J Neurol (2007) 254:1390–1394
DOI 10.1007/s00415-007-0557-0

Keiko Ohnari
Mitsuhiko Yamano
Takenori Uozumi
Tomoko Hashimoto
Sadatoshi Tsuji
Masanori Nakagawa

Received: 11 March 2006
Received in revised form:
14 December 2006
Accepted: 23 December 2006
Published online: 15 October 2007

K. Ohnari, MD (쾷) · M. Yamano ·
T. Uozumi · T. Hashimoto · S. Tsuji
Dept. of Neurology
University of Occupational
and Environmental Health
School of Medicine
Yahatanishi-ku, Kitakyushu, 807-8555,
Japan
Tel.: +81-93/6031611
Fax: +81-93/6939842
E-Mail: keiko-o@med.uoeh-u.ac.jp

ORIGINAL COMMUNICATION

An adult form of Alexander disease: a novel
mutation in glial fibrillary acidic protein

M. Nakagawa
Research Institute for Neurological
Diseases and Geriatrics
Dept. of Neurology and Gerontology
Kyoto Prefectural University of Medicine
Kawaramachi Hirokoji 465
Kamigyo-ku, Kyoto 602-0841, Japan

■ Abstract Glial fibrillary acidic
protein (GFAP) mutation has been
reported in Alexander disease. We
report a patient with the adult form
of Alexander disease who shows a
novel mutation in GFAP. This case
presented with progressive
dysarthria, dysphagia and spastic
gait on the right side. Brain and
spinal cord MRI showed marked
atrophy of the medulla oblongata
and spinal cord. Abnormal high
signal intensities in the ventral
medulla oblongata were detected

JON 2557

Introduction
Alexander disease is a leukodystrophy that is pathologically characterized by astrocytic inclusion known as
Rosenthal fibers [2]. The clinical symptoms and pathological findings differ depending on age at onset. The infantile form usually presents before 2 years of age, showing macrocephaly, motor and mental retardation, bulbar
dysfunction and seizure, and leads to death before 10
years of age. The juvenile form usually presents between
2 and 12 years of age with bulbar and pseudobulbar
palsy, spastic paresis and cerebellar ataxia. Mental function is normal, and the course progresses more slowly
than that of the infantile form. The symptoms of the

bilaterally. There were no white
matter lesions or contrast enhancing lesions. Recently, there have
been reports of patients with a juvenile form of Alexander disease
presenting with atrophy or signal
abnormalities of the medulla or
spinal cord. Atrophy of the medulla
and spinal cord have specifically
been described as suggestive of
Alexander disease [1]. Sequence
analysis of the GFAP gene of this
patient showed a heterozygous
c.221T > C mutation, predicting a
p.M74T amino acid change. In all
patients suspected of Alexander
disease on the basis of MRI findings, GFAP analysis is necessary to
confirm the diagnosis.
■ Key words Alexander disease ·
GFAP

adult form are similar to those of the juvenile form, but
palatal myoclonus is often present, and onset occurs
later. Bulbar symptoms are a prominent clinical feature
at all ages.
Rosenthal fibers contain GFAP, small heat shock protein HSP27 and β-crystallin [3]. Recently, mutations in
the GFAP gene were identified in Alexander disease [4].
These mutations are believed to cause Alexander disease, but how they produce the disease is not presently
understood. In studies on parents of affected children,
no parent has been found to have the GFAP mutation
that is present in their offspring. Thus in most cases the
disorder is not inherited, but arises spontaneously for
unknown reasons. However, not every patient with
proven Alexander disease has an identified mutation in

1391

GFAP, so that there may be other genetic or perhaps
even non-genetic causes that have yet to be identified
(http://www.waisman.wisc.edu/alexander).
We report a patient with the adult form of Alexander
disease, who showed a novel mutation in GFAP.

Patient and methods
A 53-year-old man presented with dysarthria at the age of 51. Two
years later, he developed dysphagia and gait disturbance due to spasticity in his right leg. He was referred to our hospital because of progression of dysarthria, dysphagia and gait disturbance. Neurological
examination showed a normal mental state. Dysarthria and dysphagia were noted. Tendon reflexes were increased in all extremities with
right-side dominance, but pathological reflexes were negative. He
demonstrated a spastic gait involving the right limb. There was no
muscle atrophy or fasciculations and no palatal myoclonus or cerebellar ataxia. Sensory and autonomic systems were normal. There was
no family history of similar disease.
Blood cell count, routine chemistries, syphilic serology, thyroid
function, autoantibodies and adrenal function were normal. The
cerebrospinal fluid study was normal. Central motor conduction
times recorded in the bilateral abductor pollicis brevis muscles and
abductor hallicis muscle produced by transcranial magnetic stimulation was much longer than that in normal subjects. Auditory brainstem response, peripheral nerve conduction studies and needle electromyogram were normal.
Brain and spinal cord MRI showed marked atrophy of the medulla
oblongata and spinal cord. Abnormal high intensities in the ventral
medulla oblongata were detected bilaterally. There were no white
matter lesions in the deep white matter and there were no contrast enhancing lesions (Fig. 1).
We performed a sequence analysis of the GFAP gene, because
Alexander disease was considered in the differential diagnosis due to
the clinical symptoms and brain MRI findings.

■ Genetic analysis
After informed consent was obtained from the patient, genomic DNA
was extracted from the peripheral blood. Using genomic DNA, we
screened the genes responsible for spinocerebellar ataxia (SCA) 1, 2,
6, 7, 8, Friedrich ataxia, and Machado-Joseph disease (MJD) using
polymerase chain reaction (PCR) with the primers as described in
previous reports [5–7]. The 9 exons in GFAP were sequenced by the
cycle sequence method (ABI PRISM Dye Terminator Cycle Sequencing Kit) with primer sets that were designed based on the GFAP sequence published in AceView (http://www.ncbi.nlm.nih.gov/AceView/), then analyzed using an autosequencer (ABIPRISM310).

Results
Direct sequencing of GFAP showed a heterozygous point
mutation, i. e. a transition involving a change from T to
C at nucleotide position 235, predicting an M74T amino
acid change (Fig. 2A). We made a mismatch primer (5’cgggccagtga gcgggcagagtcga –3’), which produces a new
recognition site for SalI in the mutant allele, to confirm
the substitution detected by the direct sequencing. Using this mismatch primer, we confirmed the substitution
in the patient and did not find the same substitution in
the DNAs of 100 SCA patients and 100 normal controls
(Fig. 2B). No abnormal expansion on the triplet repeats
was detected in the responsible genes of SCA 1, 2, 6, 7, 8,
Friedrich ataxia, and MJD.

Discussion
This patient presented with progressive dysarthria, dysphagia and spastic gait on the right side. The neurophysiological findings showed dysfunction of the bilateral pyramidal tract, but there was no dysfunction of the

Fig. 1 Brain and spinal cord MRI findings. MRI did not demonstrate any white
matter lesion (A), but there were bilateral abnormal high intensities in the
ventral medulla oblongata (B, C, D) and
marked atrophy of the medulla oblongata and spinal cord (E)

1392

lower motor neurons. GFAP gene analysis identified the
substitution of 221T > C, which resulted in amino acid
alteration M74T. The methionine directly next to the site
of the mutation has been found to be mutated in patients
with Alexander disease: M73T and M73R [8, 9]. The
amino acid is conserved in all species (Fig. 2C). These
findings support the presumption that M74T is a pathogenic mutation.The locations of Alexander disease-associated mutations in GFAP have been previously reported (Fig. 3). Mutations at K63, V87, E210, E223, R276
and R416 have been identified in the adult form of
Alexander disease. We reported a case presumed to be
the adult form of Alexander disease with a new heterozygous point mutation in GFAP.
Other GFAP mutations in cases showing the adult
form of Alexander disease have been reported (Table 1)
[8, 10–16]. The onset age ranged from 21 to over 50. Bulbar or pseudobulbar signs, palatal myoclonus, pyramidal sign and cerebellar ataxia are often seen and there
have been previous reports describing the adult form of
Alexander disease.Very few of the patients showed atrophy or fasciculations of the tongue. Neuropathological
examination showed well-preserved neurons in the hypoglossal nuclei and intramedullary roots despite severe
atrophy of the medulla oblongata. Therefore, the course
of dysarthria and dysphagia is considered due to
pseudobulbar signs [11]. All patients showed a normal
mental state and there were no seizures. Six patients
with the D78E mutation who were reported by Stumpf
et al. presented with dysautonomia and sleep apnea [16].
Previously, there have been reports that the genotype
is related to the phenotype [17]. Rodriguez reported that
cases showing mutations at the R239 locus present with
severe symptoms in infantile form of Alexander disease.
However, the patients with the R79C mutation did not

P. Met74 Thr
c. 22IT>C
A

T

G

A

T/C

G

G

A

G
250

c

T

c

c

A/G

T

c

A

T
180

Forward
sequence

Aa
Reverse
sequence

bp
300
200

b
B

Cc

Marker

pt

Human
Mus musculus
Rattus norvegicus
Zebrafish

C

C

C

C

C

SERAEM M ELNDRF
SERAEM M ELNDRF
SERAEM M ELNDRF
NEKVEM M GLNDRF

Fig. 2 DNA reverse sequence of GFAP exon 1 in the patient. The T-to-C transition
detected in the heterozygous patient results in conservative amino acid change
(p.M74T). A This heterozygous substitution was detected in the patient, but not in
100 normal controls using the mismatch primer that produced a new SalI recognition site in the mutant allele (B). C control. The mutation at M74 preserve in all
species (C)

Fig. 3 The locations of Alexander disease-associated mutations in GFAP [8]. The M74T we reported is
a new heterozygous point substitution in GFAP. Mutations at K63, M74, V87, E210, E223, E276 and R416
have been showed in the adult form of Alexander disease

GFAP
1

2

3

4

5

6

7

8

9

gene

200bp

Head

protein
K63Q

1
M73R
M73T
M74T
L76F
L76V
N77S
N77Y
D78E

Tail

Rod
72

L97P
R258P

L90P
V87G
R79H
R79C
R79L
R79G

R88C
R88C
R88S

E207K
E207Q
E210K
E223Q

R239H
R239C
R239P
R242D
Y242D
R244V
A244D

R276L
K279E

L331P
L352P

377

432

HL349-50ins
L352p
L359V
E362D
A364P
Y366H
E373K
E373Q
E374G

R416W
N386S

1393

Table 1 Clinical symptoms and brain MRI findings in cases of adult onset Alexander disease with GFAP mutation
Authors

Okamoto

Namekawa
Kinoshita
Brockmann
Thyagarajan
Li
Stumpf

This case

Onset
age/sex

53/F
27/F
32/F
33/M
48/M
24/M
40/M
37/F
21/F
24/F
55/F
> 50/M
> 20/F
35/F
33/M
> 20/F
51/M

Mutation

V87G
V87G
V87G
R276L
R276L
R416W
E223Q
R416W
K63Q
E210K
D78E
D78E
D78E
D78E
D78E
D78E
M74T

Clinical symptoms

MRI

Reference

Bulbar or
Pseudobulbar
sign

Palatal
Myoclonus

Pyramidal
sign

Ataxia

White
matter
lesion

+
–
–
+
–
–
+
+
+
+
+
+
+
+
+
+
+

+
+
–
–
–
+
–
+
–
+
*

+
+
+
+
+
+
+
–
+
–
+
+
–
–
+
–
+

+
+
–
–
–
+
+
+
+
+
+
+
–
+
+
–
–

+
+
+
+
+
+
–
+
–
+
+
+
+
–
+
+
–
+
–
+
–
+
–
+
not done
–
+
–
+
not done
–
+

–

Bulbar
atrophy

Spinal
cord
atrophy
+
+
+
+
+
+
–
+

–
+

10

11
12
13
14
8, 15
16

+
–
+

* Palatal myoclonus was found in only one patient in six patients reported by Stumpf et al. The patient with palatal myoclonus was not designated in the text

show macrocephaly and survived longer than those with
the R239 mutation. In several cases, however, the same
mutation produced different symptoms. For example
the R416W mutation was reported in infantile, juvenile
and adult forms of Alexander disease. Two cases of the
adult form of Alexander disease with R416W GFAP gene
mutation were reported. One case showed palatal myoclonus, pyramidal sign, cerebellar ataxia and macrocephaly [12]. The other case presented with the same
symptom along with dysarthria, but brain MRI did not
show macrocephaly [14]. These cases indicate that other
factors may also influence the phenotype.
Alexander disease is characterized pathologically by
the accumulation of intracytoplasmic inclusion within
astrocytes that are known as Rosenthal fibers. The
pathology of the infantile form involved abundant
Rosenthal fibers in the perivascular, subpial and
subependymal regions. Definitive diagnosis is established by these pathological findings. Recently five brain
MRI criteria for Alexander disease were defined: 1) extensive cerebral white matter abnormalities with a
frontal preponderance, 2) presence of a periventricular
rim, 3) abnormalities of basal ganglia and thalamus, 4)
brain stem abnormalities, 5) contrast enhancing lesion.
And four of five MRI criteria must be met for an MRIbased diagnosis of Alexander disease [18]. Signal abnormality of the white matter is due to not only hypomyelination but also hyperplasia and hypertrophy of
astrocytes and Rosenthal fiber deposition. Brain MRI

criteria were in close agreement with the histopathologic findings of the infantile form of Alexander disease.
However, MRI of the adult form of Alexander disease
does not show typical MRI findings. Adult patients presented with marked atrophy of the medulla oblongata
and spinal cord.White matter abnormalities were not always observed. The brain MRI of our patient showed
marked atrophy of the medulla oblongata and spinal
cord and abnormal high intensities in the ventral
medulla oblongata, and only one of five criteria was met.
Symmetrical abnormal intensities on the ventral
medulla oblongata may be related to the clinical symptom of pyramidal sign, but this has not been reported in
the adult form of Alexander disease. The criteria for
Alexander disease do not include spinal cord atrophy.
We consider that the above MRI criteria constitute an
unsatisfactory diagnostic tool for all forms of Alexander
disease, because later papers on MRI in Alexander disease have shown other MRI abnormalities that are not
part of the above MRI criteria, which in themselves
could be the basis of new MRI criteria. In a recent article, brain MRI of juvenile onset Alexander disease patients demonstrated signal abnormalities or atrophy of
the medulla or spinal cord [1]. One patient had only minor cerebral white matter abnormality. The conclusion
of this paper is that the presence of atrophy or signal abnormalities in the medulla or spinal cord is sufficient to
warrant analysis of GFAP.
Here we reported a patient with the adult form of

1394

Alexander disease showing a novel mutation in GFAP. A
diagnosis of Alexander disease was established by neuropathological findings of the brain. If brain biopsy cannot be performed, brain MRI findings may be useful to
diagnose the infantile form of Alexander disease. In the

adult form of Alexander disease, however, patients often
show atypical findings on brain MRI. Therefore we consider that sequence analysis of the GFAP gene was a useful method of diagnosing Alexander disease.

References
1. Van der Knaap MS, Ramesh V, et al.
(2006) Alexander disease Ventricular
garlands and abnormalities of medulla
and spinal cord. Neurology 66:494–498
2. Alexander WS (1949) Progressive fibrinoid degeneration of fibrillary astrocytes associated with mental retardation in a hydrocephalic infant. Brain
72:373–381
3. Iwaki T, Iwaki A, et al. (1993) αB-crystallin and 27-kd heat shock protein are
regulated by stress condition in the
central nerve system and nervous
system and accumulate in Rosenthal
fibers. Am J Pathol 143:487–495
4. Brenner M, Johnson AB, et al. (2001)
Mutation in GFAP, encoding glial
fibrillary acidic protein, are associated
with Alexander disease. Nat Genet 27:
117–120
5. Kawaguchi Y, Okamoto T, et al. (1994)
CAG expansions in a novel gene for
Machado-Joseph disease at chromosome 14q32.1. Nat Genet 8:221–227
6. David G, Abbas N, et al. (1997) Cloning
of the SCA7 gene reveals a highly unstable CAG repeat expansion. Nat
Genet 7:65–70

7. Sasaki H, Yabe I, et al. (2000) Prevalence of triplet repeat expansion in
ataxia patients from Hokkaido, the
northernmost island of Japan. J Neurol
Sci 175:45–51
8. Li R, Johnson A, et al. (2005) Glial
fibrillary acidic protein mutation in
infantile, Juvenile, and adult forms of
Alexander disease. Ann Neurol 57:
310–326
9. Gorospe JR, Naidu S, et al. (2002) Molecular findings in symptomatic and
pre-symptomatic Alexander disease
patients. Neurology 58:1494–1500
10. Okamoto Y, Mitsuyama H, et al. (2002)
Autosomal dominant palatal myoclonus and spinal cord atrophy.
J Neurol Sci 195:71–76
11. Namekawa M, Takiyama Y, et al. (2002)
Identification of GFAP gene mutation
in hereditary adult-onset Alexander’ s
disease. Ann Neurol 52:779–785
12. Kinoshita T, Imaizumi T, et al. (2003) A
case of adult-onset Alexander disease
with Arg416Trp human glial fibrillary
acidic protein gene mutation. Neurosci
Lett 350:169–172

13. Brockmann K, Meins M, et al. (2003) A
novel GFAP mutation and disseminated white matter lesion: adult
Alexander disease? Eur Neurol 50:
100–105
14. Thyagarajan D, Chataway T, et al.
(2004) Dominantly-inherited adultonset Leukodystrophy with palatal
tremor caused by a mutation in the
glial fibrillary acidic protein gene. Mov
Disord 19:1244–1248
15. Van der Knaap MS, Salomons G, et al.
(2005) Unusual variants of Alexander’s
disease. Ann Neurol 57:327–338
16. Stumpf E, Masson H, et al. (2003) Adult
Alexander disease with autosomal
dominant transmission. Arch Neurol
60:1307–1312
17. Rodriguez D, Gauthier F, et al. (2001)
Infantile Alexander disease: spectrum
of GFAP mutations and genotype-phenotype correlation. Am J Hum Genet
69:1134–1140
18. Van der Knaap MS, Naidu S, et al.
(2001) Alexander disease: diagnosis
with MR imaging. Am J Neuroradiol
22:541–552