-
Congenital myasthenic syndromesDaniel Hanta a, Pascale Richardb,
Jeanine Koeniga and Bruno Eymarda
Purpose of review
Congenital myasthenic syndromes are a heterogeneous group
of diseases caused by genetic defects affecting
neuromuscular
transmission. In this article, a strategy that leads to the
diagnosis of congenital myasthenic syndromes is presented,
and recent advances in the clinical, genetic and molecular
aspects of congenital myasthenic syndrome are outlined.
Recent findings
Besides the identification of new mutations in genes already
known to be implicated in congenital myasthenic syndromes
(genes for the acetylcholine receptor subunits and the
collagen
tail of acetylcholinesterase), mutations in other genes have
more
recently been discovered and characterized (genes for
choline
acetyltransferase, rapsyn, and the muscle sodium channel
SCN4A). Fluoxetine has recently been proposed as an
alternative treatment for slow channel congenital myasthenic
syndrome.
Summary
The characterization of congenital myasthenic syndromes
comprises two complementary steps: establishing the
diagnosis
and identifying the pathophysiological type of congenital
myasthenic syndrome. Characterization of the type of
congenital
myasthenic syndrome has allowed it to be classified as
caused
by presynaptic, synaptic and postsynaptic defects. A
clinically
and muscle histopathologically oriented genetic study has
identified several genes in which mutations cause the
disease.
Despite comprehensive characterization, the phenotypic
expression of one given gene involved is variable, and the
aetiology of many congenital myasthenic syndromes remains to
be discovered.
Keywords
electromyography, genetic diagnosis, microelectrophysiology,
neuromuscular junction molecules, neuromuscular
transmission,
treatment
Curr Opin Neurol 17:539551. # 2004 Lippincott Williams &
Wilkins.
aInserm U582 and Unite Clinique de Pathologie Neuromusculaire,
Institut deMyologie, and bUnite Fonctionnelle de Cardiogenetique et
Myogenetique, Hopital dela Salpetrie`re, Paris, France
Correspondence to Daniel Hanta, Inserm U582, Institut de
Myologie, Hopital de laSalpetrie`re, 47 Boulevard de lHopital,
75651 Paris Cedex 13, FranceTel: +33 1 42165706; fax: +33 1
42165700;e-mail: [email protected]
Current Opinion in Neurology 2004, 17:539551
Abbreviations
ChAT choline acetyltransferaseCMAP compound muscle action
potentialCMS congenital myasthenic syndrome
# 2004 Lippincott Williams & Wilkins1350-7540
IntroductionCongenital myasthenic syndromes (CMSs) form
aheterogeneous group of genetic diseases characterizedby a
dysfunction of neuromuscular transmission. Thisdysfunction causes
muscle weakness, which is increasedby exertion and usually starts
during childhood. Theprevalence of CMS is estimated at one in 500
000 inEurope, and CMSs are much more uncommon thanautoimmune
myasthenia [1].
Knowledge of the mechanisms underlying CMS hasincreased
considerably in the past 25 years, because ofthe pioneering work
undertaken by the group of Engel etal. [2]. Acetylcholinesterase
deficiency was the first CMSidentified, based on the lack of the
enzyme atneuromuscular junctions [2]. Progressively, the
patho-physiological heterogeneity of CMS was demonstrated:besides
synaptic CMS caused by acetylcholinesterasedeficiency, pre- and
postsynaptic CMS were described,the latter including quantitative
deficiency or kineticanomalies of the acetylcholine receptor. In
the past 15years, many gene mutations responsible for CMS
wereidentified, affecting the different acetylcholine
receptorsubunits and the collagenic tail of
acetylcholinesterase[3,4 ..]. Mutations in the genes for choline
acetyltrans-ferase (ChAT) [5], rapsyn [6], and more recently
thesodium channel SCN4A have been reported to causeCMS [7.].
Several reviews have been devoted to CMS, one of themore recent
being that of Engel et al. [8 ..]. Theobjectives of the present
review are to highlight theprincipal phenotypical and
pathophysiological character-istics of CMS, to pinpoint the more
recent advances inthe field, and to propose a strategy for the
accuratecharacterization of these disorders.
Classification of congenital myasthenicsyndromes and recent
findingsThe current classification of CMS is based on
patho-physiology, i.e. on the precise identification of
theneuromuscular transmission anomaly. The location ofthe
dysfunction of neuromuscular transmission (Fig. 1)[9], which is
specific to the different CMSs, is eitherpresynaptic (generally
caused by an anomaly of ChAT),synaptic (corresponding to an anomaly
of the acetylcho-linesterase collagen tail), or postsynaptic
(secondary to ananomaly of acetylcholine receptor or rapsyn). In
theexperience of Engels group, postsynaptic CMSs arethree times
more frequent than acetylcholinesterasedeficiency and 10 times more
frequent than presynaptic
539
-
CMS [4..]. The different classes and subclasses of CMSwill be
described below with reminders of their firstdescriptions and main
characteristics and with anemphasis on the latest findings.
Presynaptic congenital myasthenic syndromes
Among the presynaptic CMSs, only those caused bymutations in the
ChAT gene have been fully character-ized, the others remain to be
defined at the geneticlevel.
Congenital myasthenic syndromes caused by ChAT mutations
These CMSs usually manifest at birth or in the neonatalperiod
with bulbar disorders and respiratory insufficiencywith apnoea
[10,11] or even sudden death [11,12]. Theseepisodes are triggered
by fever, fatigue and overexertion.Apart from these bouts, the
myasthenic signs are oftenmodest or not present. Cholinesterase
inhibitors areeffective. Microelectrophysiology shows, after
prolonged10 Hz repetitive stimulation, a reduction in amplitude
of
the miniature endplate potentials. These anomalies
arecharacteristic of a defect in the resynthesis of acetylcho-line
or in the filling of synaptic vesicles [10]. Ultra-structural
examination shows that, when muscle is atrest, the synaptic
vesicles are of reduced size.
Ohno and collaborators [5] described the first mutationsin CHAT,
the gene encoding ChAT and located in10q11.2. ChAT is a presynaptic
protein localized in thenerve terminals, where it catalyses
acetylcholine produc-tion. As shown in knockout mice, ChAT
affectssynaptogenesis and coordinates synaptic maturation[13].
Mutations lead to a reduction or even abolition ofthe catalytic
capacity of the enzyme [5]. Fourteenmutations have been reported to
date, mostly of themissense type [5,14 .,15.]. Recent structural
studiesindicate that whereas half of the missense mutationsare
positioned in the molecule such that they affectenzyme activity
directly, the remaining mutations aredistant from the active site
and must exert indirect
Figure 1. Pathophysiological classification of congenital
myasthenic syndromes
axon terminal
basal lamina
AChR
rapsyn
muscle fibre
Na+ channel
ACh
ChAT
AChE Q, T
Presynaptic defectsDefects in ACh resynthesis (AR) CHATPaucity
of synaptic vesiclesLambert-Eaton like CMS
Synaptic defectsEndplate AChE deficiency (AR) COLQCOLQ
CHAT
Postsynaptic defectsAChR kinetic anomalies slow channel syndrome
(AD) >, . fast channel syndrome (AR)AChR deficiency (AR) AChR
>, . rapsyn (AR) RAPSNAnomaly of muscle Na+ channel -subunit
SCN4A
RAPSN
SCN4A
> , ,
> ,,
Incompletely characterized CMSCMS with plectin
deficiencyFamilial limb girdle myastheniaCMS with tubular
aggregates
No identified defects
CHRNB1CHRND
CHRNA1CHRNE
The eight genes involved in congenital myasthenic syndromes
(CMSs) are named CHRNA1, CHRNB1, CHRND, CHRNE, COLQ, CHAT, RAPSNand
SCN4A. The coded protein and the gene location are, respectively,
as follows: (1) a (2q24q32), b (17p11p12), d (2q33q34), e
(17p13)subunits of acetylcholine receptor (AChR); (2) collagenic
tail (ColQ) (3p242) of acetylcholinesterase (AChE); (3) ChAT
(10q11.2); (4) rapsyn(11p11). The endplate species of
acetylcholinesterase is composed of one, two, or three
homotetramers of T globular catalytic subunits attached to
acollagenic tail (ColQ), anchoring them in the synaptic basal
lamina. Rapsyn stabilizes the acetylcholine receptor aggregates and
links them to thepostsynaptic cytoskeleton. Heredity is either
autosomal recessive (AR) or autosomal dominant (AD). Modified from
the classification proposed by theEuropean Neuromuscular Center
(ENMC) [9].
Neuromuscular disease: muscle540
-
effects [16 ..]. The possibility that the I336T ChATmutation
found in three consanguineous Turkishfamilies was a founder was
postulated [14 .].
Other presynaptic myasthenic syndromes still incompletely
characterized
The paucity of synaptic vesicles was described in onepatient
with early-onset CMS. The density of acetylcho-line synaptic
vesicles was reduced by 80% and thenumber of quanta released was
drastically reduced [17].The exact cause of this CMS is still
unknown.
The first case of LambertEaton-like CMS was firstreported in a
child [18]. His myasthenic syndrome wascharacterized by a good
response to guanidine and byelectrophysiological anomalies
identical to those of aLambertEaton syndrome, i.e. diminished
action poten-tials markedly potentiated by tetanic stimulation.
Asecond case presented with severe hypotonia andrespiratory
distress at birth [4..]. No mutation was foundin the gene coding
for the presynaptic calcium channel.
Three patients were reported with a sporadic myasthenicsyndrome
with associated signs of attack of the centralnervous system
(cerebellar ataxia or nystagmus) [19].None presented, as in the
LambertEaton syndrome,with a reduction of the action potentials or
withpotentiation after high frequency stimulation.
Microelec-trophysiology revealed a marked reduction in
thespontaneous or nerve stimulation-induced release ofacetylcholine
quanta.
Synaptic congenital myasthenic syndrome:
acetylcholinesterase deficiency
CMSs caused by acetylcholinesterase deficiency werefirst
described in 1977 [2]. Since then, many cases ofpartial or complete
deficiency of the enzyme located inthe synaptic basal lamina have
been reported [20]. Thefirst symptoms usually arise in the neonatal
period, andthe symptoms are severe with a significant lethal
risk.However, the disease may start later, during infancy, andis
not so severe. Several observations point to thediagnosis of
acetylcholinesterase deficiency: autosomalrecessive heredity,
repetitive compound muscle actionpotential (CMAP) after single
stimulation (Fig. 2), theabsence of response to cholinesterase
inhibitors, andslowed [20] but inconstant [21] pupil responses to
light.
Diagnosis using muscle biopsies is indicated by no orpoor
visualization of acetylcholinesterase at the neuro-muscular
junction. Acetylcholinesterase deficiency isrelated to mutations in
the COLQ gene coding for thecollagenic tail of acetylcholinesterase
[2224]. At theneuromuscular synapse, acetylcholinesterase is
presentas asymmetric acetylcholinesterase, which is made up ofthree
homotetramers each comprising four globular
catalytic subunits linked together by a collagenic tail(ColQ; Q
for queue in French, which means tail) oftrimeric helicoidal
structure. The collagenic tail concen-trates and anchors the enzyme
within the synaptic basallamina.
Twenty-four recessive mutations have been described todate (Fig.
2). They are more often homozygous thanheterozygous, and nonsense
than missense [4 ..]. Thefact that the same homozygous G240X
mutation wasdetected in several Arab families and in one Iraqi
Jewishpatient suggests that it is not uncommon in Near andMiddle
East countries [25]. Depending on their localiza-tion, COLQ
mutations have different consequences: inthe N-terminal
proline-rich attachment domain, theyprevent attachment of the
acetylcholinesterase catalyticsubunits to the collagenic tail; in
the mid-part theyprevent the trimerization of the collagenic tail;
in the C-terminal domain they most often impair anchoring of
theenzyme within the synaptic basal lamina [24,26,27.,28.].This
impaired attachment of the collagenic tail to thesynaptic basal
lamina has recently been elegantly andcomprehensively tested using
purified C-terminal do-main mutant ColQ applied to frog
neuromuscularjunctions [29 ..].
To date there is no effective treatment for this type ofCMS.
Postsynaptic congenital myasthenic syndromes
Among these postsynaptic CMSs, two categories aredescribed: CMS
in connection with a kinetic anomaly ofthe acetylcholine receptor;
and CMS with a decreasednumber of acetylcholine receptors at the
neuromuscularjunction. In the latter category, besides the CMSs
withacetylcholine receptor deficiency as a result of
numerousmutations in the different acetylcholine receptor
subunitgenes, those caused by mutations in the rapsyn genewere
identified recently [6] and are far from infrequent.
Congenital myasthenic syndrome caused by acetylcholine
receptor kinetic anomalies
Slow channel syndrome is the most frequent kineticanomaly of the
acetylcholine receptor. This entity, ofautosomal dominant
inheritance, is characterized by aprolonged opening time of the
acetylcholine receptor[30]. Fifteen autosomal dominant missense
point muta-tions causing a gain of function of the
acetylcholinereceptor were identified [4..]. Although most of
themutations were found in the acetylcholine receptor asubunit
[31], other subunits are also concerned [32]. Themutations are
located in two transmembrane domainstaking part in the formation of
the acetylcholine receptorpore through which passes the sodium flux
[33], M1 formutations of the a and b subunits and M2 for those,more
frequent, affecting the a, b, d and e subunits [34];
Congenital myasthenic syndromes Hanta et al. 541
-
an area of the extracellular domain of the a subunit closeto the
acetylcholine binding site (mutations aG153S andaV156M) [4 ..].
The functional consequences of the various mutationswere studied
in intercostal muscle biopsy or byexpressing the mutation in cell
systems [35]. Theprolonged opening time of the acetylcholine
receptor isdependent either on the slowed closing of the channel
oron the increased affinity of the acetylcholine receptor forits
ligand [36]. In addition, a new mechanism thatinvolves not only
delayed closure but also delayedopening of the channel in the case
of a dS268F mutationwas recently published [37].
Clinical expression may vary from early onset andsevere to late
onset and moderate [30,38]. Thearguments in favour of the diagnosis
are autosomal
dominant heredity, no response to cholinesteraseinhibitors, and
repetitive CMAP after a single stimula-tion. The last two
characteristics are also found inacetylcholinesterase deficiency
(Fig. 2). The selectivityof muscle involvement with a prevalent
atrophic deficitof the finger extensors and of the cervical muscles
issuggestive of slow channel syndrome. Remodelling ofthe
ultrastructure of the endplate is observed withcalcium deposits,
destruction of the postsynaptic folds,vacuolizations and tubular
aggregates [30]. The diag-nosis leads to the therapeutic use of
quinidine, a blockeragent able to normalize the acetylcholine
receptoropening time [39]. Fluoxetine has recently been shownto be
an alternative to quinidine when the latter is nottolerated by the
patient [40 .].
A peculiar case has been reported of a slow channelsyndrome with
recessive transmission, occurring in a
Figure 2. Main features of slow channel syndrome and of
acetylcholinesterase deficiency
Acetylcholinesterase deficiency- autosomal recessive
- slowed pupil response
-BGT fasciculin
control
patient
AChR AChE
AChR AChE
COLQ
In common
- no response to cholinesterase inhibitors
- repetitive CMAP after single stimulation
1 2 3 4 5 6 7 8 9 102 mV 2 ms
Repetitive CMAP
Slow channel syndrome- autosomal dominant
CHRNA1
AChR
M1 M2 M3 M4
s-s
NH2COOH
In both diseases, cholinesterase inhibitors are inefficient, and
a repetitive motor response is evidenced by electrophysiological
study. Recessivelytransmitted endplate acetylcholinesterase
deficiency is demonstrated by the absence of fluorescent fasciculin
staining at the endplate. Collagenic tailgene mutations will be
determined. Slow channel syndrome is transmitted as an autosomal
dominant trait. The most common mutations involveacetylcholine
receptor a subunit in the pore region (transmembrane M1 and M2
domains) or in the vicinity of the acetylcholine binding site.
Othermutations not shown here are within the b, d and e subunits.
AChE, Acetylcholinesterase; AChR, acetylcholine receptor; a-BGT,
a-bungarotoxin;CMAP, compound muscle action potential.
Neuromuscular disease: muscle542
-
consanguineous family in connection with a homozygousmutation of
the e subunit (eL78P) located in theextramembrane region. This
mutation was pathogeniconly if present on two alleles [41]. In
addition, a slowchannel syndrome associated with a
chromosomaltranslocation 2q319p27 was described [42].
Fast channel syndromes are of autosomal recessivetransmission,
although a case of autosomal dominanttransmission was reported
recently [43.]. The diagnosisis made by microelectrophysiology
showing a shorteningof the acetylcholine receptor opening time
[44]. Clinicalseverity is variable. Arthrogryposis was reported in
onecase [45]. The patients are responsive to the combina-tion of
3,4-diaminopyridine and cholinesterase inhibitors.Eight mutations
were identified affecting a, d and esubunits and are located either
in the extracellulardomain, in the M3 transmembrane domain
(mutationaV285I), or in the cytoplasmic loop between the M3 andM4
domains (e mutations only) [8 ..]. Of the twomutations present in
the patient, one is a nonsensemutation whereas the other is
responsible for the kineticanomalies. This second mutation can
modify the kineticsof the receptor by various mechanisms that can
bedetermined on intercostal biopsy or after in-vitroexpression in
cell models. A recent article thus detailsthe detrimental effects
of a V132L mutation located inthe acetylcholine receptor a subunit
within the signaturecystine loop on acetylcholine binding and
channel gating[46 ..]. The different mechanisms underlying fast
chan-nel syndromes are the topic of a recent review [47.].
Congenital myasthenic syndromes with predominant
acetylcholine receptor deficiency (with absent or only
slight
kinetic anomalies)
These account for approximately half of CMS patients[4..]. The
majority are related to mutations of theacetylcholine receptor. No
peculiar clinical findings pointto this type of autosomal recessive
CMS, whose severityis variable. Nevertheless a founder effect in
the Gypsypopulation of the e1267delG mutation has been pro-posed
[48]. An extensive study on five disease loci in thedifferent Gypsy
groups has demonstrated a strongfounder effect and a carrier rate
of 3.74% for thismutation [49].
Cholinesterase inhibitors are most often active and
3,4-diaminopyridine can provide additional benefit.
The described mutations are numerous (60 or more),either
homozygous or heterozygous [4 ..]. They are of alltypes: missense
mutations, chromosomal deletions,insertions, deletions. The
mutations are located on thewhole gene encoding the acetylcholine
receptor esubunit, most being located in the extracellular
domainand in the cytoplasmic loop between the M3 and M4
transmembrane domains [4 ..]. Recently, a
chromosomalmicrodeletion was identified for the first time in
CHRNE[50], showing that this type of mutation may be missedby
standard screening techniques. Lately a frameshiftmutation in exon
7 of CHRNE (e553del7) was shown toprovoke skipping of the preceding
exon both in muscletissue and when expressed in COS cells [51
.].
Mutations in the promoter were also described
[52,53].Interestingly, the injection of the corresponding
recom-binant in the rat allowed the authors to demonstrate thata
mutation in an N-box of the CHRNE promoter leads toless
acetylcholine receptor synaptic expression [50].Another
experimental approach, namely the cell expres-sion of green
fluorescent tagged acetylcholine receptor,allowed others to show
that mutations affecting cysteine470 of the e subunit prevent
acetylcholine receptorsurface expression [54].
More rarely, other subunits of acetylcholine receptor a, band d
subunits are implicated [4..]. The preponderanceof mutations of the
e subunit may be caused by thepossibility of the re-expression of
the g fetal acetylcho-line receptor isoform in the case of null
mutations ofCHRNE [55,56].
Curiously, CMS not only affects humans but also SouthAfrican Red
Brahman calves. These calves suspected ofmyasthenia have been shown
to bear a homozygous 20basepair deletion mutation in bovine CHRNE.
Thismutation leads to a non-functional allele and a severephenotype
[57,58.].
Congenital myasthenic syndromes with mutations of the rapsyn
gene (RAPSN)
These were first identified in 2002 [6]. Rapsyn is a43 000 Mr
postsynaptic cytoplasmic protein, whichparticipates in
acetylcholine receptor assembly at theneuromuscular junction [59]
and allows its anchoring tothe cytoskeleton by b-dystroglycan among
other mole-cules [60]. Most mutations of this gene located in
11p11were identified in the tetratricopeptide repeat domain,and
cell expression studies revealed that the co-expression of mutant
rapsyn and acetylcholine receptorsubunits impair the recruitment of
acetylcholine recep-tors to rapsyn clusters, an essential step for
the anchoringof the acetylcholine receptor to the cytoskeleton
[6].These mutations are responsible for a reduction ofrapsyn and
consequently of the acetylcholine receptoritself at the
neuromuscular junction. The reduction inrapsyn expression is not
specific, because it is alsoobserved in primary acetylcholine
receptor deficiencies.The inheritance of this CMS is autosomal
recessive.
Since the first four cases were published, nearly 50 othercases
have been reported [6163,64.,65,66 .,67.,68]. Half
Congenital myasthenic syndromes Hanta et al. 543
-
of them bear the homozygous N88K. The other halfbears N88K on
one allele and a second mutation on theother allele. This second
mutation is localized all alongthe rapsyn molecule, and nearly 20
different mutationshave been identified to date (Fig. 3a) [6].
Missensemutations predominate (approximately two-thirds ofcases).
When the second mutation is not identified bydirect sequencing, the
search for a chromosomal micro-deletion of RAPSN is recommended
[69].
Two E-box mutations were identified in the rapsynpromoter [64.]
(Fig. 3b). Seven of the eight patientsreported originated from the
Jewish population of Iraqand Iran and had already been described
for theirpeculiar clinical phenotype: benign CMS with
facialmalformations (mandibular prognathism, elongated
face)[70].
A founder effect of the frequently identified N88Kmutation is
likely at least in the European or Indo-European population
[61,62,71..], although the exis-
tence of other founders has been proposed [72]. Thehigh
frequency of the N88K mutation may lead to casesof pseudo-dominant
inheritance.
Genotypephenotype correlation is not easy. Analysisof the corpus
of clinical observations confirms theexistence of two phenotypes:
(1) a neonatal form,even antenatal (with arthrogryposis multiplex
congeni-ta), with major respiratory disorders and severeprogression
of the disease; and (2) mild formsbeginning during childhood or in
adulthood. On thebasis of 16 cases, a distinction between early and
lateonset was proposed [66.]. The importance of theidentification
of the late-onset cases is to avoidimproper immunotherapy. Patients
with the rapsynmutation responded well to cholinesterase
inhibitors[66 .] or to the combination of cholinesterase
inhibitorsand 3,4-diaminopyridine [67.].
In summary, mutations in RAPSN and the resultingrapsyn
deficiency appear to be an important cause of
Figure 3. Diagram depicting the main domains of rapsyn and the
localization of the identified mutations
Tetratricopeptide repeats (TPR) necessaryto rapsyn
self-association
AChR
TPR1 TPR2 TPR3 TPR4 TPR5 TPR6 TPR7 coiledcoil RING
Q3K
L14P
46in
sCA
25V
F81
LY
86X
N88
KR
91L
C97
X
Q12
4X
A14
2DR
151P
V16
5M
553i
ns5
IVS
4-2A
G
A24
6V
Y26
9X
G29
1D
E33
3X
1083
_108
4dup
CT
1177
delA
A
(a)
(b)
38AG 27CG
E box E box
-d
ystr
ogly
can
(a) Seven tetratricopeptide repeat domains (TPR17) are necessary
for rapsyn to self-associate. The coiled-coil domain binds to the
large cytoplasmicloop of the acetylcholine receptor (RACh)
subunits. The RING domain binds rapsyn to b-dystroglycan. A serine
phosphorylation site is located atcodon 406. Modified from Ohno et
al. [6]. (b) Two E-box (CANNTG-type sequence, on grey background),
to which myogenic factors can bind arelocated upstream of the
transcription initiation site in the rapsyn gene promoter region.
Localization of the two different rapsyn promoter mutations.Adapted
from Ohno et al. [64.].
Neuromuscular disease: muscle544
-
CMS associated with endplate acetylcholine
receptordeficiency.
Congenital myasthenic syndrome with plectin deficiency
Plectin is a highly preserved structural protein of
thecytoskeleton expressed in several cell types, includingskeletal
muscle and the postsynaptic membrane. Plectindeficiency was
described in a patient presenting withprogressive myopathy,
associated with myasthenic syn-drome (involving facial, limb and
oculomotor muscles),and epidermolysis bullosa [73]. The
pathophysiology ofthis CMS is poorly understood.
Congenital myasthenic syndrome caused by a mutation in the
sodium channel SCN4A
The case was recently reported of a 20-year-old
patientpresenting since birth with very short bouts (330 min)of
respiratory distress and bulbar paralysis [7 .]. Thediagnosis was
made by electrophysiology of the inter-costal muscle, which
revealed the impossibility ofevoking an action potential after
nerve stimulation.Two mutations of SCN4A were identified,
includingonly one (V1442E) located in the S3/S4
extracellulardomain, which was found to be pathogenic whenexpressed
in HEK cells. The clinical aspect is quitedifferent from that
usually associated with a SCN4Amutation (dyskalemic paralysis,
congenital paramyot-ony).
Incompletely characterized congenital myasthenic
syndromes
These CMSs are described on clinical or histologicalgrounds, but
their molecular origin and more generallytheir pathophysiology
remain unknown in the absence ofan exhaustive exploration.
Familial limb girdle myasthenia
Several families have been reported [74,75]. Thispreviously
named myasthenic myopathy is of recessiveinheritance. Clinically,
the absence of oculobulbar signswas remarkable. The weakness and
fatigability involvedthe girdles. The peculiarity of this not yet
understoodentity was recently stressed with the publication of
fivecases, who all presented with tubular aggregates in theirmuscle
biopsy and who all responded favourably tocholinesterase inhibitors
[76].
Congenital myasthenic syndrome with tubular aggregates
This CMS is associated with tubular aggregates at
thehistological muscle examination. The case of threesisters
presenting with a slowly progressive myopathybeginning in early
childhood associated with cardio-myopathy was reported. A
favourable response tocholinesterase inhibitors was noted [77].
Similar char-acteristics were described in another family [75].
Asporadic case was reported recently [78]. In the absence
of thorough investigations of neuromuscular transmis-sion, the
classification of these cases remains delicate,more especially as
the presence of tubular aggregates isnot specific and can be
associated with isolatedmyopathy, painful cramps [79] and with slow
channelsyndromes [31].
Approach to the diagnosis of congenitalmyasthenic syndromesOn
the basis of these historical advances in the knowl-edge of CMS,
the diagnostic strategy includes roughlytwo successive steps: (1)
the association of a clinical-electrophysiological picture of a
myasthenic syndrome,and data in favour of a congenital origin; and
(2) therecognition of the pathophysiological type, which isbased on
clinical data, the type of hereditary transmis-sion, the response
to cholinesterase inhibitors, the resultsof electromyography, and
finally the muscle biopsy andmolecular genetics. The sequential
order of these twolast investigations depends upon the initial
clinical-electromyographical data.
Clinical presentation
The various CMSs share a common clinical presenta-tion. The
onset is in general early. Late appearance ofthe symptoms during
adolescence, or even in the adult,is more rarely reported. Some
clinical signs suggest ananomaly of neuromuscular transmission:
ophthalmople-gia and ptosis, dysphonia and swallowing
disturbance,facial paresis, and muscle fatigability. In the young
child,the ptosis is not easy to recognize because hypotonia,poor
mimicry, suction disorders, and weakness of the cryare in the
foreground. The occurrence of bouts andworsening by exertion are
characteristics of the disease.The favourable effect of
cholinesterase inhibitors is asignificant argument in favour of a
myasthenic syn-drome. However, two types of CMS are worsened
bycholinesterase inhibitors: slow channel syndrome
andacetylcholinesterase deficiency. With the propermyasthenic
signs, myopathic signs are often associated:amyotrophy, tendinous
retractions, facial malformationand scoliosis. The severity of the
CMS is variable,depending upon the severity of the walking deficit,
thebulbar disorders and the respiratory difficulties.
Acuterespiratory failure may occur, triggered by
infectiousepisodes, and is frequent in the first months of life.
Inthe absence of respiratory assistance, the risk of death ishigh
[12].
A family history of the disease is an essential argument
infavour of the genetic origin of myasthenic syndrome.Most CMSs are
of autosomal recessive inheritance. Slowchannel syndrome is the
only autosomal dominant CMScharacterized hitherto. The progression
patterns ofCMSs are highly variable, including in a given
patient,at various periods of life. Myasthenic bouts are fre-
Congenital myasthenic syndromes Hanta et al. 545
-
quently triggered by infectious episodes, pregnancy andeven
periods. Progressive aggravation of the disease maysometimes occur
late in adulthood, with the appearanceof respiratory insufficiency
[27 .]. A favourable progres-sion is possible after a severe
neonatal onset [4..].
Titration of anti-acetylcholine receptor antibodies in the
serum
The absence of antibodies against acetylcholine receptorand
muscle-specific receptor tyrosine kinase [80,81.] is aprerequisite
for the diagnosis of CMS, although anexception was reported [82
.].
Electromyography
The electrophysiology of neuromuscular transmission isthe
determinant for the diagnosis of CMS. This includessearching for
neuromuscular block, repetitive motorresponses and increments
[83,84]. The observation of aneuromuscular transmission block
affirms the myasthe-nic syndrome. The decrement can be absent in
CMS,particularly in patients who are not highly symptomatic,and in
cases of CMS caused by mutations in ChAT [5] orrapsyn [6]. In the
case of a ChAT deficiency, thedecrement may appear only after an
initial 5-min 10 Hzstimulation [14 .]. Repetitive CMAPs are
pathognomonicof two varieties of CMS: slow channel syndrome
andacetylcholinesterase deficiency (Fig. 2). The search foran
increment is imperative. An increment greater than100% in amplitude
and in area is suggestive of apresynaptic origin.
Muscle biopsy
A first role of the muscle biopsy is to eliminate thediagnosis
of myopathy (congenital myopathy or mito-chondrial cytopathy).
Although non-specific, the pre-dominance of type I fibres and the
marked atrophy oftype II fibres is suggestive of CMS. The presence
oftubular aggregates is frequent, but poses the problem ofthe group
of CMSs with tubular aggregates. Theneuromuscular junctions are
visualized for (acetyl)choli-nesterase by the histochemical
technique of Koelle,fasciculin or specific antibodies, and for
acetylcholinereceptor by fluorescent a-bungarotoxin, which binds
toit. The neuromuscular junctions frequently exhibitvariable
anomalies: reduced size, the disappearance ofsynaptic folds, all
modifications are not specific, however,to a given CMS.
Two types of information are decisive: (1) the absence
ofacetylcholinesterase at the neuromuscular junctionestablishes the
diagnosis of acetylcholinesterase defi-ciency; a study by
ultracentrifugation on sucrose gradientwill generally reveal the
absence of asymmetrical(synaptic) forms of the enzyme; and (2) a
significantreduction in the number of acetylcholine
receptors,further quantified by binding with iodinated
a-bungar-
otoxin, points to a primary anomaly of acetylcholinereceptor or
rapsyn.
The expression of the fetal g subunit of the acetylcho-line
receptor argues in favour of a primary anomaly ofthe acetylcholine
receptor e subunit [55,56]. Immuno-cytochemical study of the
expression of other markers ofthe neuromuscular synapse can also be
performed: agrin,muscle-specific receptor tyrosine kinase, rapsyn,
neur-egulin, a-dystrobrevin or utrophin [85]. It is aetiologi-cally
suggestive if there is a major and selectivereduction of the
expression of a protein, but the primarynature of the deficit is,
however, not established (adeficit in rapsyn is found in CMS with
mutations in boththe rapsyn gene and the acetylcholine receptor
subunitgenes).
Molecular genetics
The diagnosis of CMS can be confirmed by molecularanalyses in
the eight genes whose mutations are so farknown to cause CMS: four
genes encoding the variousacetylcholine receptor subunits (CHRNE,
CHRNA1,CHRNB1, CHRND), the genes encoding rapsyn(RAPSN), the
collagen tail of acetylcholinesterase(COLQ), choline
acetyltransferase (CHAT), and thesodium channel (SCN4A). With the
exception of theGypsy e1267del mutation [48] and the RAPSN
N88Kmutation [6,71 ..], a search has been made to identifyprivate
mutations. Analysis of the coding sequencesand flanking intronic
regions by direct sequencing afterpolymerase chain reaction
amplification of each fragmenton genomic DNA is required. Many
mutations havebeen described to date, and the two predominant
genesin postsynaptic CMS appear to be CHRNE and RAPSN.In
approximately half of the cases, the analysis of thesegenes does
not identify a mutation causing the disease,suggesting that other
genes could be involved. When anew mutation is identified, its
pathogenic character canbe demonstrated by expression studies of
this mutationin HEK cells, COS cells or oocytes, but other
experi-mental models can also be used.
Microelectrophysiology of neuromuscular transmission
Microelectrophysiology of the intercostal muscle can beused to
specify the pre- or postsynaptic location of thedysfunction of
neuromuscular transmission, and inpostsynaptic CMS, to find kinetic
anomalies of theacetylcholine receptor [4..]. The complexity of
thesetechniques (patch clamp) and the risks of generalanaesthesia
in a myasthenic patient limit the indicationsfor this exploration,
more especially because the expres-sion of the mutations in
experimental cell systems byitself allows the pathophysiological
characterization ofCMS. In addition, the study of other muscles
under localanaesthesia was proposed: quadriceps [86] and
ancone[15].
Neuromuscular disease: muscle546
-
Difficulties of diagnosisThe diagnosis of CMS is not always
easy. Faced withsporadic CMS beginning after the neonatal period,
thediagnosis of seronegative autoimmune myasthenia maybe difficult
to eliminate, more especially as long periodsof remission are
possible in both afflictions and bouts canoccur in the adult CMS
patient during pregnancy [38].Muscle-specific receptor tyrosine
kinase (MuSK) anti-bodies have been detected in more thanhalf of
the patients presenting with seronegative(no acetylcholine receptor
antibodies) auto-immunemyasthenia [80,81 .]. In case of
uncertainty, the absenceof MuSK antibodies must be verified before
establishinga diagnosis of CMS.
Three recent observations have stressed that it issometimes
difficult to draw clear boundaries betweenautoimmune myasthenia and
CMS [82,87,88.]. The firstreported two sisters carrying
heteroallelic mutations ofthe acetylcholine receptor a subunit,
both presentingwith neonatal myasthenic syndrome, but one
developedautoimmune myasthenia as an adult, attested by
thetransitory presence of anti-acetylcholine receptor anti-bodies
and a favourable response to plasmaphereses andcorticotherapy [82].
The authors suggested that thegenetic anomaly of the acetylcholine
receptor couldconstitute the factor triggering autoimmune
myasthenia.The second observation concerned a patient
presentingwith acquired slow channel syndrome beginning at 30years
of age. The passive transfer of the serum of thispatient to a mouse
reproduced kinetic anomalies of theacetylcholine receptor, which
demonstrated its autoim-mune origin and excluded a congenital
affliction [87].The third observation reported an acetylcholine
recep-tor-seronegative, MuSK-seropositive myasthenia gravispatient,
in whom no acetylcholine receptor or MuSKdeficiency was found in
muscle biopsies despite theelectrophysiological impairment.
Mutation analysis ofMUSK did not reveal mutations but
polymorphisms. Theauthors concluded that circulating anti-MuSK
antibodiesmay not have caused the myasthenic syndrome in
thispatient [88.].
Phenotypegenotype correlations andprognosisThe genotype and
clinical phenotype are not correlatedin CMS. Mutations in different
synaptic proteins givesimilar clinical pictures: the occurrence of
apnoeicepisodes in early childhood was reported in CMSs witha
deficit in ChAT, in those caused by primary anomaliesof the
acetylcholine receptor, of the acetylcholinesteraseor of rapsyn.
Arthrogryposis has been described in CMScaused by mutations in the
gene encoding rapsyn [66.]and the acetylcholine receptor d subunit
[45]. The samemutation could lead to very different clinical
phenotypes:for example, the homozygous N88K rapsyn mutation
leads either to a very severe neonatal form or to a late-onset
and benign form [6,66 .]. Finally, variability within afamily was
noted in certain cases of CMS.
Prognosis is difficult to assess. A favourable outcome
ispossible in cases of CMS initially thought to be severebecause of
respiratory or bulbar bouts. In contrast, motorand respiratory
degradation occurring late in adulthoodhas been reported in
patients initially only slightlyaffected [27.]. As indicated above,
knowledge of theprimary molecular anomaly of CMS does not enable
theprediction of disease progression. The response totreatments
known to ameliorate neuromuscular transmis-sion is a significant
prognostic factor: thus in acetylcho-linesterase deficiency, the
absence of amelioration bycholinesterase inhibitors or any other
drug may bealarming.
TreatmentNon-specific measures are essential: immediate
treat-ment of respiratory distress, the prevention of infectionsand
of malnutrition as a result of swallowing disorders,and orthopaedic
surveillance of spinal complications andretractions. Drug
contraindications must be respected asfor any other myasthenic
syndrome. In the case of CMS,there is no reason to apply the
immunosuppressivetherapy used for myasthenia gravis.
Cholinesteraseinhibitors are efficient in all CMSs, with the
exceptionof slow channel syndrome and
acetylcholinesterasedeficiency, which they can even worsen. They
exert apreventative effect on the respiratory decompensationsof CMS
caused by ChAT mutations [4 ..]. 3,4-Diamino-pyridine, whose mode
of action is presynaptic, issometimes effective in pre- or
postsynaptic CMSs [39].Patients suffering from slow channel
syndrome benefitfrom the regulatory action of acetylcholine
receptorblockers: quinidine is effective by correcting theprolonged
opening of the acetylcholine receptor [89],but is formally
contraindicated in all the other forms ofCMS. A favourable effect
of fluoxetine was recentlydemonstrated in some patients, and is of
interest despitethe large amount needed [40 .]. At present, there
is nospecific treatment for acetylcholinesterase deficiency.
ConclusionAlthough the epidemiology of the CMSs is
poorlyunderstood, these disorders constitute the major causeof the
myasthenic syndrome in the young child and are aminor cause of
adult myasthenic syndrome. Thediagnosis is often difficult to
ascertain because of thefrequent absence of a family history of the
disease, andbecause of the pre-eminence of the myopathic
signscompared with myasthenic signs. The early onset of thefirst
symptoms, the presence of fluctuations, thedemonstration of a
neuromuscular block, repetitiveCMAP after single stimulation, and
the cholinesterase
Congenital myasthenic syndromes Hanta et al. 547
-
inhibitor test all enable the rectification of the diagnosisand
the proposal of an effective treatment and geneticcounseling. The
numerous studies devoted to CMS overmore than 20 years have
demonstrated the patho-physiological heterogeneity of CMS.
Characterizationof the CMS is based on the mode of transmission,
thesearch for a CMAP, the positive or negative response
tocholinesterase inhibitors, the study of motor endplates,which is
easily done on a deltoid muscle biopsy, andmolecular genetics. It
will thus be possible to identifythe majority of CMSs: a primary
anomaly of one of thevarious acetylcholine receptor subunits, of
rapsyn,acetylcholinesterase, ChAT or even SCN4A. However,the origin
of a significant fraction of CMSs remainsunknown. Numerous
molecules of the neuromuscularjunction are potential candidates for
CMS and may be
tested (Fig. 4) [90]. Therefore, in non-identified cases,various
investigations will be used: genetic linkageanalysis in the case of
large families, the demonstrationin the muscle biopsy of a
selective deficit in one givensynaptic molecule, or
microelectrophysiology of theintercostal muscle. Collaboration
between clinicians,morphologists, geneticists, and neurobiologists
is essen-tial for a complete characterization of the CMSs and
forthe understanding of the fundamental mechanisms ofneuromuscular
transmission based on human pathology.
AcknowledgementsWe thank Claire Legay and Hanns Lochmuller for
critical reading of themanuscript. This study was supported by the
Direction de la RechercheClinique de lAssistance Publique, Hopitaux
de Paris (PHRC #AOM01036), Association Francaise contre les
Myopathies, GIS-MaladiesRares, and Reseau Inserm de Recherche
Clinique.
Figure 4. Neuromuscular junction molecules that are or might be
involved in congenital myasthenic syndrome
-CGRP
acetylcholineChAT
2, 4, 2, 1 laminin 3, 4, 5 type IV collagenAChE Q,
Tagrinneuregulin
ankyrindystrophin
Na+ channel
AChR- , , , 7A, B, 1 integrin , dystroglycanMuSKErbB2, ErbB4
rapsynutrophin
plectin
K+ channelCa++ channel
ErbB3
synaptobrevinsynaptotagminsynaptophysin
Molecules with proved implication in congenital myasthenic
syndromes are indicated in bold letters. The molecules, the genes
of which are potentialcandidates, are indicated in normal letters.
AChE, Acetylcholinesterase; AChR, acetylcholine receptor; MuSK,
muscle-specific receptor tyrosinekinase. Adapted from Sanes and
Lichtman [90].
Neuromuscular disease: muscle548
-
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