NEURO 410 - Lecture 5: Recent Paper

What is a major issue related to the diagnosis of AD?

the most definitive diagnosis tool is observed pathology in post-mortem tissue

Why is post-mortem being the most definitive diagnostic tool a problem?

clinical trials need to know for sure if patients have AD

What are the two tools that must be used in combination to diagnose AD?

1. cognitive function changes

2. biomarkers in the CSF or brain

What was the eligibility criteria for the clinical trial of Lecanemab (recently FDA approved AD treatment)?

1. amyloid positivity as determined by PET or CSF measurement of AB1-42

2. objective impairment in episodic memory

Why is eligibility criteria in clinical trials important?

need to prove that a drug works in AD

What does PET stand for?

positron emmission tomography

What are radiotracers? What do the radiotracers used to diagnose AD bind to?

radiolabelled small molecules - designed to bind to AB and tau aggregates (and not the soluble forms of these proteins)

How does an amyloid/tau PET scan work?

radiotracer that is designed to bind to AB or tau is injected, accumulates at binding sites, the tracer decays overtime and as it decays it generates gamma rays, scanner detects and quantifies gamma rays

If there is more decay and the radiotracer and more gamma rays, what does this indicate?

more of the molecule you wanted to target is present

What percentage of patients that have the clinical criteria for AD are amyloid PET + ?

70-90%

What are the two major disadvantages of amyloid PET as a tool for AD diagnosis?

1. short half life of radiotracer means it must be made locally

2. huge cost of PET scanner

Why do we need both clinical cognitive symptoms AND amyloid beta PET/CSF positivity for AD diagnosis?

because cognitively normal people also have an increase in amyloid beta PET and CSF positivity with age

What do amyloid beta42 levels in the CSF look like with AD?

decreased AB

What do tau levels in the CSF look like with AD?

increased tau

What is the procedure for CSF sample collected?

lumbar puncture (spinal tap)

What are the 3 disadvantages to using lumbar puncture CSF collection to diagnose/monitor AD?

1. invasive technique

2. risk of headache

3. pain at collection site

What is an immunoassay that measures protein contents in blood or CSF?

Sandwich ELISA

Describe how a Sandwich ELISA works.

one Ab binds to plate surface, antigen binds this Ab, a second Ab with a biotin tag binds antigen, streptavidin binds the biotin tag, streptavidin is covalently bound to horse radish peroxidase (HRPO), HRPO reacts with TMB to generate a colored product

Why are tau CSF levels a better diagnostic method than AB CSF levels?

have an increase in CSF tau in AD, whereas you have a decrease in CSF AB in AD so it will be more difficult to detect

Why is tau so susceptible to post-translation modifications?

tau has over 80 potential phosphorylation sites

Which phosphorylated tau site correlates with AB and tau PET as well as with a decrease in cognitive function?

pTau217

Why are they looking at tau phosphorylation site 217, and not other phosphorylation sites on tau?

because not all phosphorylated tau sites correlate with AB and tau PET and cognitive decline

What do Ab and tau levels in the blood look like in AD?

same as what is observed in the CSF:

- decrease blood AB in AD

- increase blood tau in AB

What is the problem with detecting AB and tau in the blood?

protein levels are much lower in the blood then they are in the CSF, so traditional ELISA is not sensitive enough to measure these proteins in blood samples

What are the 5 advantages of using a blood test for the diagnosis of AD?

1. cheaper

2. samples easily collected and shipped

3. easy entry test - earlier diagnosis

4. identify more candidates for clinical trials

5. identify patients suitable for early treatments that are already approved

What were the 3 sample centers used in this papers experiment?

WRAP

TRIAD

SPIN

What was the diagnostic measure done in WRAP and TRIAD centers, which they would use to compare to their new diagnostic measure?

PET to detect AB and tau positivity

What was the diagnostic measure done in the SPIN center, which they would use to compare to their new diagnostic measure?

PET to detect AB positivity, and CSF to detect tau positivity (p181tau)

What is the new diagnostic tool that they are testing in this paper?

Sandwich ELISA plus SIMOA technology

What was the capture antibody (bead) that they used for the Sandwich ELISA + SIMOA technology?

designed to bind pTau217 specifically

What are the two mechanisms by which SIMOA technology increases the sensitivity of antigen detection compared to just a normal sandwich ELISA?

1. uses fluorescence, which can be detected at lower levels than color

2. separates a single sample into many samples

What is the immunocomplex that is created in SIMOA composed of?

bead

bound protein

detection Ab which can generate a fluorescent product

How does the specific pTau217 bead make this assay more sensitive?

one bead will only bind one protein, and each well is only big enough to fit one bead, so by examining fluorescence in each individual well, you can identify the exact number of proteins in a sample

What did they study see when they used sandwich ELISA + SIMOA technology to detect pTau217 levels in the blood?

pTau217 levels in the blood are higher if brain AB and tau are detected

What does a receiver operator characteristic (ROC) curve allow you to measure?

how good your test is at predicting a given condition

What does the x-axis of a receiver operator characteristic curve represent?

true positives / sensitivity

What does the y-axis of a receiver operator characteristic curve represent?

false positives / 1-specificity

What does a completely square line on a ROC curve tell you about your test?

it has true + every time, with zero false + (test has good predictability)

What does a linear curve on a ROC curve tell you about your test?

for every true+, you also have a false+ (test has bad predictability)

What did their ROC curve for the correlation between pTau217 and AB+ tell them?

at higher levels of ptau217 in the blood, more samples were AB + (true +) than were AB - (false +)

What did their ROC curve for the correlation between pTau217 and tau+ tell them?

at higher levels of ptau217 in the blood, more samples were tau + (true +) than were tau - (false +)

What did their ROC curve for the correlation between pTau217 and A+T- vs A+T+ tell them?

pTau217 blood test has poor predictability for whether someone is A+T- or A+T+ (sometimes pTau217+ samples were A+T-, which would be a false+ in this case)

What is a major advantage to using blood pTau217 as a diagnostic tool for AD in clinical trials?

blood pTau217 is higher in AD patients, and continues to increase more in AD patients than in MCI or NA