developed sarcomas, a rare tumor that cannot be attributed to the general population.Our study cannot explain why a RB1 mutation leads or not to a SMN but demonstrated that EBRT patients with a low penetrance mutation remain at risk of SMN and should be cautiously monitored.     

Genotype-phenotype correlations of KCNJ2 mutations in Japanese patients with Andersen-Tawil syndrome

Andersen-Tawil syndrome (ATS) is a rare inherited disorder characterized by periodic paralysis, mild dysmorphic features, and QT or QU prolongation with ventricular arrhythmias in electrocardiograms (ECGs). Mutations of KCNJ2, encoding the human inward rectifying potassium channel Kir 2.1, have been identified in patients with ATS. We aimed to clarify the genotype-phenotype correlations in ATS patients. We screened 23 clinically diagnosed ATS patients from 13 unrelated Japanese families. Ten different forms of KCNJ2 mutations were identified in the 23 ATS patients included in this study. Their ECGs showed normal QTc intervals and abnormal U waves with QUc prolongation and a variety of ventricular arrhythmias. Especially, bidirectional ventricular tachycardia (VT) was observed in 13 of 23 patients (57%). Periodic paralysis was seen in 13 of 23 carriers (57%), dysmorphic features in 17 (74%), and seizures during infancy in 4 (17%). Functional assays for the two novel KCNJ2 mutations (c. 200G>A (p. R67Q) and c. 436G>A (p. G146S)) displayed no functional inward rectifying currents in a heterologous expression system and showed strong dominant negative effects when co-expressed with wild-type KCNJ2 channels (91% and 84% reduction at -50 mV respectively compared to wild-type alone). Immunocytochemistry and confocal imaging revealed normal trafficking for mutant channels. In our study, all of the clinically diagnosed ATS patients had KCNJ2 mutations and showed a high penetrance with regard to the typical cardiac phenotypes: predominant U wave and ventricular arrhythmias, typically bidirectional VT.     

Two novel severe mutations in the pancreatic secretory trypsin inhibitor gene (SPINK1) cause familial and/or hereditary pancreatitis

Mutations in the serine protease inhibitor Kazal type 1 gene (SPINK1) encoding pancreatic secretory trypsin inhibitor (PSTI) have recently been found to be associated with chronic pancreatitis. Nevertheless, knowledge of severe mutations is particularly scarce, both in terms of number and in the extent of clinical information. The aim of this study was to expand the known spectrum of such mutations. 46 unrelated families, each including at least two pancreatitis patients and carrying neither cationic trypsinogen (PRSS1) mutations nor the frequent SPINK1 N34S mutation, participated in this study. The four exons and their flanking sequences of the SPINK1 gene were screened by denaturing high performance liquid chromatography analysis (DHPLC); and mutations were identified by direct sequencing. A heterozygous microdeletion mutation (c.27delC), which occurs within a symmetric element, was identified in two families. In one family, c.27delC showed segregation with the disease across two generations, with a penetrance of up to 75%. But in the other family, however, the same mutation manifested as a low-penetrance susceptibility factor. In addition, a novel heterozygous splicing mutation, c.87+1G>A (G>A substitution at nucleotide +1 of intron 2) was found in one family with familial pancreatitis. Our results also helped to resolve the sharply differing views about PSTI's role in pancreatitis.     

Spectrum of mutations and genotype-phenotype analysis in Currarino syndrome

The triad of a presacral tumour, sacral agenesis and anorectal malformation constitutes the Currarino syndrome which is caused by dorsal-ventral patterning defects during embryonic development. The syndrome occurs in the majority of patients as an autosomal dominant trait associated with mutations in the homeobox gene HLXB9 which encodes the nuclear protein HB9. However, genotype-phenotype analyses have been performed only in a few families and there are no reports about the specific impact of HLXB9 mutations on HB9 function. We performed a mutational analysis in 72 individuals from nine families with Currarino syndrome. We identified a total of five HLXB9 mutations, four novel and one known mutation, in four out of four families and one out of five sporadic cases. Highly variable phenotypes and a low penetrance with half of all carriers being clinically asymptomatic were found in three families, whereas affected members of one family showed almost identical phenotypes. However, an obvious genotype-phenotype correlation was not found. While HLXB9 mutations were diagnosed in 23 patients, no mutation or microdeletion was detected in four sporadic patients with Currarino syndrome. The distribution pattern of here and previously reported HLXB9 mutations indicates mutational predilection sites within exon 1 and the homeobox. Furthermore, sequence homology to Drosophila homeobox genes suggest that some of these mutations located within the homeobox may alter the DNA-binding specificity of HB9 while those in sequences homologous to a recently identified NLS motif of the human homeobox gene PDX-1 may impair nuclear translocation of the mutated protein.     

Mutational screening of the RB1 gene in Italian patients with retinoblastoma reveals 11 novel mutations

Retinoblastoma (RB, OMIM#180200) is the most common intraocular tumour in infancy and early childhood. Constituent mutations in the RB1 gene predispose individuals to RB development. We performed a mutational screening of the RB1 gene in Italian patients affected by RB referred to the Medical Genetics of the University of Siena. In 35 unrelated patients, we identified germline RB1 mutations in 6 out of 9 familial cases (66%) and in 7 out of 26 with no family history of RB (27%). Using the single-strand conformational polymorphism (SSCP) technique, 11 novel mutations were detected, including 3 nonsense, 5 frameshift and 4 splice-site mutations. Only two of these mutations (1 splice site and 1 missense) were previously reported. The mutation spectrum reflects the published literature, encompassing predominately nonsense or frameshift and splicing mutations. RB1 germline mutation was detected in 37% of our cases. Gross rearrangements outside the investigated region, altered DNA methylation, or mutations in non-coding regions, may be the cause of disease in the remainder of the patients. Some cases, e.g. a case of incomplete penetrance, or variable expressivity ranging from retinoma to multiple tumours, are discussed in detail. In addition, a case of pre-conception genetic counselling resolved by rescue of banked cordonal blood of the affected deceased child is described.     

Male and female differential reproductive rate could explain parental transmission asymmetry of mutation origin in Hirschsprung disease

Hirschsprung disease (HSCR, aganglionic megacolon) is a complex and heterogeneous disease with an incidence of 1 in 5000 live births. Despite the multifactorial determination of HSCR in the vast majority of cases, there is a monogenic subgroup for which private rare RET coding sequence mutations with high penetrance are found (45% of HSCR familial cases). An asymmetrical parental origin is observed for RET coding sequence mutations with a higher maternal inheritance. A parent-of-origin effect is usually assumed. Here we show that a differential reproductive rate for males and females also leads to an asymmetrical parental origin, which was never considered as a possible explanation till now. In the case of HSCR, we show a positive association between penetrance of the mutation and parental transmission asymmetry: no parental transmission asymmetry is observed in sporadic RET CDS mutation carrier cases for which penetrance of the mutation is low, whereas a parental transmission asymmetry is observed in affected sib-pairs for which penetrance of the mutation is higher. This allows us to conclude that the explanation for this parental asymmetry is that more severe mutations have resulted in a differential reproductive rate between male and female carriers.     

RAD51C is a susceptibility gene for ovarian cancer

A homozygous mutation in the RAD51C gene was recently found to cause Fanconi anemia-like disorder. Furthermore, six heterozygous deleterious RAD51C mutations were detected in German breast and ovarian cancer families. We screened 277 Finnish familial breast or ovarian cancer patients for RAD51C and identified two recurrent deleterious mutations (c.93delG and c.837+1G>A). These mutations were further genotyped in 491 familial breast cancer patients, 409 unselected ovarian cancer patients and two series of unselected breast cancer cases (884 from Helsinki and 686 from Tampere) and population controls (1279 and 807, respectively). The mutation frequency among all breast cancer cases was not different from the controls (4 out of 2239, 0.2% versus population controls 2 out of 2086, 0.1%, P= 0.7). In the Helsinki series, each mutation was found in four cases with personal or family history of ovarian cancer. No mutations were found among cases with familial breast cancer only, four out of the eight carriers did not have family history of breast cancer. The mutations associated with an increased risk of familial breast and ovarian cancer (OR: 13.59, 95% CI 1.89-97.6, P= 0.026 compared with controls), but especially with familial ovarian cancer in the absence of breast cancer (OR: 213, 95% CI 25.6-1769, P= 0.0002) and also with unselected ovarian cancer (OR: 6.31, 95% CI 1.15-34.6, P= 0.033), with a significantly higher mutation rate among the familial cases (two out of eight, 25%) than the unselected ovarian cancer cases (4 out of 409, 1%) (OR: 33.8, 95% CI 5.15-221, P= 0.005). These results suggest RAD51C as the first moderate-to-high risk susceptibility gene for ovarian cancer.     

Epidemiological Approach to Identifying Genetic Predispositions for Atypical Hemolytic Uremic Syndrome

Atypical hemolytic uremic syndrome (aHUS) is caused by several susceptibility genes. A registry including analyses of susceptibility genes, familial occurrence and genotype-phenotype correlation should provide classification insights.
Registry data of 187 unrelated index patients included age at onset, gender, family history, relapse of aHUS and potentially triggering conditions. Mutation analyses were performed in the genes CFH , CD46 and CFI and in the six potential susceptibility genes, FHR1 to FHR5 and C4BP .
Germline mutations were identified in 17% of the index cases; 12% in CFH , 3% in CD46 and 2% in CFI . Twenty-nine patients had heterozygous mutations and one each had a homozygous and compound heterozygous mutation. Mutations were not found in the genes FHR1-5 and C4BP . In 40% of the patients with familial HUS a mutation was found. Penetrance by age 45 was 50% among carriers of any mutation including results of relatives of mutation-positive index cases. The only risk factor for a mutation was family history of HUS (p = 0.02).
P enetrance of aHUS in carriers of mutations is not complete. Occurrence of homo- and heterozygous mutations in the same gene suggests that the number of necessary DNA variants remains unclear. Among clinical information only familial occurrence predicts a mutation.