Recent progresses on p-tau as a blood- based Alzheimer’s disease biomarker

Introduction. The amyloid cascade hypothesis proposes that extracellular senile plaques largely composed of aggregated beta-amyloid (Aβ) peptides are responsible for the events that lead to neuronal death that occurs in Alzheimer's disease (AD). On the other hand, the hyperphosphorylated (p-tau) and unstructured tau protein is responsible for intracellular neurofibrillary tangles, also common in AD. Clinical diagnostic criteria for AD include Aβ and p-tau biomarker tests in cerebrospinal fluid (CSF), in addition to neuroimaging measures, clinical history, and psychometric tests. However, due to their invasive nature, side effects and need for trained personnel in a hospital environment for their collection, CSF biomarkers are not suitable for large-scale screening. Therefore, alternative blood-based biomarkers are under intense investigation. Objective. Focus on recent advances in different p-tau isoforms as blood-based AD biomarkers. Method. Review performed by searches in Medline/PubMed databases. Results. The p-tau isoforms 181 and 217 represent accessible and scalable molecules for screening and diagnosing AD, mainly due to their ability to differentiate patients with the disease from cognitively healthy participants. These results should be reproduced in

larger and more representative cohorts of population diversity. Conclusions. This review provides a more comprehensive exploration of blood p-tau as a specific molecular biomarker for AD, which could contribute not only to screening pre-symptomatic patients for clinical trials, but also to monitoring disease progression and evaluating modifying therapies. of the disease. Keywords. Alzheimer's disease; biomarkers; blood; plasma; p-tau; tau Resumen Introducción. La hipótesis de la cascada de amiloide propone que las placas seniles extracelulares, en gran parte compuestas por péptidos beta-amiloides (Aβ) agregados, son responsables de los eventos que conducen a la muerte neuronal que ocurre en la enfermedad de Alzheimer (EA). Por otro lado, la proteína tau hiperfosforilada (p-tau) y no estructurada es responsable de los ovillos neurofibrilares intracelulares, también comunes en la EA. Los criterios de diagnóstico clínico para la EA incluyen pruebas de biomarcadores Aβ y p-tau en líquido cefalorraquídeo (LCR), además de medidas de neuroimagen, historia clínica y pruebas psicométricas. Sin embargo, debido a su naturaleza invasiva, los efectos secundarios y la necesidad de personal capacitado en un entorno hospitalario para su recolección, los biomarcadores de LCR no son adecuados para el cribado a gran escala. Por lo tanto, se están investigando intensamente los biomarcadores alternativos basados en sangre. Objetivo. Concéntrese en los avances recientes en diferentes isoformas de p-tau como biomarcadores de EA en sangre. Método. Revisión realizada mediante búsquedas en bases de datos Medline/PubMed. Resultados. Las isoformas p-tau 181 y 217 representan moléculas accesibles y escalables para la detección y el diagnóstico de la EA, principalmente debido a su capacidad para diferenciar a los pacientes con la enfermedad de los participantes cognitivamente sanos. Estos resultados deben reproducirse en cohortes más grandes y representativas de diversidad de población. Conclusiones. Esta revisión proporciona una exploración más completa de la p-tau sanguínea como un biomarcador molecular específico para la EA, lo que podría contribuir no solo al cribado de pacientes presintomáticos para ensayos clínicos, sino también a monitorear la progresión de la enfermedad y evaluar las terapias modificadoras de la enfermedad.  Although the blood-brain barrier (BBB) usually prevents brain antigens to reach the blood, this barrier is compromised in AD patients, increasing the levels of CNS markers in the peripheral blood 15   NFT, together with senile plaques, comprises one of the main AD hallmarks, as already mentioned in this review. NFT isolation was first made in 1974 from frozen autopsied brains, identifying a ~50kDa protein as their main constituent 33 . In the following year, this molecule was characterized as the tau protein from microtubules 34 , and more than ten years after, abnormally hyperphosphorylated tau (p-tau) was determined as the AD-associated protein in NTF 35 from neurons' cytosol, forming oligomers that inhibit microtubule assembly 33 . Several laboratories generated polyclonal and monoclonal, leading to the discovery of elevated levels of tangle immunoreactivity in CSF from AD patients 36 . Immunohistochemical staining of AD brain sections with anti-p-tau allowed identifying the six stages of neurofibrillary pathology, known as the 'Braak stages' 37 .

Tau and p-tau CSF biomarkers and their relationship with AD pathophysiology
Further, gene cloning and primary structure identification proved that alternative splicing led to the formation of six isoforms of tau in human brains 38 and the discovery that the expression of truncated tau isoforms produced NFT that generated neurocognitive impairment 39 .
Quantification of hyperphosphorylated and nonhyperphosphorylated pools of tau revealed that total tau (ttau) levels were increased in the brains of AD people, with no significant differences of non-hyperphosphorylated tau levels between AD and normal aged brains 40 . These results revealed that affected neurons continue to synthesize tau to maintain their function in the presence of AD pathology.
Furthermore, p-tau presents higher specificity than other validated AD CSF biomarkers, t-tau and Aβ, to characterize the NFT burden in AD, probably because, unlike t-tau, it is unaffected by possible comorbidities such as brain injury or stroke. Moreover, p-tau does not correlate with other potential tauopathies, such as frontotemporal lobar degeneration, which is difficult to distinguish from AD clinically 41 . These findings provided the basis for investigating tau levels in CSF as an AD diagnostic biomarker 42 .
Following these researches, it was confirmed that CSF tau provides a valuable marker of tau pathology and can be used to monitor the efficacy of disease-modifying therapeutics 42 . Extensive evidence indicates increased CSF p-tau in patients with AD compared to controls 16   correlated with longitudinal worsening of cognition and brain atrophy. In summary, these longitudinal data shows that plasma p-tau 217 increases during early AD and, therefore, can be a powerful tool to monitor disease progression 56 . All these discoveries indicate that, as well as CSF p-tau 181 and 217, plasmatic p-tau isoforms are useful AD biomarkers, mainly due to low invasiveness and cost. The main discoveries on plasmatic p-tau as AD biomarker are summarized in Table 2.

CONCLUSIONS
This review aimed to describe the recent advances on different p-tau isoforms as a blood-based biomarker for AD, provide a more comprehensive and detailed exploration of these biomarkers, and contribute to the advance of the literature on the theme.
The results presented in this review support the two isoforms of plasma p-tau 181 and 217 have the potential to be considered simple, accessible, and scalable test for screening and AD diagnosis, mainly due to their ability to differentiate AD patients from cognitively healthy participants. Hence the advantages of more accurate and sensitive diagnostic tests for the different p-tau isoforms.
These biomarkers are important for AD diagnosis and monitoring the disease progression and treatment, especially when added to CSF tau measures.
Taken together, the data presented here pointed out that, despite more research is needed to reproduce these results in large and distinct cohorts, the ability to receive diagnostic and prognostic information using the convenience of a blood test will completely change AD research, treatment, and care from what we have nowadays.
Therefore, we emphasize the considerable ethical and social challenges that blood tests for AD will bring to the future, especially considering the absence of disease-modifying therapy for the disease.