CMT and related neuropathies exhibit all forms of mendelian inheritance—autosomal dominant, autosomal recessive, and X-linked. Autosomal dominant demyelinating CMT is the most frequent.⁴ Thirty-five linked loci (14 autosomal dominant, 12 autosomal recessive, and 3 X-linked; rarely mutations in a gene isolated as a dominant locus behave as recessive alleles in a given family) and 25 CMT-associated genes have been identified. HNPP and RLS show autosomal dominant inheritance, whereas CHN is autosomal recessive or sporadic. DSN shows both autosomal dominant and autosomal recessive inheritance. Genotype-phenotype correlation studies suggest that genetic heterogeneity, age-dependent penetrance, and variable expressivity are key characteristics of the hereditary motor-sensory neuropathies (HMSN). It is estimated that about one third of the mutations occur de novo⁵^–⁷; thus absence of family history does not preclude genetic testing.

CLASSIFICATION

The classification system for CMT and related peripheral neuropathies is based on clinical phenotype, electrophysiology, and inheritance pattern (Table 82-2). This classification was based on large pedigrees, which were invaluable tools in identifying genes responsible for certain types of CMT. Loci identified from these families were named according to this classification as well. However, it became apparent that a substantial number of cases are sporadic and do not fit this classification. Genes associated with specific loci and disease types were found to be responsible for other types of CMT or with different inheritance patterns. Thus, this classification may need revision to make it both simpler and all inclusive. For a genetic overview of CMT, we divide the genes identified into functional groups, describe the clinical phenotypes associated with mutations in those genes, and briefly describe the presumptive pathomechanism deduced from in vitro and in vivo experiments.

TABLE 82-2 Genetic Classification of Charcot-Marie-Tooth Disease and Related Peripheral Neuropathies

GENETICS

The more than two dozen genes implicated in hereditary sensorimotor neuropathies belong to various functional classes, all involved in the developmental biology and function of peripheral nerves. They include structural proteins that are important in myelination (e.g., PMP22, MPZ), radial transport proteins (e.g., Cx32), proteins of axonal transport (e.g., NEFL), transcription factors involved in Schwann cell differentiation (EGR2), members of signal transduction pathways (e.g., PRX, MTMR2, SBF2, NDGR1), proteins related to mitochondrial function (e.g., MFN2, GDAP1), proteins related to the endosome (RAB7, SIMPLE), and molecular chaperones (HSP22, HSP27). The products of one gene involved in DNA single-strand break repair (TDP1) and of other genes (e.g., LMNA, GARS, DNM2) have less clearly defined functions in nerve physiology.

Genes Associated with Peripheral Nerve Structure

Peripheral Myelin Protein 22 (PMP22)

The first molecular event discovered as responsible for the majority of CMT was the duplication of the chromosomal segment harboring PMP22.⁸ This discovery introduced a novel molecular mechanism in human mutagenesis, nonallelic homologous recombination (Fig. 82-1), and defined a new group of disorders, the genomic disorders.^9,¹⁰ The reciprocal molecular event, deletion—instead of duplication—of the same fragment was found in HNPP.^11,¹² This molecular event and the resulting diseases provided evidence for the presence of dosage-sensitive genes in the human genome.

Figure 82-1 Reciprocal recombination resulting in duplication causing Charcot-Marie-Tooth disease type 1A (CMT1A) or deletion associated with hereditary neuropathy with liability to pressure palsies (HNPP). Proximal CMT1A-REP is depicted by a red box, distal CMT1A-REP is depicted by a blue box, and the PMP22 gene encoding a peripheral myelin proteins depicted by a gray box. A, B, C, D and A′, B′, C′, D′ represent unique sequences flanking the CMT1A-REP low-copy repeats on each of the two chromosome homologues, respectively. Gray lines depict the site of crossover, and the resulting rearrangements are labeled ABCB′C′D′ for the CMT1A duplication and A′D for the reciprocal HNPP deletion. The JCT refers to specific junction fragments detected by probes that are used in diagnostics (PFGE). Homologous meiotic recombination between nonallelic copies of CMT1A-REPs leads to either the CMT1A duplication or HNPP deletion. The CMT1A duplication results in three copies (two on the duplicated chromosome and one on the normal homologue) of the dosage-sensitive PMP22 gene.

Clinical phenotypes: An extra copy of PMP22, due to the CMT1A duplication, is associated with CMT1 ^8,^13,¹⁴ and accounts for 70% of families with dominant CMT1^5,¹⁵ and 76% to 90% of sporadic CMT1.^5,⁷ Reduced compound motor and sensory nerve action potentials correlate with clinical disability, whereas motor nerve conduction velocity does not.¹⁶ A prospective study of eight patients with CMT1A¹⁷ revealed that motor nerve conduction velocities and clinical motor examinations did not change significantly over a period of 22 years. The CMT1A duplication is also associated with neuropathy in patients with wide variations in clinical phenotypes, including DSN, RLS, calf hypertrophy, and scapuloperoneal atrophy or Davidenkow syndrome.⁴

Deletion of PMP22 leads to HNPP.¹¹ In one study,¹⁸ 50% of patients diagnosed with multifocal neuropathy had the 17p11.2 deletion associated with HNPP. Point mutations in PMP22 have been seen in CMT1, HNPP, DSN, and CHN phenotypes.^19,²⁰ As anticipated, loss of function mutations²¹ including frame-shift, nonsense, and splice site mutant alleles result in HNPP; analogous to the HNPP deletion, they effectively result in PMP22 haploinsufficiency.

Function: PMP22 is expressed in the peripheral nervous system, but its role is still unclear after 15 years of research. Most of the newly synthesized PMP22 is retained in the endoplasmic reticulum, where it is degraded.²² Only a small percentage of PMP22 is transported from the endoplasmic reticulum to the Golgi apparatus, where it undergoes complex glycosylation and becomes more stable. Axonal contact appears to stimulate the redistribution of PMP22 to the Schwann cell plasma membrane as myelination occurs.²³ The ultrastructural pathology of the HNPP phenotype, tomacula, and reduced myelin compaction,²⁴ suggests that PMP22 plays a structural role in myelin formation and/or maintenance.

Strategies aimed at normalizing PMP22 expression in transgenic mice have been encouraging ²⁵ and clinical trials are under way.

Myelin Protein Zero (MPZ)

Clinical phenotypes: About 85 to 90 different myelinopathy-associated MPZ mutations have been described (http://molgen-www.uia.ac.be/CMTMutations/). Most of them are associated with CMT1, but DSN and CMT2 phenotypes are also found, together with a few cases of CHN.^26,²⁷ A patient with a severe MPZ mutant allele²⁷ presenting as a floppy baby taught us that innervation may be necessary for proper muscle differentiation and development. The original Roussy-Lévy family reported in 1926 has been shown to harbor a point mutation causing a missense amino acid substitution in the extracellular domain of MPZ.²⁸

Function: MPZ is normally expressed exclusively by myelinating Schwann cells and accounts for 50% of the total PNS myelin protein.²⁹ In vitro functional studies demonstrated that the MPZ truncating mutations associated with a severe form of peripheral neuropathy result in premature stop codons within the terminal or penultimate exons that escape nonsense-mediated decay and are stably translated into mutant proteins.³⁰ However, a subset of these mutations, also escaping nonsense-mediated decay, resulted in a mild form of peripheral neuropathy. Further in vitro experiments demonstrated that the severity of disease phenotype depends on the amount of residual function of the mutant protein. Mutations altering the cytoplasmic domain and impairing adhesion act as null alleles. If the mutations disrupt the transmembrane domain, the mutant proteins are retained in the endoplasmic reticulum, undergo aggregation, and induce apoptosis.³¹

Genes Associated With Transport Through Myelin

Connexin 32 (Cx32)

Clinical phenotypes: Mutations in Cx32 account for nearly 10% of all CMT cases and are the second most frequent cause of CMT after PMP22 duplication. Over 250 different mutations have been described (http://molgen-www.uia.ac.be/CMTMutations/) throughout the entire Cx32 protein, which, unlike the PMP22 and MPZ mutations, are not concentrated in transmembrane or extracellular domains. These mutations behave in a dominant fashion and represent 90% of CMTX. Electrophysiological studies in patients with Cx32 mutations identified three patterns of neuropathy, axonal, demyelinating, and mixed.^32,³³

Function: The Cx32 (Gap junction B1; GJB1) gene encodes a gap junction protein containing four transmembrane domains. A connexon (hemichannel) consists of six connexin subunits and two connexons, one from each apposing membrane, which form a functional channel that allows rapid transport of ions and small molecules.³⁴ Cx32 is expressed in myelinating Schwann cells and is localized to noncompact myelin in the paranode and Schmitt-Lanterman incisures, consistent with its role in providing a radial diffusion pathway between the adaxonal and perinuclear cytoplasm of the Schwann cell.^35,³⁶

Cx32-deficient mice mimic the human CMT1X phenotype ³⁷ with a slowly progressing demyelinating neuropathy. Enlarged periaxonal collars, abnormal noncompacted myelin domains, and axonal sprouts³⁸ suggest that reflexive gap junctions may be required for myelin compaction or that Cx32 may play a structural role in myelin compaction. Mice lacking Cx32 show a distinct pattern of gene dysregulation in Schwann cells,³⁹ indicating that Schwann cell homeostasis is critically dependent on the correct expression of Cx32.

Genes Associated With Axonal Transport

Neurofilament-Light (NEFL)

Clinical phenotypes: Mutations in NEFL have been identified in two independent families affected with autosomal dominant CMT2.^40,⁴¹ Studies^42,⁴³ have identified additional mutations in NEFL among CMT and DSN cases. These patients had early onset, severe CMT or DSN, and moderate to severely reduced nerve conduction velocities.⁴³

Function: NEFL encodes one of the three neurofilament subunits that are the major types of intermediate filaments found in neurons. In vitro functional studies of mutated neurofilament light chain showed defective assembly of intermediate filament networks, defective targeting of neurofilaments to processes, and altered intracellular distribution of mitochondria, suggesting defective axonal transport as the underlying pathomechanism.⁴⁴

Transcription Factors Associated With Myelination

Early Growth Response 2 (EGR2)

Clinical phenotypes: Mutations in human EGR2 are found in patients with CMT1, DSN, and CHN.^45,⁴⁶ Patients with EGR2 mutations frequently have neuropathies affecting cranial nerves III, VII, and XII. Respiratory compromise is a common problem and requires careful monitoring.

Function: EGR2, also known as KROX20, encodes a Cys₂His₂ type zinc finger-containing protein. Most mutations occur in the zinc finger domain. Functional studies have shown that most EGR2 mutations affect the DNA binding and that the amount of residual binding correlates directly with disease severity.⁴⁷ The same studies have shown that a mutation in the R1 domain of EGR2, which binds to the NAB corepressors and prevents their interaction with NAB proteins, leads to increased transcriptional activity of EGR2. Thus, failure to activate or inactivate downstream genes or deregulation of EGR2 activity could be a pathogenic mechanism.

Mouse EGR2 is implicated in the establishment of myelination and its subsequent expression is restricted to myelinating Schwann cells.^48,⁴⁹ Homozygous knockout mice for EGR2 show disruption in hindbrain segmentation^50,⁵¹ and block of Schwann cells at an early stage of differentiation.⁵²

Genes Associated with Signaling

Periaxin (PRX)

Clinical phenotype: Mutations in PRX are associated with autosomal recessive DSN and CMT4F.⁵³^–⁵⁵ PRX mutations cause early onset but slowly progressive neuropathy with marked sensory component.⁵⁶

Function: Alternative splicing of human PRX results in two forms: L-periaxin and S-periaxin.⁵⁷ L-periaxin is first expressed in the nuclei of embryonic Schwann cells and then in the plasma membrane of myelinating Schwann cells.⁵⁸ Its expression pattern in the rat sciatic nerve parallels the deposition of myelin.⁵⁹ In mature myelin, periaxin is found in the cytoplasm-filled periaxonal regions of the sheath but is excluded from compact myelin. Mice disrupted for Prx develop PNS compact myelin that degenerates as the animals age,⁶⁰ consistent with the role of periaxin in myelin stability. These mice are important models to study neuropathic pain in late onset demyelinating disease.

Myotubularin-Related Protein 2 (MTMR2)

Clinical phenotype: Mutations in MTMR2 cause a type of autosomal recessive CMT1 (CMT4B1) and CHN.^61,⁶² CMT4B1 is characterized by focally folded myelin. The mutations are distributed throughout the open reading frame.

Function: MTMR2 encodes a dual specificity phosphatase. It also contains a GRAM domain, an SET-interacting domain, and a PDZ-binding domain. MTMR2 uses the lipid second messenger, phosphoinositol 3-phosphate, as a physiological substrate. The known ⁶³ disease-associated MTMR2 mutations show reduced phosphatase activity,⁶⁴ indicating that the phosphatase activity of MTMR2 is crucial for its proper function in the peripheral nervous system.

SET Binding Factor 2 (SBF2) or Myotubularin-Related Protein 13 (MTMR13)

Clinical phenotype: A homozygous in-frame deletion encompassing exons 11 and 12 was detected in a consangious Turkish family.⁶⁵ The Japanese family from one of the clinical reports of CMT and glaucoma⁶⁶ was found to harbor a nonsense mutation in SBF2, which segregated with a phenotype characterized by markedly decreased MCV, myelin folding, and juvenile-onset glaucoma.⁶⁷

Function: SBF2 encodes an MTMR2-related protein. SBF2 is a member of the pseudo-phosphatase family of myotubularins (also known as MTMR13). It is expressed in various tissues including spinal cord and peripheral nerve. The histopathological hallmarks of the disease are focal outfoldings of myelin in nerve biopsies.