Kurt A. Jellinger. 2 nd Int. Conference BrainNet Europe, Munich, Dec , 2008 NAC A30P A53T ALPHA HELICAL HYDROPHOBIC ACID (GLU-PRO) COOH

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1 2 nd Int. Conference BrainNet Europe, Munich, Dec , 2008 NH 3 1 NAC A30P A53T ALPHA HELICAL HYDROPHOBIC ACID (GLU-PRO) COOH (Src) (GRK5, CK-1 & CK-2) Kurt A. Jellinger Institute of Clinical Neurobiology, Vienna, Austria

2 ALPHA-SYNUCLEIN 1 1' DNA (~112kb: NACP-REP1) Coding mrna amino acids Alternative splicing: No Exon 3 (amino acid residues 41-54) amino acids Alternative splicing: No Exon 5 (amino acid residues ) amino acids Tyrosine nitration sites: o,o'-dityrosine crosslinks: Protein (structural domains): NH ALPHA HELICAL HYDROPHOBIC ACID (GLU-PRO) COOH Phosphorylation sites (kinases): Familial Parkinson disease mutations: A30P A53T 87 (CK-1 & CK-2) (Src) (GRK5, CK-1 & CK-2) Sporadic PD, DLB, LBV/AD, MSA Hydrolytic fragment: Non-amyloid component of plaques (NAC): Minimal fibrillogenic domain:

3 Synucleinopathies of the human nervous system Category and name SNCA gene-linked PD Sporadic, idiopathic PD Dementia with Lewy bodies Incidental Lewy body disease Pure autonomic failure Multiple system atrophy Pantothenate kinase-assoc. neurodegeneration Parkin- and LRRK2-linked PD AD (and other tauopathies) Down's syndrome Gaucher disease Characteristic α-synuclein inclusion (site) Invariable synucleinopathies neuronal LBs, Lewy neurites (BS, BF, Ctx) neuronal LBs, Lewy neurites (BS, BF) neuronal LBs, Lewy neurites (BS, BF, Ctx) neuronal LBs (BS) neuronal LBs, Lewy neurites (PNS) glial cytoplasmic inclusions (BS, Cblm, Put, GP) neuronal LBs, axonal dystrophy (GP, BS) Variable synucleinopathies neuronal LBs, Lewy neurites (BS, BF) neuronal LBs, dystrophic neurites (BS, BF, Ctx) neuronal LBs, dystrophic neurites (Ctx) neuronal LBs (BF, hippocampus) M. Schlossmacher, in: The Dementias 2 (2007)

4 Spectrum of "Lewy body Diseases" (adapted from Ince et al, 2001) limbic cortex N.basali M. neocortex cortex lateral grey horn/ sympathetic ganglia dorsal/vagal nucleus Substantia nigra myenteric ganglia Dementia w. Lewy bodies Parkinson's disease Pure autonomic failure Lewy body dysphagia 2+/ /2+ +/3+?? +/2+ 2+/ /3+ +/2+ +/ / /+ 0 +? 3+

5 A C B D E

6 Protein distribution in cortical Lewy bodies Leverenz et al, Brain Pathol 17, 2007

7 Three-dimensional reconstruction of Lewy bodies and Lewy neurites Kanazawa, Brain Pathol 18 (2008) MSA Istanbul 2002 quer.ppt

8 Cersosimo & Benarroch, Mov Disord (2008) 23 Involvement of autonomic GI control areas in PD

9 LB pathology in the enteric NS Braak & DelTredici, Neurology 70 (2008) Adrenal medulla Fumimura et al, JNEN 66 (2007) Auerbach plexus (A,B,C) Meissner plexus (D,E)

10 αsyn-ir in peripheral nerves Ikemura et al, JNEN 67 (2008)

11 Neuropathological staging of LBD Kosaka LBD stage Brainstempredominent type Transitional (limbic) type Diffuse cortical type Braak PD stage Anatomical distribution of Lewy bodies 1 Medullary tegmentum and dorsal motor nucleus of the vagus; olfactory bulb 2 Locus ceruleus, caudal raphe, medullary and pontine reticular nucleus 3 Substantia nigra and basal forebrain 4 Medial temporal (limbic) cortices, incl. amygdala and hippocampus CA2 region 5 Multimodal association cortices, esp. frontal and temporal lobes 6 Unimodal association cortices and primary cortices

12 Olfactory bulb synucleinopathy (OBS) in LB disorders Beach et al, ANP 2008 Material: 328 elderly autopsy cases (IBD, AD, controls) Sensitivity / specificity [%] of OBS: PD 95 / 91, DLB 97 / 91, ADLS 88 / 91, ILBD 67 / 81 vs. controls ADLS 88 / 81 vs. ADNLS OBS correlated significantly with AS scores in all other brain regions, MMSE & motor UPDRS.

13 Olfactory bulb synucleinopathy (OBS) in LB disorders personal study Material: 30 autopsy cases (27 PD, 3 ILBD) Results: PD stage 5-6 severe OBS correlating with AS in CA 2-3 PD stage 3-4 mild/moderate OBS, 20% no AS in CA 2-3 2/3 ILBD no OBS, no AS in CA 2-3 In contrast to OS tau scores significantly correlating with neuritic Braak scores, no such relation was seen for OBS AS+lesions in olfactory epithelium of PD/DLB are rare suggesting that olfactory loss in LB disorders results from damage to OB or other CNS abnormalities.

14 Wakabayashi et al, Parkinsonism Relat Disord 12, Suppl 2 (2006)

15 Braak & Del Tredici, Neurology 70 (2008)

16 Control PD Neuron loss in substantia nigra Dopamine ( μ g/g) Caudate Putamen HVA ( μ g/g) Caudate Putamen Gl. pallidus

17 Simulation of the neuronal loss in the substantia nigra Greffard et al, Neurobiol Aging (2008)

18 Relations between Braak PD stages and striatal TH & VMAT2 TH immunoreactivity [%] VMAT2 immunoreactivity [%] Control ilbd PD 2 0 r 2 = 0.72 r 2 = Braak PD Stage Braak PD Stage DelleDonne et al, Arch Neurol 65 (2008)

19 a) Large neurons b) Medium-sized GABA neurons c) Glial cells d) Severity of neuritic changes Correlations between PD stages and number of αsyn+ inclusions F. Mori et al, ANP 115 (2008)

20 G.G. Kovacs et al. Mov Disord 23, 2008 Nigral burden of α-synuclein correlates with striatal dopamine deficit αsyn ensures normal functions of dopamine transporter (DAT) and tyrosine hydoxylase. The correlation of αsyn in SN with the dopaminergic deficit in the striatal target was unclear. An inverse correlation between DAT expression in the striatum and the burden of pathological αsyn in SN was demonstrated, but not with cytoplasmic αsyn aggregates. This could implicate an impairment of axonal transport or synaptic dysfunction by aggregated αsyn. Simple detection of cytoplasmic LBs may not have clinical impact, while significant cytoplamic and neuritic accumulation of pathologic αsyn is likely to have. These data may serve as rationale for the consideration of neuronal cytoplasmic and neuritic αsyn pathology in clinicopathologic studies and staging of the disease process.

21 Density of DAT IR Mean Synuclein burden Mean LB proportion % P=0.02 P=0.02 P<0.02 Contr. αsyn.- Mean Synuclein burden Mean LB proportion % opathie Kovacs et al, Mov Disord 23 (2008)

22 M.L. Kramer et al, J Neurosci 27 (2007)

23 M.L. Kramer, 2008

24 Presynaptic accumulation of αsyn aggragates Synaptic dysfunction Synapse loss Neuronal death M.L. Kramer, 2008

25 Burke et al, Ann Neurol, submitted 2008 n=96 (Braak et al, Neurobiol Aging, 2003)

26 Singleton, Trends Neurosci 28 (2005)

27 Lewy bodies develop in grafted neurons in Parkinson disease α-synuclein 16-year-old graft α-synuclein 11-year-old graft Ubiquitin 11-year-old graft Brundin et al, Nature Rev Neursci 9 (2008)

28 Mechanisms that might explain why Lewy bodies form in grafted neurons Brundin et al, Nature Rev Neursci 9 (2008)

29 Assignment of Lewy body type in brainstem, limbic, and neocortical regions Brainstem regions Basal forebrain/limbic regions Neocortical regions Lewy body type pathology IX-X LC SN nbm Amygdala Transentorhinal Cingulate Temporal Frontal Parietal Brainstempredominant Limbic (transitional) Diffuse neocortical IX-X 9 th -10 th cranial nerve nucleus; LC locus ceruleus; SN subst. nigra; nbm nucl. basalis of Meynert. Cortical Lewy body score: 0-2 Brainstem-predominant, 3-6 Limbic or 'transitional', 7-10 Neocortical In each region a count of up to 5 LBs in the cortical ribbon gives a score of 1 in the table. Counts greater than 5 score as 2. The sum of the five areas is used to derive the category of cortical spread (maximum score 10). McKeith et al, Neurology 65 (2005)

30 PD related pathology categorized as recommended by McKeith and colleagues Distribution of αs-positive structures Brainstem predominant Limbic transitional Diffuse neocortical dorsal motor nucl. of vagus locus coeruleus raphe nucleus substantia nigra nucleus basalis of Meynert amygdaloid complex (trans)entorhinal cortex cingulate gyrus n=66 n=30 n=91 temporal gyrus frontal cortex parietal cortex number with dementia (%) 8 (12) 11 (37) 52 (57) number with EPS (%) 5 (8) 10 (33) 43 (47) Parkkinen et al, ANP 115 (2008)

31 Modified criteria for categorization of Lewy-related pathology (LRP) in patients with dementia. Results from two autopsy series Predominant region LRP severity scoring with proposed criteria* SN or medulla Amygdala Cingulate gyrus Frontal cortex LADRS n (%) Results ADPR n (%) Brainstem 1+ in either (4) 20 (16) Amygdala 0 1 in both (18) 24 (19) Limbic 1+ in either (21) 22 (18) Neocortical 1+ in either (54) 55 (44) Mixed Cases not classifiable by modified criteria 4 (3) 5 (4) SN subst. nigra; LADRS Lewy Body-Associated Dementia Research Study; ADPR Alzheimer's Disease Patient Registry; AD Alzheimer's disease. * Severity of LRP was scored according to published consensus criteria as none (0), mild (1), moderate (2), severe (3) or very severe (4). Leverenz et al, Brain Pathol 18 (2008)

32 Frequency of αsyn-positive lesions 100 % dmx Form. ret. Nucl. tr.sol. CA2/3 Striat. Limb. C. Cing. C. Nppd SNc Lc NBM Amygdala Neocortex Jellinger, JNT 111 (2004) PD DLB AD Aged controls

33 LB pathology in aging human olfactory bulb R Sengoku et al, JNEN 2008, 67: 1072 Material, methods: 320 consec. autopsies (81.5±8.5 yrs). Immunohistochemistry of paraffin sections. Results: LBs in 102 (31.9%) in CNS (29.4 in amygdala). OB involved in 85 (26.6%) ant.olf.nucl. + OB n= brains with pigment loss in SN had LBs in OB. LBs did not correlate with systemic tau or Aβ. high incidence of LBs in aging brain and strong correlation of LB grade in amygdala and anterior olfact. nucleus. OB is one of the initial anatomic sites affected by LBs and useful for diagnosis of LB disease.

34 Correlation between LB stage and grade of OBS Sengoku et al, JNEN 67 (2008)

35 Epidemiological and morphological data in LBD Values: range (mean) PD no dementia n=37 m/f 23/14 PDD n=33 m/f 16/17 DLB n=20 m/f 10/10 P Age at death [yrs] (77.6) * (84.8) * (80.4) * * < 0.01 MMSE (24.7) 0-15 (12.0) * 0-18 (13.5) * * < 0.01 vs PD Duration of illness [yrs] 0-26 (14.3) * 4-14 (8.5) * (6.5) * * < 0.01 Brain weight [g] (1250), n=24 * (1127), n= (1238), n=14 * * < 0.01 vs PDD PD Braak stage 2-5 (4.0) 3-6 (4.5) 5-6 (5.4) * * < 0.05 vs PD Neuritic Braak 0-4 (2.4) 3-5 (4.2) * 0-6 (3.8) * * < 0.01 vs PD Aβ striatum 1+ (5 %) * 2/1+ (26/20 %) * 3/2+ (65/25 %) * * < 0.01 Cortical Aβ plaque load 0-3+ (1.0) 0-3+ (2.6) * 0-4+ (3.0) * * < 0.01 vs PD Cap CAA 0-2+ (0.4) 0-3+ (2.0) * 0-3+ (2.2) * * < 0.01 vs PD Gen. CAA 0-2+ (0.2) 0-3+ (1.7) * 0-3+ (1.3) * * < 0.01 vs PD

36 Severity of α-synuclein and A β pathology in striatum in DLB and PDD phenotypes 100 % 80 DLB PDD (p < 0.001) / Striatal α-synuclein pathology Striatal A β pathology

37 Amyloid load in LB disorders [11C]PiB-PET Edison et al, JNNP (2008) 13 DLB, 12 PDD, 10 PD, 41 CO Results 11/13 DLB increased Aβ load 2/12 PDD raised Aβ (uptake normal) Increased Aβ in cortical assoc. areas, cingulate, striatum PD and CO no increased Aβ Conclusion In vivo PET suggests that Aβ in DLB contributes to rapid progression of dementia. Gomperts et al, Neurology (2008) 8 DLB, 7 PDD, 11 PD, 15 AD, 37 CO Results Cortical Aβ in DLB similar to AD > PDD PDD comparable to PD and CO Striatal PiB retention in DLB > PDD related to impaired motor function increased Aβ in cortical assoc. areas, cingulate, striatum PD and CO no increased Aβ Conclusion High cortical Aβ burden in DLB > PDD contributes to cognitive impairment.

38 Hypothetic diagramm unifying pathologic processes in Alzheimer disease and Lewy body dementia Pure form DLB (neocortical) Without acceleration by Aβ Propensity for αsyn aggregation DLB (neocortical) αsyn accumulation Acceleration by Aβ Pathological process likelihood Propensity for tau aggregation tau accumulation K. Obi et al, Exp Neurol 210 (2008) 409 AD

39 α-synuclein pathology in schizophrenia Western blot studies of Brodman area 9 (lat. frontal cortex) showed significant reduction of αsyn in schizophrenia and bipolar disease vs. controls (0.89±0.18 / 0.84±0.11 / 1.01±0.18; P=0.053) (Gray et al. Bipol Disord 2006;8:133). Personal preliminary studies: 30 autopsy cases of paranoid schizophrenia or residual state (age 50-85, mean 67.0 yrs), Braak stage 0-4 (mean 1.5). Immunohistochemistry frontal, hippocampus, midbrain, pons, oblongata. 29 cases negative; female aged 80 yrs without extrapyramidal signs: incidental Lewy body disease (LBs, LNs oblongata, few in s. nigra). Incidence LBD 3.3% (10-12% among healthy people < 60 years).

40 Lewy body-related pathology An update Summary I The two most frequent synucleinopathies, PD and DLB, are neurodegenerative disorders with widespread αsyn deposits in central, peripheral and autonomic nervous system. For both staging systems based on semiquantitative assessment of LBs are considered to be linked to clinical dysfunction. In PD, six stages with ascending progression from medullary and olfactory nuclei to the cortex have been proposed, the last two stages frequently associated with cognitive impairment, also related to presynaptic synuclein aggregation or combind (limbic) LB and AD pathologies. DLB, according to revised consensus guidelines, by semi-quantitative scoring of LB density in specific brain regions, distinguishes brainstem, transitory & diffuse cortical forms. Recent studies suggest PD and DLB to be synaptopathies.

41 Lewy body-related pathology An update Summary II Retrospective clinico-pathologic studies, although partly confirming the Braak LB staging system, have shown that up to 47% (!) of autopsy-confirmed PD cases did not follow the proposed caudo-rostral progression pattern. In 7-8.3% of autopsy-confirmed PD cases with LBs in midbrain and cortex (LB stages 4-5) the medullary nuclei were spared. These facts and comparison of nigral LB counts, αsyn burden and DAT IR in striatum showed no relationship between these data and clinical severity of PD. PDD and DLB, both frequently associated with varying AD pathology, show clinical and morphologic overlap, but more severe Aβ load in striatum and hippocampal LB intensity in DLB.

42 Lewy body-related pathology An update Summary III In large autopsy series 30-55% of elderly subjects with widespread LB-pathology revealed no neuropsychiatric symptoms or were not classifiable. Modifications of both the Braak and McKeith criteria in recent autopsy studies of demented elderly patients (Leverenz et al 2008; Fujishiro et al 2008) gave much better results. In conclusion, detection and staging of Lewyrelated pathology without assessment of neuronal loss in specific areas may have no clinical impact and its predictive validity needs to be revised.

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