A biomarker is defined here as a characteristic that is objectively measured in body fluids with a level of evidence sufficient to be used in in-vitro diagnostic procedures, i.e. evaluate disease risk, guide clinical diagnosis, monitor therapeutic interventions.
In neurodegenerative diseases, the biomarkers derived from the cerebrospinal fluid (CSF) has been intensely studied. The CSF biomarkers reflect molecular events in the brain due to it being in direct contact with the extracellular space of the brain.
Nevertheless, CSF movement is not unidirectional flow and there is no evidence that the pathologic proteins can be found in the extracellular environment. The search of NDD biomarkers is a challenge because most of brain proteins are extensively modified post translationally.
Protein markers have been developed that reflect the central pathogenic processes in Alzheimer ‘s pathology (AD), i.e. the disturbance in the metabolism of β-amyloid (Aβ) and its subsequent deposition in senile plaques, the hyperphosphorylation of tau protein with subsequent formation of tangles (phosphorylated tau, P-tau) and the neuronal degeneration (total tau, T-tau);
The main component of plaques is a peptide called β-amyloid (Aβ), which is a cleavage-metabolite of the amyloid precursor protein (APP). APP is a single-transmembrane protein with the Aβ domain partly embedded in the membrane. Aβ is generated by cleavage of APP by two proteases, the β- and γ-secretases. Free Aβ is secreted into the CSF.
There are two major C-terminal variants of Aβ, a shorter form ending at amino acid 40 (Aβ40) and a longer form ending at amino acid 42 (Aβ42). The Aβ42 isoform has a high tendency for aggregation, and is also the earliest Aβ species deposited into plaques.
Tau is a normal protein located in the neuronal axons in the brain. Its function is to stabilize the microtubular network in the axons, by binding to the microtubules. There are six different isoforms of tau, depending on which exons of the tau gene are translated to the mature tau protein. There are also numerous phosphorylation sites, i.e. amino acids that can be phosphorylated on the Tau protein. Tau protein can thus be found in variants with different degrees of phosphorylation.
The CSF level of T-tau reflects the intensity of the neuronal and axonal degeneration and damage in the brain.
Hyperphosphorylated Tau protein
In AD, a phosphate group is attached to several amino acids in tau protein, and tau is thus found in variants with different degrees of phosphorylation. Phosphorylated tau has a reduced ability to bind to the microtubules in the axons, which affect the axonal stability and thus the neuronal function, and also render tau an increased tendency for aggregation into paired helical filaments which then form the larger protein aggregates that make up the tangles.
Synucleinopathies are characterized by intra-neuronal aggregates consisting mainly of α-synuclein (α-syn) are found in Lewy bodies and Lewy neurites in Parkinson’s disease (PD), PD with dementia (PDD) and Dementia with Lewy bodies (DLB) and in glial cytoplasmic inclusions in multiple system atrophy (MSA). This extracellular form of α-syn seems to be secreted from neuronal cells by exocytosis and detected in CSF as phosphorylated oligomeric α-syn.
TAR DNA-binding protein of 43kDa (TDP-43)
TDP-43 is a major component of ubiquitin-positive inclusions that are one of the neuropathological hallmarks in amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). Only 50% of FTLD patients have aggregates positive for TDP-43. Unfortunately, TDP-43 in CSF originates mainly from blood. Measurements of TDP-43 in CSF and blood are of minor importance as a diagnostic tool, but may be important for monitoring therapy effects of TDP-43 modifying drugs in the future.
Recent reports identified mutations causative of neurological disorders in the genes encoding a family of RNA-binding proteins (RBPs) named FET. RNA-binding proteins (RBPs) are involved at all stages of RNA metabolism in neurodegenerative diseases. FET proteins are highly conserved and ubiquitously expressed. Recently, it has been suggested the involvement of FET proteins in neurological diseases, such as frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS), where they have been found in cytoplasmic aggregates. Abnormal co-accumulation of FET proteins into pathological inclusions has been described in all subtypes of FTLD-FUS. FUS (fused in sarcoma) is the most examined protein; FUS is localized in dendritic granules and spines in neurons where it plays a role in mRNA transport into dendrites, which represents an essential process for local protein synthesis and synaptic plasticity. No CSF FET related-markers are available.
Prion protein associated deposits
Pathological prion protein (PrPSc)
Human prion diseases are rapidly progressive neurodegenerative disorders caused by prion protein misfolding. The sporadic Creutzfeldt-Jakob disease (sCJD) is the most common form (85–90% of cases), followed by genetic CJD (gCJD) and fatal familial insomnia (FI) (10–15% of cases), which are linked to point or insertion mutations in the prion protein gene (PRNP). Several molecular subtypes of sporadic Creutzfeldt–Jakob disease have been identified and electroencephalogram and cerebrospinal fluid biomarkers have been reported to support clinical diagnosis but with variable utility according to subtype (updated WHO criteria for the diagnosis of CJD and related disorders, 2009). The principle for detecting PrPSc is to exploit the ability of small amounts of CSF PrPSc to convert native PrP into PrPSc in a newly described protein aggregation assay known as real-time quaking-induced conversion (RT-QuIC). This technique using a recombinant PrP showed good diagnostic sensitivity (82-96%) and virtually full specificity.
Prion diseases may trigger biochemical changes similar to AD involving PrPSc, Aβ42, APOE-4 and abnormal tau. Autopsied brain of sCJD showed also Alzheimer disease (AD)-like changes (17% of cases).
Testing of 14-3-3 protein in CSF is a standard biomarker test in suspected sCJD diagnosis by established Western blot method in CJD reference laboratories. Blood-contaminated samples which may result in artificially elevated CSF levels of 14-3-3.
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