Recessive GNE Mutations in Korean Nonaka Distal Myopathy Patients with or without Peripheral Neuropathy.

Autosomal recessive Nonaka distal myopathy is a rare autosomal recessive genetic disease characterized by progressive degeneration of the distal muscles, causing muscle weakness and decreased grip strength. It is primarily associated with mutations in the GNE gene, which encodes a key enzyme of sialic acid biosynthesis (UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase). This study was performed to find GNE mutations in six independent distal myopathy patients with or without peripheral neuropathy using whole-exome sequencing (WES). In silico pathogenic prediction and simulation of 3D structural changes were performed for the mutant GNE proteins. As a result, we identified five pathogenic or likely pathogenic missense variants: c.86T>C (p.Met29Thr), c.527A>T (p.Asp176Val), c.782T>C (p.Met261Thr), c.1714G>C (p.Val572Leu), and c.1771G>A (p.Ala591Thr). Five affected individuals showed compound heterozygous mutations, while only one patient revealed a homozygous mutation. Two patients revealed unreported combinations of combined heterozygous mutations. We observed some specific clinical features, such as complex phenotypes of distal myopathy with distal hereditary peripheral neuropathy, an earlier onset of weakness in legs than that of hands, and clinical heterogeneity between two patients with the same set of compound heterozygous mutations. Our findings on these genetic causes expand the clinical spectrum associated with the GNE mutations and can help prepare therapeutic strategies.

Peripheral Neuropathy and Decreased Locomotion of a RAB40B Mutation in Human and Model Animals.

Rab40 proteins are an atypical subgroup of Rab GTPases containing a unique suppressor of the cytokine signaling (SOCS) domain that is recruited to assemble the CRL5 E3 ligase complex for proteolytic regulation in various biological processes. A nonsense mutation deleting the C-terminal SOCS box in the RAB40B gene was identified in a family with axonal peripheral neuropathy (Charcot-Marie-Tooth disease type 2), and pathogenicity of the mutation was assessed in model organisms of zebrafish and Drosophila. Compared to control fish, zebrafish larvae transformed by the human mutant hRAB40B-Y83X showed a defective swimming pattern of stalling with restricted localization and slower motility. We were consistently able to observe reduced labeling of synaptic markers along neuromuscular junctions of the transformed larvae. In addition to the neurodevelopmental phenotypes, compared to normal hRAB40B expression, we further examined ectopic expression of hRAB40B-Y83X in Drosophila to show a progressive decline of locomotion ability. Decreased ability of locomotion by ubiquitous expression of the human mutation was reproduced not with GAL4 drivers for neuron-specific expression but only when a pan-glial GAL4 driver was applied. Using the ectopic expression model of Drosophila, we identified a genetic interaction in which Cul5 down regulation exacerbated the defective motor performance, showing a consistent loss of SOCS box of the pathogenic RAB40B. Taken together, we could assess the possible gain-of-function of the human RAB40B mutation by comparing behavioral phenotypes in animal models; our results suggest that the mutant phenotypes may be associated with CRL5-mediated proteolytic regulation.

Peripheral Myelin Protein 22 Gene Mutations in Charcot-Marie-Tooth Disease Type 1E Patients.

Duplication and deletion of the peripheral myelin protein 22 (PMP22) gene cause Charcot-Marie-Tooth disease type 1A (CMT1A) and hereditary neuropathy with liability to pressure palsies (HNPP), respectively, while point mutations or small insertions and deletions (indels) usually cause CMT type 1E (CMT1E) or HNPP. This study was performed to identify PMP22 mutations and to analyze the genotype−phenotype correlation in Korean CMT families. By the application of whole-exome sequencing (WES) and targeted gene panel sequencing (TS), we identified 14 pathogenic or likely pathogenic PMP22 mutations in 21 families out of 850 CMT families who were negative for 17p12 (PMP22) duplication. Most mutations were located in the well-conserved transmembrane domains. Of these, eight mutations were not reported in other populations. High frequencies of de novo mutations were observed, and the mutation sites of c.68C>G and c.215C>T were suggested as the mutational hotspots. Affected individuals showed an early onset-severe phenotype and late onset-mild phenotype, and more than 40% of the CMT1E patients showed hearing loss. Physical and electrophysiological symptoms of the CMT1E patients were more severely damaged than those of CMT1A while similar to CMT1B caused by MPZ mutations. Our results will be useful for the reference data of Korean CMT1E and the molecular diagnosis of CMT1 with or without hearing loss.

Identification and clinical characterization of Charcot-Marie-Tooth disease type 1C patients with LITAF p.G112S mutation.

BACKGROUND: Charcot-Marie-Tooth disease type 1C (CMT1C) is a rare subtype associated with LITAF gene mutations. Until now, only a few studies have reported the clinical features of CMT1C. OBJECTIVE: This study was performed to find CMT1C patients with mutation of LITAF in a Korean CMT cohort and to characterize their clinical features. METHODS: In total, 1,143 unrelated Korean families with CMT were enrolled in a cohort. We performed whole exome sequencing to identify LITAF mutations, and examined clinical phenotypes including electrophysiological and MRI features for the identified CMT1C patients. RESULTS: We identified 10 CMT1C patients from three unrelated families with p.G112S mutation in LITAF. The frequency of CMT1C among CMT1 patients was 0.59%, which is similar to reports from Western populations. CMT1C patients showed milder symptoms than CMT1A patients. The mean CMT neuropathy score version 2 was 7.7, and the mean functional disability scale was 1.0. Electrophysiological findings showed a conduction block in 22% of affected individuals. Lower extremity MRIs showed that the superficial posterior and anterolateral compartments of the calf were predominantly affected. CONCLUSIONS: We found a conduction block in Korean CMT1C patients with p.G112S mutation and first described the characteristic MRI findings of the lower extremities in patients with LITAF mutation. These findings will be helpful for genotype-phenotype correlation and will widen understanding about the clinical spectrum of CMT1C.

Genetic and Clinical Studies of Peripheral Neuropathies with Three Small Heat Shock Protein Gene Variants in Korea.

Small heat shock proteins (sHSPs) are ATP-independent chaperones that help correct the folding of denatured proteins and protect cells from stress. Mutations in HSPB1, HSPB8, and HSPB3 are implicated in inherited peripheral neuropathies (IPNs), such as Charcot-Marie-Tooth disease type 2 (CMT2) and distal hereditary motor neuropathies (dHMN). This study, using whole exome sequencing or targeted gene sequencing, identified 9 pathogenic or likely pathogenic variants in these three sHSP genes from 11 Korean IPN families. Most variants were located in the evolutionally well conserved α-crystallin domain, except for p.P182S and p.S187L in HSPB1. As an atypical case, a patient with dHMN2 showed two compound heterozygous variants of p.R127Q and p.Y142H in HSPB1, suggesting a putative case of recessive inheritance, which requires additional research to confirm. Three HSPB8 variants were located in the p.K141 residue, which seemed to be a mutational hot spot. There were no significant differences between patient groups, which divided by sHSP genes for clinical symptoms such as onset age, severity, and nerve conduction. Early-onset patients showed a tendency of slightly decreased sensory nerve conduction values compared with late-onset patients. As a first Korean IPN cohort study examining sHSP genes, these results will, we believe, be helpful for molecular diagnosis and care of patients with CMT2 and dHMN.

Phenotypic heterogeneity in patients with NEFL-related Charcot-Marie-Tooth disease.

Charcot-Marie-Tooth disease (CMT) is the most common hereditary peripheral neuropathy. Mutations in the neurofilament light polypeptide (NEFL) gene produce diverse clinical phenotypes, including demyelinating (CMT1F), axonal (CMT2E), and intermediate (CMTDIG) neuropathies. From 2005 to 2020, 1,143 Korean CMT families underwent gene sequencing, and we investigated the clinical, genetic, and neuroimaging spectra of NEFL-related CMT patients. Ten NEFL mutations in 17 families (1.49%) were identified, of which three (p.L312P, p.Y443N, and p.K467N) were novel. Eight de novo cases were identified at a rate of 0.47 based on a cosegregation analysis. The age of onset was ≤3 years in five cases (13.5%). The patients revealed additional features including delayed walking, ataxia, dysphagia, dysarthria, dementia, ptosis, waddling gait, tremor, hearing loss, and abnormal visual evoked potential. Signs of ataxia were found in 26 patients (70.3%). In leg MRI analyses, various degrees of intramuscular fat infiltration were found. All compartments were evenly affected in CMT1F patients. The anterior and anterolateral compartments were affected in CMT2E, and the posterior compartment was affected in CMTDIG. Thus, NEFL-related CMT patients showed phenotypic heterogeneities. This study's clinical, genetic, and neuroimaging results could be helpful in the evaluation of novel NEFL variants and differential diagnosis against other CMT subtypes.

Clinical and Neuroimaging Features in Charcot-Marie-Tooth Patients with GNB4 Mutations.

Charcot-Marie-Tooth disease (CMT) is the most common inherited peripheral neuropathy. Mutations in the GNB4 gene cause dominant intermediate CMT type F (CMTDIF). The aim of this study is to investigate phenotypic heterogeneities and characteristics of CMT patients with GNB4 mutations. We enrolled 1143 Korean CMT families and excluded 344 families with a PMP22 duplication. We further analyzed the 799 remaining families to find their GNB4 mutations using whole-exome sequencing (WES). We identified two mutations (p.Gly77Arg and p.Lys89Glu) in three families, among which a heterozygous p.Gly77Arg mutation was novel. In addition, a significant uncertain variant (p.Thr177Asn) was observed in one family. The frequency of the GNB4 mutation in the Korean population is 0.38% in PMP22 duplication-negative families. All three families showed de novo mutation. Electrophysiological findings regarding the p.Lys89Glu mutation showed that the motor nerve conduction velocity (MNCV) of the median nerve was markedly reduced, indicating demyelinating neuropathy, and sural nerve biopsy revealed severe loss of myelinated axons with onion bulb formation. Lower extremity Magnetic Resonance Imaging (MRI) demonstrated relatively more severe intramuscular fat infiltrations in demyelinating type (p.Lys89Glu mutation) patients compared to intermediate type (p.Gly77Arg mutation) patients. The anterolateral and superficial posterior compartment muscles of the distal calf were preferentially affected in demyelinating type patients. Therefore, it seems that the investigated GNB4 mutations do cause not only the known intermediate type but also demyelinating-type neuropathy. We first presented three Korean families with GNB4 mutations and found phenotypic heterogeneities of both intermediate and demyelinating neuropathy. We suggest that those findings are useful for the differential diagnosis of CMT patients with unknown GNB4 variants.

Alanyl-tRNA synthetase 1 (AARS1) gene mutation in a family with intermediate Charcot-Marie-Tooth neuropathy.

BACKGROUND: Alanyl-tRNA synthetase 1 (AARS1) gene encodes a ubiquitously expressed class II enzyme that catalyzes the attachment of alanine to the cognate tRNA. AARS1 mutations are frequently responsible for autosomal dominant Charcot-Marie-Tooth disease type 2N (CMT2N). OBJECTIVE: To identify pathogenic mutation in the Korean patients with CMT and distal hereditary motor neuronopathy (dHMN). METHODS: We screened AARS1 mutations in 373 unrelated CMT families including 318 axonal CMT, 36 dHMN, and 19 intermediate CMT (Int-CMT) who were negative for 17p12 (PMP22) duplication or deletion using whole exome sequencing and targeted sequencing of CMT-related genes. RESULTS: This study identified an early onset Int-CMT family harboring an AARS1 p.Arg329His mutation which was previously reported as pathogenic in French and Australian families. The mutation was located in the highly conserved tRNA binding domain and several in silico analyses suggested pathogenic prediction of the mutations. The patients harboring p.Arg329His showed clinically similar phenotypes of the early onset and electrophysiological intermediate type as those in Australian patients with same mutation. We also found a novel c.2564A>G (p.Gln855Arg) in a CMT2 patient, but its' pathogenic role was uncertain (variant of uncertain significance). CONCLUSION: This study suggests that the frequency of the AARS1 mutations appears to be quite low in Korean CMT. This is the first report of the AARS1 mutation in Korean CMT patients and will be helpful for the exact molecular diagnosis and treatment of Int-CMT patients.

Compound heterozygous mutations of SH3TC2 in Charcot-Marie-Tooth disease type 4C patients.

Charcot-Marie-Tooth disease type 4C (CMT4C) is an autosomal recessive neuropathy caused by SH3TC2 mutations, characterized by spine deformities and cranial nerve involvement. This study identified four CMT4C families with compound heterozygous SH3TC2 mutations from 504 Korean demyelinating or intermediate CMT patients. The frequency of the CMT4C was calculated as 0.79% in demyelinating and intermediate patients (n = 504), but it was calculated as 2.02% in patients without PMP22 duplication (n = 198). The CMT4C frequency was similar to patients in Japan, but it was relatively low compared to those patients in other populations. The symptom was less severe and slowly progressed compared to the other AR-CMT. A patient harboring an intermediate neuropathy showed cranial nerve involvement but did not have scoliosis. This study will be helpful in making molecular diagnoses of demyelinating or intermediate CMT due to SH3TC2 mutations.