LRRK2 P755L variant in sporadic Parkinson’s disease

Parkinson's disease (PD) is a neurodegenerative disorder of unknown etiology with probable involvement of genetic-environmental factors. The majority of PD cases (approximately 90-95%) are sporadic, while familial cases account for approximately 5-10% of PD. In a recent report, a heterozygous LRRK2 P755L mutation within LRRK2 exon 19 was found in 2% of Chinese sporadic PD patients and in 0% of normal controls or Caucasians, suggesting that the mutation is disease-associated with ethnic specificity. To further evaluate the role of LRRK2 P755L variant in sporadic PD, we performed direct sequencing of LRRK2 exon 19 in 501 Japanese sporadic PD patients (male 249, female 252, aged 28-92 years, mean 65.0 years) and 583 controls of the Japanese general population as an extended association study. In this group, we found six patients (6/501 = 1.2%) and eight controls of the general population (8/583 = 1.6%) with a heterozygous P755L variant (P = 0.80, chi(2) = 0.064). No other variants were found in exon 19. Together with previous reports, our extended case-controlled study of large sample size suggests that LRRK2 P755L is a non-disease-associated polymorphism in PD patients.     

PLA2G6 mutations in PARK14-linked young-onset parkinsonism and sporadic Parkinson's disease

Mutations of PLA2G6 gene have been lately proposed to be the causative gene for PARK14 in patients with autosomal recessive young-onset parkinsonism (YOPD). The role of PLA2G6 mutations as a risk factor for Parkinson's disease is not clear. To study the PLA2G6 mutations in PARK14-linked patients and its association with the onset of sporadic Parkinson's disease (sPD), sequencing and gene dosage analyses were carried out in 25 patients (onset age ≦30 years) then the identified variants were assessed in 956 sporadic PD (sPD) patients and 802 age-matched healthy controls. Four genetic variants were identified; one patient had homozygous c.991G > T (p.Asp331Tyr) mutation, two had compound heterozygous c.991G > T/c.1077G > A (p.Met358IlefsX) mutation, one had single c.1976A > G (p.Asn659Ser) mutation, and one patient had an exon 1 hetero-deletion. The c.1077G > A mutation resulted in a 4-bp deletion in leukocyte mRNA by activating a cryptic splice site in exon 7. Only p.Asp331Tyr was identified in four sPD patients and four controls. The onset age for PLA2G6 mutation carriers was younger than that for sPD (29.86 ± 8.59 vs. 56.84 ± 11.33 years, P = 0.0002). The analysis of previously reported PARK14 patients revealed that those who carried a truncated mutation tended to have a complicated phenotype and atrophies of cortex and cerebellum. In conclusion, PLA2G6 mutation was the second common genetic cause after PRKN mutation in our YOPD patients and might be a risk factor for early-onset PD in Han Chinese. Additionally, mutation data should be interpreted carefully because even a synonymous mutation could cause abnormal mRNA splicing.     

Comprehensive mutational analysis of LRRK2 reveals variants supporting association with autosomal dominant Parkinson's disease

Parkinson's disease (PD) is a neurodegenerative disorder characterized by neurodegeneration, most notably of dopaminergic neurons in the substantia nigra. To date, six causative genes have been identified including LRRK2, whose mutations are the most frequent in autosomal dominant PD (Ad-PD). We conducted a comprehensive mutational analysis of LRRK2 in 30 Ad-PD (11 Japanese and 19 Caucasian) families employing a DNA microarray-based resequencing system and direct nucleotide sequence analysis, and identified 23 variants including two known mutations, p.G2019S and p.I1371V, in three Caucasian families and one Caucasian family, respectively, a novel putative pathogenic mutation, p.N1221K, in one Japanese family, and a known nonsynonymous variant, p.G2385R, in two Japanese families. Detailed analysis of the frequency of p.G2385R among 100 Japanese Ad-PD, 73 sporadic PD (sPD) and 238 controls revealed that the frequency of the p.G2385R variant was significantly higher in Ad-PD than in controls (allele frequency, 9.0 vs 2.1%) (χ(2)=16.32, P=5.34 × 10(-5)). The p.G2385R variant, however, did not show complete cosegregation with PD. In addition, the frequency of p.G2385R was also higher in sPD than in controls, although not significant (allele frequency, 3.4 vs 2.1%) (χ(2)=0.76, P=0.38). These observations support the possibility that p.G2385R is associated with an increased risk of PD.     

Analysis of PLA2G6 gene mutation in sporadic early-onset parkinsonism patients from Chinese population

Recent studies have shown that PLA2G6 is a causative gene for PARK14-linked autosomal recessive early-onset complicated dystonia-parkinsonism, early-onset parkinsonism with frontotemporal dementia and autosomal recessive early-onset Parkinsonism without added complicated clinical features. In order to investigate the characteristics of PLA2G6 gene mutations in Chinese sporadic early-onset parkinsonism (EOP) patients, we performed polymerase chain reaction and DNA direct sequencing on a cohort of sporadic EOP patients from Chinese population. In this study, we found a novel heterozygous varient (p.G679V). Bioinformatics demonstrates that p.G679V exhibits highly conserved residues across species, which hints it might be a pathogenic mutation. Our result indicated that PLA2G6 mutations might not be a main cause of Chinese sporadic EOP.     

LRRK2 G2019S is a common mutation in Spanish patients with late-onset Parkinson's disease

Mutations in the leucine-rich repat kinase 2 (LRRK2) gene have been shown to cause both autosomal dominant and sporadic Parkinson's disease (PD). The common G2019S mutation shows wide geographical distribution while R1441G has been only reported in Northern Spain. The overall frequency of these mutations remains to be established. To determine the prevalence of G2019S and R1441G mutations in our population of Cantabria (Northern Spain), we recruited 105 consecutive PD patients and 310 controls and conducted genetic analysis of these mutations. G2019S was detected in eight late-onset patients (7.6%). Five of them had no relevant family history. R1441G was not detected in any of our study subjects. The prevalence of G2019S mutation in unselected late-onset PD patients might be higher than previously reported: 3/16 (18.7%) of familial PD and 5/82 (6.1%) of sporadic PD.     

Identification of a Japanese family with LRRK2 p.R1441G-related Parkinson's disease

Leucine-rich repeat kinase 2 (LRRK2) is a causative gene of autosomal dominant familial Parkinson's disease (PD). We screened for LRRK2 mutations in 3 frequently reported exons (31, 41, and 48) in our cohort of 871 Japanese patients with PD (430 with sporadic PD and 441 probands with familial PD). Direct sequencing analysis of LRRK2 revealed 1 proband (0.11%) with a p.R1441G mutation, identified for the first time in Asian countries, besides frequently reported substitutions including, the p.G2019S mutation (0.11%) and p.G2385R variant (11.37%). Several studies have suggested that the LRRK2 p.R1441G mutation, which is highly prevalent in the Basque country, is extremely rare outside of northern Spain. Further analysis of family members of the proband with the p.R1441G mutation revealed that her mother and first cousin shared the same mutation and parkinsonism. Haplotype analysis revealed a different haplotype from that of the original Spanish families. Our patients demonstrated levodopa-responsive parkinsonism with intrafamilial clinical heterogeneity. This is the first report of familial PD because of the LRRK2 p.R1441G mutation in Asia.     

Low frequency of common LRRK2 mutations in Mexican patients with Parkinson's disease

Mutations in leucine-rich repeat kinase 2 gene (LRRK2) account for as much as 5–6% of familial Parkinson's disease (PD) and 1–2% of sporadic PD. These mutations represent the most frequent cause of autosomal dominant PD, particularly in certain ethnic groups. In this first report concerning LRRK2 mutations in Mexican-mestizos, we screened 319 consecutive PD patients (186 males; 133 females; mean age at onset: 52.4 years) for LRRK2 mutations in exons 31 and 41 and for the mutation in exon 35, which produces the Y1699C substitution. Three (0.94%) patients, two with sporadic PD and one with familial PD (disease mean age at onset, 53.3 years), were heterozygous for LRRK2 mutations. Of these three, two patients had one of two different mutations in exon 31 (R1441G and R1441H, respectively); the other patient carried the G2019S mutation in exon 41. The Y1699C mutation was absent from this PD sample. Four additional subjects, unaffected relatives of one PD patient with a mutation in LRRK2, were subsequently genetically tested. None of the three LRRK2 mutations identified was present in 200 neurologically healthy Mexican control individuals. These findings have important implications for molecular testing of LRRK2 mutations in Mexican PD patients.     

No evidence for pathogenic role of GIGYF2 mutation in Parkinson disease in Japanese patients

Grb10-Interacting GYF Protein-2 (GIGYF2) is a candidate gene for PARK11 locus. To date, seven different GIGYF2 missense mutations have been identified in patients with familial Parkinson disease (PD) of European descent. To clarify the pathogenic role of GIGYF2 in PD, we analyzed the frequency of GIGYF2 mutations in 389 Japanese patients with PD (including 93 patients with late-onset familial PD, 276 with sporadic PD, and 20 with a single heterozygous mutation in the PD-associated genes), and 336 Japanese normal controls, by direct sequencing and/or high-resolution melting analysis. None of the reported GIGYF2 mutations or digenic mutations were detected. Two novel non-synonymous variants were identified (p.Q1211delQ and p.H1023Q), however, we could not determine their roles in PD. In summary, we found no evidence for PD-associated roles of GIGYF2 mutations. Our data suggest that GIGYF2 is unlikely to play a major role in PD in Japanese patients, similar to other populations.     

Differential orexin/hypocretin expression in addiction-prone and -resistant rats selectively bred for high (HiS) and low (LoS) saccharin intake

Recent studies have shown an association between Parkinson disease (PD) and mutations in the gene encoding the lysosomal enzyme glucocerebrosidase (GBA), which is deficient in patients with Gaucher disease. In Asian populations, 2 mutational analysis studies have been performed in all exons of GBA; one study in a Japanese population showed the highest odds ratio among all ethnic groups, whereas the other study in ethnic Chinese observed a trend of a higher frequency of GBA mutation in PD patients without statistical significance. To investigate whether there is an association between PD and mutations of GBA in a Korean population, we analyzed mutations of GBA and compared mutation frequencies between Korean PD patients and a control population. We analyzed mutations in GBA by sequencing exons of GBA in 277 Korean PD patients and 291 control subjects. All exons of GBA were sequenced in all PD cases and 100 control subjects. Exon 2 and exons 5-11, where mutations of GBA were found in our PD patients, were analyzed in an additional 191 control subjects. Five different pathogenic heterozygous GBA mutations, including N188S, P201H, R257Q, S271G, and L444P, were identified in 9 PD cases (3.2%), whereas there were no GBA mutations found in control subjects (p<0.01, OR 20.6, 95% CI 1.2-356.4). The mean age-at-onset of heterozygous GBA variants carriers was younger than that of non-carriers (48.6±11.9 versus 57.9±13.5, p<0.05, Mann-Whitney test). Our results suggest that heterozygous mutations of GBA represent a risk factor for PD in Koreans.     

A novel mutation of MSX1 inherited from maternal mosaicism causes a severely affected child with nonsyndromic oligodontia

Mutations of MSX1 have been associated with nonsyndromic hypodontia. To seek the causal gene mutation sites in a family with nonsyndromic oligodontia, whole-exome sequencing (WES) was performed to seek the causative locus of the family. The candidate mutation was further identified by Sanger sequencing afterward. Two mutations of MSX1 were found both in the proband and her mother. One novel heterozygous missense mutation (c.C667G, p.R223G) of MSX1 inherited from the asymptomatic mother with mosaic mutation was located in the highly conserved fragment of exon 2. The other was a synonymous mutation (c.C348T, p.G116G) in exon 1, which had been reported. The novel maternal heterozygous missense mutation (c.C667G, p.R223G) was likely to be the major reason for nonsyndromic oligodontia in the family. This is the first mosaic variant that has been recorded of the MSX1 gene. Our study expands the phenotype–genotype correlation associated with MSX1 variants. Our study also suggests that the determination of the mosaicism is essential for precise genetic counseling if a disease appears to arise de novo.     

Variants at the secretory phospholipase A2 (PLA2G2A) locus: analysis of associations with familial adenomatous polyposis and sporadic colorectal tumours

The Min mouse is a model for human familial adenomatous polyposis (FAP), an autosomal dominant disease characterised by multiple adenomatous gastrointestinal polyps. The severity of the Min phenotype is modified by a locus ( Mom1 ) on mouse chromosome 4, at a position syntenic with human chromosome 1p35-p36. The secretory phospholipase A2 ( Pla2s ) gene is a candidate for this modifier locus and there is evidence that a locus on human chromosome 1p35-p36 acts to modify the severity of human duodenal FAP. We have analysed the human secretory phospholipase A2 locus ( PLA2G2A ) for variants that could directly influence the FAP phenotype. We found no PLA2G2A variants predicted to result in functional variation in the phospholipase A2 protein. Two PLA2G2A polymorphisms were, however, discovered, one a 'silent' base change in exon 3 and another in a non-coding region. Three other variants (possible mutations) were found in non-coding regions. In 70 FAP patients from 20 families, no associations were found between the severity of duodenal polyposis and any PLA2G2A variant. One allele at the exon 3 polymorphic site did, however, occur more often then expected in patients with relatively severe colonie FAP. Although of borderline statistical significance, this association, if genuine, is likely to result from linkage disequilibrium between the PLA2G2A alleles studied and undetected genetic variation at a closely linked locus. The frequency of the alleles at both polymorphic sites has also been determined in the germ line of patients with sporadic colorectal adenomas and carcinomas and in random controls, but no differences were found among these groups. Our results suggest that PLA2G2A variants do not influence inherited or sporadic colonie tumours. A linked locus may be a modifier of human FAP, but does not influence the risk of colorectal tumours in the general population.     

视网膜色素变性隐性遗传致病基因PDE6B的突变分析

目的 了解常染色体隐性遗传视网膜色素变性(autosomal recessive retinitis pigmentosa，ARRP)致病基因磷酸二酯酶β亚单位(phosphodiesterase β subunit,PDE6B)基因在中国视网膜色素变性(retinitis pigmentosa，RP)患者中的突变谱及突变率。方法 应用聚合酶链反应—单链构象多态性，对收集的35个常染色体隐性RP家系38例患者和55例散发RP患者进行PDE6B基因的22个外显子和5’端非翻译区突变筛选；对有变异条带者进行DNA序列分析。结果 测得一个常染色体隐性家系患者PDE6B基因第11外显子5’端上游第19位碱基(第10内含子内)发生G→A转换。1例散发RP患者同时检测到第6外显子第2492位点碱基T颠换为C和第10外显子5’端上游(第9内含子内)第27—28碱基之间有两个碱基TG插入。另两例散发RP患者分别发现第4外显子5’端上游30—31碱基处两个碱基GT插入和第18外显子3’端下游第15个碱基发生G→C颠换。结论 发现1名中国人的散发RP患者携带RP致病基因PDE6B基因的一种复合杂合突变。中国人的PDE6B基因内含子有多种变异。     

Multiple LRRK2 variants modulate risk of Parkinson disease: a chinese multicenter study

We and others found two polymorphic LRRK2 (leucine‐rich repeat kinase 2) variants (rs34778348:G>A; p.G2385R and rs33949390:G>C; p.R1628P) associated with Parkinson disease (PD) among Chinese patients, but the common worldwide rs34637584:G>A; p.G2019S mutation, was absent. Focusing exclusively on Han Chinese, we first sequenced the coding regions in young onset and familial PD patients and identified 59 variants. We then examined these variants in 250 patients and 250 control subjects. Among the 17 polymorphic variants, five demonstrated different frequency in cases versus controls and were considered in a larger sample of 1,363 patients and 1,251 control subjects. The relative risk of an individual with both p.G2385R and p.R1628P is about 1.9, and this is reduced to 1.5–1.6 if the individual also carries rs7133914:G>C; p.R1398H or rs7308720:C>A: p.N551K. The risk of a carrier with p.R1628P is largely negated if the individual also carries p.R1398H or p.N551K. In dopaminergic neuronal lines, p.R1398H had significantly lower kinase activity, whereas p.G2385R and p.R1628P showed higher kinase activity than wild type. We provided the first evidence that multiple LRRK2 variants exert an individual effect and together modulate the risk of PD among Chinese. Hum Mutat 31:561–568, 2010. © 2010 Wiley‐Liss, Inc.     

Analysis of the PINK1 gene in a cohort of patients with sporadic early-onset parkinsonism in Taiwan

Mutations in the PINK1 gene have been shown to cause autosomal recessive Parkinson's disease (PD) and/or early onset sporadic PD in Italy, Spain, North America, Ireland, and Asia. However, there are limited data on PINK1 mutations in sporadic early onset Asian PD patients. To determine the prevalence of PINK1 mutation in Taiwanese population, we conducted genetic analysis of PINK1 mutation in 73 early onset sporadic PD and 94 normal control subjects. We only identified a novel single heterozygous mutation R 407Q mutation in exon 6 of this gene in one patient at the age onset of 54. Overall, these data indicate that PINK1 mutations are rare in our population. Based on our results, unless common mutational hotspots are identified, routine testing for this mutation at least in our population may not be cost-effective.     

Association between GBA L444P mutation and sporadic Parkinson's disease from Mainland China

Mutations in GBA gene have been reported to be in patients with Parkinson's disease (PD) from different ethnic populations, including Taiwanese Chinese. To explore whether mutation in GBA is also associated with PD in Mainland China, we have now a case control study. The occurrence of the GBA L444P mutation was analyzed in an independent cohort of PD patients and controls from Mainland China. This mutation was present in 20/616 (3.2%) of PD compared with 1/411 (0.2%) of controls (odds ratio, OR = 13.76, 95% Confidence interval, CI: 1.84–102.92, p = 0.001). All carriers harbored the heterozygous genotype. In a subset analysis, the frequency of this mutation was higher both in early onset (EOPD) and late onset PD (LOPD) than in controls. However, no difference in clinical characteristics, such as gender, age at onset, onset symptoms, Hoehn–Yahr stage and UPDRS, was found between L444P carriers and non-carriers. In addition, we also explored the potential relationship between GBA L444P mutation and LRRK2 G2385R and R1628P variants in patients with PD. But no association was found, either. In conclusion, our data suggest that the GBA L444P mutation plays an important role in the development of PD also in Han-Chinese patients from Mainland China.     

De novo FUS gene mutations are associated with juvenile-onset sporadic amyotrophic lateral sclerosis in China

Juvenile amyotrophic lateral sclerosis (ALS) is a rare form of motor neuron disease and occurs before 25 years of age. Only very few sporadic cases of juvenile-onset ALS have been reported. Rare SOD1 mutations and several FUS mutations have been identified in juvenile-onset ALS patients. To define the genetics of juvenile-onset sporadic ALS (SALS) of Chinese origin, we sequenced all 5 exons of SOD1, exons 3–6 and 12–15 of FUS in 11 juvenile-onset SALS patients, 105 adult-onset ALS patients (including 6 familial ALS [FALS] pedigrees), and 245 healthy controls. For the 11 juvenile-onset SALS and 6 FALS cases, the other 7 exons of FUS were also screened. A heterozygous de novo missense mutation c.1574C>T (p.P525L), a heterozygous de novo 2-base pair deletion c.1509_1510delAG (p.G504Wfs*12), and a nonsense mutation c.1483C>T (p.R495X) was each identified in 1 juvenile SALS patient. A heterozygous missense mutation c.1561C>G (p.R521G) was identified in a FALS proband. In the Chinese population, the frequency of FUS mutation in FALS is 11.4% (95% confidence interval [CI], 0.9%–22.0%), higher than the Japanese (10%; 95% CI, 0.7%–19.3%), and Caucasians (4.9%; 95% CI, 3.9%–6.0%). The frequency of FUS mutation in SALS patients is 1.5% (95% CI, 0.2%–2.9%), which is similar to Koreans (1.6%; 95% CI, 0%–3.2%), but higher than in Caucasians (0.6%; 95% CI, 0.4%–0.8%). Our findings suggest that de novo FUS mutations are associated with juvenile-onset SALS of Chinese origin and that this gene should be screened in ALS patients with a young age of onset, aggressive progression, and sporadic occurrence.     

A novel peripherin gene (PRPH) mutation identified in one sporadic amyotrophic lateral sclerosis patient

Motor neurons in amyotrophic lateral sclerosis (ALS) are characterized by the presence of inclusion bodies composed of intermediate filament (IF) proteins. Peripherin protein is as components of these inclusions and rare mutations in peripherin gene (PRPH) were identified in sporadic ALS cases. The aim of this study was to further define the spectrum of PRPH mutations in a cohort of 122 Italian ALS patients. We screened the coding sequence, the exon/intron boundaries, and the 5′-3′ un-translated regions (UTRs) in 122 ALS patients. Eighteen sequence variations were detected. Seven variants were not identified in a panel of at least 245 matched controls, including 2 missense variations, namely p.R133P and p.D141Y, each identified in one heterozygous patient. p.R133P was newly identified whereas p.D141Y was previously described in one homozygous sporadic ALS patient. These 2 variants were predicted to have a deleterious effect on protein structure or function. This work contributes to determine the role of PRPH gene variants in ALS. Further studies are necessary to define the mechanisms through which the mutant peripherin could cause ALS phenotype.     

Clinical features and genetic analysis of seven patients with congenital hyperinsulinismCSCD

Objective: To analyze clinical characteristics, genetic mutation and therapeutic effect of seven patients diagnosed with congenital hyperinsulinism(CHI). Methods: Clinical data for the patients was retrospectively analyzed. Results: All patients presented with hyperinsulinism(serum insulin:2. 0-58. 4 mU/L),even after hypoglycemia (blood glucose: 0. 7-2. 39 mmol/L)has developed. Mutations were identified in 4 patients (57. 1%), which included a heterozygous c. 262C>T(p. R88C) mutation in exon 4 of the UCP2 gene, a heterozygous c. 1495C>A(p. G499C) mutation in exon 12 of the GLUD1 gene, a heterozygous c. 1493C>T(p. S498L) mutation in exon 1 of the GLUD1 gene, and a heterozygous c. 4432G > A (p. G1478R) mutation in exon 37 of the ABCC8 gene. The patient carrying a maternally inherited ABCC8 mutation was treated with cornstarch and had his blood glucose kept normal. All other patients responded well to diazoxide. Conclusion: A genetic diagnosis was attained for 51. 7% of patients in this study. Mild CHI patients can have their blood glucose controlled by giving cornstarch. Diazoxide is safe and effective for most CHI patients. © 2018 MeDitorial Ltd. All rights reserved.     

Absence/rarity of commonly reported LRRK2 mutations in Indian Parkinson's disease patients

Recent discovery of pathogenic mutations in the leucine-rich repeat kinase 2 (LRRK2) gene in Parkinson's disease (PD) patients in different ethnic groups have raised a hope of diagnostic screening and genetic counseling. We investigated the six most commonly reported mutations in LRRK2 gene among Indian PD patients, using PCR-RFLP method. Mutations G2019S, R1441C, R1441G, and R1441H were screened in 1012 individuals (PD, 800; controls, 212) while mutations I2012T and I2020T were screened in 748 PD patients. We did not observe any of these six mutations in this study sample except in a single female young onset PD patient who showed a heterozygous G2019S mutation. The absence of mutations was reconfirmed by sequencing of probands from several autosomal dominant PD families. Our observations suggest that these mutations may be a rare cause of PD among Indians and therefore of little help for diagnostic screening and genetic counseling for Indian PD patients.     

BRCA1 germline mutations in Indian familial breast cancer

Germline mutation analysis of BRCA1 gene has demonstrated significant allelic heterogeneity. These differences represent historical influences of migration, population structure and geographic or cultural isolation. To date, there have been no reports of Indian families with mutations in BRCA1. We have screened for mutations in selected coding exons of BRCA1 and their flanking intron regions in three breast or breast and ovarian cancer families with family history of three or more cases of breast cancer under age 45 and/or ovarian cancer at any age. We have also analyzed 10 female patients with sporadic breast cancer regardless of age and family history, as well as 50 unrelated normal individuals as controls. Thus a total of 90 samples were analyzed for BRCA1 mutations using polymerase chain reaction-mediated site directed mutagenesis (PSM) and single stranded conformation polymorphism (SSCP) analysis for various selected exons followed by sequencing of variant bands. Eight point mutations were identified. Two deleterious pathogenic, protein truncating non-sense mutations were detected in exon 11 (E1250X) and exon 20 (E1754X) and six novel and unique amino acid substitutions (F1734S, D1739Y, V1741G, Q1747H, P1749A, R1753K). One complex missense mutation of exon 20 [V1741G; P1749A] was seen in two out of three families and another complex combination of missense and non-sense mutations of the same exon [V1741G; E1754X] was observed in only one family. These complex mutations exist only in breast cancer families but not in control populations of women. Three splice site variants (IVS20+3A>C, IVS20+4A>T, IVS20+5A>T) and two intronic variants (IVS20+21_22insG, IVS20+21T>G) were also detected. In the group of 10 sporadic female patients no mutations were found.