Heterozygous Variants in KCNJ10 Cause Paroxysmal Kinesigenic Dyskinesia Via Haploinsufficiency.

OBJECTIVE: Most paroxysmal kinesigenic dyskinesia (PKD) cases are hereditary, yet approximately 60% of patients remain genetically undiagnosed. We undertook the present study to uncover the genetic basis for undiagnosed PKD patients. METHODS: Whole-exome sequencing was performed for 106 PRRT2-negative PKD probands. The functional impact of the genetic variants was investigated in HEK293T cells and Drosophila. RESULTS: Heterozygous variants in KCNJ10 were identified in 11 individuals from 8 unrelated families, which accounted for 7.5% (8/106) of the PRRT2-negative probands. Both co-segregation of the identified variants and the significantly higher frequency of rare KCNJ10 variants in PKD cases supported impacts from the detected KCNJ10 heterozygous variants on PKD pathogenesis. Moreover, a KCNJ10 mutation-carrying father from a typical EAST/SeSAME family was identified as a PKD patient. All patients manifested dystonia attacks triggered by sudden movement with a short episodic duration. Patch-clamp recordings in HEK293T cells revealed apparent reductions in K+ currents of the patient-derived variants, indicating a loss-of-function. In Drosophila, milder hyperexcitability phenotypes were observed in heterozygous Irk2 knock-in flies compared to homozygotes, supporting haploinsufficiency as the mechanism for the detected heterozygous variants. Electrophysiological recordings showed that excitatory neurons in Irk2 haploinsufficiency flies exhibited increased excitability, and glia-specific complementation with human Kir4.1 rescued the Irk2 mutant phenotypes. INTERPRETATION: Our study established haploinsufficiency resulting from heterozygous variants in KCNJ10 can be understood as a previously unrecognized genetic cause for PKD and provided evidence of glial involvement in the pathophysiology of PKD. ANN NEUROL 2024.

Adenine base editing-mediated exon skipping restores dystrophin in humanized Duchenne mouse model.

Duchenne muscular dystrophy (DMD) affecting 1 in 3500-5000 live male newborns is the frequently fatal genetic disease resulted from various mutations in DMD gene encoding dystrophin protein. About 70% of DMD-causing mutations are exon deletion leading to frameshift of open reading frame and dystrophin deficiency. To facilitate translating human DMD-targeting CRISPR therapeutics into patients, we herein establish a genetically humanized mouse model of DMD by replacing exon 50 and 51 of mouse Dmd gene with human exon 50 sequence. This humanized mouse model recapitulats patient's DMD phenotypes of dystrophin deficiency and muscle dysfunction. Furthermore, we target splicing sites in human exon 50 with adenine base editor to induce exon skipping and robustly restored dystrophin expression in heart, tibialis anterior and diaphragm muscles. Importantly, systemic delivery of base editor via adeno-associated virus in the humanized male mouse model improves the muscle function of DMD mice to the similar level of wildtype ones, indicating the therapeutic efficacy of base editing strategy in treating most of DMD types with exon deletion or point mutations via exon-skipping induction.

Disease Progression and Multiparametric Imaging Characteristics of Spinocerebellar Ataxia Type 3 With Spastic Paraplegia.

Background and Objectives: Spinocerebellar ataxia type 3 (SCA3) is a hereditary ataxia that occurs worldwide. Clinical patterns were observed, including the one characterized by marked spastic paraplegia. This study investigated the clinical features, disease progression, and multiparametric imaging aspects of patients with SCA3. Methods: We retrospectively analyzed 249 patients with SCA3 recruited from the Organization for Southeast China for cerebellar ataxia research between October 2014 and December 2020. Of the 249 patients, 145 were selected and assigned to 2 groups based on neurologic examination: SCA3 patients with spastic paraplegia (SCA3-SP) and SCA3 patients with nonspastic paraplegia (SCA3-NSP). Participants underwent 3.0-T brain MRI examinations, and voxel-wise and volume-of-interest-based approaches were used for the resulting images. A tract-based spatial statistical approach was used to investigate the white matter (WM) alterations using diffusion tensor imaging, neurite orientation dispersion, and density imaging metrics. Multiple linear regression analyses were performed to compare the clinical and imaging parameters between the 2 groups. The longitudinal data were evaluated using a linear mixed-effects model. Results: Forty-three patients with SCA3-SP (mean age, 37.58years ± 11.72 [SD]; 18 women) and 102 patients with SCA3-NSP (mean age, 47.42years ± 12.50 [SD]; 39 women) were analyzed. Patients with SCA3-SP were younger and had a lower onset age but a larger cytosine-adenine-guanine repeat number, as well as higher clinical severity scores (all corrected p < 0.05). The estimated progression rates of the Scale for the Assessment and Rating of Ataxia (SARA) and International Cooperative Ataxia Rating Scale scores were higher in the SCA3-SP subgroup than in the SCA3-NSP subgroup (SARA, 2.136 vs 1.218 points; ICARS, 5.576 vs 3.480 points; both p < 0.001). In addition, patients with SCA3-SP showed gray matter volume loss in the precentral gyrus with a decreased neurite density index in the WM of the corticospinal tract and cerebellar peduncles compared with patients with SCA3-NSP. Discussion: SCA3-SP differs from SCA3-NSP in clinical features, multiparametric brain imaging findings, and longitudinal follow-up progression.

Characteristics of tandem repeat inheritance and sympathetic nerve involvement in GAA-FGF14 ataxia.

BACKGROUND: Intronic GAA repeat expansion ([GAA] ≥250) in FGF14 is associated with the late-onset neurodegenerative disorder, spinocerebellar ataxia 27B (SCA27B, GAA-FGF14 ataxia). We aim to determine the prevalence of the GAA repeat expansion in FGF14 in Chinese populations presenting late-onset cerebellar ataxia (LOCA) and evaluate the characteristics of tandem repeat inheritance, radiological features and sympathetic nerve involvement. METHODS: GAA-FGF14 repeat expansion was screened in an undiagnosed LOCA cohort (n = 664) and variations in repeat-length were analyzed in families of confirmed GAA-FGF14 ataxia patients. Brain magnetic resonance imaging (MRI) was used to evaluate the radiological feature in GAA-FGF14 ataxia patients. Clinical examinations and sympathetic skin response (SSR) recordings in GAA-FGF14 patients (n = 16) were used to quantify sympathetic nerve involvement. RESULTS: Two unrelated probands (2/664) were identified. Genetic screening for GAA-FGF14 repeat expansion was performed in 39 family members, 16 of whom were genetically diagnosed with GAA-FGF14 ataxia. Familial screening revealed expansion of GAA repeats in maternal transmissions, but contraction upon paternal transmission. Brain MRI showed slight to moderate cerebellar atrophy. SSR amplitude was lower in GAA-FGF14 patients in pre-symptomatic stage compared to healthy controls, and further decreased in the symptomatic stage. CONCLUSIONS: GAA-FGF14 ataxia was rare among Chinese LOCA cases. Parental gender appears to affect variability in GAA repeat number between generations. Reduced SSR amplitude is a prominent feature in GAA-FGF14 patients, even in the pre-symptomatic stage.

Nomogram and calculator for predicting the prognosis of patients with giant hepatocellular carcinoma.

OBJECTIVES: This study aimed to explore the factors affecting the overall survival (OS) of giant hepatocellular carcinoma (G-HCC) patients and establish a nomogram and an Internet-based OS calculator for evaluating the OS of G-HCC patients. RESEARCH DESIGN AND METHODS: A total of 2445 G-HCC patients were searched in the SEER database. The independent variables affecting OS of G-HCC patients were determined by univariate and multivariate analyses, and a nomogram and Internet-based OS calculator were established. The accuracy of the nomogram was evaluated by the C-index, the AUC curve, and calibration curve. RESULTS: Grade, surgery, radiotherapy, chemotherapy, T-staging, M-staging, AFP, and fibrosis were identified as independent variables affecting OS. These variables were included in the nomogram model and Internet-based OS calculator to evaluate OS in G-HCC patients. The C-indices and AUC of the nomogram are better than AJCC-staging system. Similarly, the calibration curves revealed that the actual survival was consistent with nomogram-based survival. CONCLUSION: The nomogram and Internet-based OS calculator are superior to the traditional AJCC-staging system in the reliability and convenience of prognosis assessment for G-HCC patients, which is more conducive for clinicians to predict the survival of G-HCC patients and make the best treatment strategy.

Expression and Purification of Mammalian NMDA Receptor Protein for Functional Characterization.

N-methyl-D-aspartate (NMDA) receptors are critical for brain function and serve as drug targets for the treatment of neurological and psychiatric disorders. They typically form the tetrameric assembly of GluN1-GluN2 (2A to 2D) subtypes, with their diverse three-dimensional conformations linked with the physiologically relevant function in vivo. Purified proteins of tetrameric assembled NMDA receptors have broad applications in the structural elucidation, hybridoma technology for antibody production, and high-throughput drug screening. However, obtaining sufficient quantity and monodisperse NMDA receptor protein is still technically challenging. Here, we summarize a paradigm for the expression and purification of diverse NMDA receptor subtypes, with detailed descriptions on screening constructs by fluorescence size-exclusion chromatography (FSEC), generation of recombinant baculovirus, expression in the eukaryotic expression system, protein purification by affinity chromatography and size-exclusion chromatography (SEC), biochemical and functional validation assays.

A Post-Marketing Surveillance Study of Nusinersen for Spinal Muscular Atrophy in Routine Medical Practice in China: Interim Results.

INTRODUCTION: Spinal muscular atrophy (SMA) is a rare, autosomal recessive, neuromuscular disease that leads to progressive muscular weakness and atrophy. Nusinersen, an antisense oligonucleotide, was approved for SMA in China in February 2019. We report interim results from a post-marketing surveillance phase 4 study, PANDA (NCT04419233), that collects data on the safety, efficacy, and pharmacokinetics of nusinersen in children with SMA in routine clinical practice in China. METHODS: Participants enrolled in PANDA will be observed for 2 years following nusinersen treatment initiation. The primary endpoint is the incidence of adverse events (AEs)/serious AEs (SAEs) during the treatment period. Efficacy assessments include World Health Organization (WHO) Motor Milestones assessment, the Hammersmith Infant Neurological Examination (HINE), and ventilation support. Plasma and cerebrospinal fluid (CSF) concentrations of nusinersen are measured at each dose visit. RESULTS: Fifty participants were enrolled as of the January 4, 2023, data cutoff: 10 with infantile-onset (≤ 6 months) and 40 with later-onset (> 6 months) SMA. All 50 participants have received at least one dose of nusinersen; 6 have completed the study. AEs were experienced by 45 (90%) participants and were mostly mild/moderate; no AEs led to nusinersen discontinuation or study withdrawal. Eleven participants experienced SAEs, most commonly pneumonia (n = 9); none were considered related to study treatment. Stability or gain of WHO motor milestone was observed and mean HINE-2 scores improved in both subgroups throughout the study. No serious respiratory events occurred, and no permanent ventilation support was initiated during the study. Pre-dose nusinersen CSF concentrations increased steadily through the loading-dose period, with no accumulation in plasma after multiple doses. CONCLUSION: Nusinersen was generally well tolerated with an acceptable overall safety profile, consistent with the known safety of nusinersen. Efficacy, safety, and nusinersen exposure are consistent with prior observations. These results support continuing PANDA and evaluation of nusinersen in Chinese participants with SMA. TRIAL REGISTRATION: ClinicalTrials.gov identifier, NCT04419233.

Detection of pTDP-43 via routine muscle biopsy: A promising diagnostic biomarker for amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease, pathologically characterized by TDP-43 aggregates. Recent evidence has been indicated that phosphorylated TDP-43 (pTDP-43) is present not only in motor neurons but also in muscle tissues. However, it is unclear whether testing pTDP-43 aggregation in muscle tissue would assist in the diagnosis of ALS. We propose three key questions: (i) Is aggregation of pTDP-43 detectable in routine biopsied muscles? (ii) Can detection of pTDP-43 aggregation discriminate between ALS and non-ALS patients? (iii) Can pTDP-43 aggregation be observed in the early stages of ALS? We conducted a diagnostic study comprising 2 groups: an ALS group in which 18 cases underwent muscle biopsy screened from a registered ALS cohort consisting of 802 patients and a non-ALS control group, in which we randomly selected 54 muscle samples from a biospecimen bank of 684 patients. Among the 18 ALS patients, 3 patients carried pathological GGGGCC repeats in the C9ORF72 gene, 2 patients carried SOD1 mutations, and 7 patients were at an early stage with only one body region clinically affected. The pTDP-43 accumulation could be detected in routine biopsied muscles, including biceps brachii, deltoid, tibialis anterior, and quadriceps. Abnormal aggregation of pTDP-43 was present in 94.4% of ALS patients (17/18) compared to 29.6% of non-ALS controls (16/54; p < 0.001). The pTDP-43 aggregates were mainly close to the sarcolemma. Using a semi-quantified pTDP-43 aggregates score, we applied a cut-off value of 3 as a diagnostic biomarker, resulting in a sensitivity of 94.4% and a specificity of 83.3%. Moreover, we observed that accumulation of pTDP-43 occurred in muscle tissues prior to clinical symptoms and electromyographic lesions. Our study provides proof-of-concept for the detection of pTDP-43 accumulation via routine muscle biopsy which may serve as a novel biomarker for diagnosis of ALS.

Multidimensional features of sporadic Creutzfeldt-Jakob disease in the elderly: a case report and systematic review.

Background: As a rare neurodegenerative disease, sporadic Creutzfeldt-Jakob disease (sCJD) is poorly understood in the elderly populace. This study aims to enunciate the multidimensional features of sCJD in this group. Methods: A case of probable sCJD was reported in a 90-year-old Chinese man with initial dizziness. Then, available English literature of the elderly sCJD cases (aged 80 years and over) was reviewed and analyzed. Patients (15 cases) were subdivided and compared geographically. Results: In the elderly sCJD cohort, the onset age was 84.9 ± 4.5 years and the median disease duration was 6.8 months, with respiratory infection/failure as the commonest death cause. Various clinical symptoms were identified, with cognitive disorder (86.7%) as the commonest typical symptom and speech impairment (66.7%) as the most atypical one. Restricted hyperintensities were reported in 60.0% cases on DWI, periodic sharp wave complexes in 73.3% cases on electroencephalogram, and cerebral hypoperfusion/hypometabolism in 26.7% cases on molecular imaging. The sensitive cerebrospinal fluid biomarkers were total tau (83.3%), 14-3-3 protein (75.0%), and PrP RT-QuIC (75.0%). Neuropathological profiles in the cerebral cortex revealed vacuolar spongiosis, neuronal loss, gliosis, and aging-related markers, with synaptic deposit as the commonest PrP pattern (60.0%). The polymorphic PRNP analysis at codon 129 was M/M (90.9%), with MM1 and MM2C as the primary molecular phenotypes. Latency to first clinic visit, hyperintense signals on DWI, and disease duration were significantly different between the patient subgroups. Conclusion: The characteristics of sCJD are multidimensional in the elderly, deepening our understanding of the disease and facilitating an earlier recognition and better care for this group.

Correction of human nonsense mutation via adenine base editing for Duchenne muscular dystrophy treatment in mouse.

Duchenne muscular dystrophy (DMD) is the most prevalent herediatry disease in men, characterized by dystrophin deficiency, progressive muscle wasting, cardiac insufficiency, and premature mortality, with no effective therapeutic options. Here, we investigated whether adenine base editing can correct pathological nonsense point mutations leading to premature stop codons in the dystrophin gene. We identified 27 causative nonsense mutations in our DMD patient cohort. Treatment with adenine base editor (ABE) could restore dystrophin expression by direct A-to-G editing of pathological nonsense mutations in cardiomyocytes generated from DMD patient-derived induced pluripotent stem cells. We also generated two humanized mouse models of DMD expressing mutation-bearing exons 23 or 30 of human dystrophin gene. Intramuscular administration of ABE, driven by ubiquitous or muscle-specific promoters could correct these nonsense mutations in vivo, albeit with higher efficiency in exon 30, restoring dystrophin expression in skeletal fibers of humanized DMD mice. Moreover, a single systemic delivery of ABE with human single guide RNA (sgRNA) could induce body-wide dystrophin expression and improve muscle function in rotarod tests of humanized DMD mice. These findings demonstrate that ABE with human sgRNAs can confer therapeutic alleviation of DMD in mice, providing a basis for development of adenine base editing therapies in monogenic diseases.

Diagnosis of Challenging Spinal Muscular Atrophy Cases with Long-Read Sequencing.

Spinal muscular atrophy (SMA) is an autosomal recessive neuromuscular disorder primarily caused by the deletion or mutation of the survival motor neuron 1 (SMN1) gene. This study assesses the diagnostic potential of long-read sequencing (LRS) in three patients with SMA. For Patient 1, who has a heterozygous SMN1 deletion, LRS unveiled a missense mutation in SMN1 exon 5. In Patient 2, an Alu/Alu-mediated rearrangement covering the SMN1 promoter and exon 1 was identified through a blend of multiplex ligation-dependent probe amplification, LRS, and PCR across the breakpoint. The third patient, born to a consanguineous family, bore four copies of hybrid SMN genes. LRS determined the genomic structures, indicating two distinct hybrids of SMN2 exon 7 and SMN1 exon 8. However, a discrepancy was found between the SMN1/SMN2 ratio interpretations by LRS (0:2) and multiplex ligation-dependent probe amplification (0:4), which suggested a limitation of LRS in SMA diagnosis. In conclusion, this newly adapted long PCR-based third-generation sequencing introduces an additional avenue for SMA diagnosis.

ESCRT-I protein UBAP1 controls ventricular expansion and cortical neurogenesis via modulating adherens junctions of radial glial cells.

Intricate cerebral cortex formation is orchestrated by the precise behavior and division dynamics of radial glial cells (RGCs). Endocytosis functions in the recycling and remodeling of adherens junctions (AJs) in response to changes in RGC activity and function. Here, we show that conditional disruption of ubiquitin-associated protein 1 (UBAP1), a component of endosomal sorting complex required for transport (ESCRT), causes severe brain dysplasia and prenatal ventriculomegaly. UBAP1 depletion disrupts the AJs and polarity of RGCs, leading to failure of apically directed interkinetic nuclear migration. Accordingly, UBAP1 knockout or knockdown results in reduced proliferation and precocious differentiation of neural progenitor cells. Mechanistically, UBAP1 regulates the expression and surface localization of cell adhesion molecules, and ß-catenin over-expression significantly rescues the phenotypes of Ubap1 knockdown in vivo. Our study reveals a critical physiological role of the ESCRT machinery in cortical neurogenesis by regulating AJs of RGCs.

Cognitive impairment associated with cerebellar volume loss in spinocerebellar ataxia type 3.

BACKGROUND: Many neuroscience and neurology studies have forced a reconsideration of the traditional motor-related scope of cerebellar function, which has now expanded to include various cognitive functions. Spinocerebellar ataxia type 3 (SCA3; the most common hereditary ataxia) is neuropathologically characterized by cerebellar atrophy and frequently presents with cognitive impairment. OBJECTIVE: To characterize cognitive impairment in SCA3 and investigate the cerebellum-cognition associations. METHODS: This prospective, cross-sectional cohort study recruited 126 SCA3 patients and 41 healthy control individuals (HCs). Participants underwent a brain 3D T1-weighted images as well as neuropsychological tests. Voxel-based morphometry (VBM) and region of interest (ROI) approaches were performed on the 3D T1-weighted images. CERES was used to automatically segment cerebellums. Patients were grouped into cognitively impaired (CI) and cognitively preserved (CP), and clinical and MRI parameters were compared. Multivariable regression models were fitted to examine associations between cerebellar microstructural alterations and cognitive domain impairments. RESULTS: Compared to HCs, SCA3 patients showed cognitive domain impairments in information processing speed, verbal memory, executive function, and visuospatial perception. Between CI and CP subgroups, the CI subgroup was older and had lower education, as well as higher severity scores. VBM and ROI analyses revealed volume loss in cerebellar bilateral lobule VI, right lobule Crus I, and right lobule IV of the CI subgroup, and all these cerebellar lobules were associated with the above cognitive domain impairments. CONCLUSIONS: Our findings demonstrate the multiple cognitive domain impairments in SCA3 patients and indicate the responsible cerebellar lobules for the impaired cognitive domain(s).

Biallelic COQ4 Variants in Hereditary Spastic Paraplegia: Clinical and Molecular Characterization.

BACKGROUND: Hereditary spastic paraplegias (HSP) are neurologic disorders characterized by progressive lower-extremity spasticity. Despite the identification of several HSP-related genes, many patients lack a genetic diagnosis. OBJECTIVES: The aims were to confirm the pathogenic role of biallelic COQ4 mutations in HSP and elucidate the clinical, genetic, and functional molecular features of COQ4-associated HSP. METHODS: Whole exome sequences of 310 index patients with HSP of unknown cause from three distinct populations were analyzed to identify potential HSP causal genes. Clinical data obtained from patients harboring candidate causal mutations were examined. Functional characterization of COQ4 variants was performed using bioinformatic tools, single-cell RNA sequencing, biochemical assays in cell lines, primary fibroblasts, induced pluripotent stem cell-derived pyramidal neurons, and zebrafish. RESULTS: Compound heterozygous variants in COQ4, which cosegregated with HSP in pedigrees, were identified in 7 patients from six unrelated families. Patients from four of the six families presented with pure HSP, whereas probands of the other two families exhibited complicated HSP with epilepsy or with cerebellar ataxia. In patient-derived fibroblasts and COQ4 knockout complementation lines, stable expression of these missense variants exerted loss-of-function effects, including mitochondrial reactive oxygen species accumulation, decreased mitochondrial membrane potential, and lower ubiquinone biosynthesis. Whereas differentiated pyramidal neurons expressed high COQ4 levels, coq4 knockdown zebrafish displayed severe motor dysfunction, reflecting motor neuron dysregulation. CONCLUSIONS: Our study confirms that loss-of-function, compound heterozygous, pathogenic COQ4 variants are causal for autosomal recessive pure and complicated HSP. Moreover, reduced COQ4 levels attributable to variants correspond with decreased ubiquinone biosynthesis, impaired mitochondrial function, and higher phenotypic disease severity. © 2023 International Parkinson and Movement Disorder Society.

CGG repeat expansion in LOC642361/NUTM2B-AS1 typically presents as oculopharyngodistal myopathy.

CGG repeat expansions in LOC642361/NUTM2B-AS1 have recently been identified as a cause of oculopharyngeal myopathy with leukoencephalopathy. However, since only three patients from a single family were reported, it remains unknown whether their clinicopathological features are typical for CGG repeat expansions in LOC642361/NUTM2B-AS1. Here, using repeat-primed-polymerase chain reaction and long-read sequencing, we identify 12 individuals from 3 unrelated families with CGG repeat expansions in LOC642361/NUTM2B-AS1, typically presenting with oculopharyngodistal myopathy. The CGG repeat expansions range from 161 to 669 repeat units. Most of the patients present with ptosis, restricted eye movements, dysphagia, dysarthria, and diffuse limb muscle weakness. Only one patient shows T2-weighted hyperintensity in the cerebellar white matter surrounding the deep cerebellar nuclei on brain magnetic resonance imaging. Muscle biopsies from three patients show a myopathic pattern and rimmed vacuoles. Analyses of muscle biopsies suggest that CGG repeat expansions in LOC642361/NUTM2B-AS1 may deleteriously affect aggrephagic capacity, suggesting that RNA toxicity and mitochondrial dysfunction may contribute to pathogenesis. Our study thus expands the phenotypic spectrum for the CGG repeat expansion of LOC642361/NUTM2B-AS1 and indicates that this genetic variant typically manifests as oculopharyngodistal myopathy with chronic myopathic changes with rimmed vacuoles and filamentous intranuclear inclusions in muscle fibers.

Alu Retrotransposition Event in SPAST Gene as a Novel Cause of Hereditary Spastic Paraplegia.

OBJECTIVES: To diagnose the molecular cause of hereditary spastic paraplegia (HSP) observed in a four-generation family with autosomal dominant inheritance. METHODS: Multiplex ligation-dependent probe amplification (MLPA), whole-exome sequencing (WES), and RNA sequencing (RNA-seq) of peripheral blood leukocytes were performed. Reverse transcription polymerase chain reaction (RT-PCR) and Sanger sequencing were used to characterize target regions of SPAST. RESULTS: A 121-bp AluYb9 insertion with a 30-bp poly-A tail flanked by 15-bp direct repeats on both sides was identified in the edge of intron 16 in SPAST that segregated with the disease phenotype. CONCLUSIONS: We identified an intronic AluYb9 insertion inducing splicing alteration in SPAST causing pure HSP phenotype that was not detected by routine WES analysis. Our findings suggest RNA-seq is a recommended implementation for undiagnosed cases by first-line diagnostic approaches. © 2023 International Parkinson and Movement Disorder Society.