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antagonist Onapristone, on the other hand, reduced transgenic PMP22overexpression (see below) (Sereda et al., 2003). This provides strong evidence that the.
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Journal of Molecular Neuroscience Copyright © 2006 Humana Press Inc. All rights of any nature whatsoever reserved. ISSN0895-8696/06/28:77–88/$30.00 JMN (Online)ISSN 1559-1166 DOI 10.1385/JMN/28:01:77

ORIGINAL ARTICLE

Myelin Disorders Causes and Perspectives of Charcot-Marie-Tooth Neuropathy

Gerd Meyer zu Hörste,1 Thomas Prukop,1 Klaus-Armin Nave,1 and Michael W. Sereda*,1,2 1Max-Planck-Institute

of Experimental Medicine, Göttingen, Germany; and 2Center of Neurology, University of Göttingen, Germany Received July 16, 2005; Accepted August 8, 2005

Abstract Charcot-Marie-Tooth (CMT) disease is a common hereditary neuropathy that causes progressive distally pronounced muscle weakness and can lead to life-long disability in patients. In most cases, the disorder has been associated with a partial duplication of human chromosome 17 (CMT1A), causing 1.5-fold overexpression of the peripheral myelin protein 22 kDa (PMP22). Increased PMP22 gene dosage results in demyelination, secondary axonal loss, and neurogenic muscle atrophy. Experimental therapeutic approaches based on the role of progesterone and ascorbic acid in myelin formation recently have reached preclinical proof-of-principle trials in rodents. It was shown that progesterone receptor antagonists can reduce PMP22 overexpression and clinical severity in a CMT1A rat model. Furthermore, ascorbic acid treatment reduced premature death and demyelination in a CMT1A mouse model. Thus, basic research has opened up new vistas for the understanding and treatment of hereditary neuropathies. DOI 10.1385/JMN/28:01:77 Index Entries: Myelin; PMP22; Charcot-Marie-Tooth; CMT1A; hereditary neuropathy; animal model.

Introduction The adequate motor and sensory function of peripheral nerves relies on long neuronal processes (axons) engulfed by glial cells (Schwann cells). The latter provide support and form myelin sheaths, which facilitate fast, saltatory signal propagation along the axon. Peripheral neuropathies are defined as disorders disrupting peripheral nervous system (PNS) function and are common neurological diseases with various known causes. Diabetes and alcoholism frequently cause acquired neuropathies. Hereditary neuropathies defined by genetic mutations resulting in defects of the PNS are less common. However, great advances have recently been

achieved in unraveling disease mechanisms by identifying underlying mutations in the responsible disease genes. Hereditary neuropathies form a heterogeneous group of diseases. Before genetic diagnosis was possible, these disorders were classified by their clinical characteristics and mainly affected nerve fiber quality (Dyck et al., 1993). The most common hereditary neuropathy is Charcot-Marie-Tooth (CMT) disease. Among clinicians it is synonymously referred to as hereditary motor and sensory neuropathy (HMSN). Its prevalence ranges between 1 and 4 in 10,000 individuals (Skre, 1974; Mostacciuolo et al., 1991), and an increasing number of genetic mutations

*Author to whom all correspondence and reprint requests should be addressed. E-mail: [email protected]

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Meyer zu Hörste et al. Table 1 Simplified Genetic Classification of Common CMT Disease Forms

CMT-subtype CMT1 CMT1A HNPP CMT1B CMT1C CMT1D CMTX CMTX or CMT1X CMT2X CMT3X CMT4X CMT2 CMT2A CMT2A CMT2B CMT2D CMT2E CMT2F CMT2

Typical features Demyelinating CMT (mNCV < 38 m/s) Typical CMT1, childhood — typically onset in second decade Recurrent painless pressure palsies, entrapment syndroms, histology: tomacula, onset variable Often more severe than CMT1A, congenital to second-decade onset Typical CMT1, childhood — onset in second decade Often more severe than CMT1A, congenital or first-decade onset X-linked forms of CMT, often intermediate mNCV Classical X-linked CMT, females often less severely affected than males Severe distal weakness, deafness, mental retardation, congenital or early childhood onset Mental retardation, infantile onset Pyramidal signs, onset in second decade Axonal CMT (NCV > 38 m/s or normal) Similar to classical CMT, adult onset Similar to classical CMT, adult onset Mainly sensory neuropathy, acral ulcerations, onset in second or third decade Predominant upper limbs and motor neuropathy, onset in second or third decade Typical CMT, intermediate NCVs, onset in the second or third decade Similar to classical CMT but additional trophic changes, onset in second or third decade Hearing loss, pupillary dysfunction in some pat., onset in fourth or fifth decade

Inheritance

Locus

Gene

AD AD

17p11.2

AD

17p11.2

AD

1q22-q23

AD

16p13.1-p12.3

LITAF

AD

10q21.1-q22.1

EGR2

PMP22 duplic./ point mut. PMP22 deletion MPZ/P0

XR and XD XR/XD

Xq13.1

Cx32/GJB1

XR

Xq24-q26

Unknown

XR XR AD AD AD AD

Xp22.2 Xq26-q28

Unknown Unknown

1p35-p36 1p35-p36 3q13-q22

KIF1B (β) MFN2 RAB7

AD

7p14

GARS

AD

8p21

NEFL

AD

7q11–21

HSP27

AD

1q22-q23

MPZ/P0

Infrequent and autosomal recessive subtypes were excluded for reasons of simplification. (Reprinted, with permission, from Kuhlenbaumer et al., 2002.) AD, autosomal dominant inheritance; XR, X-chromosomal recessive inheritance; XD, X-chromosomal dominant inheritance; NCV, nerve conduction velocity; PMP22, peripheral myelin protein 22 kDa; MPZ/P0, myelin protein zero; LITAF, lipopolysaccharide-induced TNF factor; EGR2, early growth response 2; KIF1Bβ, kinesin family member 1Bβ; MFN2, mitofusin 2, RAB7, member RAS oncogene family 7; GARS, glycyl-tRNA synthetase; NEFL, neurofilament light polypeptide 68 kDa; HSP27, heat shock protein 27 kDa; LMNA, lamin A/C transcript variant 1; Cx32/GJB1, Connexin 32 kDa.

have been associated with different subtypes of CMT disease (Table 1).

Clinical Symptoms of CMT Disease The main clinical feature of CMT disease is a symmetric and distally pronounced muscle weakness of the lower limb and, to a lesser extent, of the upper limb, which slowly progresses proximally. Initially, the muscle atrophy affects the distal leg and intrin-

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sic foot muscles, which clinically results in steppage gait and foot deformities. Sensory deficits are usually not as prominent but are a common feature (Dyck et al., 1993) (Fig. 1A–C). These CMT disease symptoms are characterized by a high degree of interindividual variability. The age at which patients notice first symptoms varies between 10 and 40 yr (Dyck et al., 1993), and disease severity varies greatly. In some cases, a genetically proven disease does not cause any perceivable symptoms,

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Fig. 1. CMT1 patients display atrophy of peroneal (A) and hand muscles (B,C). (D) In a PMP22 transgenic rat model of CMT1A, neurographic analysis demonstrates reduced NCV attributable to peripheral demyelination. (E) Furthermore, CMT rats show spontaneous activity in EMG recordings indicative of muscle denervation and an axonal phenotype. (F) Histologically, CMT rats show a CMT1 typical onion-bulb formation. These findings imitate human CMT1A with the same genetic defect. (A–C were reprinted, with permission, from Kuhlenbaumer et al., 2002; D–F were reprinted, with permission, from Sereda et al., 1996.)

whereas other patients become wheel chair dependent (Birouk et al., 1997). This clinical variability has even been described in identical twins (Garcia et al., 1995), most probably attributable to nongenetic (epigenetic) causes and, in other cases, to modifier genes.

Electrophysiology and Histology in CMT Disease In CMT type 1 the genetic defect primarily affects the myelin-forming Schwann cell, causing the

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degeneration of myelin sheaths (demyelination). The loss of myelin leads to reduced nerve conduction velocity (NCV). In CMT type 2 mutations primarily affect axonal function and cause axonal loss without alterations of NCV. Although genetic testing is increasingly available, the measurement of NCVs remains the most important first-line diagnostic tool for the classification of CMT disease. It allows differentiation between patients with significantly reduced (38 m/s) NCV (CMT2) (Harding and

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80 Thomas, 1980). The CMT1 subtype is more common than CMT2 and will be described in more detail. In CMT1 patients NCV is regularly reduced, but the degree of reduction does not correlate with the clinical picture (Killian et al., 1996) (Fig. 1D). NCV reductions can even be demonstrated before clinical handicaps become apparent and can be used for the early diagnosis of CMT1 (Birouk et al., 1997). Electromyography (EMG) shows spontaneous activity as signs of chronic muscle denervation and axonal loss. Interestingly, the severity of EMG alterations correlates directly with the clinical phenotype (Darciano et al., 2000; Krajewski et al., 2000) (Fig. 1E). This indicates that axonal loss and consecutive muscle atrophy cause CMT1 symptoms, even though the primary genetic defect affects the Schwann cell. Histologically, CMT1 is characterized by a segmental demyelination of predominantly largecaliber axons and so-called onion-bulb formations (Fig. 1F). This pictorial term describes axons that are not surrounded by a single myelinating Schwann cell but rather by several concentric layers of cells and their processes. These cells show characteristics of promyelinating Schwann cells (Guenard et al., 1996), presumably representing the process of overlapping de- and remyelination. With further progression of the disease, mainly large-caliber motor axons degenerate, resulting in muscle atrophy and clinical symptoms (Lewis et al., 2003).

Genetics and Pathomechanisms of CMT Disease Numerous mutations have been described that cause different CMT disease forms, and the rather crude clinical classification of CMT disease subtypes was extended by a genetic classification (Table 1). It became obvious that identical clinical symptoms are caused by different mutations. In contrast, mutations in the same gene can result in distinct phenotypes (Kuhlenbaumer et al., 2002). Mutations are located in myelin protein genes like peripheral myelin protein of 22 KDa (PMP22), myelin protein zero (MPZ), and Connexin 32 (Cx32). Genetic defects in the transcription factor gene early growth response 2 (EGR2) and axonal proteins genes (NEFL, Gigaxonin, Kif1Bβ ) cause different CMT disease subtypes. An updated list of CMT mutations is available at www.molgen.ua.ac.be/CMTMutations/. Recently, different genes have been found responsible for axonal subtypes of CMT (CMT2). The Dynactin protein is involved in retrograde axonal

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Meyer zu Hörste et al. transport. Mutations in this gene cause lengthdependent axonal degeneration and hereditary neuropathy (Puls et al., 2003). Point mutations in the Mitofusin gene (MFN2) and in a Kinesin family gene (Kif1Bβ) are associated with CMT2A (Zhao et al., 2001; Zuchner et al., 2004). Both genes are expressed in neurons. The MFN2 mutation impairs mitochondrial morphology and intracellular transport along the axonal cytoskeleton (Chen et al., 2003). This mitochondrial movement is part of the fast anterograde axonal transport. The intracellular motor protein Kif1Bβ transports mitochondria and precursors of synaptic vesicles within the axon (Zhao et al., 2001). Thus, for some of the axonal CMT disease subforms (CMT2) it is thought that the impaired transport of mitochondria affects the axonal energy supply, ultimately leading to its premature degeneration. In this context it is important to point out that primary Schwann cell defects (CMT1) also cause (secondary) axonal damage that is responsible for the clinical phenotype (Lewis et al., 2003). Peripheral nerves of trembler mice, carrying a naturally occurring PMP22 mutation, show an increased axonal neurofilament density and impaired axonal transport (de Waegh et al., 1992). Xenotransplantation experiments demonstrated that Schwann cells from CMT1A patients carrying the PMP22 duplication change the intracellular structure of healthy axons. Axons surrounded by pathological Schwann cells show increased neurofilament density and distal axonal degeneration (Sahenk et al., 1999). Axonal defects caused by glial defects were also described in the central nervous system (CNS). Mice overexpressing the CNS myelin proteolipid protein (PLP) display demyelination and axonal degeneration. They are regarded as an animal model for the demyelinating CNS disorder Pelizaeus–Merzbacher disease (Kagawa et al., 1994; Readhead et al., 1994; Anderson et al., 1998; Edgar et al., 2004). Thus, glial defects might disturb axonal structure and transport. Obviously, axons are especially sensitive to a reduced energy supply. This might explain the length-dependent axonal loss in CMT1 and could also constitute a disease mechanism in nonhereditary neuropathies.

The PMP22 Protein and CMT1A The most common genetic defect (60–70%) causing CMT disease (Ionasescu, 1995) is a DNA duplication of about 1.5 million base pairs on human chromosome 17p11.2–p12. The resulting neuropathy is then

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Fig. 2. The respective gene dosage of PMP22 determines the clinical symptoms of patients affected by PMP22-related hereditary neuropathies. Genomic deletion of PMP22 causes HNPP. Duplication of PMP22 results in CMT1A symptoms.

termed CMT1A (Lupski et al., 1991; Raeymaekers et al., 1991). In the majority of cases, the duplication results from unequal crossing over during spermatogenesis. The affected region is flanked by highly homologous repeat sequences favoring unequal crossing over (Palau et al., 1993). Thus, this mutational hot spot is the reason for the high prevalence of genomic duplication. The gene for PMP22 is contained in this region and was identified as the responsible disease gene (Matsunami et al., 1992; Patel et al., 1992; Timmerman et al., 1992; Valentijn et al., 1992). Peripheral myelin protein of 22 KDa (PMP22) is a hydrophobic protein with four transmembrane domains and contributes 3–5% to the protein mass of peripheral myelin (Suter and Scherer, 2003). The biological function of this protein in Schwann cells is not fully understood.

CMT1A is a Gene-Dosage Disease In CMT1Apatients the altered number of genomic PMP22 copies determines the type of neuropathy and correlates with disease severity. Loss of one functional PMP22 allele results in a reduced gene dosage (0.5-fold) and in hereditary neuropathy with liability to pressure palsies (HNPP) (Stogbauer et al., 2000) (Fig. 2). A 1.5-fold increased copy number of PMP22 causes CMT1A. The homozygous PMP22 duplication (2.0-fold gene dosage) leads to an aggravated disease course (Lupski et al., 1991). Nerve biopsies from CMT1A patients show an overexpression of

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PMP22 mRNA and protein (Yoshikawa et al., 1994; Vallat et al., 1996). Different hypotheses have been developed to explain the gene-dosage phenomenon. An interaction between PMP22 and the most abundant protein in the myelin sheath, MPZ, was described (D’Urso et al., 1999; Hasse et al., 2004). Possibly, an altered ratio between both interaction partners, caused by overexpression of PMP22, destabilizes the myelin sheath. Alternatively, it was proposed that PMP22 overexpression is associated with abnormal intracellular sorting processes. In cell culture experiments the overexpressed protein accumulates in late endosomes and disturbs cellular membrane transport (Chies et al., 2003). Furthermore, PMP22-overexpressing cells form intracellular protein aggregates, so-called aggresomes, containing PMP22 and ubiquitin immunoreactivity (Notterpek et al., 1999; Fortun et al., 2005). These aggresomes were initially described as a general cellular response to an overload of misfolded proteins (Johnston et al., 1998). Therefore, PMP22 overexpression might disturb Schwann cell and peripheral nerve function by interfering with the intracellular sorting of proteins and membranes and possibly by overloading the protein degradation machinery of the cells. Interestingly, in a recent study, transgenic PMP22 mice showed reduced expression of genes involved in cholesterol biosynthesis (Giambonini-Brugnoli et al., 2005). This may indicate that disturbed lipid composition underlies demyelinating diseases.

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Animal Models of CMT1A Animal models of human diseases help to understand the pathomechanisms involved and allow developing and testing of new therapeutic approaches. Different PMP22 transgenic mouse models (Magyar et al., 1996; Huxley et al., 1998) and one rat model (Sereda et al., 1996) have been designed to mimic the CMT1A disease phenotype in rodents. Especially animal models with moderate PMP22 overexpression present symptoms resembling human CMT1A. Mice with severalfold increased PMP22 expression do not form any peripheral myelin at all, mimicking human Dejerine-Sottas syndrome (Magyar et al., 1996). In the PMP22 transgenic rat model (CMT rat), 1.5-fold overexpression of PMP22 in Schwann cells leads to peripheral demyelination with onion-bulb formations as histological characteristics of human CMT1A. Axonal loss and distally pronounced muscle atrophy similarly match human symptoms (Sereda et al., 1996). Therefore, the CMT rat is particularly suited as an experimental CMT1A model.

Steroids in the Nervous System Although steroids were originally discovered as products and regulators of endocrine tissues and gonads, steroid-based signaling systems of the nervous system have been described and characterized more recently. Various steroids are synthesized in the CNS and PNS. The key enzymes for steroidogenesis are found on mRNA and protein levels in multiple brain areas. The ability to synthesize certain steroids, however, varies greatly between different brain regions and between different time points in development (for review, see Compagnone and Mellon, 2000; Mellon and Griffin, 2002). After removal of all steroidogenic tissues, steroid concentrations remain high in the PNS (Koenig et al., 1995) and CNS (Corpechot et al., 1981). The term neurosteroid was introduced to designate steroids synthesized in the nervous system (Corpechot et al., 1981). Among others, dehydroepiandrosterone (DHEA) (Corpechot et al., 1981), progesterone (Cheney et al., 1995), and allopregnanolone (Cheney et al., 1995), as well as metabolites of the respective steroids, feature brain intrinsic synthesis (for review, see Baulieu et al., 2001). The independence of neurosteroid production from endocrine tissues indicated a distinct role of these substances in the nervous system apart from wellknown functions in the regulation of reproduction, immune system, and water–electrolyte balance.

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Meyer zu Hörste et al. Neurosteroids modulate numerous nervous system functions via either genomic or nongenomic mechanisms. Genomic signaling denotes steroid signal transduction via intracellular receptors. These receptors act as ligand-activated transcription factors (for review, see Tsai and O’Malley, 1994; McKenna and O’Malley, 2002). Furthermore, steroids modulate various neurotransmitter receptors. Among others, this holds true for GABAA receptors (for review, see Lambert et al., 2003), NMDA receptors (Wu et al., 1991; Maciejak et al., 2004), nicotinic receptors (Bullock et al., 1997), glycine-activated chloride channels (Prince and Simmonds, 1992; Wu et al., 1997), and 5-HT3 receptors (Wetzel et al., 1998). Because of these pharmacological properties, neurosteroids influence different complex brain functions. Neurosteroids have sleep-inducing anxiolytic and anticonvulsive effects (for review, see Rupprecht, 2003). Progesterone, for instance, has sedative-like effects in humans (Soderpalm et al., 2004). Altered neurosteroid concentrations in the CNS were demonstrated in animal models of stress (Dubrovsky, 2005; Higashi et al., 2005). Allopregnanolone exerted anxiolytic effects in rats (Akwa et al., 1999) and alleviated the neurodegenerative Niemann-Pick type-C disease (Griffin et al., 2004), indicating a possible neuroprotective function of this neurosteroid in the CNS.

Neurosteroids in the PNS In contrast to the CNS complexity of numerous neurotransmitter receptors and brain functions being modulated by different neurosteroids, peripheral neurosteroids feature relative simplicity. Progesterone and its metabolites are of special interest for the PNS, whereas a major role for other steroids has not been demonstrated (for review, see Schumacher et al., 2001). The enzymes required for progesterone synthesis are expressed in Schwann cells (Chan et al., 1998), and local synthesis occurs independent of endocrine tissues (Koenig et al., 1995). Furthermore, the progesterone receptor itself is expressed in Schwann cells and is functionally active (Jung-Testas et al., 1996), suggesting autocrinic signal transduction. Myelination in cell culture experiments leads to an induction of progesterone-synthesizing enzymes and the progesterone receptor. The time course of myelination closely correlates with the expression of progesterone-synthesizing enzymes in Schwann cells (Robert et al., 2001), supporting a function of endogenous progesterone in the regulation of myelination.

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CMT1A is a Gene-Dosage Disease Exogenously added progesterone increased the expression of myelin genes, like PMP22 and MPZ, in cell culture experiments. The promoters of these myelin genes were activated by progesterone (Desarnaud et al., 1998). Rats treated with progesterone displayed an increased expression of MPZ and, to a lesser degree, of PMP22 (Melcangi et al., 1999). On one hand, addition of progesterone increased PMP22 expression in the PNS of wild-type and transgenic rats (Sereda et al., 2003). The selective progesterone antagonist Onapristone, on the other hand, reduced transgenic PMP22 overexpression (see below) (Sereda et al., 2003). This provides strong evidence that the intracellular progesterone receptor at least partially mediates peripheral PMP22 expression. It has been discussed, whether PMP22 is also regulated via a GABAA receptor-dependent signaling mechanism (Magnaghi et al., 2001). Furthermore, progesterone stimulates remyelination in experimental nerve lesion models. The speed of myelin reformation after peripheral nerve crush injuries was increased by the addition of this neurosteroid (Koenig et al., 1995). In conclusion, progesterone is an important regulator of myelin gene expression and peripheral myelination.

Neurosteroids as Experimental Therapies of CMT Disease Several different approaches to treatment of CMT disease have been proposed, whereas none has reached the stage of clinical trials in humans yet (Shy, 2004; Grandis and Shy, 2005). However, important advances have been achieved recently in animal models of the most common hereditary neuropathy, CMT1A (Sereda et al., 2003; Passage et al., 2004). In an attempt to combine the findings that progesterone increases PMP22 expression and that PMP22 overexpression causes CMT1A, it was hypothesized that the progesterone receptor might be an interesting target to modulate the CMT1A phenotype (Fig. 3). In normal signal transduction, progesterone binds progesterone receptor monomers in the cytosol, which in consequence dimerize and translocate to the nucleus (Beato, 1989; McKenna and O’Malley, 2002). Progesterone receptor complexes bind specific DNA sequences and enhance or repress transcription, depending on the cellular composition of coregulators (McKenna and O’Malley, 2002; Wu et al., 2005). Two different types of progesterone receptor antagonists have been designed (Edwards et al., 1995). Type-I antagonists like Mifepristone (RU 486)

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83 activate nuclear translocation and DNA binding of antagonist receptor complexes but prevent transcriptional initiation (Edwards et al., 1995). However, type-II antagonists like Onapristone (ZK 98299) impede DNA binding of progesterone receptors in general (Edwards et al., 1995). This mechanism is the basis for differences in pharmacological behavior of both antagonist classes in certain experimental settings. High intracellular activity of protein kinase A causes a functional switch of type-I antagonists to partial agonists (Sartorius et al., 1993; Nordeen et al., 1995), which explains PMP22 promoter activation by Mifepristone, the type-I antagonist, in Schwann cell cultures in the presence of high cAMP levels (Desarnaud et al., 1998). Therefore, the selective progesterone receptor antagonist Onapristone (type-II antagonist exerting no agonistic activity) was applied in a transgenic rat model of CMT1A to influence PMP22 overexpression and resulting peripheral neuropathy. In this first proof-of-principle study, the progesterone antagonist reduced PMP22 overexpression, ameliorated axonal loss, and improved the clinical neuropathic phenotype. Progesterone treatment exerted the opposite effect as it further increased PMP22 overexpression and accelerated disease progression (Sereda et al., 2003). These results demonstrate that PMP22 expression is coregulated by the nuclear progesterone receptor. Onapristone or other anti-progesterone drugs might be considered as future therapeutic options in CMT1A, whereas additional progesterone (as in some oral contraceptives) should be avoided (Sereda et al., 2003; Shy, 2004; Grandis and Shy, 2005). Onapristone was originally developed for the treatment of hormone-sensitive tumors like breast cancer or meningeomas (Neef et al., 1984; Jang and Benet, 1997). The drug was tested in phase-II clinical studies (Robertson et al., 1999) and caused transient liver enzyme elevations (Robertson et al., 1999). Further clinical development was abandoned. However, newly characterized progesterone antagonists display fewer adverse effects and allow for oral administration (Fuhrmann et al., 2000). This new compound has already been tested in phase-II clinical studies and could offer new therapeutic perspectives for CMT1A.

Other Experimental Therapy of CMT Disease A second interesting therapeutic approach was recently reported in a CMT1A mouse model overexpressing the PMP22 gene. Ascorbic acid (vitamin C)

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Fig. 3. Exogenously administered or locally synthesized progesterone acts on Schwann cells via the intracellular progesterone receptor. The progesterone receptor complex activates the transcription of myelin genes presumably via transcription factors like EGR2. Progesterone receptor antagonists prevent the binding of progesterone to its receptor, thereby modifying myelin gene expression.

is required for extracellular matrix formation and collagen synthesis (Carey and Todd, 1987; Podratz et al., 2001) and is therefore necessary for myelination in cell cultures (Podratz et al., 2004). It was proposed that increased application of ascorbic acid might enhance myelination in a mouse model overexpressing PMP22 at high levels. Demyelination and premature death in this mouse strain were reduced by high-dose ascorbic acid treatment (Passage et al., 2004). The mechanism of action of ascorbic acid in this experimental model needs to be determined, particularly the vitamin effect on peripheral myelin gene expression. As this vitamin is not restricted by drug legislation, clinical trials in human CMT patients are intended to start shortly.

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Recently, neuroprotective agents have been shown to reduce neuronal degeneration after energy depletion in experimental stroke models (Siren et al., 2001). Even attenuated axonal loss has been described in cell culture models of HIV-induced neuropathy (Keswani et al., 2004). Therefore, neuroprotection might also present a future treatment option for hereditary neuropathies. In summary, different animal models of hereditary neuropathies have helped to extend our knowledge of underlying disease mechanisms and possible future treatment strategies. Schwann cell defects, as well as axonal protein mutations, ultimately alter axonal ultrastructure and intra-axonal transport. It appears that axons are particular sensitive to reduced intracellular transport and energy supply, which

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CMT1A is a Gene-Dosage Disease might provide a common pathomechanism for different neuropathies. Recently, proof-of-principle studies performed in transgenic animal models have identified drugs that might improve the disease course of CMT1A. The progesterone antagonist Onapristone ameliorated the clinical phenotype and axonal loss of CMT rats. Ascorbic acid prevented premature death and demyelination in a CMT1A mouse model. Clinical studies are now needed to confirm a clinical benefit for CMT1A patients.

Acknowlegdments We thank G. Endo for organizational help; J. R. Lupski, R. Melcangi, and the Nave lab for discussion. This work was supported by the Max-Planck Society and by grants from the Euopean Union (to K. A. N.). M. W. S. was supported in part by the Del Marmol Foundation.

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