Screening for SH3TC2 gene mutations in a series of demyelinating recessive Charcot‐Marie‐Tooth disease (CMT4)

Charcot‐Marie‐Tooth disease type 4C (CMT4C) is an autosomal recessive (AR) demyelinating neuropathy associated to SH3TC2 mutations, characterized by early onset, spine deformities, and cranial nerve involvement. We screened 43 CMT4 patients (36 index cases) with AR inheritance, demyelinating nerve conductions, and negative testing for PMP22 duplication, GJB1 and MPZ mutations, for SH3TC2 mutations. Twelve patients (11 index cases) had CMT4C as they carried homozygous or compound heterozygous mutations in SH3TC2. We found six mutations: three nonsense (p.R1109*, p.R954*, p.Q892*), one splice site (c.805+2T>C), one synonymous variant (p.K93K) predicting altered splicing, and one frameshift (p.F491Lfs*32) mutation. The splice site and the frameshift mutations are novel. Mean onset age was 7 years (range: 1–14). Neuropathy was moderate‐to‐severe. Scoliosis was present in 11 patients (severe in 4), and cranial nerve deficits in 9 (hearing loss in 7). Scoliosis and cranial nerve involvement are frequent features of this CMT4 subtype, and their presence should prompt the clinician to look for SH3TC2 gene mutations. In our series of undiagnosed CMT4 patients, SH3TC2 mutation frequency is 30%, confirming that CMT4C may be the most common AR‐CMT type.     

Autosomal recessive Charcot‐Marie‐Tooth disease: from genes to phenotypes

The prevalence of Charcot‐Marie‐Tooth (CMT) disease or hereditary motor and sensory neuropathy (HMSN) varies in different populations. While in some countries of Western Europe, the United States and Japan the dominant form of HMSN is the most frequent, in other countries such as those of the Mediterranean Basin, the autosomal recessive form (AR‐CMT) is more common. Autosomal recessive CMT cases are generally characterized by earlier onset, usually before the age of 2 or 3 years, and rapid clinical progression that results in severe polyneuropathy and more marked distal limb deformities such as pes equino‐varus, claw‐like hands, and often major spinal deformities. Recent clinical, morphological and molecular investigations of CMT families with autosomal recessive inheritance allowed the identification of many genes such as GDAP1, MTMR2, SBF2, NDRG1, EGR2, SH3TC2, PRX, FGD4, and FIG4, implicated in demyelinating forms (ARCMT1 or CMT4), and LMNA, MED25, HINT1, GDAP1, LRSAM1, NEFL, HSPB1 and MFN2 in axonal forms (ARCMT2). However, many patients remain without genetic diagnosis to date, prompting investigations into ARCMT families in order to help discover new genes and common pathways. This review summarizes recent advances regarding the genotypes and corresponding phenotypes of AR‐CMT.     

A novel NDRG1 mutation in a non‐Romani patient with CMT4D/HMSN‐Lom

Charcot‐Marie‐Tooth disease type 4D (CMT4D), also known as hereditary motor and sensory neuropathy Lom type (HMSNL), is an autosomal recessive, early onset, severe demyelinating neuropathy with hearing loss, caused by N‐Myc downstream‐regulated gene 1 (NDRG1) mutations. CMT4D is rare with only three known mutations, one of which (p.Arg148Ter) is found in patients of Romani ancestry and accounts for the vast majority of cases. We report a 38‐year‐old Italian female with motor development delay, progressive neuropathy, and sensorineural deafness. Magnetic resonance imaging showed slight atrophy of cerebellum, medulla oblongata, and upper cervical spinal cord. She had a novel homozygous NDRG1 frameshift mutation (c.739delC; p.His247ThrfsTer74). The identification of this NDRG1 mutation confirms that CMT4D is not a private Romani disease and should be considered in the differential diagnosis of recessive demyelinating CMT.     

Sh3tc2 deficiency affects neuregulin‐1/ErbB signaling

Mutations in SH3TC2 trigger autosomal recessive demyelinating Charcot‐Marie‐Tooth type 4C (CMT4C) neuropathy. Sh3tc2 is specifically expressed in Schwann cells and is necessary for proper myelination of peripheral axons. In line with the early onset of neuropathy observed in patients with CMT4C, our analyses of the murine model of CMT4C revealed that the myelinating properties of Sh3tc2‐deficient Schwann cells are affected at an early stage. This early phenotype is associated with changes in the canonical Nrg1/ErbB pathway involved in control of myelination. We demonstrated that Sh3tc2 interacts with ErbB2 and plays a role in the regulation of ErbB2 intracellular trafficking from the plasma membrane upon Nrg1 activation. Interestingly, both the loss of Sh3tc2 function in mice and the pathological mutations present in CMT4C patients affect ErbB2 internalization, potentially altering its downstream intracellular signaling pathways. Altogether, our results indicate that the molecular mechanism for the axonal size sensing is disturbed in Sh3tc2‐deficient myelinating Schwann cells, thus providing a novel insight into the pathophysiology of CMT4C neuropathy.     

A cryptic splicing mutation in the INF2 gene causing Charcot‐Marie‐Tooth disease with minimal glomerular dysfunction

Heterozygous mutations in the inverted formin‐2 (INF2) gene provoke focal segmental glomerulosclerosis (FSGS) and intermediate Charcot‐Marie‐Tooth (CMT) disease with FSGS. Here, we report four patients from a three‐generation family with a new cryptic splicing INF2 mutation causing autosomal dominant intermediate CMT with minimal glomerular dysfunction. Three males and one female with a mean age of 51 years (26‐87) presented with a slowly progressive sensorimotor polyneuropathy, pes cavus, and kyphoscoliosis. Mean age at CMT disease onset was 11.5 years (3‐17), and electrophysiological studies showed demyelinating and axonal features consistent with intermediate CMT. Plasma albumin and creatinine were normal in all four cases, and urine protein was normal in one case and mildly raised in three patients (mean: 0.32 g/L [0.18‐0.44], N < 0.14). Genetic analysis found a c.271C > G (p. Arg91Gly) variation in INF2 exon 2, and in vitro splicing assays showed the deletion of the last 120 nucleotides of INF2 exon 2 leading to a 40 amino acids in‐frame deletion (p. Arg91_p. Gln130del). This report expands the genetic spectrum of INF2‐associated disorders and demonstrates that INF2 mutations may provoke isolated CMT with no clinically relevant kidney involvement. Consequently, INF2 mutation analysis should not be restricted to individuals with coincident neuropathy and renal disease.     

Charcot‐Marie‐Tooth disease: frequency of genetic subtypes in a Southern Italy population

The objective of this study is to assess the genetic distribution of Charcot‐Marie‐Tooth (CMT) disease in Campania, a region of Southern Italy. We analyzed a cohort of 197 index cases and reported the type and frequency of mutations for the whole CMT population and for each electrophysiological group (CMT1, CMT2, and hereditary neuropathy with susceptibility to pressure palsies [HNPP]) and for familial and isolated CMT cases. Genetic diagnosis was achieved in 148 patients (75.1%) with a higher success rate in HNPP and CMT1 than CMT2. Only four genes (PMP22, GJB1, MPZ, and GDAP1) accounted for 92% of all genetically confirmed CMT cases. In CMT1, PMP22 duplication was the most common mutation while the second gene in order of frequency was MPZ in familial and SH3TC2 in isolated cases. In CMT2, GJB1 was the most frequent mutated gene and GJB1 with GDAP1 accounted for almost 3/4 of genetically defined CMT2 patients. The first gene in order of frequency was GJB1 in familial and GDAP1 in isolated cases. In HNPP, the majority of patients harbored the PMP22 gene deletion. The novelty of our data is the relatively high frequency of SH3TC2 and GDAP1 mutations in demyelinating and axonal forms, respectively. These epidemiological data can help in panel design for our patients' population.     

POLG mutations presenting as Charcot‐Marie‐Tooth disease

We report on two patients, with different POLG mutations, in whom axonal neuropathy dominated the clinical picture. One patient presented with late onset sensory axonal neuropathy caused by a homozygous c.2243G>C (p.Trp748Ser) mutation that resulted from uniparental disomy of the long arm of chromosome 15. The other patient had a complex phenotype that included early onset axonal Charcot‐Marie‐Tooth disease (CMT) caused by compound heterozygous c.926G>A (p.Arg309His) and c.2209G>C (p.Gly737Arg) mutations.     

Early onset demyelinating Charcot‐Marie‐Tooth disease caused by a novel in‐frame isoleucine deletion in peripheral myelin protein 2

Peripheral myelin protein 2 (PMP2) is a small protein located on the cytoplasmic side of compact myelin, involved in the lipids transport and in the myelination process. In the last years few families affected with demyelinating Charcot‐Marie‐Tooth neuropathy (CMT1), caused by PMP2 mutations, have been identified. In this study we describe the first case of a PMP2 in‐frame deletion. PMP2 was analyzed by direct sequencing after exclusion of the most frequent CMT‐associated genes by using a next generation sequencing (NGS) genes panel. Sanger sequencing was used for family's segregation analysis. Molecular modeling analysis was used to evaluate the mutation impact on the protein structure. A novel PMP2: p.I50del has been identified in a child with early onset CMT1 and in three affected family members. All family members show an early onset demyelinating neuropathy without other distinguish features. Molecular modeling analysis and in silico evaluations do not suggest a strong impact on the overall protein structure, but a most likely altered protein function. This study suggests the importance to add PMP2 in CMT NGS genes panels or, at most, to test it after major CMT1 genes exclusion, due to the lack of diagnostic‐addressing additional features.     

The phenotype of Charcot–Marie–Tooth disease type 4C due to SH3TC2 mutations and possible predisposition to an inflammatory neuropathy

Charcot–Marie–Tooth (CMT) disease is a heterogeneous group of inherited peripheral motor and sensory neuropathies. The locus responsible for CMT4C was previously assigned to the chromosome 5q23 region by homozygosity mapping and mutations in the SH3TC2 (KIAA1985) gene have been subsequently identified mainly in families around the Mediterranean basin but also frequently in European Gypsies. No English families have been reported to date. To determine the frequency, phenotype and neuropathology of CMT due to SH3TC2 mutations we screened 23 English autosomal recessive (AR) demyelinating CMT families. Five families with AR demyelinating CMT and SH3TC2 mutations were identified, four families were homozygous for the R954X mutation and the fifth family was compound heterozygous for the R954X and E657K mutations. There was significant clinical variation between these families with some cases presenting with a severe childhood onset neuropathy with respiratory and cranial nerve involvement, compared to other families with mild scoliosis and foot deformity. Characteristic sural nerve neuropathology was seen in three families with frequent demyelinating fibres surrounded by excess Schwann cell lamellae forming basal lamina onion bulbs and abnormally long and attenuated Schwann cell processes. One patient homozygous for the R954X mutation had a 20-year history of an inflammatory neuropathy that was superimposed onto the hereditary form, indicating that structural alterations to the SH3TC2 gene could possibly predispose to peripheral nerve inflammation.     

Diabetes mellitus exacerbates motor and sensory impairment in CMT1A

Abstract Charcot‐Marie‐Tooth disease type 1A (CMT1A) is caused by a duplication of PMP22 on chromosome 17 and is the most commonly inherited demyelinating neuropathy. Diabetes frequently causes predominantly sensory neuropathy. Whether diabetes exacerbates CMT1A is unknown. We identified 10 patients with CMT1A and diabetes and compared their impairment with 48 age‐matched control patients with CMT1A alone. Comparisons were made with the Charcot‐Marie‐Tooth disease (CMT) neuropathy score (CMTNS) and by electrophysiology. The CMTNS was significantly higher in patients with diabetes (20.25 ± 2.35) compared with controls (15.19 ± 0.69; p = 0.01). Values were particularly higher for motor signs and symptoms. Seven of the 10 diabetic patients had CMTNS >20 (severe CMT), while only 7 of the 48 age‐matched controls had scores >20. There was a trend for CMT1A patients with diabetes to have low compound muscle action potentials and sensory nerve action potentials, although nerve conduction velocities were not slower in diabetic patients compared with controls. Diabetes was associated with more severe motor and sensory impairment in patients with CMT1A.     

Clinical characterization and genetic analysis of Korean patients with X‐linked Charcot‐Marie‐Tooth disease type 1

Mutations in the gap junction protein beta 1 gene (GJB1) cause X‐linked Charcot‐Marie‐Tooth disease type 1 (CMTX1). CMTX1 is representative of the intermediate type of CMT, having both demyelinating and axonal neuropathic features. We analyzed the clinical and genetic characterization of 128 patients with CMTX1 from 63 unrelated families. Genetic analysis revealed a total of 43 mutations including 6 novel mutations. Ten mutations were found from two or more unrelated families. p.V95M was most frequently observed. The frequency of CMTX1 was 9.6% of total Korean CMT family and was 14.8% when calculated within genetically identified cases. Among 67 male and 61 female patients, 22 females were asymptomatic. A high‐arched foot, ataxia, and tremor were observed in 87%, 41%, and 35% of the patients, respectively. In the male patients, functional disability scale, CMT neuropathy score, and compound muscle action potential of the median/ulnar nerves were more severely affected than in the female patients. This study provides a comprehensive summary of the clinical features and spectrum of GJB1 gene mutations in Korean CMTX1 patients.     

CMT4D (NDRG1 mutation): genotype–phenotype correlations

Charcot‐Marie‐Tooth (CMT) disease is a heterogeneous condition with a large number of clinical, electrophysiological and pathological phenotypes. More than 40 genes are involved. We report a child of gypsy origin with an autosomal recessive demyelinating phenotype. Clinical data, familial history, and electrophysiological studies were in favor of a CMT4 sub‐type. The characteristic N‐Myc downstream‐regulated gene 1 (NDRG1) mutation responsible for this CMT4D phenotype was confirmed: p.R148X. The exact molecular function of the NDRG1 protein has yet to be elucidated.     

Genotype‐phenotype correlation in a novel ABHD12 mutation underlying PHARC syndrome

PHARC syndrome is a rare neurodegenerative disorder caused by mutations in the ABHD12 gene. It is a genetically heterogeneous and clinically variable disease, which is characterized by demyelinating polyneuropathy, hearing loss, cerebellar ataxia, retinitis pigmentosa, and early‐onset cataract and can easily be misdiagnosed as other neurologic disorders with a similar clinical picture, such as Charcot‐Marie‐Tooth disease and Refsum disease. We describe the genotype‐phenotype correlation of two siblings with a novel genotype underlying PHARC syndrome. The genotype was identified using next‐generation sequencing. We examined both patients by means of thorough history taking and clinical examination, nerve conduction studies (NCS), brain imaging, and optical coherence tomography to establish a genotype‐phenotype correlation. We identified a novel homozygous point mutation (c.784C > T, p.Arg262*) in the ABHD12 gene. This mutation was detected in both siblings, who had bilateral hearing loss and cataracts, signs of cerebellar ataxia, and neuropathy with a primarily demyelinating pattern in NCS. In one case, retinitis pigmentosa was also evident. As PHARC syndrome is a rare autosomal recessive disorder, our findings highlight the importance of an interdisciplinary diagnostic workup with clinical and molecular genetic testing to avoid a misdiagnosis as Charcot‐Marie‐Tooth disease or Refsum disease.     

Exome sequencing reveals mutations in MFN2 and GDAP1 in severe Charcot–Marie–Tooth disease

The aim of our study was to characterize electrophysiologically and explain the genetic cause of severe Charcot–Marie–Tooth (CMT) in a 3.5‐year‐old with asymptomatic parents and a maternal grandfather with a history of mild adult‐onset axonal neuropathy. Severity of neuropathy was assessed by Charcot–Marie–Tooth neuropathy score (CMTNS). Whole‐exome sequencing was performed using an Illumina TruSeq Exome Enrichment Kit on the HiSeq 1500 with results followed up by Sanger sequencing on an ABI Prism 3500XL (Applied Biosystems, Foster City, CA, USA). Paternity was confirmed using a panel of 15 hypervariable markers. Electrophysiological studies demonstrated severe axonal sensory‐motor neuropathy in the proband, mild motor neuropathy in his mother, and mild sensory‐motor neuropathy in his grandfather. CMTNS in the proband, his mother, and grandfather was 21, 1, and 12, respectively. On genetic analysis, the boy was found to carry a heterozygous dominant MFN2 T236M mutation transmitted via the maternal line and a de novo GDAP1 H123R mutation. Our findings emphasize the need to search for more than one causative mutation when significant intrafamilial variability of CMT phenotype occurs and underline the role of whole‐exome sequencing in the diagnosis of compound forms of CMT disease.     

Novel mutations in the PRX and the MTMR2 genes are responsible for unusual Charcot-Marie-Tooth disease phenotypes

Autosomal recessive Charcot-Marie-Tooth diseases, relatively common in Algeria due to high prevalence of consanguineous marriages, are clinically and genetically heterogeneous. We report on two consanguineous families with demyelinating autosomal recessive Charcot-Marie-Tooth disease (CMT4) associated with novel homozygous mutations in the MTMR2 gene, c.331dupA (p.Arg111LysfsX24) and PRX gene, c.1090C>T (p.Arg364X) respectively, and peculiar clinical phenotypes. The three patients with MTMR2 mutations (CMT4B1 family) had a typical phenotype of severe early onset motor and sensory neuropathy with typical focally folded myelin on nerve biopsy. Associated clinical features included vocal cord paresis, prominent chest deformities and claw hands. Contrasting with the classical presentation of CMT4F (early-onset Dejerine-Sottas phenotype), the four patients with PRX mutations (CMT4F family) had essentially a late age of onset and a protracted and relatively benign evolution, although they presented marked spine deformities. These observations broaden the spectrum of clinical phenotypes associated with these two CMT4 forms.     

Increased mitochondrial fusion in a autosomal recessive CMT2A family with mitochondrial GTPase mitofusin 2 mutations

Charcot‐Marie‐Tooth type 2A disease (CMT2A) is an inherited peripheral neuropathy mainly caused by mutations in the MFN2 gene coding for the mitochondrial fusion protein mitofusin 2. Although the disease is mainly inherited in a dominant fashion, few cases of early‐onset autosomal recessive CMT2A (AR‐CMT2A) have been reported in recent years. In this study, we characterized the structure of the mitochondrial network in cultured primary fibroblasts obtained from AR‐CMT2A family members. The patient‐derived cells showed an increase of the mitochondrial fusion with large connected networks and an increase of the mitochondrial volume. Interestingly, fibroblasts derived from the two asymptomatic parents showed similar changes to a lesser extent. These results support the hypothesis that AR‐CMT2A‐related MFN2 mutations acts through a semi‐dominant negative mechanism and suggest that other biological parameters might show mild alterations in asymptomatic heterozygote AR‐CMT2A patients. Such alterations could be useful biomarkers helping to distinguish MFN2 mutations from variants, a growing challenge with the advent of next generation sequencing into routine clinical practice.     

Whole‐exome sequencing reveals a novel missense mutation in the MARS gene related to a rare Charcot‐Marie‐Tooth neuropathy type 2U

Charcot‐Marie‐Tooth (CMT) is a heterogeneous group of progressive disorders, characterized by chronic motor and sensory polyneuropathy. This hereditary disorder is related to numerous genes and varying inheritance patterns. Thus, many patients do not reach a final genetic diagnosis. We describe a 13‐year‐old girl presenting with progressive bilateral leg weakness and gait instability. Extensive laboratory studies and spinal magnetic resonance imaging scan were normal. Nerve conduction studies revealed severe lower limb peripheral neuropathy with prominent demyelinative component. Following presumptive diagnosis of chronic inflammatory demyelinating polyneuropathy, the patient received treatment with steroids and intravenous immunoglobulins courses for several months, with no apparent improvement. Whole‐exome sequencing revealed a novel heterozygous c.2209C>T (p.Arg737Trp) mutation in the MARS gene (OMIM 156560). This gene has recently been related to CMT type 2U. In‐silico prediction programs classified this mutation as a probable cause for protein malfunction. Allele frequency data reported this variant in 0.003% of representative Caucasian population. Family segregation analysis study revealed that the patient had inherited the variant from her 60‐years old mother, reported as healthy. Neurologic examination of the mother demonstrated decreased tendon reflexes, while nerve conduction studies were consistent with demyelinative and axonal sensory‐motor polyneuropathy. Our report highlights the importance of next‐generation sequencing approach to facilitate the proper molecular diagnosis of highly heterogeneous neurologic disorders. Amongst other numerous benefits, this approach might prevent unnecessary diagnostic testing and potentially harmful medical treatment.     

Histopathological features of a patient with Charcot‐Marie‐Tooth disease type 2U/AD‐CMTax‐MARS

Charcot‐Marie‐Tooth (CMT) disease is a complex of peripheral nervous system disorders. CMT type 2U (CMT2U) is an autosomal dominant (AD) disease caused by mutations in the MARS gene encoding methionyl‐tRNA synthetase; this disease has thus been newly called AD‐CMTax‐MARS. A few families with mutations in the MARS gene have been reported, without detailed histopathological findings. We describe a 70‐year‐old woman who had bilateral dysesthesia of the soles since the age of 66 years. Sural nerve biopsy showed a decrease in the density of large myelinated nerve fibers. Increased clusters of regenerating myelinated nerve fibers were noted. Electron microscopic analyses revealed degeneration of unmyelinated nerves. There was no vasculitis or inflammatory cell infiltration. Genetic analysis identified a heterozygous p.P800T mutation, a reported mutation in the MARS gene. We report the detailed histopathological findings in a patient with CMT2U/AD‐CMTax‐MARS. The findings are similar to those found in CMT2D caused by mutations in the GARS gene, encoding glycyl‐tRNA synthetase.     

Rigid spine syndrome associated with sensory‐motor axonal neuropathy resembling Charcot–Marie‐Tooth disease is characteristic of Bcl‐2‐associated athanogene‐3 gene mutations even without cardiac involvement

Introduction: Bcl‐2‐associated athanogene‐3 (BAG3) mutations have been described in rare cases of rapidly progressive myofibrillar myopathies. Symptoms begin in the first decade with axial involvement and contractures and are associated with cardiac and respiratory impairment in the second decade. Axonal neuropathy has been documented but usually not as a key clinical feature. Methods: We report a 24‐year‐old woman with severe rigid spine syndrome and sensory‐motor neuropathy resembling Charcot–Marie–Tooth disease (CMT). Results: Muscle MRI showed severe fat infiltration without any specific pattern. Deltoid muscle biopsy showed neurogenic changes and discrete myofibrillar abnormalities. Electrocardiogram and transthoracic echocardiography results were normal. Genetic analysis of a panel of 45 CMT genes showed no mutation. BAG3 gene screening identified the previously reported c.626C>T, pPro209Leu, mutation. Discussion: This case indicates that rigid spine syndrome and sensory‐motor axonal neuropathy are key clinical features of BAG3 mutations that should be considered even without cardiac involvement. Muscle Nerve, 57 : 330–334, 2018