Targeting M1 muscarinic and sigma-1 receptors in Alzheimer ... · Blue: Aβ(McSA1) Brown: microglia (MHCII) Green: plaques (ThioS) Pre-plaque (early): Accumulation of intraneuronal

Targeting M1 muscarinic and sigma-1 receptors in Alzheimer's disease

Reversal of pathological hallmarks and associated cognitive dysfunction in McGill-R-Thy1-APP rats

Hélène Hall, PharmD, PhD

Postdoctoral fellow | Cuello labMcGill universityMontreal, Canada

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March 30 - April 2, 2014

Sheraton Sonoma County

Petaluma, California

Off-Label Product Use

Will you be presenting or referencing off-label or investigational use of a therapeutic product?

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Fisher, 2012, Journal of Neurochemistry 120(suppl 1)-22

M1 muscarinic receptors as a therapeutic target for AD

Stimulation of M1 muscarinic receptors

reduces amyloid pathology decreases tau hyperphosphorylation

Hélène Hall | McGill University | Cuello lab

AF710B (aka ANAVEX 3-71): a novel M1/sigma-1 agonist

AF710B drug (aka ANAVEX 3-71)

Agonist of sigma 1 receptor: enhanced cognition, neuroprotection(Senda et al, 1996, Eur J Pharmacol, Meunier et al, 2006, J Pharmacol Exp Ther.)

Allosteric modulator of M1 muscarinic receptor: modulation of the APP processing(Nitsch et al, 1992, Science; Caccamo et al, 2006, Neuron)AF710B

In 3xtg mice:


AF710B attenuates cognitive deficits

Reduces AD-like hallmarks (amyloid pathology, tau hyperphosphorylation, inflammation)

Fisher et al, 2016, Neurodegener Dis 16(1-2)-95

Express human APP, carrying mutations associated with familial AD, under Thy1 promoter

One single transgene insertion per allele, minimal genetic charge

McGill-R-Thy1-APP rats: a model of AD-like amyloid pathology

Leon et al, 2010, JAD

Post-plaque (late, >9mo):Deposition of Aβ into extracellular plaques

Wilson et al, 2016Cerebral Cortex

Leon et al, 2010, JAD20(1)-113

15 months 20 months

Blue: Aβ (McSA1)Brown: microglia (MHCII)

Green: plaques (ThioS)

Pre-plaque (early):Accumulation ofintraneuronal Aβ

Wilson et al, 2016Cerebral Cortex

Leon et al, 2010, JAD20(1)-113

Cortex

Aβ staining

Cognitive decline

Inflammation

Oxidative stress

Impaired LTP

4 6 9 12 months31 week

IntraneuronalAβ

Plaques

Hypometabolism

Hippocampal shrinkage

Mitochondrial dysfunction


1 week 6 months

Experimental design

FOR 5 MONTHS:AF710Bdaily treatment10µg/kg per os

Age 13 mo Age 19 mo

5 months 1 monthSTART END

Long-term treatment of McGill transgenic rats with AF710B, at an advanced stage of pathology

Cognitive decline

Inflammation

Oxidative stress

Impaired LTP

4 6 9 12 mo31 week

IntraneuronalAβ

Plaques

Hypometabolism

Hippocampal shrinkage

Mitochondrial dysfunction

13 mo


0.00

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crim

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*****

vehicle drug

AF710B fully reverts the cognitive deficit in aged McGill tg rats

**p<0.01, ***p<0.001two-way ANOVA

Wilson et al, 2017submitted

Day 2

2 min

30 min

3 min

Day 1

2 min

Novel Object Recognition

Hall et al, 2017, in preparation

Based on spontaneous differential exploration of familiar and new objects

Rodents will naturally spend more time exploring novel objects over familiar objects



**p<0.01, ***p<0.001two-way ANOVA

Wilson et al, 2017submitted

Day 2

2 min

30 min

3 min

Day 1

2 min

Based on spontaneous differential exploration of familiar and new objects

Rodents will naturally spend more time exploring novel objects over familiar objects

Novel Object Recognition

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Hall et al, 2017, in preparationHélène Hall | McGill University | Cuello lab

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Morris Water Maze


5 days of training to locate hidden platform

4 trials/day

Probe trial on day 6


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Representative swim paths

******

Performance on trial 4 - day 5

vehicle

drug

Morris Water Maze

5 days of training to locate hidden platform

4 trials/day

Probe trial on day 6

*p<0.05, **p<0.01, ***p<0.001; two-way ANOVA

Hall et al, 2017, in preparationHélène Hall | McGill University | Cuello lab

0

5

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CA1

DGsubiculum

tg drug

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Decrease in mature plaques (ThioS+)

AF710B reduces AD-like amyloid pathology in McGill tg rats

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Increase in CSF Aβ42

*p<0.05; two-tailed t-test

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Decrease in cortical Aβ42 (ELISA)

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Decrease in Aβ immunoreactivity (McSA1 staining)


vehicle

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Increase in cytokines and chemokines

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Microglia recruitment to Aβ-burdened neurons in CA1

vehicle

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AF710B reduces inflammation in McGill tg rats


19 months

Blue: NeuNBrown:Iba1

Early and late inflammation in McGill rats

Hanzel et al, 2014, Neurobiol Aging35(10)-2249

tg 13 monthstg 4 monthswt 4 months


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exp

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leve

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Hippocampal levels of Iba1

Iba1 (17 kDa)

GAPDH (35 kDa)

0

1

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Iba1

/GA

PD

H

**

Cortical IL 10 mRNA

transgene - - ++drug - -+ +

vehicle

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aver

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a1+c

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icle

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Microglia recruitment to Aβ-burdened neurons in CA1

vehicle

drug

vehicle

drug

Early and late inflammation in McGill rats AF710B has anti-inflammatory properties

Hall et al, 2017, in preparation*p<0.05, **p<0.01; two-way ANOVA

AF710B reduces inflammation in McGill tg rats

Increase in cytokines and chemokines

tg 13 monthstg 4 monthswt 4 months

Hanzel et al, 2014, Neurobiol Aging35(10)-2249


AF710B has synaptogenic properties

Synaptophysin (34 kDa)

Tubulin (50 kDa)

transgene - - ++drug - -+ +

At the postsynaptic levelAF710B restores synaptic spines

At the presynaptic levelAF710B restores synaptophysin levels


0.0

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Synaptophysin mRNA levels

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Synaptophysin protein levels


*p<0.05; two-way ANOVA

Fisher et al, Neurodegener Dis 16(1-2)-95

APP-KI

veh

icle

AF7

10

B

Iulita, Bistue et al, 2017, submitted

wt 20 month-old tg 20 month-old

Decrease in cholinergic boutons in McGill tg rats

VAChT immunostainingtgwt

AF710B has synaptogenic properties

0.0

0.5

1.0

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NA

exp

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leve

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vehicle

drug

*

*

Synaptophysin mRNA levels

Fisher et al, Neurodegener Dis 16(1-2)-95

0.0

0.5

1.0

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wt tg

syn

apto

ph

ysin

/tu

bu

lin

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vehicle

drug

Synaptophysin protein levels

At the postsynaptic levelAF710B restores synaptic spines

At the presynaptic levelAF710B restores synaptophysin levels


1mm

20µm

ONGOING

VAChT immunostaining

APP-KI


veh

icle

AF7

10

B

AF710B (a dual M1/Sigma-1 agonist) has long-lasting procognitive effectsat the late stages of the amyloid neuropathology in McGill-R-Thy1-APP rats.

Disease-modifying effects:

- Decrease in AD-like amyloid pathology

- Anti-inflammatory effect

- Synaptogenic activity

Effects maintained following a 5- week interruption in the treatment, suggesting true disease-modifying properties.

Conclusions


Acknowledgements

Dr A. Claudio Cuello

Adriana Ducatenzeiler

Dr M. Florencia Iulita, PhD

Lisi Flores Aguilar

Palma Gubert

Dr Mark Hancock

Dr Abraham Fisher

Richard and Edith Strauss Canada Foundation

ANAVEX is a trademark of Anavex Life Sciences

Documents

Targeting M1 muscarinic and sigma-1 receptors in Alzheimer ... · Blue: Aβ(McSA1) Brown: microglia (MHCII) Green: plaques (ThioS) Pre-plaque (early): Accumulation of intraneuronal