Plasma Biomarkers: Alzheimer’s Disease Detection
Alzheimer’s disease (AD) is a prevalent disease that continues to perplex the scientific community and devastate individuals and families. Researchers are committed to unraveling the complex pathways of this disease and providing tools and insights to improve AD diagnostics and quality of life for patients and caregivers.
Accurate and reliable detection of plasma biomarkers indicative of early Alzheimer’s disease pathology allows researchers to run more efficient clinical trials, monitor disease progression, and increase the likelihood of finding a cure. Learning about the biomarkers under investigation and the available technologies will help you identify the most promising options to target and explore for your research.
- AD diagnostic paradigm: How it’s changing
- Plasma biomarkers: Alzheimer’s disease detection
- Today’s most promising biomarkers
- Emerging biomarkers to explore
- Additional promising biomarkers to consider
- The ATN classification system
- The need for ultrasensitive assays for detecting plasma biomarkers in Alzheimer’s disease patients
AD diagnostic paradigm: How it’s changing
For most of the past century, an examination during autopsy was the only way to reach a definitive Alzheimer’s diagnosis. The advent of positron emission tomography (PET) brain imaging and cerebral spinal fluid (CSF) sampling made it possible to identify the disease in patients before the initial symptom appeared. However, those tests are invasive, expensive, and generally limited to research settings as opposed to being part of routine care.
Blood tests examining plasma biomarkers of Alzheimer’s disease could make early detection a reality and pave the way for reliable prevention and treatment
An overwhelming majority of primary care physicians believe it is crucial to diagnose mild cognitive impairment (MCI) and that early intervention is critical to managing AD. Blood tests sensitive enough to detect AD biomarkers could be the path forward — in the clinic and the research lab.
Benefits of a blood test for clinical diagnosis
- Routine screening for at-risk populations
- Repeat testing to monitor disease progression
- Less invasive
- Easy to administer
- More cost-effective than alternatives
Current screening techniques for pre-clinical AD research studies are expensive and invasive or symptom-based and imprecise
The two major hallmarks of Alzheimer’s disease are the accumulation of beta-amyloid protein (called beta-amyloid plaques) outside neurons and the accumulation of abnormal tau protein (called tau tangles) inside neurons. However, since these are not yet easy to measure, Alzheimer’s dementia is frequently diagnosed based on symptoms.
The problem arises when individuals are classified as having Alzheimer’s dementia without having the biological changes associated with the disease. PET scans and CSF sampling reveal that 15% to 30% of patients who meet the criteria for Alzheimer’s dementia based on symptoms do not have Alzheimer’s-related brain changes. Enrolling these participants in AD trials could lead to data artifacts and dramatically skew research findings, derailing treatment insights. Unfortunately, PET scans and CSF sampling are not viable tools for clinical trial enrollment because they are expensive, invasive, and often inaccessible. A blood test able to discern between AD and non-AD individuals could dramatically reduce costs, speed up study enrollment, and ensure high levels of diagnostic accuracy.
However, before a blood test enters the clinic, we need to know what biomarkers indicate AD and which can be reliably measured. AD research gives us some leads and insights into these critical questions.
Today’s most promising biomarkers: Amyloid Beta, pTau, GFAP, Neurofilament Light
Amyloid beta (Aβ) peptides are one of the most widely researched biomarkers for Alzheimer’s, MCI, dementia, and other cognitive disorders. In a diseased state, Aβ accumulates in the brain and forms extracellular plaques, which likely play a crucial role in the neurodegenerative processes.
Did you know? Aβ deposition may begin two decades before clinically noticeable cognitive impairment (Selkoe et al., 2016), at which point Aβ levels in the surrounding fluid decline before eventually reaching a plateau.
Of particular importance are amyloid beta-40 (Aβ40) and amyloid beta-42 (Aβ42). Both are proteolytic products from the amyloid precursor protein (APP). Beta-secretase cleavage of APP initially results in producing an APP fragment that is further cleaved by gamma-secretase at residues 40-42 to generate the two primary forms of amyloid beta.
Aβ40 is considered an initiating factor of AD plaques. In healthy and disease states, Aβ40 is the most abundant form of the amyloid peptides in cerebrospinal fluid (CSF) and plasma, with an average concentration of 10–20X higher than Aβ42.
In addition to measuring Aβ40 and Aβ42 individually, researchers commonly look at the Aβ42/Aβ40 ratio to distinguish AD from non-AD patients (Lehmann et al., 2018). Furthermore, the Aβ42/Aβ40 ratio is uniquely useful as an ongoing measurement indicative of AD progression.
Although cells throughout the body produce Aβ peptides, the expression is exceptionally high in the brain. Concentrations of Aβ in the blood are over 100-fold lower than in cerebrospinal fluid, requiring high analytical sensitivity for its reliable measurement.
Measuring Aβ in plasma presents unique challenges due to the low concentrations but has strong diagnostic potential and can accurately differentiate between amyloid positive and amyloid negative patients (West et al., 2021, Benedet et al., 2022).
Tau levels are elevated in the CSF and plasma of patients with neurodegenerative disease and head injuries, suggesting its extracellular release during neuronal damage and its role as a brain injury biomarker. In Alzheimer’s disease, tau is abnormally phosphorylated and aggregated into bundles of filaments.
Tau is a microtubule-stabilizing protein primarily localized in central nervous system neurons but also expressed at low levels in astrocytes and oligodendrocytes. It consists of six isoforms in the human brain, with molecular weights of 48,000 to 67,000 daltons, depending on the isoform.
The potential movement of elevated CSF tau across the blood-brain barrier presents the possibility that measurements of tau protein in blood could offer a convenient peripheral window into brain/CSF status. Unfortunately, studying tau in serum and plasma is challenging since its abundance is low. However, the advent of ultrasensitive assays has opened the door for investigating these highly informative biomarkers.
Plasma tau phosphorylated at residue 181 (p-tau181), tau phosphorylated at residue 217 (p-tau217), and tau phosphorylated at residue 231 (p-tau231) are of particular interest in AD research and have been shown to demonstrate the strongest overall sensitivity and specificity for AD neuropathological change (Smirnov et al., 2022).
Studies show that p-tau181 is increased 3.5-fold in AD patients and could distinguish AD from other neurodegenerative disorders almost as accurately as PET scans and CSF measurements (Thijssen, 2020). Additionally, higher p-tau181 is associated with a steeper cognitive decline and gray matter loss in temporal regions (Simrén et al., 2021).
Cerebrospinal fluid p-tau217 may perform better than p-tau181 as a biomarker of Alzheimer’s disease (Janelidze et al., 2020). This biomarker also shows promise when detected in plasma. Mattsson-Carlgren et al. showed that plasma p-tau217 increases during early Alzheimer’s disease and can be used to monitor disease progression (Mattsson-Carlgren et al, 2020).
Until recently, p-tau231 hadn’t been studied in plasma. Using Quanterix’s ultrasensitive single molecule array (Simoa®) technology, Ashton et al. developed custom assays and found that plasma p-tau231 could be used to differentiate the clinical stages of AD and neuropathology earlier than plasma p-tau181. Plasma p-tau231 increased earlier than plasma p-tau181 and showed measurable changes before Aβ positivity could be identified on a PET scan (Ashton et al., 2021).
Glial Fibrillary Acidic Protein (GFAP) is a class-III intermediate filament majorly expressed in astrocytic glial cells in the central nervous system. Astrocytes play critical roles in supporting, guiding, nurturing, and signaling neuronal architecture and activity. In addition to forming homodimers and heterodimers, GFAP can polymerize with other type III proteins or neurofilament proteins such as Nf-L. GFAP is involved in many important CNS processes, including cell communication and the functioning of the blood-brain barrier.
High plasma GFAP concentration is a relatively strong indicator of AD pathology. Furthermore, it may accurately predict the clinical progression of AD dementia (Cicognola et al., 2021), making it a promising biomarker to assess in blood-based panels.
- 2021 Nature: Plasma glial fibrillary acidic protein is elevated in cognitively normal older adults at risk of Alzheimer’s disease
- 2021 JAMA: Differences Between Plasma and Cerebrospinal Fluid Glial Fibrillary Acidic Protein Levels Across the Alzheimer Disease Continuum
- 2022 BMJ: Serum GFAP differentiates Alzheimer’s disease from frontotemporal dementia and predicts MCI-to-dementia conversion
Neurofilament light (Nf-L) is an axonal cytoskeletal intermediate filament protein expressed only in neurons. It associates with neurofilament medium (Nf-M) and neurofilament heavy (Nf-H) to form neurofilaments. Together they are significant components of the neuronal cytoskeleton and provide structural support for the axon and regulate axon diameter. Of the subtypes, Nf-L is the most abundant.
Since neurofilaments can be released in significant quantities following axonal damage or neuronal degeneration, elevated plasma levels of Nf-L are associated with neurodegeneration and dementia in Alzheimer’s patients. Thus, plasma Nf-L may serve as a non-invasive biomarker for tracking neurodegeneration in AD patients and monitoring the effects of disease-modifying drugs in clinical trials (Mattsson et al., 2019).
Publication Highlight: Plasma Tau, Neurofilament Light Chain and Amyloid-β Associated With Risk of AD Dementia
Using Quanterix’s Simoa® NF-light™ and Simoa® Human Neurology 3-Plex A assay, Wolf et al. concluded that high plasma levels of Nf-L and low plasma levels of amyloid-β42 are each associated with an increased risk of developing all-cause as well as Alzheimer’s disease dementia, regardless of age, sex, education, and risk factors (Read the publication).
While Aβ, tau protein, GFAP, and Nf-L have gained attention as promising biomarkers, other plasma biomarkers implicated in Alzheimer’s disease require further investigation.
VAMP-2 (vesicle-associated membrane protein 2), beta synuclein, and, in particular, SNAP-25 (synaptosomal- associated protein, 25kDa) are gaining support and a body of evidence that they are a promising AD biomarkers.
SNAP-25 is one of the major proteins involved in the formation of the SNARE (Soluble N-ethylmaleimidesensitive factor attachment protein receptors) protein complex.
Recent research demonstrates the potential for SNAP-25 to distinguish Alzheimer’s disease patients from non-demented controls and as a biomarker for early diagnosis and as a disease progression predictor, suggesting the value of this biomarker as a tool to further advance the understanding of Alzheimer’s disease.
The TAR DNA binding protein of 43 kDa (TDP43 or TARDBP) plays a fundamental role in exon skipping, nuclear transcription, splicing and stability of RNA transcripts, micro-RNA processing, and other cellular functions. It is also the major component of ubiquitin-positive cytoplasmic inclusions found in the brains of patients with frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS). Many individuals with Alzheimer’s disease and other neuromuscular disorders exhibit TDP43-containing aggregates.
There is an increased level of TDP-43 in plasma neuronal-derived exosomes (NDEs) in 57% of AD patients, and AD patients with TDP-43 pathology have more severe cognitive impairment than AD patients without TDP-43 pathology (Zhang et al., 2020, Meneses et al., 2021).
- Neurology 2-Plex B (Nf-L, GFAP)
- Neurology 3-Plex A (TAU, Aβ42, Aβ40)
- Neurology 4-Plex E (Nf-L, GFAP, AB40, AB42)
The ATN (A: amyloid pathology, T: tau pathology, N: neurodegeneration/neurological injury) classification system provides a common diagnostic framework for researchers and clinicians to categorize individuals based on biomarker evidence of pathology (Jack et al., 2016).
In this system, biomarkers are divided into binary categories and rated as positive or negative, with an individual’s score presented as A+/T+/N+ or A-/T-/N-, etc.
Most studies examining plasma use one or more of the following amyloid isoforms, tau biomarkers, and measures of neurodegeneration:
A: Amyloid pathology
- Aβ42/Aβ40 ratio
T: Tau pathology
Although the framework was developed using PET and CSF sampling, recent studies have adopted the framework and applied it to plasma proteins. In the original permutations of the system, neurodegeneration (N) was measured through [18F]-fluorodeoxyglucose–PET, structural MRI, or CSF total tau (Jack et al., 2016). When applied to plasma sampling, however, many researchers use Nf-L to measure neurodegeneration.
Some studies conclude that combining several biomarkers with basic demographics best predicts changes in cognition in unimpaired populations (Cullen et al., 2021). In contrast, others report that the combination of all markers does not outperform p-tau181 alone when discriminating A+T+ from A-T- participants (Alcolea et al., 2021). These results are not unexpected given that biomarkers change throughout the disease course and in relation to genetic and other demographic information.
Research and clinical settings could benefit from developing a reliable and repeatable assay for detecting, diagnosing, and monitoring AD, allowing us to change how we treat the disease.
The number of adults living with Alzheimer’s disease is increasing significantly as the baby-boomer generation ages. Thus, there is an urgent need for early detection and diagnostic tests.
PET scans and CSF sampling are sensitive and reliable but have substantial limitations that exclude their use as first-line diagnostic and predictive tools. On the other hand, blood tests are widely available, easier to administer, repeatable, and cost-effective.
Unfortunately, blood is a more complex system than CSF. Molecules from the CNS cannot freely cross the blood-brain barrier resulting in much lower concentrations of CSF biomarkers in the blood (Gabelli, 2020). We need ultrasensitive assays.
Quanterix gives researchers the power to detect even the most elusive biomarkers in neurology. Using our Simoa® technology, scientists have the tools they need to unlock the future potential of AD diagnostics for promoting health and treating disease.
How will you examine plasma biomarkers in Alzheimer’s disease?
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