Susen Winkler

α-Synuclein and Parkinson disease susceptibility

Background: Mutations in the α-synuclein (SNCA) gene have been shown to be responsible for a rare familial form of Parkinson disease (PD). Furthermore, polymorphic variants in multiple regions of the gene have been associated with susceptibility to idiopathic PD in different populations. Objective: To evaluate and to confirm the role of SNCA variants in PD pathogenesis. Methods: We included 667 subjects (397 cases with idiopathic PD and 270 healthy, ethnically matched controls) of Northern Central and Southeastern European origin. We analyzed genotypes at 14 markers spanning the SNCA locus and its major haplotype blocks and conducted a haplotype analysis for four promoter markers including the microsatellite marker Rep1. Results: The three single nucleotide polymorphisms (SNPs) of the promoter region (rs2583988, rs2619364, rs2619363) and a SNP in the 3′UTR (rs356165) of the SNCA gene showed the greatest evidence for an association with PD (p ≤ 0.003), with significant pairwise values for linkage disequilibrium (D′ ≥ 0.74, r 2 ≥ 0.29). The promoter haplotype “261-T-G-T” (Rep1-rs2583988-rs2619364-rs2619363) was associated with disease (p = 0.032). The most significant association with PD was generated by excluding Rep1 (p = 0.008). This association remained significant when analyzing the Serbian patients separately and was of borderline significance for the German patients. Conclusions: Our findings confirm that genetic variability within the SNCA locus is associated with susceptibility to idiopathic Parkinson disease (PD). We found evidence for disease association with single nucleotide polymorphisms at both the 5′ and the 3′ end of the gene with pairwise linkage disequilibrium between them. The association was independent of the Rep1 status, and one major SNCA promoter haplotype class seems to be associated with PD susceptibility. GLOSSARY: SNCA = α-synuclein; CI = confidence interval; HT = haplotype-tagging; LD = linkage disequilibrium; OR = odds ratio; PD = Parkinson disease; SNP = single nucleotide polymorphism.

PINK1 , Parkin , and DJ-1 mutations in Italian patients with early-onset parkinsonism

Recessively inherited early-onset parkinsonism (EOP) has been associated with mutations in the Parkin, DJ-1, and PINK1 genes. We studied the prevalence of mutations in all three genes in 65 Italian patients (mean age of onset: 43.2±5.4 years, 62 sporadic, three familial), selected by age at onset equal or younger than 51 years. Clinical features were compatible with idiopathic Parkinsons disease in all cases. To detect small sequence alterations in Parkin, DJ-1, and PINK1, we performed a conventional mutational analysis (SSCP/dHPLC/sequencing) of all coding exons of these genes. To test for the presence of exon rearrangements in PINK1, we established a new quantitative duplex PCR assay. Gene dosage alterations in Parkin and DJ-1 were excluded using previously reported protocols. Five patients (8%; one woman/four men; mean age at onset: 38.2±9.7 (range 25–49) years) carried mutations in one of the genes studied: three cases had novel PINK1 mutations, one of which occurred twice (homozygous c.1602_1603insCAA; heterozygous c.1602_1603insCAA; heterozygous c.836G>A), and two patients had known Parkin mutations (heterozygous c.734A>T and c.924C>T; heterozygous c.924C>T). Family history was negative for all mutation carriers, but one with a history of tremor. Additionally, we detected one novel polymorphism (c.344A>T) and four novel PINK1 changes of unknown pathogenic significance (−21G/A; IVS1+97A/G; IVS3+38_40delTTT; c.852C>T), but no exon rearrangements. No mutations were found in the DJ-1 gene. The number of mutation carriers in both the Parkin and the PINK1 gene in our cohort is low but comparable, suggesting that PINK1 has to be considered in EOP.

Whispering dysphonia (DYT4 dystonia) is caused by a mutation in the TUBB4 gene

A study was undertaken to identify the gene underlying DYT4 dystonia, a dominantly inherited form of spasmodic dysphonia combined with other focal or generalized dystonia and a characteristic facies and body habitus, in an Australian family.

Bilateral subthalamic stimulation in Parkin and PINK1 parkinsonism

Objectives: To study the frequency of different gene mutations in patients with early-onset parkinsonism and bilateral subthalamic nucleus deep brain stimulation (STN-DBS) and the short- and long-term surgical outcome in mutation-positive (MUT+) and -negative (MUT−) patients. Methods: Eighty patients with disease onset at age ≤ 45 years and bilateral STN-DBS were screened for mutations in the Parkin gene and PINK1 gene and for the recurrent p.G2019S mutation in the LRRK2 gene. The Unified Parkinson’s Disease Rating Scale (UPDRS) and Hoehn and Yahr (H-Y) scale were used to compare the on- and off-medication conditions preoperatively and in the off-medication/on-stimulation condition postoperatively. Results: We identified 12 mutation carriers (11 Parkin [6 with 2 mutated alleles, 5 with 1 mutated allele], 1 homozygous PINK1). There were no clinical differences between the MUT− and MUT+ patients preoperatively, except for more severe H-Y stage and postural and gait scores in the on-medication state in the MUT+ group. During the first year after surgery, MUT− patients showed better clinical improvement (56% motor UPDRS improvement) compared with MUT+ patients (36%). However, in the long-term follow-up (3–6 years), both groups presented with the same degree of clinical improvement (MUT−: 44% vs MUT+: 42%). Although the MUT+ group showed more severe axial signs preoperatively, MUT− patients developed levodopa– and deep brain stimulation–resistant axial signs within the first 3 to 6 years postoperatively, which diminished the initial benefit soon after surgery. Conclusions: Patients with Parkin or PINK1 mutations benefit from subthalamic nucleus deep brain stimulation. However, the clinical response is not superior to non–mutation carriers and might be limited by more advanced axial motor symptoms at a relatively early disease stage.

ATP13A2 variants in early-onset Parkinson's disease patients and controls.

Four genes responsible for recessively inherited forms of Parkinsons disease (PD) have been identified, including the recently discovered ATP13A2 (PARK9) gene. Our objective was to investigate the role of this gene in a large cohort of PD patients and controls. We extensively screened all 29 exons of the ATP13A2 coding region in 112 patients with early‐onset PD (EOPD; <40 years) of mostly European ethnic origin and of 55 controls. We identified four carriers (3.6%) of novel single heterozygous ATP13A2 missense changes that were absent in controls. Interestingly, the carrier of one of these variants also harbored two mutations in the Parkin gene. None of the carriers had atypical features previously described in patients with two mutated ATP13A2 alleles (Kufor–Rakeb syndrome). Our data suggest that two mutated ATP13A2 alleles are not a common cause of PD. Although heterozygous variants are present in a considerable number of patients, they are—based on this relatively small sample—not significantly more frequent in patients compared to controls.

Childhood-onset restless legs syndrome: clinical and genetic features of 22 families.

Restless legs syndrome (RLS) is a sensory‐motor disorder that is underdiagnosed in children and often misclassified as attention deficit hyperactivity disorder. Five different gene loci (RLS1‐5) and three susceptibility loci have been identified in adult‐onset RLS. We included 23 children with RLS (age at onset ≤14 years) from 22 families. In 14 families, we performed linkage and genotype analyses. Of the 23 RLS patients, only seven (30.4%) were admitted for a suspected diagnosis of RLS. Five patients had a retrospectively established onset at an age as early as 1 year. The most frequent complaint in patients were sleep problems (21 of 23; 91%) resulting in fatigue in 14 children (60.9%). Twelve of the 19 tested cases (63.2%) exhibited an index of periodic limb movements in sleep greater than 5. Dopaminergic therapy was successful in 12 of 14 treated patients (85.7%). Family history for RLS was positive in 20 of 23 children (87.0%) and compatible with an autosomal dominant inheritance pattern. Linkage analysis excluded all five loci in two families. A trend for an association at two of the three reported susceptibility regions was observed. RLS symptoms can occur in early childhood. The positive family history suggests a genetic cause in most families with at least one additional RLS gene locus.

Myoclonus-Dystonia Due to Maternal Uniparental Disomy

BACKGROUND Myoclonus-dystonia is a movement disorder often associated with mutations in the maternally imprinted epsilon-sarcoglycan (SGCE) gene located on chromosome 7q21. Silver-Russell syndrome is a heterogeneous disorder characterized by prenatal and postnatal growth restriction and a characteristic facies, caused in some cases by maternal uniparental disomy of chromosome 7. OBJECTIVES To describe and investigate the combination of a typical myoclonus-dystonia syndrome and Silver-Russell syndrome. DESIGN Clinical and neurophysiological examination as well as cytogenetic and molecular analyses. SETTING Movement disorder clinic. Patient A 36-year-old man with typical myoclonus-dystonia and Silver-Russell syndrome. MAIN OUTCOME MEASURES Clinical description of the disease and its genetic cause. RESULTS Cytogenetic analysis revealed mosaicism for a small chromosome 7 marker chromosome. Microsatellite analysis indicated loss of the paternal allele and maternal uniparental disomy of chromosome 7. In keeping with the maternal imprinting mechanism, no unmethylated allele of SGCE was detected after bisulfite treatment of the patients DNA, and reverse transcription-polymerase chain reaction demonstrated loss of SGCE expression. Molecular analysis ruled out mutations in the SGCE gene. CONCLUSIONS We identified a new genetic alteration-maternal chromosome 7 disomy-that can cause myoclonus-dystonia. This alteration results in repression of both alleles of the maternally imprinted SGCE gene and suggests SGCE loss of function as the disease mechanism.

Recurrent LRRK2 (Park8) mutations in early-onset Parkinson's disease.

Mutations in LRRK2 (leucine‐rich repeat kinase 2) have been associated with autosomal dominant Parkinsons disease (PD) and cluster in several 3′ exons of the gene. The majority of mutations have been detected in late‐onset cases (age at onset >50 years). We screened 5 of the 51 exons of LRRK2 that previously have been reported to harbor mutations in 98 early‐onset and 42 late‐onset PD patients. We identified two mutations (c.4321C>T, c.6055G>A) in three early‐onset patients. Screening of an additional 220 early‐onset PD patients for these mutations revealed another mutation carrier. In conclusion, LRRK2 mutations need to be considered also in early‐onset PD.

Whispering dysphonia in an Australian family (DYT4): a clinical and genetic reappraisal.

The designation, DYT4, was assigned to an Australian family with whispering dysphonia. The role of known causes of dystonia has not been comprehensively investigated in this family, nor has the possible relationship with Wilson disease (WND) in 2 siblings. Eighteen family members were neurologically examined, and DNA samples were obtained. Linkage analysis was performed to DYT1, DYT6, DYT7, DYT11, DYT13, DYT15, and ATP7B with microsatellite markers and the THAP1 (DYT6), PRKRA (DYT16), and ATP7B (WND) genes were sequenced. Reevaluation of the family identified 9 living affected family members, 6 of whom are newly affected. Phenotypic expression was variable, ranging from isolated spasmodic dysphonia (often with mild craniocervical dystonia) to severe generalized dystonia. Two newly described features included an extrusional tongue dystonia and a unique “hobby horse gait.” Genetic analyses excluded all tested loci. Haplotype analysis of the ATP7B region resulted in three different combinations of the two parental alleles in the 8 investigated siblings of the 2 deceased WND patients, indicating that the fourth combination (of two mutated alleles) had occurred only in the deceased WND patients. On these two alleles, we identified a missense (c.2297C>G; p.T766R) and a splice‐site mutation (IVS5+1G>T). The c.2297C>G mutation was detected in 3 affected and 4 unaffected family members, whereas the IVS5+1G>T mutation was detected in 1 affected and unaffected family member. Five DYT4 patients carried neither mutation. DYT4 is a familial form of dystonia unrelated to known dystonia genes and loci. ATP7B mutations do not segregate with the dystonia phenotype, indicating two independent genetic diseases in this family.

Identification and functional analysis of novel THAP1 mutations

Mutations in THAP1 have been associated with dystonia 6 (DYT6). THAP1 encodes a transcription factor that represses the expression of DYT1. To further evaluate the mutational spectrum of THAP1 and its associated phenotype, we sequenced THAP1 in 567 patients with focal (n=461), segmental (n=68), or generalized dystonia (n=38). We identified 10 novel variants, including six missense substitutions within the DNA-binding Thanatos-associated protein domain (Arg13His, Lys16Glu, His23Pro, Lys24Glu, Pro26Leu, Ile80Val), a 1bp-deletion downstream of the nuclear localization signal (Asp191Thrfs*9), and three alterations in the untranslated regions. The effect of the missense variants was assessed using prediction tools and luciferase reporter gene assays. This indicated the Ile80Val substitution as a benign variant. The subcellular localization of Asp191Thrfs*9 suggests a disturbed nuclear import for this mutation. Thus, we consider six of the 10 novel variants as pathogenic mutations accounting for a mutation frequency of 1.1%. Mutation carriers presented mainly with early onset dystonia (<12 years in five of six patients). Symptoms started in an arm or neck and spread to become generalized in three patients or segmental in two patients. Speech was affected in four mutation carriers. In conclusion, THAP1 mutations are rare in unselected dystonia patients and functional analysis is necessary to distinguish between benign variants and pathogenic mutations.