Transcript Prion Diseases

Prion Diseases
Kuru
Kuru figures
• Incidence 1 % (population aprox.
15,000)
total F : M = 10 : 1
• < 20 years of age F : M = 3 : 1
(20 % of cases)
• 20-40 years of age F : M = 15 : 1
Etiology of Kuru
• Primary infectious: no fever, no CSF cell raise
• Genetical disease:
– Pro: in families, in patients who moved out of Fore
region.
– Con: also in patients who came to live in Fore region,
as well in young as in old patients.
• Toxic/deficiency: no toxic compound isolated,
balanced diet.
• Endocanibalism
Sporadic CJD
1920/1921
Sporadic Creutzfeldt-Jakob disease diagnosis
•
•
•
•
•
•
Rare disease: 1/106
Duration: short (months)
Neurological signs and symptoms:
Rapidly progressive dementia, myoclonus, ataxia, central
visual disturbances, extrapyramidal signs, pyramidal signs,
akinetic mutism (variant: chorea, dysesthesias, psychiatric
disturbances)
EEG and MRI
Neuropathology
WHO criteria:
– Type of CJD (sporadic, genetic, iatrogenic, variant)
– Strength of diagnosis (definite, probable, possible)
MRI
Proton density
Weighted T2
EEG abnormalities
normal
sCJD
Gross sCJD
Micro sCJD
Prion Diseases/Transmissible Spongiform Encephalopathies
Prion diseases
sporadic
genetic
acq. by infection
TSE's in animals
TSE's in man
Scrapie
sCJD
ex-Scr
fCJD
CWD
GSS
TME
FFI
BSE
Kuru
FSE
iCJD
ex-TSE
vCJD
85%
}
}
14%
1%
Etiology
Possibilities (ca 1980):
• Degenerative/Hereditary
•
(slow)Virus
Againts virus hypothesis
Very resistant agent:
• resists dry heat ( > 200ºC)
• resists steam autoclaving (up to134ºC, 18 mins.)
• resists formaldehyde
• resists UV-radiation
• resists Gamma-radiation ( > 0.3 MGray)
• etc.
Prion docterine (Prusiner)
In biochemical separation-infection studies:
• Infectivity is not present in a DNA/RNA fraction
• Infectivity is present in a protein fraction
In conclusion:
• A protein (and that protein only) causes a prion
disease
From Prion gene to Prion protein
1
codon
22
51
91
-8
polymorph.
5 OR
PRNP
sign. peptide
E/K
M/V
ß1
α1 ß2
Glycin - prolin
rich sequency
5 OR
PrPc/PrPSc
(33 - 35 kD)
219 231 254
129
PrPc-fragm.
α3
hydrophob. seq.
CHO CHO
ß1
α1 ß2
α2
α3
S
S S GPI
CHO CHO
Proteinase K
PrPSc
(27- 30 kD)
α2
ß1
α1 ß2
α2
S
α3
S S GPI
C
PrP
Octa-repeat
H1
H3
N-terminus
GPI
H2
Sh1
129
106-126
Sh2
PrPC to PrPSc
Abnormal folding of a protein protein (and that
protein only) causes a prion disease
Conversion models for PrPC to PrPSc
Dimerisation
PrPSc
PrPC
PrP*
PrP* PrPSc
PrPSc PrPSc
Polymerisation
PrPC
PrP*
PrP*
PrPSc
…and subsequent breaking and seeding.
PrPSc
Structural assembly of
Sc
PrP
Aggregation
Crystal /Scrapie associated fibril
IHC for PrP in sCJD-neocortex
The “Z” family
I
II
III
IV
Clin: dementia, ataxia, hypokinesia
Example of gCJD in cerebellum
Dominant inheritance with dementia and ataxia mostly
Familial CJD vs Familial Fatal Insomnia
178Asn-129V-f-CJD
178Asn-129M-FFI
Prion diseases acquired through infection
• Kuru
• iatrogenic CJD
• new variant CJD
iCJD
number
Intracerebral
cornea
neurosurgery
stereotactic EEG
Cerebral surface
dura mater*
Peripheral blood
growth hormone
gonadotrophin
incubation
time
clinical presentation
4
5
2
17 mo
17 mo
18 mo
dementia/ataxia
dementia/ataxia
dementia/ataxia
174
6-12 y
ataxia
180
4
12 y
13 y
ataxia
ataxia
* Mostly Lyodura, now 2 cases with 0.1 N NaOH treated Tutoplast
40000
35000
30000
25000
20000
15000
10000
5000
0
1982
1985
1988
1991
1994
1997
Bovine Spongiform
Encephalopathy
1985
1988
1991
2000
1994
2003
2006
1997
2000
2003
2006
BSE medulla oblongata
BSE-microscopy
Clinical history in v-CJD
65% of cases onset with ‘psychiatric’ complaints:
•Behavioural disturbance
•Attention deficit
•Personality changes
•Depression
Later weeks - months:
•Dysaesthesia (20 % onset with dysaesth)
•Ataxia
•Myoclonus/choreo-athetosis
•Progressive dementia
No typical EEG
14-3-3 protein 50% of the cases positive
bilateral Pulvinar sign on MRI
MRI
Flair sCJD
Flair vCJD
Age Comparison with sp-CJD
v-CJD
sp-CJD
0
10
20
30
40
50
60
70
80
vCJD autopsy
Tonsil biopsies in vCJD
Variant CJD
vCJD caused by BSE?
Strain typing
•
•
•
•
Glycoform pattern
Genotype/polymorphisms
Clinical profile
Inoculation in experimental
animals:
– Incubation time
– Pathology profile
CH sidechains of PrP
Glycoform analysis
sCJD
vCJD/BSE
Survival after inoculation
% surviving mice
100
80
spCJD
spCJD-farmers
60
vCJD
40
BSE
FSE
20
0
0
100 200 300 400 500 600 700
days after inoculation
Lesion profiling example
3
2
1
0
1
2
3
4
5
6
7
8
9
Lesion profiling
vCJD
3
2
2
1
1
0
0
1
2
FSE, exTSE
3
3
4
5
6
7
8
9
3
4
5
6
7
8
9
1
2
3
4
5
6
7
8
9
4
5
6
7
8
9
BSE
3
2
1
0
1
2
spCJD
3
2
2
1
1
0
0
1
2
Scrapie
3
3
4
5
6
7
8
9
1
2
3
What’s the problem with Prion
diseases
• Infectious material (whatever the origin:
sporadic, genetic or infectious derived)
• Poorly disinfectable:
– formic acid 96% 1 hr
– 2 N NaOH 1 hr
– 20,000 ppm Chlorine 1 hr
Smallest infectious dose
ID50/kD
0
250
500
750
1000
1250
1500
kD
Smallest infectious dose = 300-600 kD = 14-28 PrPSc molec.
Pathogenesis of infection
1. Direct brain contact
2. Vascular inoculation
3. Oral (intestinal) inoculation
Role for autonomic nervous system
• 1997: infectivity after oral challenge in
hamster first seen at T7 level of spinal cord
(Diringer)
• 2002: PrPSc seen in vagus nerve and
splanchnic nerve before reaching spinal
cord after oral challenge (Beekes)
Role for hematologic system
• 1996: immunodeficient mice develop no prion
disease after peripheral inoculation (Bruce)-spleen
dependency
• 1991: FDCs found positive for PrPSc in mice
(Kitamoto)
• 1997: crucial role for B-cells in CJD pathogenesis
(Aguzzi)
• 1998: PrP expression in B-cells not necessarry for
CJD pathogenesis (Aguzzi) –exit B-cells
Proposed route of peripheral infection
GALT
M-cell
Gut
Macroph.
FDC
Xth CN
Brainstem
Brain
>
+
B-cell
lymphatics
+
Macroph.
FDC
Symp NS
Spinal cord
Spleen
PrPSc pathogenesis: loss or toxic gain
of function?
PrP properties:
• Proposed function: ligand
• Cu2+ binding (metallo-protein)
• Hydrophobic section: PrPcmt
PrP protein-copper binding sites
Manlgcwmlvlfvatwsdlglckkrpkpggwnt
ggsrypgqgspggnryppqggggwgqphgggwg
qphgggwgqphgggwgqphgggwgqgggthsqw
nkpskpktnmkhmagaaaagavvgglggymlgs
amsrpiihfgsdyedryyrenmhrypnqvyyrp
mdeysnqnnfvhdcvnitikqhtvttttkgenf
tetdvkmmervveqmcitqyeresqayyqrgss
mvlfssppvillisfliflivg
Cu binding motiv: hgggw or ggth
Octarepeats
R1 R2 R2 R3 R4
R1
R2
R2
R3
R4
CCT
pro
CCT
pro
CCT
pro
CCC
pro
CCT
pro
(5 x 8) + 1 codon = 123 bp
CAG GGC GGT GGT GGC TCG GGG CAG
gln gly gly gly gly trp gly gln
CAT
GGT GGT GGC TCG GGG CAG
his
gly gly gly trp gly gln
CAT
GGT GGT GGC TCG GGG CAG
his
gly gly gly trp gly gln
CAT
GGT GGT GGC TCG GGA CAG
his
gly gly gly trp gly gln
CAT
GGT GGT GGC TCG GGT CAA
his
gly gly gly trp gly gln
Normal
Pathology
Generation of H2O2
Cu 2+ + 2PrP
H20
H0·
PrP2-Cu
Shiraishi et al Biochem J 2005;387: 247-255
Built in of
Sc
PrP
in lipid rafts
• ‘Transformation of PrPC to PrPSc occurs
preferentially in cholesterol rich lipid rafts’
(Hooper Biochem Soc Trans 2005; 32: 335-338)
• ‘Cell membranes have a role in
transformation of PrPC to PrPSc’ (Kazlauskaite
& Pinheiro Biochem Soc Symp 2005; 72: 211-222)
• ‘Prion replication alters the distribution of
synaptophysin in membrane’ (RusselakisCarneiro et al Am J Pathol 2004; 165: 1839-1848)
Protein misfolding to neuronal
dysfunction pathogenesis
Protein misfolding & accumulation diseases:
• Alzheimer’s disease
• Parkinson’s disease
• Tauopathies
– Progressive Supranuclear palsy
– Fronto-temporal dementias
– Cortico-basal degeneration
Side step: Alzheimer’s disease
• Non infective disease
• Leading to dementia
Amyloid Precursor Protein
Membrane
Extra cellular
Glycosylation
Signal peptide
Aβ
671-713
Cystein rich area
Phosphorylation
daefrhdsgyevhhqklvffaedvgsnkgaiiglmvggvvia
β - secretase
α - secretase
γ - secretase
ABeta generation
Toxic oligomers in ABeta
• Cu 2+ leads to H2O2 production
• A Beta density in AD is related to synapse
loss (lower level of synaptofysin)
• Small oligomers more toxic than large
aggregates
• Sounds familiar???
Protein folding diseases
Protein-normal folded
+
Mutation in protein
Chaperones/Ubiquitin/
Hydrophobic interactions +
- age Clipping mechanisms
Protein-abnormal folded – small aggregates (toxic)
Protein aggregated (non toxic?)
amyloid
H2O2 lipid raft:
synapse dysf
membrane damage
Break down of referred patients
April 1, 1998-December 1, 2006
1322 referred
595 ‘pending file’
727 ‘true’ referrals
32 info refused
17 ‘pending classification’
43 alive
635 referrals at study end-point
13 possible prion disease
332 Prion cases (279 definite + 53 probable)
Mean mortality 1.09 /106.yr
289 non Prion (164 definite + 125 probable)
Incidence of CJD Deaths in Canada
(per Million Population by Year)
Year of Death
Average incidence rate of CJD in Canada = 1.09 per million Canadians
1994
2
1995
3
1996
13
1997
18
1998
24
1999
31
2000
35
2001
30
2002
36
2003
29
2004
42
0.07
0.10
0.44
0.60
0.79
1.02
1.14
1
1.15
0.92
1.31
1.22
2005 40
0.84
2006 23
0
0.2
0.4
0.6
0.8
Incidence of CJD Cases
1
1.2
1.4
Classification of Referrals
Definite CJD
279 Definite sporadic CJD (sCJD)
Definite familial CJD (fCJD)
Probable CJD
Possible CJD
256
5
Definite GSS
11
Definite FFI
1
Definite sporadic GSS
1
Definite iatrogenic CJD (iCJD)
4
Definite variant CJD (vCJD)
1
53 Probable sporadic CJD (sCJD)
49
Probable familial CJD (fCJD)
4
13 Possible sporadic CJD (sCJD)
13
Non-CJD
289
TOTAL
635
CJD Cases (n=332)
Sporadic
Genetic
Iatrogenic
Variant
92.1%
6.3%
0.3%
1.2%
sCJD Cases by Province/Territory
YUKON
NUNAVUT
0
NORTHWEST TERRITORIES
BRITISH
COLUMBIA
40
99%
NEWFOUNDLAND
4
83%
ALBERTA
27
84%
SASKATCHEWAN
MANITOBA
15
161%
16
144%
ONTARIO
QUEBEC
115
96%
68
97%
PRINCE EDWARD ISLAND
0
NOVA SCOTIA
Actual sCJD
% of expected CJD
(based on 2006 population)
NEW BRUNSWICK
10
141%
11
124%
Canadian iCJD cases (dura mater)
Incubation Period
Duration of Illness
M 19
F 49
M 14
F 59
1980
1985
1995
1990
Year
2000
2005
vCJD criteria in Canadian case
I
II
III
A
Progressive neuropsychiatric disorder
B
Duration of illness > 6 months
C
No alternative diagnosis
D
No iatrogenic exposure
A
Early psychiatric symptoms
B
Painful dysaesthesias
C
Ataxia
D
Myoclonus or chorea or dystonia
E
Dementia
A
No EEG/No ‘classical’* EEG
*Generalized triphasic periodic complexes at approx. 1 per second
B
IV
MRI pulvinar sign positive
Positive tonsil biopsy for PrP












Possible vCJD only
(without pathological confirmation)
Occipital Neocortex H&E of Canadian Case
Travel history
• Sep ’87-Aug ’90: UK education + 2
weeks visit France.
• Feb ’91-Mar ’91: visit UK
• Feb ’92-Mar ’92: visit UK
• June 2000:
visit UK
Total 38 months risk exposure 1-14 years
prior to symptoms
Survival all types of CJD and non-CJD
1.0
duration of disease (mo)
mean
0.9
0.8
0.7
median
sCJD
6.3 ± 0.3
4
fCJD
18.0 ± 6.0
10
GSS
69.3 ± 18.6
35
non-CJD
22.8 ± 2.3
16
0.6
0.5
0.4
0.3
0.2
0.1
0.0
0
12
24
36
48
60
72
84
96
108
120
132
144
156
sCJD
GSS
fCJD
iCJD
vCJD
Total
Definite
Probable
Definite
Probable
Definite
Probable
Definite
Probable
Definite
Probable
Cases
256
49
11
0
5
4
4
0
1
0
of which unsuspected cases
22 (8.6 %)
332
30 (9.0 %)
8 ( 73 %)
0
0
0
Why where they missed?
•
•
•
•
•
Lack of experience?
Different age of onset/duration?
Different clinical signs and symptoms?
Different subtype?
Different auxiliary investigations findings?
Conclusions unsuspected Prion
disease cases-1
• 9 % of Prion disease cases were clinically
unsuspected in Canada in last 8½ years.
• GSS poorly recognized (73% of GSS are
clinically unsuspected in this series)
Conclusions unsuspected Prion
disease cases-2
• sCJD and GSS can be missed clinically
• Missing is due to the fact that these cases are
atypical:
– Less usual subtypes (MM2, MV2)
– Missing symptoms in the presentation
– Longer duration of disease
• Or (occasionally) due to missed clues:
 MRI T2/FLAIR attenuation
 EEG
•
Missing is not due to ‘academic status’ of clinician
...but...are we missing much more cases??
‘Back of the envelope’
Minimum True Mortality rate of CJD for Canada = 1.09/106.yr
Maximum True Mortality rate of CJD for Canada = ?
•
General autopsy rate in Canada: 9.5 % = 1:10.5 = 1:(9.5 + 1)
means 9.5x(30/332)x0.91x1.09/106.yr unnoticed CJD as they
are not autopsied: actual CJD frequency in Canada could be
max 1.09 /106.yr + 0.85/106.yr = 1.94/106.yr
•
Comparison:
European surveillance figures for Switzerland, Spain, Italy,
France and Austria have been last 2 or more years well over 1/
106.yr, and even over 2 in selected case.
True mortality figure is probably somewhere between
1.09 and 1.94.
Prion diseases
- regional distribution in the Netherlands until 2000 -
Is sporadic CJD really sporadic?
• Concept of spontaneous generation of
abnormal prion protein is difficult to grasp
• But no relation with food, surgery or other
possible cause found
• However in UK geographical clustering
seen 15-25 years before onset