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Parkinson's disease (PD) is a genetically heterogeneous disorder and new putative disease genes are discovered constantly. Therefore, whole-exome sequencing could be an efficient approach to genetic testing in PD. To evaluate its performance in early-onset sporadic PD, we performed diagnostic exome sequencing in 80 individuals with manifestation of PD symptoms at age 40 or earlier and a negative family history of PD. Variants in validated and candidate disease genes and risk factors for PD and atypical Parkinson syndromes were annotated, followed by further analysis for selected variants. We detected pathogenic variants in Mendelian genes in 6.25% of cases and high-impact risk factor variants in GBA in 5% of cases, resulting in overall maximum diagnostic yield of 11.25%. One individual was compound heterozygous for variants affecting canonical splice sites in VPS13C, confirming the causal role of protein-truncating variants in this gene linked to autosomal-recessive early-onset PD. Despite the low diagnostic yield of exome sequencing in sporadic early-onset PD, the confirmation of the recently discovered VPS13C gene highlights its advantage over using predefined gene panels.

Wilson's disease (WD) is a hereditary disorder caused by ATP7B mutations resulting in systemic copper accumulation. WD may manifest as early-adulthood parkinsonism; and atypical cases may be difficult to distinguish from early-onset Parkinson's disease (EO-PD), a neurodegenerative disorder with onset ≤40 years of age. The aim of our study was to compare transcranial sonography (TCS)-magnetic resonance fusion imaging in WD and EO-PD and examine whether TCS can provide clinically useful information.We examined 22 WD, 16 EO-PD, and 24 healthy control subjects. We measured echogenicity and determined presence of MRI signal changes in T2-weighted images in the substantia nigra (SN) and lentiform nucleus (NL). TCS with the capability of magnetic resonance fusion and Virtual Navigator was used. The echogenicity indices of SN and NL were processed using digital image analysis to eliminate subjective evaluation errors.Mean SN echogenicity index in EO-PD (39.8 ± 5.9 [SD]) was higher compared to WD (28.0 ± 4.6, p 0.0001) and control subjects (28.8 ± 4.9, p 0.0001). Mean NL echogenicity index was higher in WD (117.5 ± 37.0) compared to EO-PD (61.6 ± 5.4, p 0.0001) and control subjects (54.9 ± 11.2, p 0.0001). The SN hyperechogenicity had sensitivity 93.8%, and specificity 90.9%, while the NL hyperechogenicity had sensitivity 95.5% and specificity 93.8% for differentiation of WD and EO-PD. NL hyperechogenicity was more pronounced in WD subjects with putaminal MRI T2 hyperintensity (p 0.05) but was also present in subjects without MRI abnormality.There are distinct TCS findings in WD and EO-PD complementary to MRI that can be utilized as highly sensitive and specific biomarkers of these disorders.

Rett syndrome (RTT) is an X-linked neurodevelopmental disorder characterized by developmental regression with loss of motor, communication and social skills, onset of stereotypic hand movements and often seizures. RTT is primarily caused by de novo mutations in the methyl-CpG-binding protein 2 gene (MECP2). We established a high-resolution melting (HRM) technique for mutation scanning of the MECP2 gene and performed analyses in Czech patients with RTT, autism spectrum conditions and intellectual disability with Rett-like features. In the cases with confirmed MECP2 mutations, we determined X-chromosome inactivation (XCI), examined the relationships between genotype and clinical severity and evaluated the modifying influence of XCI. Our results demonstrate that HRM analysis is a reliable method for the detection of point mutations, small deletions and duplications in the MECP2 gene. We identified 29 pathogenic mutations in 75 girls, including four novel mutations: c.155_1189del1035;909_932inv;insC, c.573delC, c.857_858dupAA and c.1163_1200del38. Skewed XCI (ratio >75%) was found in 19.3% of the girls, but no gross divergence in clinical severity was observed. Our findings confirm a high mutation frequency in classic RTT (92%) and a correlation between the MECP2 mutation type and clinical severity. We also demonstrate limitations of XCI in explaining all of the phenotypic differences in RTT.

Rett syndrome is a neurodevelopmental disorder mainly caused by de novo mutations in the MECP2 (methyl-CpG-binding protein 2) gene. There is considerable variation in the severity of clinical features among Rett syndrome patients, even among patients with the same MECP2 mutation. In addition to X-chromosome inactivation pattern, the genetic background of the affected individual might also have a role in determining the severity of the disorder. We suggest that APOE is one of the genetic modulating factors. We analyzed clinical phenotypes of 46 patients with Rett syndrome, with confirmed MECP2 mutation. We discovered that among ε4 carriers, some clinical features were more severe, and the developmental regression occurred 4 months earlier on average than in those without the ε4 allele. Earlier onset of regression suggests a possible trend; however, it did not achieve distinctive statistical significance. Nevertheless, the ε4 allele of APOE may serve as a candidate modulation factor for the Rett syndrome phenotype.

Rett syndrome (RTT), an X-linked dominant neurodevelopmental disorder in females, is caused mainly by de novo mutations in the methyl-CpG-binding protein 2 gene (MECP2). Here we report mutation analysis of the MECP2 gene in 87 patients with RTT from the Czech and Slovak Republics, and Ukraine. The patients, all girls, with classical RTT were investigated for mutations using bi-directional DNA sequencing and conformation sensitive gel electrophoresis analysis of the coding sequence and exon/intron boundaries of the MECP2 gene. Restriction fragment length polymorphism analysis was performed to confirm the mutations that cause the creation or abolition of the restriction site. Mutation-negative cases were subsequently examined by multiple ligation-dependent probe amplification (MLPA) to identify large deletions. Mutation screening revealed 31 different mutations in 68 patients and 12 non-pathogenic polymorphisms. Six mutations have not been previously published: two point mutations (323T>A, 904C>T), three deletions (189_190delGA, 816_832del17, 1069delAGC) and one deletion/inversion (1063_1236del174;1189_1231inv43). MLPA analysis revealed large deletions in two patients. The detection rate was 78.16%. Our results confirm the high frequency of MECP2 mutations in females with RTT and provide data concerning the mutation heterogeneity in the Slavic population.

In this chapter, we describe genetic movement disorders arising from endogenous dysregulation of calcium, copper, and manganese homeostasis. In general, dysregulation of metal homeostasis may lead to regional brain metal accumulation, systemic metal accumulation affecting the whole brain and other organs, or brain metal deficiency.

Objective The aim of the study is to determine the frequency of parkin allelic variants in Czech early-onset Parkinson's disease patients and healthy controls. Methods A total of 70 early-onset Parkinson's disease patients (age at onset ≤40 years) and 75 controls were screened for the sequence variants and exon rearrangements in the parkin gene. Results Parkin mutations were identified in five patients (7.1%): the p.R334C point mutation was present in one patient, four patients had exon deletions. The detected mutations were observed in the heterozygous state except one homozygous deletion of the exon 4. No mutations were obtained in control subjects. A novel sequence variant p.V380I (c.1138G>A) was identified in one control. Non-pathogenic polymorphisms p.S167N and p.D394N were seen in similar percentage in patients and controls, polymorphism p.V380L was almost twice as frequent in controls as in patients. Conclusions Our study contributes to the growing body of evidence on the low frequency of the parkin mutations in the early-onset Parkinson's disease suggesting the potential role of other genes in the pathogenesis of the disease.

Essential tremor (ET) is one of the most common neurological movement disorders. In more than half of the cases, ET is inherited in an autosomal dominant manner, but no causative ET gene has been identified. However, several candidate loci have been reported, including the ETM2 locus that was originally found in a large American family of Czech descent.To ascertain the association with ETM2, we performed a genetic analysis of three polymorphic loci, etm1231, etm1234, and etm1240, located within the ETM2 candidate region in 61 Czech patients with a family history of ET and 68 healthy controls.The allele frequencies were not significantly different between the patients and the controls, and we did not observe any haplotype specifically associated with ET.This is the first report on the genetics of familial essential tremor in Czech patients. Our findings provide further evidence of genetic heterogeneity for ET.

Rett syndrome (RTT) is an X-linked disorder caused by mutations in the methyl-CpG-binding protein 2 gene (MECP2). The incidence is 1:10,000-1:15,000 females worldwide. To date, the mutational spectrum of MECP2 in the Ukrainian population is not known. Here we present first Ukrainian girl with classic clinical signs of RTT, in whom mutation of MECP2 gene was detected. Total genomic DNA was extracted from a dry blood spot using the QIAamp DNA Mini Kit (Qiagen) according to the manufacturer's protocol. Genomic DNA was used to amplify coding sequence and exon/intron borders of MECP2 gene. Products were examined by restriction analysis and automatic direct sequencing. The sequencing analysis of our patient revealed a small deletion of 4 bases AAAG at position 856-859 in exon 4 of MECP2 gene (856-859del4). This mutation leads to a frameshift (K286fs) and a premature stop codon. The creation of premature stop codon results in synthesis of truncated MeCP2 protein. Localization of the mutation into the transcription repression domain (TRD) probably affects the function of MECP2 protein in the process of transcriptional repression. To our knowledge this is the first case from Ukraine, in whom clinical diagnosis of RTT was confirmed by mutation analysis of MECP2 gene. Mutation analyses of further patients are needed to establish the spectrum of MECP2 mutations in the Ukrainian population. (Tab. 1, Fig. 3, Ref. 22.)