Genetic prion disease: the EUROCJD experience

Abstract

A total of 10-15% of human transmissible spongiform encephalopathies (TSEs) or prion diseases are characterised by disease-specific mutations in the prion protein gene (PRNP). We examined the phenotype, distribution, and frequency of genetic TSEs (gTSEs) in different countries/geographical regions. We collected standardised data on gTSEs between 1993 and 2002 in the framework of the EUROCJD collaborative surveillance project. Our results show that clinicopathological phenotypes include genetic Creutzfeldt-Jakob disease (gCJD), fatal familial insomnia (FFI), and Gerstmann-Sträussler-Scheinker disease (GSS). Genetic TSE patients with insert mutation in the PRNP represent a separate group. Point and insertional mutations in the PRNP gene varies significantly in frequency between countries. The commonest mutation is E200K. Absence of a positive family history is noted in a significant proportion of cases in all mutation types (12-88%). FFI and GSS patients develop disease earlier than gCJD. Base pair insertions associated with the Creutzfeldt-Jakob disease (CJD) phenotype, GSS, and FFI cases have a longer duration of illness compared to cases with point mutations and gCJD. Cerebrospinal fluid 14-3-3 immunoassay, EEG, and MRI brain scan are useful in the diagnosis of CJD with point mutations, but are less sensitive in the other forms. Given the low prevalence of family history, the term "gTSE" is preferable to "familial TSE". Application of genetic screening in clinical practice has the advantage of early diagnosis and may lead to the identification of a risk of a TSE.

Full text

ORIGINAL INVESTIGATION
Ga
´
bor G. Kova
´
cs Æ Maria Puopolo Æ Anna Ladogana
Maurizio Pocchiari Æ Herbert Budka
Cornelia van Duijn Æ Steven J. Collins Æ Alison Boyd
Antonio Giulivi Æ Mike Coulthart
Nicole Delasnerie-Laupretre Æ Jean Philippe Brandel
Inga Zerr Æ Hans A. Kretzschmar
Jesus de Pedro-Cuesta Æ Miguel Calero-Lara
Markus Glatzel Æ Adriano Aguzzi Æ Matthew Bishop
Richard Knight Æ Girma Belay Æ Robert Will
Eva Mitrova
Genetic prion disease: the EUROCJD experience
Received: 18 March 2005 / Accepted: 15 June 2005 / Published online: 27 September 2005
Ó Springer-Verlag 2005
Abstract A total of 10–15% of human transmissible
spongiform encephalopathies (TSEs) or prion diseases
are characterised by disease-specific mutations in the
prion protein gene (PRNP). We examined the pheno-
type, distribution, and frequency of genetic TSEs
(gTSEs) in different countries/geographical regions. We
collected standardised data on gTSEs between 1993 and
2002 in the framework of the EUROCJD collaborative
I. Zerr
Department of Neurology, Georg-August-Universita
¨
tGo
¨
ttingen,
Robert-Koch Strasse 40, 37075 Gottingen, Germany
H. A. Kretzschmar
Institute of Neuropathology, University of Munich,
Marchioninistr. 17, 81377 Munich, Germany
J. de Pedro-Cuesta
Departamento de Epidemiologia Aplicada, Instituto de Salud
Carlos III, Centro Nacional de Epidemiologia, Calle Sinesio
Delgado 6, 28029 Madrid, Spain
M. Calero-Lara
Centro National de Microbiologia Unidad de Encefalopatias
Espongiformes, Ctra Majadakonda—Pozuelo km2,
28220 Majadakonda, Madrid, Spain
M. Glatzel Æ A. Aguzzi
Swiss National Reference Centre for Prion Diseases, University
Hospital of Zurich, Schmelzbergstrasse 12,
CH-8091 Zurich, Switzerland
M. Bishop Æ R. Knight
National CJD Surveillance Unit, Western General Hospital,
Edinburgh, EH4 2XU UK
G. Belay Æ R. Will Æ E. Mitrova (&)
Institute of Preventive and Clinical Medicine, Research Base of
Slovak Medical University, National Reference Centre of prion
Diseases, Limbova 14, 833 01 Bratislava, Slovakia
E-mail: eva.mitrova@szu.sk
Tel.: +421-2-59-36-9564
Fax: +421-2-59-36-9585
Ga
´
bor G. Kova
´
cs and Maria Propolo Contributed equally
G. G. Kova
´
cs Æ H. Budka
Austrian Reference Centre for Human Prion Diseases (OERPE)
and Institute of Neurology, Medical University AKH 4J,
Waehringer Guertel 18-20, 1097 Vienna, Austria
M. Puopolo Æ A. Ladogana Æ M. Pocchiari
Department of Cell Biology and Neurosciences,
Istituto Superiore di Sanita
`
, Viale Regina Elena 299, 00161
Rome, Italy
C. van Duijn
Department of Epidemiology and Biostatistics, Erasmus MC,
PO Box 1738, 3000 DR, Rotterdam, The Netherlands
S. J. Collins Æ A. Boyd
Department of Pathology, The University of Melbourne, Parkville,
Victoria, 3052 Australia
A. Giulivi
Blood Safety Surveillance and Health Care Acquired Infections
Division, The Centre for Infectious Disease Prevention
and Control, LCDC Building, PL 0601E2,
Tunney’s Pasture, Ottawa, ON,
K1A 0L2 Canada
M. Coulthart
National Laboratory for Host Genetic and Prion Diseases,
NML, PHAC, Health, Winnipeg, MB, Canada
N. Delasnerie-Laupretre Æ J. P. Brandel
U.360 INSERM, Hopital de la Salpetriere, 75651 Paris,
Cedex 13, France
Hum Genet (2005) 118: 166–174
DOI 10.1007/s00439-005-0020-1

surveillance project. Our results show that clinicopath-
ological phenotypes include genetic Creutzfeldt–Jakob
disease (gCJD), fatal familial insomnia (FFI), and
Gerstmann–Stra
¨
ussler–Scheinker disease (GSS). Genetic
TSE patients with insert mutation in the PRNP repre-
sent a separate group. Point an d insertional mutations in
the PRNP gene varies significantly in frequency between
countries. The commonest mutation is E200K. Absence
of a positive family history is noted in a significant
proportion of cases in all mutation types (12–88%). FFI
and GSS patients develop disease earlier than gCJD.
Base pair insertions associated with the Creutzfeldt–Ja-
kob disease (CJD) phenotype, GSS, and FFI cases have
a longer duration of illness compared to cases with point
mutations and gCJD. Cerebrospinal fluid 14-3-3
immunoassay, EEG, and MRI brain scan are useful in
the diagnosis of CJD with point mutations, but are less
sensitive in the other forms. Given the low prevalence of
family history, the term ‘‘gTSE’’ is preferable to
‘‘familial TSE’’. Application of genetic screening in
clinical practice has the advantage of early diagnosis and
may lead to the identification of a risk of a TSE.
Keywords Prion protein gene Æ Creutzfeldt-Jakob
disease Æ Fatal familial insomnia Æ Gerstmann-
Stra
¨
ussler-Scheinker disease Æ Point
mutation Æ Insertional mutation
Introduction
Human transmissible spongiform encephalopathies
(TSEs) or prion diseases are characterised by neuro log-
ical and psychiatric symptoms and a progressive fatal
course. Accumulation in the central nervous system
(CNS) of the pathological prion protein (PrP
Sc
)isa
common disease marker (Prusiner 2001). The most fre-
quent human TSE is Creutzfeldt–Jakob disease (CJD).
The majority of cases (85%) present as a sporadic dis-
order (sCJD) without defined aetiology (Masters et al.
1979; WHO 2003). Acquired forms, including iatrogenic
CJD due to transmission of infection in the course of
medical or surgical treatment, and variant CJD (vCJD),
which has been linked to infection wi th bovine spongi-
form encephalopathy, are less frequent (Masters et al.
1979; Prusiner 2001;WHO2003). The familial occur-
rence of cases has been reported with a broad range of
frequency: 6% of TSE cases in France (Brown et al.
1979), 25.5% in Israel (Kahana et al. 1974), 26% in Chile
(Galvez et al. 1980), 53.6% in Slovakia (Mitrova and
Belay 2002), and world-wide 15% (Masters et al. 1979).
Current clinical and neuropathological diagnostic
criteria distinguish familial or genetic TSEs (gTSEs),
including familial/genetic CJD (gCJD), fatal familial
insomnia (FFI), and Gerstmann–Stra
¨
ussler–Scheinker
disease (GSS) (Budka et al. 1995; Prusiner 2001; WHO
2003). Experimental transmissibility of all the major
subtypes has been established, albeit not with all muta-
tions (Masters et al. 1981; Tateishi et al. 1979, 1995). In
gTSEs, disease-specific point or insertional mutations in
the prion protein gene (PRNP) have been demonstrated
(Goldfarb et al. 1990a, b, 1992; Goldgaber et al. 1989;
Haltia et al. 1991; Owen et al. 1989), with individual
PRNP mutations showing variable geographical distri-
bution and frequency. While certain mutations are rare
(Kovacs et al. 2002), the E200K mutation has been re-
ported not only in Europe but also in Chile, Israel, Japan,
and USA (Goldfarb et al. 1990a, b; Goldgaber et al.
1989; Miyakawa et al. 1998). Geographic or ethnic clus-
ters of cases of gTSEs have been found in Israel, Slovakia,
Chile, and Italy (Chapman et al. 1994; D’Alessandro
et al. 1998; Kahana et al. 1974; Mayer et al. 1977;
Mitrova and Belay 2002; Mitrova and Bronis 1991).
In parallel with the increasing numbers of PRNP
mutations, it has been recognised that, unexpectedly, not
all patients with PRNP mutations appear to have affected
family members (Chapman et al. 1994; D’Alessandro
et al. 1998; EuroCJD group 2001; Goldman et al. 2004;
Mitrova and Belay 2002). Therefor e the terms ‘‘familial’’,
‘‘hereditary’’, or ‘‘inherited’’ TSE may not be appropriate
in cases without a family history and the inclusive term
‘‘genetic TSE’’ may be preferable for cases associated
with a mutation, whether or not there is a family history.
The negative family history in some gTSE cases and
the identification of ‘‘healthy’’ carriers has drawn
attention to the issue of penetrance, i.e., the proportion
of carriers who will eventually develop the disease
(Chapman et al. 1994; D’Alessandro et al. 1998; Gold-
farb et al. 1990a, b; Mitrova and Belay 2002). Pene-
trance of the E200K mutation shows considerable
variability. While in Israeli carriers, penetrance appears
to be almost complete (89%), in Slovakian and Italian
E200K carriers, penetrance is partial (54–59%) (Chap-
man et al. 1994; D’Alessandro et al. 1998; Goldfarb
et al. 1990a, b; Mitrova and Belay 2002 ).
The EUROCJD project, funded by the European
Commission, started in 1993 and compares data from
national registries in Australia, Austria, Canada,
France, Germany, Italy, The Netherlands, Slovakia,
Spain, Switzerland, and the UK. The present study
analyses the EUROCJD data on gTSE cases. Recently
clinicopathological data of more than 500 published
genetic cases of the literature has been reviewed (Kovacs
et al. 2002) and some of the data on genetic epidemiol-
ogy of CJD in Europe has been published (EuroCJD
group 2001). The present paper includes standardised
data from a large number of cases not previously re-
ported in the context of a study carried out over a period
of yea rs in a defined population. Our aims are: (1) to
describe the distribution and frequency of gTSEs, (2) to
define and describe specific features of subgroups of
gTSEs, (3) to contribute to a better understanding on
how mutations and polymorphisms of PRNP influence
clinical phenotypes, and (4) to assess the terms genetic
and familial according to epidemiological and molecular
biological data.
167

Patients and methods
The EUROCJD database contains data on sporadic,
variant, iatrogenic, and gTSE cases collected between
1993 and 2002 (for detailed methodology see Ladogana
et al. 2005a). In this paper we have analysed data of 455
gTSE cases from the following groups: gCJD cases
(including patients with PRNP analysis and those with
no available PRNP analysis but positive family history
for TSE), FFI, GSS, and gTSE patients carrying base
pair insertions (insert gTSE): referred to in previous
publications as BPI (Kovacs et al. 2002) because these
mutations are associated with repeat base pair insertions
in the octarepeat region. No deletions in this region were
identified in this study. We evaluated E200K gCJD and
V210I gCJD separately as there were sufficient numbers
of cases to allow statistical analyses. Cases of gCJD
without either an E200K or V210I mutation are classi-
fied as ‘‘other CJD’’. Cases were classified either as
definite or probable according to recent surveillance cri-
teria (WHO 2003), which for the diagnosis of ‘‘familial’’
CJD require definite or probable CJD in the index case
and a first-degree relative or a neuropsychiatric disorder
in association with a PRNP mutation.
Statistical analysis
Differences among distinct forms of gTSE cases with
respect to age at onset or clinical duration were assessed
by Mann–Whitney test; variation in clinical signs by the
chi-square test or the Fisher’s exact probability test. The
Bonferroni correction for multiple testing was ad opted
within the six subgroups of gTSE (E200K, V210I, and
other forms of gCJD, GSS, FFI, and insert mutations)
at an experimental probability type 1 error of 0.05.
Age at onset was given as mean, standard deviation
(SD), and range; clinical duration as median and half
interquartile range (IQR) because of highly skewed
distribution of data. Box-plots were used for the graphic
representations of continuous variables. Statistical
analyses were performed using BMDP and STATA.
Crude and sex-specific mortality rates were calculated
using as denominator populations data for 1998 pro-
vided from National or Federal Statistics Bureau of
participating countries. For Canada, population data
(2001) were taken from the Canadian Statistics website.
The annual mortality rates were calculated using data
collected from 1999–2002 when the surveillance system
was well established in all countries.
Results
Distribution of cases and mutations
The distribution and frequency of PRNP mutations
among countries participating in the study are sum-
marised in Table. 1 and 2. The proportion of all gTSE
cases with respect to the total number of TSE cases
(including sporadic, iatrogenic, and variant CJD) was
10.2%, but, this varied significantly among participating
countries ranging from 69.5% in Slo vakia to 1.2% in
Switzerland. The overall annual mortality rate of gTSE
cases was 0.17 patients per million people for the period
1999–2002. It is of note that Slovakia had an overall
mortality rate for gTSE diseases 3.5 times that of the
second highest country (Italy). However, while the
E200K mutation was the only mutation present in the
Slovak population, several mutations were present in the
Italian population, including the E200K and V210I
gCJD, FFI, and GSS. Switzerland reported a single
Table 1 EUROCJD 1993–2002: number of reported cases in each EUROCJD country
All genetic TSE diseases gCJD GSS FFI Insert
n gTSE (%) gTSE (%)
with respect
to all TSE
in each
country
Mortality rates
(million people)
a
Australia 22/215 4.8 10.2 0.14 14 3 4 1
Austria 13/90 2.9 14.4 0.28 9 0 3 1
Canada 16/189 3.5 8.5 0.12 7 9 0 0
France 84/938 18.5 9.0 0.18 68 5 6 5
Germany 68/900 14.9 7.6 0.13 31 8 17 12
Italy 115/662 25.3 17.4 0.30 94 8 10 3
Netherlands 3/142 0.7 2.1 0.02 1 0 0 2
Slovakia 41/59 9.0 69.5 1.07 41 0 0 0
Spain 44/429 9.7 10.3 0.23 18 0 25 1
Switzerland 1/85 0.2 1.2 0.04 1 0 0 0
UK 48/732 10.5 6.6 0.07 11 19 1 17
Total 455/4,441 – – 0.17 295 52 66 42
% Total – 100 10.2 – 64.9 11.4 14.5 9.2
Abbreviations: gTSE, genetic transmissible spongiform encephalopathy; gCJD, genetic Creutzfeldt–Jakob disease; FFI, fatal familial
insomnia; GSS, Gerstmann–Stra
¨
ussler–Scheinker disease; Insert, gTSE patients with insert mutations.
a
For the period 1999–2002.
168

gTSE patient for the period 1996–2002 while the Neth-
erlands had the lowest mortality rate for gTSE diseases.
More than 90% of FFI and 75% of GSS patients
underwent post-mortem examination and were classified
as definite gTSE cases, while 52% of V210I gCJD pa-
tients did not undergo autopsy (see Table 3) and were
classified as probable gTSE based on clinical and labo-
ratory features. There was an excess of females in gCJD
(more pronounced for E200K than V210I or ‘‘other’’
gCJD patients) and, to a lesser extent, in GSS or insert
gTSE cases, and an excess of males in FFI cases (Ta-
ble 3). Data on the polymorphic codon 129 of the PRNP
gene were available in 87% of all cases (95% in E200K,
97% in V210I, 75% in ‘‘other’’ gCJD, 95% in FFI, 56%
in GSS, and 79% in insert gTSE). The distribution of the
polymorphic codon 129 in all forms of gTSE is shown in
Table 3. Differences between codon 129 distribution in
controls (39% methionine homozygous (MM), 50%
heterozygous (MV), 11% valine homozygous (VV), Al-
perovitch et al. 1999) and in individual gTSE groups
have been evaluated by chi-sqared test with the follow-
ing results: E200K gCJD, V210I gCJD and FFI,
P<0.0001, ‘‘other’’ gTSE, P=0.0010, Insert gTSE,
P=0.0577 and GSS, P=0.3825. The majority of PRNP
mutations co-segregate with methionine at the poly-
morphic codon 129 (see Table 2). Thus, the majority of
gTSE patients overall were either MM (67.9%) or MV
(25.8%) with only a few valine homozygotes (6.3%).
Family history
About 47% of all gTSE cases were reported to have no
TSE or other neurological disorder in family members.
Almost 90% of V210I gCJD patients had a negative
family histor y implying that a correct classification of
these cases would not have been possible without PRNP
genetic analysis. Interestingly, a positive family history
for TSE was reported in only about two-thirds of GSS
patients (Table 3).
Age at onset
There were significant differences in the age at onset in
gTSE forms (Fig. 1a). Patients with FFI (mean
51.2 years, SD 12.3, range 19–83) and GSS (51.6, 12.8,
26–87) developed disease significantly earlier than
E200K gCJD (60.4, 10.2, 33–84, p<0.0001 ), V210I
gCJD (59.3, 9.8, 39–82, p=0.0001 and 0.0009, respec-
tively), and ‘‘other’’ gCJD cases (60.4, 14.7, 31–87,
p=0.0004, 0.0036). There was no significant difference
in the age at onset between FFI and GSS patients and
among distinct forms of gCJD. Age at onset in patients
with insert mutations (57.2, 14.8, 32–85) did not signif-
icantly differ from that of gCJD, GSS, or from FFI
(p=0.0193) after Bonferroni’s correction. The youngest
age at onset was 19 years (FFI, MM at codon 129),
while the two oldest were 87 years (gCJD with missing
information on PRNP mutation, and A117V GSS, VV
at codon 129). The age at onset was earlier in valine
homozygotes in comparison to methionine homozygotes
(P=0.0001) and, to a lesser extent, in MV vs MM
(P=0.0059, not significant after Bonf erroni’s correction)
in ‘‘other’’ gCJD patients. The codon 129 polymorphism
did not significantly influence the age at onset in all the
other forms of gTSEs (Table 4). Gender did not have
any significant effect on the age at onset (data not
shown).
Duration of illness
The duration of disease varied between different forms
of gTSE (Fig. 1b). Clinical durations of E200K (median
5.0 months, IQR 2.5) and V210I (4.0, 1.5) gCJD
patients were shorter than ‘‘other’’ gCJD (7.0, 7.0,
p=0.0031 and 0.0012, respectively), FFI (12.4, 4.2,
P<0.0001 and 0.0001), GSS (40.0, 25.0, P<0 .0001 and
0.0001) and insert gTSE (13.0, 35.5, P=0.0001 and
0.0001). GSS patients had a longer survival than FFI
(P<0.0001) and ‘‘other’’ gCJD (P<0.0001) patients.
Table 2 EUROCJD 1993–2002: number of reported cases and distribution of PRNP mutations in each EUROCJD country
Country gCJD
P105T-129? N171S-129V D178N-129V V180I-129M T188A-129M E196K-129M/V E200K-129M/V V203I-129M R208H-129M V210I-129M E211Q-129M NS
Australia 1 – – – 1 – 8 – – 2 – 2
Austria – – 1 – – 1 5 – – 1 – 1
Canada – – 2 – – – 4 1 – – – –
France – 1 8 1 – 1 46 3 – 6 2 –
Germany – – – – 2 3 15 – 1 9 1 –
Italy – – – – – – 35 1 1 50 1 6
Netherlands – – 1 – – – – – – – – –
Slovakia – – – – – – 40 – – – – 1
Spain – – 4 – – – 14 – – – – –
Switzerland – – – – – – 1 – – – – –
UK––––––7––1–3
Total 1 1 16 1 3 5 175 5 2 69 4 13
% Total 0.2 0.2 3.5 0.2 0.7 1.1 38.5 1.1 0.4 15.2 0.9 2.9
Abbreviations: NS, not specified; others as in Table 1.
169

The longest duration of illness was 216 months in an
insert gTSE case, the group with the highest variability
in clinical duration. However, in all other groups there
were a few cases with an exceptional long clinical
duration for their group (outliers in Fig. 1b). This in-
cluded 11 cases in E200K gCJD (five MM at codon 129:
16, 19, 20 and two 24 months; 6 MV: 17, two 18, 19,
and two 36 months), four cases in V210I gCJD (MM:
11, 15, 31, and 34 months), four in ‘‘other’’ gCJD
(43 months, D178N-129VV, 46 months, V180I-129MV,
52 months, V203I-129MV, 59 months, missing muta-
tion and codon 129), and one in FFI (MV: 97 months),
and two in insert gTSE (216 months 120 bp insert-
129MV, and 192 months 168 bp insert-129 MV).
Investigations
Data on CSF 14-3-3 immunoassay were available in
57% of cases (60% in E200K, 72% in V210I, 59% in
‘‘other’’ gCJD, 58% in FFI, 31% in GSS, and 50% in
insert gTSE), EEG in 82% of cases (90% in E200K,
96% in V210I, 86% in ‘‘other’’ gCJD, 77% in FFI, 50%
in GSS, and 67% in insert gTSE), and MRI brain scan
in 43% of cases (38% in E200K, 67% in V210I, 31% in
‘‘other’’ gCJD, 52% in FFI, 33% in GSS, and 36% in
insert gTSE). In the various forms of gTSEs, there were
significant differences in the frequency in positivity of
the 14-3-3 test in the CSF (P<0.0001, Fisher’s exact
test), the EEG (P<0.0001, chi-squared test) and the
MRI brain scan (P=0.0038, Fisher’s exact test) (WHO
2003). A positive 14-3-3 test was present in the majority
of gCJD patients and insert gTSE patients (Fig. 2). A
typical EEG (WHO 2003) was more frequent in gCJD
than in other forms of gTSE diseases. About 50% of
GSS cases had a positive 14-3-3 test in the CSF, while
the EEG showed typical pseudoperiodic activity in only
2 of 26 patients with available information (Fig. 2). In
GSS and FFI, neither the EEG nor the 14-3-3 tests were
of help in the clinical diagnosis of disease. MRI brain
scan was performed in a small proportion of cases and
was positive (WHO 2003) in about 50% of E200K and
‘‘other’’ gCJD and in about 30% of GSS and insert
gTSE cases. In FFI and V210I gCJD patients, the MRI
brain scan was positive in only 18 and 15%, respectively.
Classification of protease-resistant PrP
Results were available in only 43 cases. Twenty-two
E200K, 6 V210I gCJD, 9 ‘‘other’’ gCJD, 1 FFI, 2 GSS,
and 3 insert gTSE cases (Table 5). Thirty-four patients
had type 1 protease-resistant prion protein (according to
the system described by Parchi et al.) (Parchi et al. 1999)
in brain samples, six had type 2A, and three patients had
both type 1 and type 2A.
Discussion
EUROCJD has data on 23 specified PRNP mutations.
Currently more than 30 mutations have been reported in
the world literature, but many of these mutations are
very rare or are restricted to specific populat ions (Kov-
acs et al. 2002). Comparison of genetic and sporadic
TSEs shows striking differences in their frequency and
geographic distribution. While sCJD has a similar inci-
dence in all participating countries (Ladogana et al.
2005a), PRNP mutations show significant variability.
Some mutations (P105L, N171S, V180I, T188A, E196K,
R208H, V203I, 168 BPI) are extremely rare, while
E200K is recognised in 9 out of 10 reporting countries.
The high absolute and proportionate incidence of the
V210I mutation in Italy is striking; most affected fami-
lies lived in three adjac ent areas (Ladogana et al. 2005b).
It is a pertinent point that a single report on the N171S
mutation suggested a psychiatric phenotype, whereas in
our series this mutation was associated with CJD
(Samaia et al. 1997). A high proportion of all cases in
each category underwent genetic analysis. The three
lowest percentages of PRNP tests were 42.3% (Canada),
32% (Australia) and 31.7% (Netherlands). The vari-
ability in the frequency of mutations countrywise is
unexpected and is not related to variation countrywise in
GSS FFI Insert
P102L-129M A117V-129V G131V-129M NS D178N-129M NS 24-129M 48-129M 72-129? 96-129M/V 120-129M/V 144-129M 168-129M 192-129M/V NS
1 – 1 13 1––––––1––
– – – –3 ––––––1–––
1 – – 8– ––––––––––
3 2 – –6 –2––1–1–1–
3 5 – –17–1–128––––
8 – – –10–2––1–––––
– – – –– –1––––––1–
– – – –– ––––––––––
– – – –241–1–––––––
– – – –– ––––––––––
8 5 – 61 ––––1491–2
24121 1564261151211222
5.3 2.6 0.2 3.3 14.1 0.4 1.3 0.2 0.2 1.1 2.6 2.4 0.4 0.4 0.4
170

the availability of genetic data. Focal accumulations of
genetic patients (Mayer et al. 1977) may be due to ge-
netic isolation but the explanation for the overall inter-
country variability in this study is uncertain.
The ratio of female to male mortality rates in Table 3
indicates that there is an excess of females in all mutations
except FFI. Taking account of varying distribution by
age, the weighted female to male ratio (1.59 for E200K,
1.62 for V2 10I, 1.90 for other gCJD, 1.43 for GSS, 1.62
for insert gTSE, and 0.62 for FFI) strongly confirms the
excess of female cases in all forms of gCJD except FFI.
Although consistent with similar findings in sporadic
CJD (Ladogana et al. 2005a), the explanation for this
gender bias in mortality is uncertain. Possible explana-
tion include sex-linked genetic factors influencing disease
expression, bias in case ascertainment linked to gender or
differential susceptibility/exposure to a co-factor.
Clinical and laboratory parameter s show consider-
able similarity between the most frequent gCJD (e.g.,
E200K mutation) and sCJD cases. Although gCJD cases
with point mutations have an earlier mean age at death,
there is no difference between gCJD cases with point
mutations and sCJD in the mean duration of the disease
(Alperovitch et al. 1999; Pocchiari et al. 2004). As in
sCJD (Zerr et al. 2000), analysis of CSF 14-3-3 protein,
EEG, and MRI is helpful in the diagnosis of gTSEs and
Table 3 Characteristics of genetic transmissible spongiform encephalopathies
Forms of gTSE n Percentage of
definite cases
(n)
Gender Codon 129 polymorphism Percentage of
positive family
history for TSE
(available data)
F M F/M ratios
of mortality
rates
a
MM % (n)MV%(n)VV%(n)
E200K gCJD 175 68.0 (119) 107 68 1.69 78.3 (130) 19.9 (33) 1.8 (3) 49.1 (114)
V210I gCJD 69 47.8 (33) 39 30 1.42 73.1 (49) 26.9 (18) – 12.3 (57)
Other gCJD 51 58.8 (30) 28 23 1.38 36.8 (14) 34.2 (13) 29.0 (11) 75.9 (29)
FFI 66 92.4 (61) 29 37 0.62 71.4 (45) 28.6 (18) – 88.0 (50)
GSS 52 76.9 (40) 29 23 1.17 51.7 (15) 34.5 (10) 13.8 (4) 69.7 (33)
Insert gTSE 42 69.0 (29) 21 21 1.23 48.5 (16) 30.3 (10) 21.2 (7) 48.0 (25)
Abbreviations: gTSE, genetic transmissible spongiform encephalopathy; gCJD, genetic Creutzfeldt–Jakob disease; FFI, fatal familial
insomnia; GSS, Gerstmann–Stra
¨
ussler–Scheinker disease; Insert, gTSE patients with insert mutations.
a
For the period 1999–2002.
Fig. 1 Box- and whisker-plots of age at onset (a) and clinical
duration (b) in different forms of genetic TSE diseases. The line in
the middle of the box represents median. The box extends from the
25th percentile (x
[25]
) to the 75th percentile (x
[75]
), the so-called
interquartile range (2IQR). The lines emerging from the box are
called the whiskers and they extend to the upper and lower adjacent
values. The upper adjacent value is defined as the largest data point
less than or equal to x
[75]
+1.52IQR. The lower adjacent value is
defined as the smallest data point greater than or equal to x
[25]
1.52IQR. Filled circles represent values more extreme than the
adjacent values (referred to as outliers)
Table 4 Influence of codon 129 polymorphism of the PRNP gene
in determining the age at onset
Forms of gTSE Codon 129
Met/ Met
mean (SD)
Met/Val
mean (SD)
Val/Val
mean (SD)
E200K gCJD 60.4 (10.8) 60.7 (8.6) 55.7 (9.0)
V210I gCJD 59.3 (9.7) 57.9 (9.8) –
Other gCJD 70.9 (7.5) 56.2 (14.2) 48.1 (12.6)
FFI 50.8 (13.4) 52.5 (10.0) –
GSS 54.0 (11.1)
a
49.0 (12.1) 66.2 (17.9)
Insert gTSE 60.9 (14.0)
a
56.2 (15.2) 62.7 (11.3)
Abbreviations: gTSE, genetic transmissible spongiform encepha-
lopathy; gCJD, genetic Creutzfeldt–Jakob disease; FFI, fatal
familial insomnia; GSS, Gerstmann–Stra
¨
ussler–Scheinker disease;
Insert, gTSE patients with insert mutations.
a
Two patients had missing data on age at onset.
171

in particular in gCJD cases with point mutations, while
in other gTSE cases these laboratory examinations are
less sensi tive. Our data confirm previous studies that
insertional mutations often present atypically with a
relatively protracted duration of illness (although less
protracted than a previous study which included larger
numbers of cases and with a unique type of PrP
immunoreactivity in the cerebellum (Kovacs et al. 2002;
Vital et al. 1999).
We also demonstrate that, except for some specific
mutations like D178N-129M, protease-resistant PrP
may be variable within the same brain in some gTSEs as
in sCJD (Puoti et al. 1999).
It is of great importance that a positive family
history of a human TSE is absent in a high proportion
of cases overall and with all the mutations. Although
this has been reported previously by this group and
others (EuroCJD group 2001; Goldman et al. 2004;
Mitrova and Belay 2002), this is the first study to
provide a detailed analysis of the frequency of a po-
sitive family history in a range of PRNP mutations.
Possible explanations include lack of knowledge of the
family history in the relatives of index cases, premature
death in antecedents prior to the development of a
TSE, and non-paternity. However, there must be some
doubt that the very high frequency of a negative family
history in this study can be explained by these mech-
anisms and there remains the possibility that a
minority of mutations arise de novo (Dagvadorj et al.
2003) or that the mutations are not fully penetrant
(Mitrova and Belay 2002).
As cases considered as sCJD often lack full-length
PRNP analysis, accurate diagnosis of gTSEs may de-
pend on more widespread genetic screening, particularly
as some mutations, e.g., E200K and V2 10I, are associ-
ated with a phenotype indistinguishable from sCJD. The
negative family history in a high proportion of cases
indicates that mutations may be identified unexpectedly
in cases thought to have sCJD or indeed some other
neurological disorder and this may have important
implications for other family members.
At present gTSEs are the only sub-group of human
TSEs in which the diagnosis can be supported by
genetic screening in life. There is the potential for
early diagnosis and the risk of developing a TSE can
be identified before the onset of disease in asymp-
tomatic carriers. This may be an important advantage
in the application of potential therapies as treatment
could be instituted before severe and irreversible
damage to the CNS occurs, or perhaps, as a means to
prevent iatrogenic transmission of CJD from at-risk
patients.
Fig. 2 Percentage of cases
showing positive 14-3-3 protein
in the CSF, typical EEG
pattern, and positive MRI brain
scan in different forms of
genetic TSE disease. The
number of patients with
available data is shown on top
of each column
Table 5 Type of the pathological PrP in the brain of genetic TSE cases
PrP type gCJD
a
FFI
a
GSS
a
Insert
a
N171S D178N E196K E200K V203I R208H V210I E211Q D178N P102L
Type 1 – 2 VV 1 MM 17 MM 1 MM 1 MM 5 MM 1 NA – 2 MV 1 MV
1 MV 1 MV 1 MV
Type 2A 1 VV – – 1 MV 1 MV – – – – – 1 VV
1VV
Type 1–2A – – – 2 MM – – – – – – 1 MV
Type 2B – – – – – – – – 1 MV – –
Abbreviations: gTSE, genetic transmissible spongiform encepha-
lopathy; gCJD, genetic Creutzfeldt–Jakob disease; FFI, fatal
familial insomnia; GSS, Gerstmann–Stra
¨
ussler–Scheinker disease;
Insert, gTSE patients with insert mutations; MM, methionine
homozygous; VV, valine homozygous; MV, heterozygous patients;
NA, not available.
a
Number of cases of codon 129 polymorphism.
172

The occurrence of asymptomatic carriers of PRNP
mutations (Mitrova and Belay 2002) und erlines the
importance of genetic testing in all suspect cases of hu-
man TSEs and, in positive cases, relatives may be offered
genetic testing, provided appropriate ethical protocols
are followed. Although there may be incomplete pene-
trance and an absence of presymptomatic neuropatho-
logical changes or accumulation of disease-associated
PrP (Sasaki et al. 2003), healthy carriers of PRNP
mutations are at greater risk of developing a TSE and
may be subject to restrictions, such as donating blood
and tissues (e.g., cornea, dura mater), because of the risk
of iatrogenic transmission.
It is not known why there is partial penetrance in
some mutations, or why there is marked variability in
clinical phenotype, including age at onset, both between
and within pedigrees. There is a need to analyse events
preceding the onset of clinical manifestation in carriers
of PRNP mutations as possible triggering factors. In
E200K, psychological stress (divorce, death of a close
relative, retirement, loss of a job), complicated surgery
with prolonged anaesthesia, serious accidents, or curre nt
infectious dis ease have been frequently noted (Brandel
and Delasnerie-Laupretre 1997; Mitrova and Belay
2002).
In conclusion, the term gTSE, which includes gCJD,
FFI, and GSS, is more appropriate than familial TSE
for all patients with TSE-specific genetic marker,
regardless of the number of affected family members,
while familial TSE designa tes only genetic cases with
other (one or more) TSE-affected relative. The distri-
bution of gCJD is geographically heterogeneous. The
codon 129 polymorphism influences phenotype. In
gTSEs, PRNP analysis may allow early or pre-symp-
tomatic diagnosis and this may be important if specific
therapies become available. Further studies are required
to identify whether PRNP mutations are sufficient in
themselves to cause disease.
Acknowledgements This study was funded through an EU Con-
certed Action (BIOMED2 Contract No. BMH4-CT97-2216).
Australia: The Australian National CJD Registry is funded by the
Commonwealth Department of Health and Ageing. We are
grateful to the following people involved in the Australian National
CJD Registry: C.L. Masters, A. Boyd, G. Klug, and J. Lee. Aus-
tria: The Austrian Reference Centre for Human Prion Diseases
(O
¨
ERPE, Head: Prof. Herbert Budka) acknowledges the help of
Drs Christa Jarius, Ellen Gelpi, Christine Haberler, Thomas
Stro
¨
bel, and Till Voigtla
¨
nder; DI Dita Drobna; and Ms. Helga
Flicker, Brigitte Millan-Ruiz, and Monika Richter. Canada: The
Canadian Surveillance System is funded by Health Canada. Other
collaborators on the project are Dr C Bergeron, neuropathologist
(University of Toronto), Dr N Cashman, neurologist and one of
the principal investigators for CJD-SS, Dr D Westaway, consulting
scientist (University of Toronto). France: We would like to
acknowledge all reporting physicians and the members of the Re-
seau National de surveillance de maladies de Creutzfeldt-Jakob et
maladies apparentees. Germany: The German surveillance system is
funded by the Federal Ministry of Health 9BMG, 325-4471-02/15.
We are grateful to all reporting physicians throughout Germany
who contributed to the German surveillance system and especially
to Maja Schneider-Dominico for her excellent support in the co-
ordination of surveillance. We also acknowledge the help of Drs
Otto Windl and Walter Sculz-Schaeffer. Italy: We would like to
acknowledge the Ministry of Health and the Istituto Superiore di
Sanita
`
for supporting the surveillance of CJD in Italy and S. Al-
monti, V. Mellina, and L. Ingrosso for help in collecting data and
advice. The Netherlands: CJD surveillance in the Netherlands is
funded by the Dutch Ministry of Health, Welfare, and Sports. We
acknowledge the help of colleagues at the Department of Neurol-
ogy at the Academic Medical Centre, Amsterdam and the
Department of Pathology at the University Medical Centre, Utr-
echt. Slovakia: The Slovak Surveillance System was supported by
the Slovak Ministry of Health and by grants from the European
Union. We would like to acknowledge the help of Dr. Dana Sliv-
arichova
´
, Dr. Vladimı
´
ra Verchovodkova
´
, all reporting physicians
and collaborating pathologists. Spain: We are grateful to all
reporting physicians and to members of the Spanish TSE study
group at Consejo Interterritorial and co-workers at CNE and
ISCIII and laboratories, particularly to N. Cuadrado and J.Yague.
Switzerland: This work was supported by the Kanton of Zurich and
by grants from the European Union. The Swiss Reference Center
for Prion Diseases is being funded by the Swiss Federal Office of
Public Health. UK: The UK CJD Surveillance System is funded by
the Department of Health and the Scottish Executive Health
Department. We are grateful to all the members of staff at the
National CJD Surveillance Unit and in particular to James Iron-
side for neuropathological expertise and to clinicians throughout
the UK for their co-operation with the study.
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