Genome-wide meta-analyses of restless legs syndrome yield insights into genetic architecture, disease biology and risk prediction.

Restless legs syndrome (RLS) affects up to 10% of older adults. Their healthcare is impeded by delayed diagnosis and insufficient treatment. To advance disease prediction and find new entry points for therapy, we performed meta-analyses of genome-wide association studies in 116,647 individuals with RLS (cases) and 1,546,466 controls of European ancestry. The pooled analysis increased the number of risk loci eightfold to 164, including three on chromosome X. Sex-specific meta-analyses revealed largely overlapping genetic predispositions of the sexes (rg = 0.96). Locus annotation prioritized druggable genes such as glutamate receptors 1 and 4, and Mendelian randomization indicated RLS as a causal risk factor for diabetes. Machine learning approaches combining genetic and nongenetic information performed best in risk prediction (area under the curve (AUC) = 0.82-0.91). In summary, we identified targets for drug development and repurposing, prioritized potential causal relationships between RLS and relevant comorbidities and risk factors for follow-up and provided evidence that nonlinear interactions are likely relevant to RLS risk prediction.

From brain to spinal cord: neuromodulation by direct current stimulation and its promising effects as a treatment option for restless legs syndrome.

Neuromodulation is a fast-growing field of mostly non-invasive therapies, which includes spinal cord stimulation (SCS), transcranial direct current stimulation (tDCS), vagal nerve stimulation (VNS), peripheral nerve stimulation, transcranial magnetic stimulation (TMS) and transcutaneous spinal direct current stimulation (tsDCS). This narrative review offers an overview of the therapy options, especially of tDCS and tsDCS for chronic pain and spinal cord injury. Finally, we discuss the potential of tsDCS in Restless Legs Syndrome as a promising non-invasive, alternative therapy to medication therapy.

Case report: An OSAS patient with comorbid RLS/PLMS - a closer look at the causes of PAP intolerance.

Since 2017, hypoglossal nerve stimulation has been included in the S3-guidelines on restorative sleep/sleep disorders as an alternative treatment for patients with obstructive sleep related breathing disorders who cannot tolerate conventional PAP-therapy. Under certain conditions, some of these patients have the option to have a tongue pacemaker implanted during a surgical procedure to regain a restful night's sleep. However, in some cases it does not solve the problem. In this case report, we present a patient who continued to have restless sleep despite implantation of a hypoglossus nerve stimulator. We provide a closer look at the underlying causes of PAP intolerance and emphasize the importance of a combined pneumological and neurological approach to sleep medicine in sleep-specific therapy evaluation.

ExomeChip-based rare variant association study in restless legs syndrome.

Restless legs syndrome (RLS) is a common sleep-related movement disorder in populations of European descent and disease risk is strongly influenced by genetic factors. Common variants have been assessed extensively in several genome-wide association studies, but the contribution of rarer genetic variation has not been investigated at this scale. We therefore genotyped a case-control set of 9246 individuals for mainly rare and low frequency exonic variants using the Illumina ExomeChip. However, standard single variant and gene-level association tests were negative. This does not preclude a role of rare variants in RLS, but is likely due to the small sample size and the limited selection of rare genetic variation captured on the array. Therefore, exome or whole genome sequencing should be performed rather than increasing the sample size of ExomeChip studies in order to identify rare risk variants for RLS.

Identification of Restless Legs Syndrome Genes by Mutational Load Analysis.

OBJECTIVE: Restless legs syndrome is a frequent neurological disorder with substantial burden on individual well-being and public health. Genetic risk loci have been identified, but the causatives genes at these loci are largely unknown, so that functional investigation and clinical translation of molecular research data are still inhibited. To identify putatively causative genes, we searched for highly significant mutational burden in candidate genes. METHODS: We analyzed 84 candidate genes in 4,649 patients and 4,982 controls by next generation sequencing using molecular inversion probes that targeted mainly coding regions. The burden of low-frequency and rare variants was assessed, and in addition, an algorithm (binomial performance deviation analysis) was established to estimate independently the sequence variation in the probe binding regions from the variation in sequencing depth. RESULTS: Highly significant results (considering the number of genes in the genome) of the conventional burden test and the binomial performance deviation analysis overlapped significantly. Fourteen genes were highly significant by one method and confirmed with Bonferroni-corrected significance by the other to show a differential burden of low-frequency and rare variants in restless legs syndrome. Nine of them (AAGAB, ATP2C1, CNTN4, COL6A6, CRBN, GLO1, NTNG1, STEAP4, VAV3) resided in the vicinity of known restless legs syndrome loci, whereas 5 (BBS7, CADM1, CREB5, NRG3, SUN1) have not previously been associated with restless legs syndrome. Burden test and binomial performance deviation analysis also converged significantly in fine-mapping potentially causative domains within these genes. INTERPRETATION: Differential burden with intragenic low-frequency variants reveals putatively causative genes in restless legs syndrome. ANN NEUROL 2020;87:184-193.

Identification of novel risk loci for restless legs syndrome in genome-wide association studies in individuals of European ancestry: a meta-analysis.

BACKGROUND: Restless legs syndrome is a prevalent chronic neurological disorder with potentially severe mental and physical health consequences. Clearer understanding of the underlying pathophysiology is needed to improve treatment options. We did a meta-analysis of genome-wide association studies (GWASs) to identify potential molecular targets. METHODS: In the discovery stage, we combined three GWAS datasets (EU-RLS GENE, INTERVAL, and 23andMe) with diagnosis data collected from 2003 to 2017, in face-to-face interviews or via questionnaires, and involving 15 126 cases and 95 725 controls of European ancestry. We identified common variants by fixed-effect inverse-variance meta-analysis. Significant genome-wide signals (p≤5 × 10-8) were tested for replication in an independent GWAS of 30 770 cases and 286 913 controls, followed by a joint analysis of the discovery and replication stages. We did gene annotation, pathway, and gene-set-enrichment analyses and studied the genetic correlations between restless legs syndrome and traits of interest. FINDINGS: We identified and replicated 13 new risk loci for restless legs syndrome and confirmed the previously identified six risk loci. MEIS1 was confirmed as the strongest genetic risk factor for restless legs syndrome (odds ratio 1·92, 95% CI 1·85-1·99). Gene prioritisation, enrichment, and genetic correlation analyses showed that identified pathways were related to neurodevelopment and highlighted genes linked to axon guidance (associated with SEMA6D), synapse formation (NTNG1), and neuronal specification (HOXB cluster family and MYT1). INTERPRETATION: Identification of new candidate genes and associated pathways will inform future functional research. Advances in understanding of the molecular mechanisms that underlie restless legs syndrome could lead to new treatment options. We focused on common variants; thus, additional studies are needed to dissect the roles of rare and structural variations. FUNDING: Deutsche Forschungsgemeinschaft, Helmholtz Zentrum München-Deutsches Forschungszentrum für Gesundheit und Umwelt, National Research Institutions, NHS Blood and Transplant, National Institute for Health Research, British Heart Foundation, European Commission, European Research Council, National Institutes of Health, National Institute of Neurological Disorders and Stroke, NIH Research Cambridge Biomedical Research Centre, and UK Medical Research Council.

Central and peripheral nervous system excitability in restless legs syndrome.

Neurophysiological techniques have been applied in restless legs syndrome (RLS) to obtain direct and indirect measures of central and peripheral nervous system excitability, as well as to probe different neurotransmission pathways. Data converge on the hypothesis that, from a pure electrophysiological perspective, RLS should be regarded as a complex sensorimotor disorder in which cortical, subcortical, spinal cord, and peripheral nerve generators are all involved in a network disorder, resulting in an enhanced excitability and/or decreased inhibition. Although the spinal component may have dominated in neurophysiological assessment, possibly because of better accessibility compared to the brainstem or cerebral components of a hypothetical dysfunction of the diencephalic A11 area, multiple mechanisms, such as reduced central inhibition and abnormal peripheral nerve function, contribute to the pathogenesis of RLS similarly to some chronic pain conditions. Dopamine transmission dysfunction, either primary or triggered by low iron and ferritin concentrations, may also bridge the gap between RLS and chronic pain entities. Further support of disturbed central and peripheral excitability in RLS is provided by the effectiveness of nonpharmacological tools, such as repetitive transcranial magnetic stimulation and transcutaneous spinal direct current stimulation, in transiently modulating neural excitability, thereby extending the therapeutic repertoire. Understanding the complex interaction of central and peripheral neuronal circuits in generating the symptoms of RLS is mandatory for a better refinement of its therapeutic support.

Targeted resequencing and systematic in vivo functional testing identifies rare variants in MEIS1 as significant contributors to restless legs syndrome.

Restless legs syndrome (RLS) is a common neurologic condition characterized by nocturnal dysesthesias and an urge to move, affecting the legs. RLS is a complex trait, for which genome-wide association studies (GWASs) have identified common susceptibility alleles of modest (OR 1.2-1.7) risk at six genomic loci. Among these, variants in MEIS1 have emerged as the largest risk factors for RLS, suggesting that perturbations in this transcription factor might be causally related to RLS susceptibility. To establish this causality, direction of effect, and total genetic burden of MEIS1, we interrogated 188 case subjects and 182 control subjects for rare alleles not captured by previous GWASs, followed by genotyping of ∼3,000 case subjects and 3,000 control subjects, and concluded with systematic functionalization of all discovered variants using a previously established in vivo model of neurogenesis. We observed a significant excess of rare MEIS1 variants in individuals with RLS. Subsequent assessment of all nonsynonymous variants by in vivo complementation revealed an excess of loss-of-function alleles in individuals with RLS. Strikingly, these alleles compromised the function of the canonical MEIS1 splice isoform but were irrelevant to an isoform known to utilize an alternative 3' sequence. Our data link MEIS1 loss of function to the etiopathology of RLS, highlight how combined sequencing and systematic functional annotation of rare variation at GWAS loci can detect risk burden, and offer a plausible explanation for the specificity of phenotypic expressivity of loss-of-function alleles at a locus broadly necessary for neurogenesis and neurodevelopment.

Blood cis-eQTL analysis fails to identify novel association signals among sub-threshold candidates from genome-wide association studies in restless legs syndrome.

Restless legs syndrome (RLS) is a common neurologic disorder characterized by nightly dysesthesias affecting the legs primarily during periods of rest and relieved by movement. RLS is a complex genetic disease and susceptibility factors in six genomic regions have been identified by means of genome-wide association studies (GWAS). For some complex genetic traits, expression quantitative trait loci (eQTLs) are enriched among trait-associated single nucleotide polymorphisms (SNPs). With the aim of identifying new genetic susceptibility factors for RLS, we assessed the 332 best-associated SNPs from the genome-wide phase of the to date largest RLS GWAS for cis-eQTL effects in peripheral blood from individuals of European descent. In 740 individuals belonging to the KORA general population cohort, 52 cis-eQTLs with pnominal<10(-3) were identified, while in 976 individuals belonging to the SHIP-TREND general population study 53 cis-eQTLs with pnominal<10(-3) were present. 23 of these cis-eQTLs overlapped between the two cohorts. Subsequently, the twelve of the 23 cis-eQTL SNPs, which were not located at an already published RLS-associated locus, were tested for association in 2449 RLS cases and 1462 controls. The top SNP, located in the DET1 gene, was nominally significant (p<0.05) but did not withstand correction for multiple testing (p = 0.42). Although a similar approach has been used successfully with regard to other complex diseases, we were unable to identify new genetic susceptibility factor for RLS by adding this novel level of functional assessment to RLS GWAS data.

Effectiveness and tolerability of rotigotine transdermal patch for the treatment of restless legs syndrome in a routine clinical practice setting in Germany.

OBJECTIVE: We aimed to assess effectiveness and tolerability of rotigotine in patients with moderate to severe idiopathic restless legs syndrome (RLS) under daily practice conditions in Germany. METHODS: In this 3-month noninterventional study, effectiveness was assessed using RLS-6 (primary variables were symptom severity when falling asleep [item 2] and during the night [item 3]). Data were collected at baseline and at the end of treatment. Safety assessments included adverse events (AEs). RESULTS: Six hundred and eighty-four patients were treated with rotigotine and 418 (61%) completed the study. The full analysis set (FAS) comprised 564 patients (106 de novo; 458 pretreated [454 had complete rotigotine dosing data]). Mean rotigotine dose of longest duration was 2.4±1.4 mg/24 h. Rotigotine improved all RLS-6 items (mean change from baseline [item 2], -2.4±3.6; [item 3], -2.7±3.4), with the most pronounced improvement observed in daytime symptoms while at rest (item 4, -2.9±3.2). AEs were typical of dopaminergic treatment and transdermal administration. De novo patients generally started rotigotine on 1 mg/24 h (85% [90/106]) and pretreated patients on 1 (50% [227/454]) or 2 mg/24 h (40% [183/454]). Most patients who were pretreated with levodopa (57%), pramipexole (84%), or ropinirole (78%) monotherapy discontinued these medications on initiation of rotigotine. CONCLUSIONS: Rotigotine was effective and well-tolerated when used in routine clinical practice.

Effects of transcranial direct current stimulation of the primary sensory cortex on somatosensory perception.

BACKGROUND: Transcranial direct current stimulation (tDCS) is able to modify cortical excitability and activity in humans. OBJECTIVE: The aim of the present study was to analyze the effects of tDCS of the primary sensory cortex (SI) on thermal and mechanical perception, assessed by quantitative sensory testing (QST). METHODS: The comprehensive QST protocol encompassing thermal and mechanical detection and pain thresholds as devised by the German Research Network on Neuropathic Pain (DFNS) was applied to skin areas innervated by the radial and median nerve of 12 healthy subjects, who were examined before and after each tDCS stimulation type. Anodal, cathodal, and sham tDCS was applied at a 1 mA current intensity with the active electrode placed over the left primary sensory cortex (SI) and the reference electrode above the right orbit for 15 minutes. RESULTS: After cathodal tDCS cold detection threshold (CDT) significantly increased in the contralateral (P < .01) and ipsilateral hand (P < .05) as compared to baseline condition and sham stimulation, after cathodal stimulation significantly increased warm detection threshold (WDT) was observed in the contralateral hand when compared with the baseline condition (P < .05) but not with sham stimulation. Thermal pain as well as mechanical detection and pain thresholds remained unaltered. CONCLUSIONS: Cathodal tDCS of the primary sensory cortex significantly reduced the sensitivity to Aδ-fiber-mediated cold sensation, C-fiber-mediated warm sensation was reduced only compared with baseline, whereas Aß-fiber-mediated somatosensory inputs were less affected. Our results correspond with our previous observations of primary motor cortex tDCS effects on QST parameters.

Genome-wide association study identifies novel restless legs syndrome susceptibility loci on 2p14 and 16q12.1.

Restless legs syndrome (RLS) is a sensorimotor disorder with an age-dependent prevalence of up to 10% in the general population above 65 years of age. Affected individuals suffer from uncomfortable sensations and an urge to move in the lower limbs that occurs mainly in resting situations during the evening or at night. Moving the legs or walking leads to an improvement of symptoms. Concomitantly, patients report sleep disturbances with consequences such as reduced daytime functioning. We conducted a genome-wide association study (GWA) for RLS in 922 cases and 1,526 controls (using 301,406 SNPs) followed by a replication of 76 candidate SNPs in 3,935 cases and 5,754 controls, all of European ancestry. Herein, we identified six RLS susceptibility loci of genome-wide significance, two of them novel: an intergenic region on chromosome 2p14 (rs6747972, P = 9.03 × 10(-11), OR = 1.23) and a locus on 16q12.1 (rs3104767, P = 9.4 × 10(-19), OR = 1.35) in a linkage disequilibrium block of 140 kb containing the 5'-end of TOX3 and the adjacent non-coding RNA BC034767.

Individual voxel-based subtype prediction can differentiate progressive supranuclear palsy from idiopathic Parkinson syndrome and healthy controls.

Voxel-based morphometry (VBM) shows a differentiated pattern in patients with atypical Parkinson syndrome but so far has had little impact in individual cases. It is desirable to translate VBM findings into clinical practice and individual classification. To this end, we examined whether a support vector machine (SVM) can provide useful accuracies for the differential diagnosis. We acquired a volumetric 3D T1-weighted MRI of 21 patients with idiopathic Parkinson syndrome (IPS), 11 multiple systems atrophy (MSA-P) and 10 progressive supranuclear palsy (PSP), and 22 healthy controls. Images were segmented, normalized, and compared at group level with SPM8 in a classical VBM design. Next, a SVM analysis was performed on an individual basis with leave-one-out cross-validation. VBM showed a strong white matter loss in the mesencephalon of patients with PSP, a putaminal grey matter loss in MSA, and a cerebellar grey matter loss in patients with PSP compared with IPS. The SVM allowed for an individual classification in PSP versus IPS with up to 96.8% accuracy with 90% sensitivity and 100% specificity. In MSA versus IPS, an accuracy of 71.9% was achieved; sensitivity, however, was low with 36.4%. Patients with IPS could not be differentiated from controls. In summary, a voxel-based SVM analysis allows for a reliable classification of individual cases in PSP that can be directly clinically useful. For patients with MSA and IPS, further developments like quantitative MRI are needed.

Transcranial direct current stimulation of the motor cortex induces distinct changes in thermal and mechanical sensory percepts.

OBJECTIVE: The aim of this single-blinded, complete crossover study was to evaluate the effects of tDCS on thermal and mechanical perception, as assessed by quantitative sensory testing (QST). METHODS: QST was performed upon the radial part of both hands of eight healthy subjects (3 female, 5 male, 25-41years of age). These subjects were examined before and after cathodal, anodal or sham tDCS, applied in a random order. TDCS was administered for 15min at a 1mA current intensity, with the active electrode placed over the left primary motor cortex and the reference electrode above the right orbit. RESULTS: After cathodal tDCS, cold detection thresholds (CDT), mechanical detection thresholds (MDT), and mechanical pain thresholds (MPT) significantly increased in the contralateral hand, when compared to the baseline condition. CONCLUSIONS: Cathodal tDCS temporarily reduced the sensitivity to A-fiber mediated somatosensory inputs. SIGNIFICANCE: Impairment of these somatosensory percepts suggests a short-term suppression of lemniscal or suprathalamic sensory pathways following motor cortex stimulation by cathodal tDCS.

Thermal hypoaesthesia differentiates secondary restless legs syndrome associated with small fibre neuropathy from primary restless legs syndrome.

This study aimed to assess thermal and mechanical perception and pain thresholds in primary idiopathic restless legs syndrome and secondary restless legs syndrome associated with small fibre neuropathy. Twenty-one patients (age: 53.4 + or - 8.4, n = 3, male) with primary restless legs syndrome and 13 patients (age: 63.0 + or - 8.2, n = 1, male) with secondary restless legs syndrome associated with small fibre neuropathy were compared with 20 healthy subjects (age: 58.0 + or - 7.0; n = 2, male). Differential diagnosis of secondary restless legs syndrome associated with small fibre neuropathy was based on clinical symptoms and confirmed with skin biopsies in all patients. A comprehensive quantitative sensory testing protocol encompassing thermal and mechanical detection and pain thresholds, as devised by the German Research Network on Neuropathic Pain, was performed on the clinically more affected foot between 2 pm and 1 am when restless legs syndrome symptoms were present in all patients. Patients with primary restless legs syndrome showed hyperalgesia to blunt pressure (P < 0.001), pinprick (P < 0.001) and vibratory hyperaesthesia (P < 0.001). Patients with secondary restless legs syndrome associated with small fibre neuropathy showed thermal hypoaesthesia to cold (Adelta-fibre mediated) and warm (C-fibre mediated) (all P < 0.001) and hyperalgesia to pinprick (P < 0.001). Static mechanical hyperalgesia in primary and secondary restless legs syndrome is consistent with the concept of central disinhibition of nociceptive pathways, which might be induced by conditioning afferent input from damaged small fibre neurons in secondary restless legs syndrome.

Modulatory effects of transcranial direct current stimulation on laser-evoked potentials.

OBJECTIVE: Invasive stimulation of the motor cortex has been used for years to alleviate chronic intractable pain in humans. In our study, we have investigated the effect of transcranial direct current stimulation (tDCS), a noninvasive stimulation method, for manipulating the excitability of cortical motor areas on laser evoked potentials (LEP) and acute pain perception. DESIGNS AND SETTINGS: The amplitude of the N1, N2, and P2 LEP components of 10 healthy volunteers were evaluated prior to and following anodal, cathodal, and sham stimulation of the primary motor cortex. In a separate experiment subjective, pain rating scores of 16 healthy subjects in two perceptual categories (warm sensation, mild pain) were also analyzed. RESULTS: Cathodal tDCS significantly reduced the amplitude of N2 and P2 components compared with anodal or sham stimulation. However, neither of the tDCS types modified significantly the laser energy values necessary to induce moderate pain. In a separate experiment, cathodal stimulation significantly diminished mild pain sensation only when laser-stimulating the hand contralateral to the side of tDCS, while anodal stimulation modified warm sensation. CONCLUSIONS: The possible underlying mechanisms of our findings in view of recent neuroimaging studies are discussed. To our knowledge this study is the first to demonstrate the mild antinociceptive effect of tDCS over the primary motor cortex in healthy volunteers.

Effect of thermode application pressure on thermal threshold detection.

Studies using quantitative sensory testing (QST) often present incongruent results due to intra- and intersubject as well as interobserver variability which limit widespread use of the technique. Eliminating or reducing the factors responsible for this variability is of great interest, as it increases reliability and reproducibility of QST. Thermal sensory threshold determination is a crucial part of QST. It was previously suggested that the pressure of the thermode on the skin could influence measurements. To verify this, we developed a new thermode with a built-in pressure sensor. Thresholds obtained with this thermode were compared to those obtained with a commercially available thermotesting device (Medoc TSA-II). Heat detection and heat pain detection thresholds were higher, and cold detection thresholds were lower when measured with our thermode than they were with the Medoc thermode. Cold pain detection thresholds did not differ between the thermodes. Analysis of the heat transfer capacity of the thermodes indicated that the material of the skin contact surface of the thermode may play a role in these shifts in threshold values. Altering the thermode pressure on the skin did not affect the thermal thresholds. Furthermore, the intrasubject variability of the measurements (minimal-to-maximal range of measured threshold values in individual subjects) was also not influenced by the pressure with which the thermode was attached to the skin. Our results suggest that the pressure with which the thermode is attached to the skin does not significantly affect the intra- and intersubject reproducibility of the thermal sensory threshold measurements.

PTPRD (protein tyrosine phosphatase receptor type delta) is associated with restless legs syndrome.

We identified association of restless legs syndrome (RLS) with PTPRD at 9p23-24 in 2,458 affected individuals and 4,749 controls from Germany, Austria, Czechia and Canada. Two independent SNPs in the 5' UTR of splice variants expressed predominantly in the central nervous system showed highly significant P values (rs4626664, P(nominal/lambda corrected) = 5.91 x 10(-10), odds ratio (OR) = 1.44; rs1975197, P(nominal/lambda corrected) = 5.81 x 10(-9), OR = 1.31). This work identifies PTPRD as the fourth genome-wide significant locus for RLS.

Homocysteine in restless legs syndrome.

BACKGROUND AND PURPOSE: Total plasma homocysteine (tHcy) may be a risk factor for vascular diseases and is associated with renal failure or deficiency of vitamin B12 or folate. Recently, elevated tHcy concentrations were observed in patients with Parkinson's disease (PD), particularly those under levodopa treatment. Our objective was to determine whether changes in tHcy are also found in patients with restless legs syndrome (RLS) in relation to levodopa treatment and whether folate and vitamins B6 and B12 play a role in RLS. METHODS: In a total of 228 subjects, tHcy and B vitamin status (vitamins B6 and B12, folate) were studied: 97 patients with idiopathic RLS (40 under levodopa therapy), 39 with PD (25 under levodopa therapy), and 92 healthy controls adjusted for age and gender. RESULTS: No significant differences were observed in tHcy levels between RLS patients and controls or between the RLS groups without treatment or with levodopa or dopamine agonist treatment. Mean tHcy was significantly higher in PD patients (13.8 micromol/l) than in either RLS patients (11.7 micromol/l) or controls (11.0 micromol/l; p<0.001). There was an inverse association between tHcy and vitamin B12 in each group. CONCLUSIONS: RLS and, in particular, levodopa treatment in RLS are not associated with hyperhomocysteinemia. Elevated tHcy could, however, be confirmed in PD patients.

Genome-wide association study of restless legs syndrome identifies common variants in three genomic regions.

Restless legs syndrome (RLS) is a frequent neurological disorder characterized by an imperative urge to move the legs during night, unpleasant sensation in the lower limbs, disturbed sleep and increased cardiovascular morbidity. In a genome-wide association study we found highly significant associations between RLS and intronic variants in the homeobox gene MEIS1, the BTBD9 gene encoding a BTB(POZ) domain as well as variants in a third locus containing the genes encoding mitogen-activated protein kinase MAP2K5 and the transcription factor LBXCOR1 on chromosomes 2p, 6p and 15q, respectively. Two independent replications confirmed these association signals. Each genetic variant was associated with a more than 50% increase in risk for RLS, with the combined allelic variants conferring more than half of the risk. MEIS1 has been implicated in limb development, raising the possibility that RLS has components of a developmental disorder.