2/5/2015

1

Jeffrey H. Kordower, Ph.D..

Department of Neurological SciencesRush University Medical Center

� Sensitive:

� Discriminative: � Simple

� Inexpensive

� Example of a good biomarker:

� blood tests, CSF test but they are not discriminative for PD

Control 1 years PD 4 years PD

5 years PD 11 years PD 15 years PD 21 years PD

TH immunohistochemistry in the Putamen of human brain

3

2/5/2015

2

Premotor symptoms� olfactory dysfunction� autonomic dysfunction� constipation� depression, anxiety� REM behavior disorder� executive dysfunction

Signs/findings� presynaptic ↓ DA� ↑echogenicity of the SN� hyposmia� slight motor signs

Berg. Neurodegenerative Diseases. 2008;5:133

50% ↓DAneurons atsymptom onset; rapiddecline over 5y

� Parkinson’s disease is caused by exposure to

lipopolysaccharide (bacterial endotoxin), which promotes α-synuclein

overexpression/aggregation in neurons of the

submucosal plexus of the gastrointestinal system, then spreads via neural networks, to

the medulla, then rostally, with cumulative

neurological signs.

medulla oblongata

pontine tegmentum

basal, mid and forebrain

amygdala, hypothalamus, thalamus

cerebral cortex, mesocortex

cerebral cortex, neocortex

SN, IV

DMN X, II raphe, II LC, II

SN, IV

Aurbach, V

Braak & Del Tredici. Neurology 2008;70:1916

2/5/2015

3

Braak & Del Tredici. Neurology. 2008;70:1916.

Braak et al. Neuroscience Lettters 2006;396:67-72.

“…a putative environmental pathogen…might

induce α-synuclein misfolding and aggregationin specific cell types of the submucosal plexus and reach

the brain via a consecutive series of projection neurons.”

Grafts of dopamine cells placed into the striatum with viral over-expression of alpha synuclein

Note the physical segregation of the graft (brown) and gene delivery (black)

Kordower et al., Neurobiology of Disease, in press.

2/5/2015

4

A small percentage (5%) of grafted neurons retrogradely transported host-derived alpha synuclein

Kordower et al., Neurobiology of Disease, in press.

Synuclein Ubiquitin

Synuclein Ubiquitin

Host

nig

raG

raft

ed n

euro

ns

Kordower et al. Nat Med 2009;34:254.

1965-1968Honolulu Heart Program

8,006 Japanese ♂

1991-1993Honolulu-Asia Aging Study

3,741 Japanese ♂(80% of survivors of HHP)

2/5/2015

5

Incidence/10K person-years

Bowel

movements

Sample size Incident PD

cases*

Unadjusted Age-

adjusted

< 1 289 10 79.6 18.9

1 4371 66 8.0 7.9

2 1704 17 5.2 5.4

>2 426 3 3.8 3.9

Test for trend P=0.002 P=0.005

Overall 6790 96 7.5

Honolulu Health & Aging Study. Abbott et al. 2001;57(3):456.

•Appears highest in men with <1BM/day and laxatives > 2/week: 51.6/10,000 person years

� Can we demonstrate early GI involvement in PD?� biopsy distal colon in early PD

� Is the gut “leaky” in PD subjects?� intestinal permeability studies

� evidence for endotoxin exposure▪ Systemic

▪ local

� Do PD subjects have “dysbiosis? (alterations in intestinal flora)”� DNA “fingerprinting”

� Early untreated “classical PD”� no coagulopathy, alcoholism, occupational

exposure to microbes, primary GI disease

� Unprepped flexible sigmoidoscopy with biopsy� immunohistochemistry (synuclein, nitrotyrosine)

� Differential sugar absorption� sugar ingestion

� 24-hour urine collection with assays

� 16S rRNA “fingerprinting” for dysbiosis

2/5/2015

6

Age/sex Disease

Duration

Onset /dx (y)

tUPDRS HY #BM/d

55M 4/.25 28 2 1

66M 1/.5 27 2 1

56M 1/.75 15 1.5 <1

78M 1/.5 20 2 1

75M 4/.1 28 2 <1

68F 0.5/.33 24 2 2

46F 2/1 16 1 <1

49M 8/12 18 2 1

61M 1/.5 18 2 1

51M 2/1.5 28 2 1-2

58.5 1.5/.75 22 2Median

Controls (N=26) Crohn’s (N=14) Ulcerative colitis (N=11)

Age, median(range) 54 (36-71) 40 (24-72) 43 (21-62)

Gender, male 12 (46%) 6 (55%) 4 (36%)

Parkinson disease

Crohn’s disease

Control

2/5/2015

7

Alpha synuclein Nitro-tyrosine

A

C

E

D

F

B

Parkinson’s

Ulcerative

Colitis

Aged-Matched Control

A

C

B

D

α-synuclein N-tyrosine� 85-y/o woman

� Psychotic depression 2002→ECT

� MCI

� Rest tremor 2/2010

Colonic polyp biopsied 2005.

MPTP MPTP

Aged Aged

Young

control

Young

control

low

low

low

high

high

high

2/5/2015

8

Alpha syn TH Merged

Sugar Site

Sucrose Gastric

Lactose/mannitol Enteric

Sucralose colonic

24-hour sucralose excretion

0

0.5

1

1.5

2

2.5

control PD

*p=.013

2/5/2015

9

LBP (ng/ml)

0

50000

100000

150000

200000

250000

300000

control PD

*p=0.016

� 1 kg bacteria� most unknown identity� most anaerobic� luminal population & mucosal biofilm

� 1012 microbes/ml luminal content� Established by age 2; stable� Important because…

� genetic component (100x as many genes…)� metabolic efficiency� energy homeostasis� important in systemic inflammation▪ HIV, coronary artery disease, insulin resistance, EtOH liver disease…

� Gram (–) rods

� bacteroides

� desulfovibrios

� escherichia

� fusobacteria

� Gram (+) c0cci

� ruminococci

� peptostreptococci

� peptococci

� streptococci

� Gram (+) rods

� eubacteria

� bifidobacteria

� clostridia

� lactobacilli

� propioibacteria

� actinomyces

� Gram (+) coccobacilli

� methanobrevibacter

Salminen et al. Br J Nutr 1998;80:S147.

2/5/2015

10

Method Advantages Disadvantages

morphology/biochemistry straightforward, cheap subjective, culturable only

specific biomarkers may not require culture can’t help with unknown

species, requires unique biomarker

ribotyping (RNA

polymorphisms)

reliable, high

discriminatory power

culturable only

16S ribosomal RNA typing high fidelity, reliable,

cumulative database, culturable & non-

culturable

costly

Salminen et al. Br J Nutr 1998;80:S147.

Parkinson disease

(N=10)

Crohn’s disease

(N=11)

Ulcerative colitis

(N=13)

Controls

(N=26)

Age, median

(range)

57 (46-79) 40 (24-72) 42 (21-62) 55 (36-71)

Gender, male 70% 55% 31% 46%

Duration PD (y)

Median (range)

1.5 (0.5-8)

Total UPDRS 22 (15-28)

HY Stage

III

28

Healthy Lumen

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

1 2 3 4 50%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

1 2 3 4 5 6 7 8

Parkinson lumen

Healthy sigmoid

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

1 2 3 4 5 6

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

1 2 3 4 5 6 7 8 9

Parkinson's sigmoid

2/5/2015

11

PCO case scores (Bray Curtis)

Control lumen

Control mucosa

Parkinson mucosa

Parkinson lumen

Axi

s 2

Axis 1

-0.1

-0.2

-0.3

-0.4

-0.5

0.1

0.2

0.3

0.4

-0.1-0.2-0.3-0.4-0.5 0.1 0.2 0.3 0.4

All Parkinson

Age/sex Disease

Duration

(y)

tUPDRS HY #BM/d α-synuclein nitrotyrosine Abn mucosa Abn

lumen

55M 4 28 2 1 ++ ++ √ √

57M 1 27 2 1 ++ ++

48M 1 15 1.5 <1 ++ ++ √ √

77M 1 20 2 1 ++++ ++++

74M 4 28 2 <1 ++ ++ √ √

68F 0.5 24 2 2 ++++ √ √44F 2 16 1 <1 ++++ √48M 8 18 2 1 ++++ ++++

60M 1 18 2 1 ++++ ++++ √

57M 2 28 2 1-2 ++ ++++ √

2/5/2015

12

2/5/2015

13

GI changes, such as a leaky gut or alpha synuclein in thecolon demonstrates excellent sensitivity

Colonic biopsies can be a sensitive and inexspensive biomarker

Still to be proven is how specific this potential biomnarker is and studies in PSP and MSA patients are currently underway

Kathleen Shannon, M.D.Ali ]Keshavarzian, M.D.

Christopher Forsyth

Hemraj DodiyaLeo Kelly