Anticipation in a unique family with Charcot-Marie-Tooth syndrome and deafness: delineation of the clinical features and review of the literature

Charcot-Marie-Tooth disease (CMT) is a clinically and genetically heterogeneous group of polyneuropathies characterized by degeneration of peripheral nerves, resulting in distal muscle atrophy, sensory loss, and deformities of hands and feet. We have studied 34 individuals in a large 84-member four-generation central Illinois family with autosomal dominant Charcot-Marie-Tooth and deafness. Nerve conduction velocities are consistent with type 1 CMT. Audiological evaluation revealed both auditory neuropathy and cochlear involvement in affected individuals. There is increasing clinical severity and younger age of onset of CMT and deafness with each progressive generation, suggestive of anticipation (P < 0.05). The proband, a female diagnosed at birth with hypotonia, bilateral vocal cord palsy, swallowing incoordination, and hearing impairment, died at age 18 months. Another individual died at the age of 3 months from hypotonia later attributed to CMT. Genetic analysis indicated that affected individuals in this family do not have the common 1.4 Mb duplication associated with type 1A CMT; however, all affected individuals have a unique G to C transversion at position 248 in coding exon 3 of the peripheral myelin PMP22 gene located on chromosome 17p11.2-p12. This mutation is predicted to cause an Ala67Pro substitution in the second transmembrane domain of PMP22, consistent with the molecular cause of the CMT phenotype. However, it does not explain the cochlear component of the deafness, the clinical observation of anticipation, and other features in this family.     

DGAT2 Mutation in a Family with Autosomal‐Dominant Early‐Onset Axonal Charcot‐Marie‐Tooth Disease

Charcot‐Marie‐Tooth disease (CMT) is the most common inherited peripheral neuropathy and is a genetically and clinically heterogeneous disorder. We examined a Korean family in which two individuals had an autosomal‐dominant axonal CMT with early‐onset, sensory ataxia, tremor, and slow disease progression. Pedigree analysis and exome sequencing identified a de novo missense mutation (p.Y223H) in the diacylglycerol O‐acyltransferase 2 (DGAT2) gene. DGAT2 encodes an endoplasmic reticulum‐mitochondrial‐associated membrane protein, acyl‐CoA:diacylglycerol acyltransferase, which catalyzes the final step of the triglyceride (TG) biosynthesis pathway. The patient showed consistently decreased serum TG levels, and overexpression of the mutant DGAT2 significantly inhibited the proliferation of mouse motor neuron cells. Moreover, the variant form of human DGAT2 inhibited the axonal branching in the peripheral nervous system of zebrafish. We suggest that mutation of DGAT2 is the novel underlying cause of an autosomal‐dominant axonal CMT2 neuropathy. This study will help provide a better understanding of the pathophysiology of axonal CMT and contribute to the molecular diagnostics of peripheral neuropathies.     

Genetics of the Charcot‐Marie‐Tooth disease in the Spanish Gypsy population: the hereditary motor and sensory neuropathy‐Russe in depth

Four private mutations responsible for three forms demyelinating of Charcot‐Marie‐Tooth (CMT) or hereditary motor and sensory neuropathy (HMSN) have been associated with the Gypsy population: the NDRG1 p.R148X in CMT type 4D (CMT4D/HMSN‐Lom); p.C737_P738delinsX and p.R1109X mutations in the SH3TC2 gene (CMT4C); and a G>C change in a novel alternative untranslated exon in the HK1 gene causative of CMT4G (CMT4G/HMSN‐Russe). Here we address the findings of a genetic study of 29 Gypsy Spanish families with autosomal recessive demyelinating CMT. The most frequent form is CMT4C (57.14%), followed by HMSN‐Russe (25%) and HMSN‐Lom (17.86%). The relevant frequency of HMSN‐Russe has allowed us to investigate in depth the genetics and the associated clinical symptoms of this CMT form. HMSN‐Russe probands share the same haplotype confirming that the HK1 g.9712G>C is a founder mutation, which arrived in Spain around the end of the 18th century. The clinical picture of HMSN‐Russe is a progressive CMT disorder leading to severe weakness of the lower limbs and prominent distal sensory loss. Motor nerve conduction velocity was in the demyelinating or intermediate range.     

Impaired Function is a Common Feature of Neuropathy‐Associated Glycyl‐tRNA Synthetase Mutations

Charcot–Marie–Tooth disease type 2D (CMT2D) is an autosomal‐dominant axonal peripheral neuropathy characterized by impaired motor and sensory function in the distal extremities. Mutations in the glycyl‐tRNA synthetase (GARS) gene cause CMT2D. GARS is a member of the ubiquitously expressed aminoacyl‐tRNA synthetase (ARS) family and is responsible for charging tRNA with glycine. To date, 13 GARS mutations have been identified in patients with CMT disease. While functional studies have revealed loss‐of‐function characteristics, only four GARS mutations have been rigorously studied. Here, we report the functional evaluation of nine CMT‐associated GARS mutations in tRNA charging, yeast complementation, and subcellular localization assays. Our results demonstrate that impaired function is a common characteristic of CMT‐associated GARS mutations. Additionally, one mutation previously associated with CMT disease (p.Ser581Leu) does not demonstrate impaired function, was identified in the general population, and failed to segregate with disease in two newly identified families with CMT disease. Thus, we propose that this variant is not a disease‐causing mutation. Together, our data indicate that impaired function is a key component of GARS‐mediated CMT disease and emphasize the need for careful genetic and functional evaluation before implicating a variant in disease onset.     

Genetic basis of inherited peripheral neuropathies

Progress in the elucidation of the genetic basis for inherited peripheral neuropathies has been remarkable over the last years. In particular, the molecular mechanisms underlying the autosomal dominantly inherited disorders Charcot–Marie–Tooth disease type 1A (CMT1 A), Charcot–Marie–Tooth disease type 1B (CMT1B), and hereditary neuropathy with liability to pressure palsies (HNPP) have been determined. While mutation in the gene encoding the major myelin protein, P_o has been associated with CMT1B, CMT1A and HNPP have been shown to be associated with reciprocal recombination events leading either to a large submicroscopic duplication in CMT1 A, or the corresponding DNA deletion in HNPP. Available evidence is consistent with the hypothesis that one or more genes within the relevant rearranged segment of 1.5 Mb on chromosome 17 is sensitive to gene dosage providing a novel mechanism for inherited human disorders. It is likely that the gene encoding the peripheral myelin protein PMP22 is at least one of the genes involved since the PMP22 gene maps within the CMT1A duplication (or HNPP deletion), and point mutations within it have been shown to cause a CMT phenotype in humans and comparable neuropathies in rodents (trembler and trembler^J). The mechanism(s) by which gene dosage and point mutations affecting the same gene might lead to a similar phenotype are currently unknown but recent transgenic mouse experiments suggest that similar mechanisms may also underlie other genetic diseases. © 1994 Wiley-Liss, Inc.     

Charcot‐Marie‐Tooth disease type I and related demyelinating neuropathies: Mutation analysis in a large cohort of Italian families

Charcot‐Marie‐Tooth neuropathy type 1 (CMT1), the most common hereditary neurological disorder in humans, is characterized by clinical and genetic heterogeneity. It is caused mainly by a 1.5 Mb duplication in 17p11.2, but also by mutations in the myelin genes PMP22 (peripheral myelin protein 22), MPZ (myelin protein zero), Cx32 (connexin 32; also called GJB1), and EGR2 (early growth response 2). In this study, we have screened 172 index cases of Italian families in which there was at least one subject with a CMT1 diagnosis for the duplication on 17p11.2 and mutations in these genes. Among 170 informative unrelated patients, the overall duplication frequency was 57.6%. A difference could be observed between the duplication frequency in familial cases (71.6%) and that observed in non‐familial cases (36.8%). Among the non‐duplicated patients, 12 were mutated in Cx32, four in MPZ, two in PMP22, and none in the EGR2. In the non‐duplicated cases, the overall point mutation frequency for these genes was 25.0%. We describe the mutations identified, and consider possible genotype–phenotype correlation. Hum Mutat 18:32–41, 2001. © 2001 Wiley‐Liss, Inc.     

ATP1A1 mutations cause intermediate Charcot‐Marie‐Tooth disease

Intermediate Charcot‐Marie‐Tooth (CMT) disease is a heterogeneous group of inherited neuropathies characterized by progressive muscle weakness and atrophy of the distal extremities, distal sensory loss. There were still a large proportion of causative genes for intermediate CMT failed to be identified. Here, using whole‐exome sequencing technique, we identified two novel missense mutations in ATP1A1 gene, c.620C>T (p.S207F) and c.2629G>A (p.G877S), in two Chinese CMT families. Further functional analysis revealed that these mutations led to the loss function of the ATP1A1 protein. The two mutations did not affect the levels of messenger RNA but possessed a damaging effect on ATP1A1 protein expression and they downregulated the protein levels of ATP1A1 by promoting its proteasome degradation. Taken together, we confirmed ATP1A1 as a novel causative gene for intermediate CMT.     

Diagnostic algorithms in Charcot–Marie–Tooth neuropathies: experiences from a German genetic laboratory on the basis of 1206 index patients

We present clinical features and genetic results of 1206 index patients and 124 affected relatives who were referred for genetic testing of Charcot–Marie–Tooth (CMT) neuropathy at the laboratory in Aachen between 2001 and 2012. Genetic detection rates were 56% in demyelinating CMT (71% of autosomal dominant (AD) CMT1/CMTX), and 17% in axonal CMT (24% of AD CMT2/CMTX). Three genetic defects (PMP22 duplication/deletion, GJB1/Cx32 or MPZ/P0 mutation) were responsible for 89.3% of demyelinating CMT index patients in whom a genetic diagnosis was achieved, and the diagnostic yield of the three main genetic defects in axonal CMT (GJB1/Cx32, MFN2, MPZ/P0 mutations) was 84.2%. De novo mutations were detected in 1.3% of PMP22 duplication, 25% of MPZ/P0, and none in GJB1/Cx32. Motor nerve conduction velocity was uniformly <38 m/s in median or ulnar nerves in PMP22 duplication, >40 m/s in MFN2, and more variable in GJB1/Cx32, MPZ/P0 mutations. Patients with CMT2A showed a broad clinical severity regardless of the type or position of the MFN2 mutation. Out of 75 patients, 8 patients (11%) with PMP22 deletions were categorized as CMT1 or CMT2. Diagnostic algorithms are still useful for cost‐efficient mutation detection and for the interpretation of large‐scale genetic data made available by next generation sequencing strategies.     

Continuum of phenotypes in hereditary motor and sensory neuropathy with proximal predominance and Charcot–Marie–Tooth patients with TFG mutation

Charcot–Marie–Tooth (CMT) is a common neuropathy, and hereditary motor and sensory neuropathy with proximal predominance (HMSN‐P) is a recently described rare neuromuscular disease. Although many genes have been implicated for CMT, TFG is the only known HMSN‐P‐causing gene. Within the framework of diagnostic criteria, clinical variation is evident among CMT‐diagnosed and also HMSN‐P‐diagnosed individuals. Mutations that cause p.(Pro285Leu) and p.(Gly269Val) in TFG were earlier reported as cause of HMSN‐P in two Iranian pedigrees. Here, we report the identification of p.(Gly269Val) in TFG as cause of CMT in a large Iranian pedigree. The clinical features of patients of the three pedigrees are presented and critically compared. Similarities between the two HMSN‐P‐diagnosed pedigrees with different TFG mutations, and differences between the two differentially diagnosed pedigrees with the same p.(Gly269Val) mutation were evident. The clinical features of the HMSN‐P pedigree with the p.(Pro285Leu) and the CMT pedigree with the p.(Gly269Val) mutation were clearly congruent with the respective diagnoses, whereas the features of the HMSN‐P‐diagnosed pedigree with the p.(Gly269Val) were intermediate between the other two pedigrees. It is therefore suggested that the clinical features of the three Iranian pedigrees with TFG mutations and diagnosed with HMSN‐P or CMT represent a continuum.     

Charcot‐Marie‐Tooth Disease Type 4H Resulting from Compound Heterozygous Mutations in FGD4 from Nonconsanguineous Korean Families

Charcot‐Marie‐Tooth disease type 4H (CMT4H) is an autosomal recessive demyelinating subtype of peripheral enuropathies caused by mutations in the FGD4 gene. Most CMT4H patients are in consanguineous Mediterranean families characterized by early onset and slow progression. We identified two CMT4H patients from a Korean CMT cohort, and performed a detailed genetic and clinical analysis in both cases. Both patients from nonconsanguineous families showed characteristic clinical manifestations of CMT4H including early onset, scoliosis, areflexia, and slow disease progression. Exome sequencing revealed novel compound heterozygous mutations in FGD4 as the underlying cause in both families (p.Arg468Gln and c.1512‐2A>C in FC73, p.Met345Thr and c.2043+1G>A (p.Trp663Trpfs*30) in FC646). The missense mutations were located in highly conserved RhoGEF and PH domains which were predicted to be pathogenic in nature by in silico modeling. The CMT4H occurrence frequency was calculated to 0.7% in the Korean demyelinating CMT patients. This study is the first report of CMT4H in Korea. FGD4 assay could be considered as a means of molecular diagnosis for sporadic cases of demyelinating CMT with slow progression.     

Charcot‐Marie‐Tooth 1A: A narrative review with clinical and anatomical perspectives

Charcot–Marie‐Tooth 1A (CMT1A) is regarded as the most common hereditary peripheral neurodegenerative disorder. This narrative review highlights perspectives around the historically well‐established and characteristic anatomical manifestations of CMT1A seen in the feet, legs and hands, in addition to a clinical diagnosis that may be confirmed by electrophysiology, genetic or molecular markers together with the presence of a typical family history. A less well‐known perspective is the potential for systemic manifestations and wider complication. The condition is characterised by a progressive clinical picture with unmistakable anatomical and neurological features that have been described since the late 19th century. There remains no cure although supportive, rehabilitative, and surgical regimes may provide helpful management or amelioration of symptoms. Most recently, the emergence of a pleotherapeutic approach suggests distinct promise. Future research focused on a detailed elucidation of the underlying molecular mechanisms underpinning myelin and axonal function may eventually hold the key to successful treatment of CMT1A. Genetic modification would potentially present a cure. Clin. Anat. 29:547–554, 2016. © 2015 Wiley Periodicals, Inc.     

A Novel Mutation in FGD4/FRABIN Causes Charcot Marie Tooth Disease Type 4H in Patients from a Consanguineous Tunisian Family

Charcot‐Marie‐Tooth (CMT) disease constitutes a clinically and genetically heterogeneous group of hereditary neuropathies characterized by progressive muscular and sensory loss in the distal extremities with chronic distal weakness, deformation of the feet, and loss of deep tendon reflexes. CMT4H is an autosomal recessive demyelinating subtype of CMT, due to mutations in FGD4/FRABIN, for which nine mutations are described to date. In this study, we describe three patients from a consanguineous Tunisian family, presenting with severe, early onset, slowly progressive, autosomal recessive demyelinating CMT, complicated by mild to severe kyphoscoliosis, consistent with CMT4H. In these patients, we report the identification of a novel homozygous frameshift mutation in FGD4: c.514_515insG; p.Ala172Glyfs*27. Our study reports the first mutation identified in FGD4 in Tunisian patients affected with CMT. It further confirms the important clinical heterogeneity observed in patients with mutations in FGD4 and the lack of phenotype/genotype correlations in CMT4H. Our results suggest that FGD4 should be screened in other early‐onset CMT subtypes, regardless of the severity of the phenotype, and particularly in patients of consanguineous descent. In Tunisians, as in other populations with high consanguinity rates, screening of genes responsible for rare autosomal recessive CMT subtypes should be prioritized.     

Massively Parallel Sequencing Detected a Mutation in the MFN2 Gene Missed by Sanger Sequencing Due to a Primer Mismatch on an SNP Site

We describe a patient with early onset severe axonal Charcot‐Marie‐Tooth disease (CMT2) with dominant inheritance, in whom Sanger sequencing failed to detect a mutation in the mitofusin 2 (MFN2) gene because of a single nucleotide polymorphism (rs2236057) under the PCR primer sequence. The severe early onset phenotype and the family history with severely affected mother (died after delivery) was very suggestive of CMT2A and this suspicion was finally confirmed by a MFN2 mutation. The mutation p.His361Tyr was later detected in the patient by massively parallel sequencing with a gene panel for hereditary neuropathies. According to this information, new primers for amplification and sequencing were designed which bind away from the polymorphic sites of the patient's DNA. Sanger sequencing with these new primers then confirmed the heterozygous mutation in the MFN2 gene in this patient. This case report shows that massively parallel sequencing may in some rare cases be more sensitive than Sanger sequencing and highlights the importance of accurate primer design which requires special attention.     

Identification and functional characterization of two missense mutations in NDRG1 associated with Charcot‐Marie‐Tooth disease type 4D

Charcot‐Marie‐Tooth disease type 4D (CMT4D) is an autosomal‐recessive demyelinating form of CMT characterized by a severe distal motor and sensory neuropathy. NDRG1 is the causative gene for CMT4D. To date, only four mutations in NDRG1 —c.442C>T (p.Arg148*), c.739delC (p.His247Thrfs*74), c.538‐1G>A, and duplication of exons 6–8—have been described in CMT4D patients. Here, using targeted next‐generation sequencing examination, we identified for the first time two homozygous missense variants in NDRG1, c.437T>C (p.Leu146Pro) and c.701G>A (p.Arg234Gln), in two Chinese CMT families with consanguineous histories. Further functional studies were performed to characterize the biological effects of these variants. Cell culture transfection studies showed that mutant NDRG1 carrying p.Leu146Pro, p.Arg148*, or p.Arg234Gln variant degraded faster than wild‐type NDRG1, resulting in lower protein levels. Live cell confocal microscopy and coimmunoprecipitation analysis indicated that these variants did not disrupt the interaction between NDRG1 and Rab4a protein. However, NDRG1‐knockdown cells expressing mutant NDRG1 displayed enlarged Rab4a‐positive compartments. Moreover, mutant NDRG1 could not enhance the uptake of DiI‐LDL or increase the fraction of low‐density lipoprotein receptor on the cell surface. Taken together, our study described two missense mutations in NDRG1 and emphasized the important role of NDRG1 in intracellular protein trafficking.     

Substrate interaction defects in histidyl‐tRNA synthetase linked to dominant axonal peripheral neuropathy

Histidyl‐tRNA synthetase (HARS) ligates histidine to cognate tRNA molecules, which is required for protein translation. Mutations in HARS cause the dominant axonal peripheral neuropathy Charcot‐Marie‐Tooth disease type 2W (CMT2W); however, the precise molecular mechanism remains undefined. Here, we investigated three HARS missense mutations associated with CMT2W (p.Tyr330Cys, p.Ser356Asn, and p.Val155Gly). The three mutations localize to the HARS catalytic domain and failed to complement deletion of the yeast ortholog (HTS1). Enzyme kinetics, differential scanning fluorimetry (DSF), and analytical ultracentrifugation (AUC) were employed to assess the effect of these substitutions on primary aminoacylation function and overall dimeric structure. Notably, the p.Tyr330Cys, p.Ser356Asn, and p.Val155Gly HARS substitutions all led to reduced aminoacylation, providing a direct connection between CMT2W‐linked HARS mutations and loss of canonical ARS function. While DSF assays revealed that only one of the variants (p.Val155Gly) was less thermally stable relative to wild‐type, all three HARS mutants formed stable dimers, as measured by AUC. Our work represents the first biochemical analysis of CMT‐associated HARS mutations and underscores how loss of the primary aminoacylation function can contribute to disease pathology.     

A Novel Mutation (D305V) in the early growth response 2 gene is associated with severe Charcot‐Marie‐Tooth type 1 disease

Hereditary motor and sensory neuropathies (HMSN) comprises a wide clinical spectrum of related disorders with defects in peripheral nerve myelination. Charcot‐Marie‐Tooth type 1 (CMT1) is the most common form and is usually a mild disease with onset in the first or second decade; however there is a interfamilial and intrafamilial clinical variation, ranging from asymptomatic expression to severe muscular weakness and atrophy. Recently point mutations in the early growth response 2 gene (EGR2/Krox‐20) have been associated with hereditary myelinopathies. We investigated for mutations at the EGR2 gene a patient with severe CMT1 phenotype. Direct sequencing of EGR2 gene showed a heterozygous A T transversion at nucleotide 1064 that predicts an Asp305Val substitution within the first zinc‐finger domain. The finding of a novel EGR2 mutation associated with a different phenotype confirms that peripheral neuropathies represent a continuum spectrum of related disorders due to an underlying defect in myelination. Hum Mutat 14:353–354, 1999. © 1999 Wiley‐Liss, Inc.     

Missense variants in the X‐linked gene PRPS1 cause retinal degeneration in females

Retinal dystrophies are a heterogeneous group of disorders of visual function leading to partial or complete blindness. We report the genetic basis of an unusual retinal dystrophy in five families with affected females and no affected males. Heterozygous missense variants were identified in the X‐linked phosphoribosyl pyrophosphate synthetase 1 (PRPS1) gene: c.47C > T, p.(Ser16Phe); c.586C > T, p.(Arg196Trp); c.641G > C, p.(Arg214Pro); and c.640C > T, p.(Arg214Trp). Missense variants in PRPS1 are usually associated with disease in male patients, including Arts syndrome, Charcot–Marie–Tooth, and nonsyndromic sensorineural deafness. In our study families, affected females manifested a retinal dystrophy with interocular asymmetry. Three unrelated females from these families had hearing loss leading to a diagnosis of Usher syndrome. Other neurological manifestations were also observed in three individuals. Our data highlight the unexpected X‐linked inheritance of retinal degeneration in females caused by variants in PRPS1 and suggest that tissue‐specific skewed X‐inactivation or variable levels of pyrophosphate synthetase‐1 deficiency are the underlying mechanism(s). We speculate that the absence of affected males in the study families suggests that some variants may be male embryonic lethal when inherited in the hemizygous state. The unbiased nature of next‐generation sequencing enables all possible modes of inheritance to be considered for association of gene variants with novel phenotypic presentation.     

Patient with large 17p11.2 deletion presenting with Smith‐Magenis syndrome and Joubert syndrome phenotype

We report on a 22‐year‐old woman carrying a del(17)(p11.2p12) and presenting with the clinical manifestations of both Smith‐Magenis syndrome (SMS) and Joubert syndrome (JS). Her facial anomalies, brachydactyly, severe mental retardation, and self‐injuring behavior could be attributed to SMS, whereas the cerebellar vermis hypoplasia, hypotonia, ataxic gait, developmental delay, and abnormal respiratory pattern were suggestive of JS. By fluorescent in situ hybridization analyses with Yeast Artificial Chromosomes (YAC) mapping to the 17p11.2 region, as well as locus‐specific probes generated through a novel procedure, we could establish that the deletion encompasses a 4‐Mb interval with centromeric and telomeric breakpoints at loci D17S793 and D17S953, the latter close to the locus Charcot Marie Tooth 1A (CMT1A)‐REP. The deletion differs from that commonly found in SMS in its telomeric boundary, which is more distal than usually observed. The presence of JS phenotype in our patient and the detection of an unusual SMS deletion might suggest the presence of a JS gene in close proximity to the SMS locus. Am. J. Med. Genet. 95:467–472, 2000. © 2000 Wiley‐Liss, Inc.     

伴有脑干听觉诱发电位异常的腓骨肌萎缩症发现连接蛋白32基因新突变

目的：报告一个脑干听觉诱发电位有异常改变的腓骨肌萎缩症（Charcot-Marie-Tooth disease,CMT）家系，并探讨与连接蛋白32(connexin32，Cx32)基因突变的关系。方法：对整个家系进行临床检查，对先证者进行肌电图及脑干听觉诱发电位检查，并应用聚合酶链式反应-单链构象多态（polymerase chain reaction -single strand conformation polymouphism，PCR-SSCP）技术结合DNA序列分析方法检测了先证者、家系内8人及家系外50名无血缘关系的正常人。结果：先证者肌电图检查示神经传导速度明显减慢，脑干听觉诱发电位示中枢传导延长，该家系中先证者及另3人均出现异常SSCP条带，经测序证实为392T→C(Leu131Pro)突变。结论：Leu131Pro是未报道过的突变，腓骨肌萎缩症患者可以出现脑干听觉诿发电位异常。