AD

Question 1

outline and describe the main histopathological features of AD

Answer 1

Severe atrophy of neurons. looking at a brain i postmortem it will have large sulci and enlarged lateral ventricle.
severe atrophy of the HPC is also a sign related to early spatial memory deficits

Extracellular depostions of amyloid beta plaques. Large insoluble aggregates fromed from the abberant proteolysis of precursor APP.
A-beta deposition is also visualised in cerebral amyloid angiopathy (CAA) where plaques form in the walls of blood vessels.

Intracelular inclusions of NeuroFibillary Tangles. These consist of hyper phosphorylated microtuble associated protein Tau. (MAPT). look like Whispy Flames.

Neuroinflamation. This can be seen by the surrounding of plaques by microglia that have undergone hypertrophy to enter a Ramified state.

Question 2

briefly outline the Thal phases of AD compare this to BRAAK staging of AD.

Answer 2

THAL phases tracks the progression of AD through the depostion of AMYLOID BETA. spread being anterograde transmission to regions innervated by infected regions.
1- neocortical depostion
2- inclusions of allocortical regions
3- involvement of striatum, cholinergic nuclei of basal forebrain, diencephallic nuclei
4- inclusion of several brain stem nuceli
5- the final phase with inclusion of the cerebellum.
Phases 1-3 are associated with the preclinical phases and phase 4+5 with the clinical symptoms.

Braak staging tracks the deposition of tau pathology (neuro fibilliary tangles)
1+2- these involve primarily the entorhinal cortex and hippocampus.
3+4- these involve the inclusion of limbic structures
5+6- nemerous regions of the cortex now included.
1+2 confer a non symptomatic stae with the onset of symptoms in stages 3+4 and 5+6 conferring severe stages of AD.

Tau progression has been shown to correlate with the progression of AD. However, Amyloid beta depostion has not with many dying with abundant amyloid beat deposits and no symptoms. Hence, Braak staging may be more reliable in AD.

Question 3

provide a breif descripion of the clinical progression of AD and its correlation pathological changes.

Answer 3

The progression of AD follows a predictable pattern of atrophy.

atrophy can be visualised by MRI far before the symptomapric phase in a preclinical phase. early on here atrophy can be seen in the EC and HPC, are associated with spatial cognition, something that can be seen in ealry phase AD patients.

soon atrophy in limbic and later cortical areas is observed. conferring with the onset of symptoms like memory loss and later confusion, and the detrioation of intelectual capacity associated with dementia.

Although a hall mark of AD histopathology this does not correlate with the spread of AB pathology. However, it does correlate with the predictable spreading pattern of tau pathology described in Braak staging.
1+2- this being the preclinical phase where there is accumualtion in EC and HPC areas, confers with spatial cognition deficits.
3+4- There is inclusion of limbic areas and the onset of symptoms like memory loss confer with this.

5+6 this is where multiple cortical regiosn begin to be involved, this is seen in late stage AD and correlates with the progressive severity of dementia.

Question 4

what are the main therapeutic targets in AD?

Answer 4

stop amyloid plaque formation
stop the formation of hyperphosphorylated Tau tangles
reduce neuroinflamation (thought to be a cause of issues with synaptic efficacy, a early contributon to neurodegenerative cascades)
modulate neurotransmission
use genetics to identify novel targets

Question 5

what are the early behvaioural warning signs of AD?

Answer 5

memory loss, in particular issues with spatial memory as the hippocampus is degenerated early on, we can assess this through the 3 peaks study which test both spatial memory and perception (perception is normally fine)

changes in personality
issues with pallning and decision making.
misplacing things,
change in mood
disorientiation is space an time.
difficutly performing familiar tasks.

Question 6

what risk of bias can be associated with randomised studies of neruodegenrative disease like AD, what is the issue of the currently known markers?

Answer 6

In neruodegenrative conditions the rate of progression can vary alot.

Randomised smapling runs the risk of having fast progressor and slow porgressors dominantly in placebo vs test trials. this can cause false positive/negative results when analysing the progression od symptoms in tertiary studies or disease markers in secondar trials.

The issue with current biomarkers is there are alot of RETROSPECTIVE markers that provde inof on risk of onset but very few that tell uss how a disease will progress. making it hard to distinguish between fast and slow progressors.

Question 7

Tertiary efficacy outline a test used to measure the change in AD patient cognition? Give an example of a clinically meaningful exam.

Answer 7

ADAS-COG (Alzheimers disease assessment scale cognitive subscale) this is used to assess the changes in cognitive symptoms. the score ranges between 1-70 with normally elderly individuals scoring 0-1 and AD patients score usually increasing yearly. (score is number of errors

A clinically meaniful exam would be the Clinical interview based impression of change with carer input (CBIC-plus). this is a score of improvement between 1-7

either halting of worsening or improvement is considered successful.

Question 8

importance of placebo efficacy on AD trials for tertiary cures

Answer 8

Here improvement in test drug and decline in placebo is a sign of efficacy.

However, improvement with drug and no decline in placebo is not strng evidence of efficacy. as this could simply mean that trial drug is beinmg tested on slow progressors.

Question 9

In secondary studies what is seen as a sign of effectiveness, what do you need to assess this.

Answer 9

Secondary studies a sign of effectiveness in the halting of progression of a dynamic biomarker (cannot uno damage)

In order to assess this you need a dynamic biomarker who changes correlate with the progression of symptoms. in PD this is MRI to investigate atrophy of neural tissue that racks decline in cognition.

Question 10

Why is amyloid plaque accumualtion not a good biomarker of disease progression, How can we visualise it and what is it used for?

Answer 10

amyloid plaques do accumualte throughout the disease and indeed accumualtion can be seen even 15 years before onset as a toxic threshold is need for disease.

This can be visusalised using postiron emisiion tomography using a Amyloid specific radioactive marker.

Question 11

Give the best cuurent exmaple of a dynamic marker that can be used to track changes in disease pogression for secondary trials of AD?

Answer 11

currently the best marker is magnetic resonance imaging of the brain.
There used to be some issues as movement of the head could make images appear to be shifted and the images never where done with the exact same point of reference.

Now days a computer program can precisely sperimpose several images alllwoing the visualisation and calculation of the prgressive atrophy. we now Know in a healthy 40 year old this is 0.2% a yar and rises to 2.8% a year in AD hence this is a rnage of reference for reductions in progression.

severe atrophy is is seen far before the onset of symptoms and Atrophy does correlate with the progression of cognitve deffiencies.

Question 12

How do you overcome issues of bias in secondary efficacy trials?

Answer 12

In secondary trial the risk of fast and slow progressos still exists so instead STRATIFIED SAMPLES are used.
The take individuals once they have been analysed and are confirmed fast or slow progressors.

Although logic suggest using fast progressors as the change in disease progression will be most visible.
It is also importnat to test in slow progressors as fats progressors once identified may have already passed a irreversible threshold and thus may produce false negatives.

Question 13

issues with using Event related pottentials (ERP) as a marker of impairment and progression in AD, equally wat are the weakness of EGG.

Answer 13

ERP work as declines have been both correlated with cognitve deffiencies and disease progression in AD. usually tested using a constat repeated tone with infrequent changes to investigate the neuronal reponse. this is reduced in amplitude and latency

Issue is this is aslo observed in PD, schizophrenia and dyslexia and so is not a specific AD test.

simmilarly testing for EEG in which in AD patients we observe a progressive INCREASE in slow wAVE THETA AND DELTA WAVE AND reduction IN FAST BETA AND ALPHA WAVES. this is also observed with normal aging in elederly and several othe rpathological isues.

Question 14

Benefits of double blind

Answer 14

In these trial neither the patient o experimenter Know who has the placebo and who has the dru. ence, ths can reduce psychological or experimental bias, improving the validity of findings.

Question 15

Can we diagnose AD in life?

Answer 15

NO given the biomarkers today we can predict the likelihood of AD onset and the probability that somebody has AD but we cannot diagnose it as AD until postmortem studes are done.

Question 16

outline the process by which Amyloid beta plaques from APP and how Neurofibilliary tangles form from TAU.

Answer 16

Amyloid beta is produced by the from th precursor APP.
APP is usually cleaved by alpha-secretase and then gamma-secretase this produces the normal peptides.
However, it can be alternatively cleaved by Beta-secretase and them gamma secretase froming the toxic amyloid beta peptides. gamma secretase can cleave at multiple points primrily producing amyloid beta 40 or 42. These will then form plaques. 42 has MORE hydrophobic amino acids and further promotes aggregation. (Idea is that then mutations in APP or subunits of gamma-secretase will increase the cleaving to produce amyloid beta)

NFTs- Tau is a microtuble associated proteina (MAPT) stabilises microtubules. It binds via c-terminal repeats of which there are 2 (3R+4R). The Reverisble binding is regulated by phophorylation.However, in pathogenic state there is hyperphosphorylation of MAPT resulting in the formation of Paired helical filaments of MTs and TAU. These are deposited intracelluarly as NFTS.

Note there are normally equall amounts of 3 and 4 but in AD taus express the C-terminal repeat 4 more than 3.

Question 17

what is the social importance of treating alzheimers?

Answer 17

We have an aging population and the greatest risk f AD is age. hence, the AD population is growing.
forms of dementia like this are evermore prevalent with 1:3 being ascoiated with someone with dementia.

The estimates cost to society is 26billion a year.

Hence is is of great soceital importance to treat this condition.

Question 18

what are the main risk factors of AD out side of genetics?

Answer 18

Age is the main risk factor of AD.

Question 19

outline and decribe some of the identified genetic causes and risk factors (not including GWAS) of AD. (3)+(2)

Answer 19

Monogenic EOAD: less tan 5% of cases are fammilial.

APP mutations- this is the precursor of amyloid beta and muations here promote cleavage via the beta-gamma pathway and thus increase the production of AMyloid beta.

Trisonomy 21- this is the cause of down syndrome and chromosome 21 in the location of the APP gene. Hence, this is a dose dependat increase in APP and subsequently in Amyloid beta.

PSEN1+PSEN2- these are subunits of gamma secretase and mutations promote the lesion to produce the aggreagation-enhanced Amyloid beta 42.

• Lee et al 2010 report effects besides this showing that PS1 KO in cells prevented the provision of a proton transporter to lysosomes causing dysfucntion with their acididification and disrupting autophagy. Studies reproted that this led to axonal swelling filled with autophagic vacuoles containg incompletely degraded proteins.
• Issues with autophagy have been previously related to increased Plaques and boosting it has been related to partial recovery of cognitive deficits in mouse models.
• It may be that the lifelong reduction in the efficacy of Autophagy reproted by CUERVO ET AL 2008 may expalin largely the increased risk with age.

Risk factors (removal)

APOE4- there are 4 allels of this gene, APOE2 asscoaited with protection against AD, APOE3 which is AD neutral and APOE4 whcih increases the risk of AD. These are involved with the transport of cholesterol to neuron and APOE4 is thought to slow A-BETA removal.

TREM2- triggering receptor on myeloid cells 2. This is associated with the response to amyloid beta by microglia.Thus mutations inhibit the immune removal (phagocytosis) of toxic amyloid beta.

• There is also evidence that APOE4 and TREM2 intercat to this same effect.
• This suggests that dysfucntion of microglia is not a secondary fetaure of AD but is instead a primary factor.
• Alteration in TREM2 levels in the CSF are now being investoigated as a biomarker of AD.

Question 20

what is dementia

Answer 20

Dementia is caused by several diseases and is defined by the detrioration of intelectual capacity. (Judgement , memory e.t.c)

Question 21

outline the amyloid beta theory and the brief evidence behind it. include varied roles of tau an amyloid in pathology.

Answer 21

The amyloid beta theory states that abberant proteolysis of APP froms toxic species of amyloid beta. These then act ia 3 mechanisms.

1- they intefear with the kinases asscoiates with MAPT an drive the formations of hyperphosphorylated tau and NFTs, leading to neruonal death.

2- AB plaques form and drive the subsequent dysfunction of neruons and contributes to neuronal death.

3- the fromation of soluble AB oligomer causes synaptic dysfunction (shown in mouse models) and contributes to neuronal death.

hence within this theory AB acts upstream of NFT formation and neuronal death.

evidnece:
Mutations related to the fromation of AB can cause familial AD suggesting it is the driver of the disease.

Crossing APP mutant mice with a tau KO was unable to cause pathology, neuronal death or cognitive deficits. But APP cross with WT did. This suggests Tau pathology is key to the neurotoxic aspects of the disease and AB is key to the onset. (Hence need to target both)

MAPT mutations do not cause AD (although they do cause deeneration asscoiated with FTD shown by P301 mutation mouse models)

Clavaguera et al 2015
Injection of Mutant Tau fibrils homogenates from human tissue into mouse models was able to induce the misfolding of endogenous tau creating simmilar inclusions to those injected. Using human specific and murine specific antibodies they showed that murine Tau has indded misfolded. This suggestsed that ince misfolded Tau pathology an become self maintaing.

They also reported that the crossing of APP muatant mice with the ALZMice model. The APP mutation could pottentiate th spread of tau.

Question 22

Do Tau mutations have varied impacts of AD an FTD.

Answer 22

MAPT muations do not cause AD. However, they have identified roles in FTD.

Question 23

discuss the efficacy and usefullness of early mouse line trials in targetting Tau kinases in therpeutics?

Answer 23

GSK3beta and CDK5 are both kinases that have been associated with the phophoryaltion of tau. trials have been run in mice attemtping to stop this to stop the formation of NFTs.

This was able to reduce progression of cognitive defects.

But these are not clinnically useful because the fucntion of Tau phosphryaltion and the function of these kinases besides tau is important and thus this has severe off target effects.

Question 24

discuss the importance of critical periods of actions to therapeutics. Imapact of likely hood of certain targets being effective.

Answer 24

amyloid beta accumualtion and atrophy can be seen 15+20 years, respectively before symptoms onset. hence this offers a critical period of action in which we can stop AB action efore it is too late.

aafter a period tau pathology became self-maintaining. thus there is a periodin which inhibtiing the formation of AB would not stop disease progression and thus stopping the action of tau is also important.

Question 25

How has GWAS been used in AD and what is the next step?

Answer 25

GWAS has been used to idnetifiy several common variants that contribute to the risk of AD. there are now 20.
However, these all confer relatively small risks.

The next step is to investigate the effcts of cummlative risk. would it increase rsk to have 5 muattions in different aspects of a pathway as compared to 1 muatation.

Question 26

outline the investigation of mechanism of how tau pathology spreads.

Answer 26

A indepth study has been performed. in neuronal cell cultures pulled from the MAPT mutant P301 mouse line

In Braak staging we see that the pathology spreads from the EC and then to limbic regions and thus it appears to spread between connected regions.

Expressing the MAPT mutant neuron in culture with a WT neuron the MAPT acts as a donor. After 2 WEEKS intracellular Tau inclusions are seen in the wild type neurons.

They investigated whether direct contact was needed:
In IPSC cells they placed mutant tau neurons and tau KO neurons in a medium and then tested the medium for phosphoryalted tau and the tau-KO neurons using tau specific antibodies in an ELISA test. This showed that the TAU-KO neuron took up tau from the medium and thus cell to cell contact is NOT neccesary.

Finally the investigated the role neural activity.
They expressed the optogenetic channel; CHR2 in tau expressing neurons and stimulated them whilst assessing thelevel of tau in the medium. they reported a 250% increase after 30 mins of stimulation.

Hence, it would appear that mutant tau can spread between cells and thus neural regions connected by active synaptic connections.

Many believe that NFTs induce the abberant folding of other tau molecules

Question 27

What is the theory on how Tau dysfunction may cause neuronal dysfucntion and death? (evidence)

Answer 27

Some studies have linked this LOF of tau.

Ittner et al (2010), showed that tau has a key role in synaptic targetting. She found that the expression of truncated forms of tau or KO led to a decline in dendritic Src kinase Fyn. Thus indicating a decrease in synaptic targetting.
The NMDA receptor is a substrate of Fyn and thus this uncouples this synaptic link.
They showed expressing truncated Tau or KO tau in APP mutant mice was able to attenuate memory deficits and neurodegeneration, Suggesting is may prevent NMDAR mediated toxicity stemming fron AB action.

Hence, Tau is neccesary fro AB toxicity.

Question 28

who are possibly a underused population of patients for AD therapy trialing

Answer 28

own syndrome patients, due to trisonomy 21 they have an extremely high likelyhood of AD and thus act as a good sample who can be treated in the preclinical phase to investigate the efficacy of stopping AB formation. This is an issue with normal trialling and you dont know if participants have already progressed too far.

Question 29

what are the links of amyloid beta oligomers instead of aggreagtaes to neuronal dysfunctions?

Answer 29

studies have indicated that plaques may be an attempt to sequester toxic soluble AB oligomers.

studie have suggested these oligomers disrupt synaptic function in mouse models. The showed DECREASED SYNAPSE NUMBER, DECREASED LTP in rodent hippocampus. SHANKAR 2008, these were features he found were specific to soluble species and no AB cores. He sourced these directly from AD human brains.

In human Post-mortem they found a distinguishable difference between mildly and severely demented patients was the level of AB oligomer, identified by ELISA, higher when severe.
This suggests in worsens of pottentiates the severity of the pathology and thus may cause toxicity.

Question 30

Discuss the heterogeneity of Amyloid beta

Answer 30

The heterogenitiy of AB peptides mainly 40 and 42 makes it hard to target them with therpeutics

in addition their are forms like AB 43 that aggeragate much later.

• Receently this proved pottentially beneficial with the idnentification of the late agreagting P3E form. Tey found targetting this specifically causes a phenoemena of bystander clearance in which other forms were cleared along with it. Offers a pottentail therpeutic target.

There is growing research to devlop specific antibodies fro eaach to better undertsand the contribtion of each to patholgy, these would also be able to drive nearby microglia to clear the AB build up.

Question 31

what is the prion-like spread theorised in AD

Answer 31

There is an argument over whether the spread of patholpogy is temporally dependant, I.e mre vulnerable neurons go first and more resistant later.

Or is this linked to physiological spreading of pathology. This is what would be suggested by Braak staging and optogenetic study on the spread of tau.

the prion like theory is that misfolded protein act as templates to drive the misfolfing of endogenous proteins through permissive templating thus spreadinmg apthology and dysfucntion.

studies have now shown that the use of surgical tools used in AD can transfer AB pathology to new patients. This Iatrogenic inheritance suggets The may spread in a prion like fashion.

Question 32

Pottential biomarker

Answer 32

CSF AB this is elevated and then begins to decline concomittantly with deposition.
Simmilarly tau CSF is thought to track disease. Recenetly CCL11 is believed th=o reliably track Ta levels in th eCSF and is being invetsigated in CTE.

Cognitive impairment is often too late. by things like the 3 peak test can be sen to spot early signs of spatial cogntive defects.

PET imaging for AB fibrilialy amyloid plauqes is falry robust. Using the like of Thioflavin T deriviative.`

Question 33

Therapeutics imunotherpay

Answer 33

Been looking at AB imune therpaies. It would seem tragetting AB oligomers is best.

issues with recent trial is some show a larger affinity for oligomers and thus high doses are required to taregt oligomers.

• Tragetting the N-terminal sites of aggeragtaes has been shown to cause diaagregation and actually release oligomers cause increased toxicity.
• Seems mid plaque recognises looking at C-terminal recognition are less detrimental.