BIOMARCATORI NEUROCHIMICI - PowerPoint Presentation

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BIOMARCATORI NEUROCHIMICI NELLE FASI INIZIALI DI DEMENZA

Dr. Filippo Baldacci

5 -7 Aprile 2019, Viareggio

III Meeting delle Neuroscienze Toscane

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Abbreviations: A

β42,

amyloid‑

β42; ALS, amyotrophic lateral sclerosis; APOE, apolipoprotein E; APP, amyloid precursor protein; bvFTD, behavioural variant frontotemporal dementia; DJ1, parkinson protein 7; DLB, dementia with Lewy bodies; EOAD, early-onset Alzheimer disease; FUS, fused in sarcoma; GBA, glucocerebrosidase; GRN, granulin; HP‑tau, hyperphosphorylated-tau; LOAD, late-onset Alzheimer disease; LRRK2, leucine-rich repeat kinase 2; MAPT, microtubule-associated protein tau; PARK2, parkin RBR E3 ubiquitin protein ligase; PCA, posterior cortical atrophy; PD, Parkinson disease; PINK1, PTEN induced putative kinase 1; PPA, primary progressive aphasia; PS, presenilin; PSP, progressive supranuclear palsy; SNCA, α‑synuclein; SOD1, superoxide dismutase 1; TARDBP/TDP‑43, TAR DNA-binding protein 43; TOMM40, translocase of outer mitochondrial membrane 40 homologue; TREM2, triggering receptor expressed on myeloid cells 2; UPS, ubiquitin proteasome system; VCP, valosin containing protein.

Pievani M, Nat Rew Neurol 2014, modified

OVERLAPPING AMONG DEMENTIAS AND NEURODEGENERATIVE DISEASES (NDD)

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AD HETEROGENEITY

(Rabinovici,

Alzheimers Dement (NY), 2017)

Pathological spectrum of AD

AD: Alzheimer's disease; VaD: vascular dementia; LBD: Lewy body disease; HS: hippocampal sclerosis;

(McCann, Parkinsonism and Related Disorders, 2014)

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AD: DEFINITION OF A PATHOPHYSIOLOGICAL ENTITY

IWG: 2007

IWG: 2010

NIA-AA: 2011

IWG: 2014

NIA-AA: 2018

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DIAGNOSTIC CRITERIA AD, IWG-2 2014

Dubois

, Lancet

Neurol 2014

“pathophysiological-diagnostic biomarkers (PET amyloide tracer, CSF, genetic analysis) and topographic biomarkers of progression (FDG -PET, RM volumetry hippocampal)”

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CSF AND BLOOD BIOMARKERS FOR THE DIAGNOSIS OF AD

CSF core biomarkers and NFL differentiated AD and prodromal AD from controls with good performance

. CSF NSE, VLP-1, HFABP, and YKL-40 were moderate.Plasma T-tau had large effect sizes when differentiating between controls and AD

The upper and lower values delimiting the gray zone indicate the 95% confidence interval (95% CI) values calculated using the ADNI cohort (

n

=116 for controls and

n=100 for AD)

Coart E, Alzh&Dem 2015

T-tau, P-tau, Aβ42, and NFL in CSF should be used in clinical practice and clinical research

Olsson B, Lancet Neurol 2016

NFL=neurofilament light

chain

, marker of large

myelinated

axonal

fibers

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CSF NEUROGRANIN (Ng) CONCENTRATIONS ARE ASSOCIATED WITH AD PATHOPHYSIOLOGY IN AUTOPSY CONFIRMED CASES

Higher CSF Ng levels were positively associated with greater Aβ neuritic plaque and tau tangle pathology scores. In the hippocampus and amygdala CSF Ng was associated with plaque but not with tangle, α-synuclein, or TAR DNA-binding protein 43 loads.It correlates with longitudinal decline of MMSE

Portelius E, Acta Neuropathologica 2018

116

pts

Ng

= marker of

dendritic

synpatic

damage

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Abbreviations: Aβ1-42, 42-amino acid-long amyloid beta peptide; ADD, Alzheimer’s disease dementia; AUROC, area under the receiver operating characteristic curve; C.I., confidence intervals; CSF, cerebrospinal fluid; FTD, frontotemporal dementia; HC, cognitively healthy controls; NFL, neurofilaments light chain protein; p-tau, hyperphosphorylated tau; t-tau, total tau. NOTE. The AUROC curves result from fitting a logistic regression model within a 10-fold cross validation scheme to biomarkers data adjusted for age, sex, and site.

The first 10 combinations of CSF biomarkers with the best diagnostic accuracy in discriminating HC versus ADD and ADD versus FTD (the results were displayed by the highest AUROC value to the lowest).

NOVEL BIOMARKERS NG, NFL,YKL-40 ALONE AND IN COMBINATION DID NOT IMPROVE DIAGNOSTIC ACCURACY IN NDD COMPARING WITH CORE BIOMARKERS

Hampel H , Alzheimer and Dementia 2018

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CSF total α-syn is lower in PD than in controls (α-syn sequestration in Lewy bodies) CSF α-syn is correlated to t-tau and p-tau in PD; CSF t-tau and p-tau are lower in PD than in controls CSF oligomers of α-synuclein and α-synuclein phophorylated are increased in PDIncrease of CSF total α-syn in AD patients when compared with controls, PD and atypical parkinsonismsConflicting results in prognosis and staging of PD: Lower CSF total α-syn levels or higher oligomeric α-syn correlated with higher HYahr scores. Correlation between phosphorylated a-syn and UPDRS III scores. Higher CSF total α-syn levels at the baseline predict cognitive decline in PDThe measurement of CSF total α-syn has not a diagnostic accuracy satisfactory (also in the early phase)A high variability of the total α­syn in the different laboratories Contamination of CSF with blood increases the levels of α-syn (CSF hemoglobin levels are therefore measured in order to exclude contaminated samples)

Magdalinou

N, JNNP 2014; Blennow K, Mov Disorders 2016; Kim D, Journal of clinical Neurology 2016; Parnetti L, Biomark Med 2016

CSF α-SYN AND OTHER CSF BIOMARKERS IN PD AND PARKINSONISMS

CSF A

β

42

is

reduced

in PDD in

comparison

to PD and

is

predictive

of

dementia

in PD

CSF NFL

and YKL-40 are

increased in atypical park

.

compared

to PD

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FTLD-TDP43

(types A, B, and C)

[50% of FTD]FTLD-tau 35-50% (Pick-disease 30%, CBD 35%, PSP 30%)FTLD-FUSA family history in up to 40% of cases, a clear autosomal dominant history accounts for 10% of cases. Mutations in C9orf72, MAPT, and GRN genes account for about 60% of all familiar casesBiomarkers strongly indicative of AD (or other NDDs) are exclusionary

Piguet, Lancet Neurol 2010; Gorno-Tempini, Neurology 2011; Bang, Lancet Neurol 2015; Oecke Biochimica et Biophysica Acta 2015; Teunissen Alz&Dem 2016; Kuiperij, JAD 2016

bvFTLD

: Imaging results consistent with frontal and/or anterior temporal atrophy on CT or MRI or frontal hypoperfusion or hypometabolism on SPECT or PET Non fluent/Agrammatic variant PPA: predominant left posterior fronto-insular atrophy on MRI or predominant left posterior fronto-insular hypoperfusion or hypometabolism on SPECT or PET Semantic variant PPA: predominant anterior temporal lobe atrophy and/or predominant anterior temporal hypoperfusion or hypometabolism on SPECT or PETLogopenic variant PPA (??): predominant left posterior perisylvian or parietal atrophy on MRI and/or predominant left posterior perisylvian or parietal hypoperfusion or hypometabolism on SPECT or PETStructural changes on imaging reflect phenotypes rather than pathology

CSF and plasma progranulin screening of genetic FTD with GRN mutation CSF t-tau, p-tau, t-TDP-43 and p-TDP-43, NFL, AgRP, YKL-40, somatostatin, reelin, enzyme activity of catalase  may distinguish FTLD from other NDDs and/or different subtypes of FTLD

THE FTLD SPECTRUM AND ITS POTENTIAL BIOMARKERS

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BLOOD BIOMARKERS AS SCREENING TOOL IN A MULTI-STAGE NDD DIAGNOSTIC STEPS

Hampel H, JAD 2018 (modified); Hampel H, Nat Rev Neurol 2018; Baldacci F, Exp Mol Diagn 2019

Predictive Value (PV) of a screening test

, besides being related to its intrinsic sensitivity and specificity,

is critically dependent on disease prevalence

. As prevalence of the disease increases, the positive PV (PPV) of a test increases, while the negative PV (NPV) decreases. PV of a screening test for NDD would be subject to variations, based on the prevalence of NDDAD and dementia disorders are quite frequent in the general population, (5-6 % in western countries) Amyotrophic lateral sclerosis (ALS) is a rare disease with a prevalence of 2/3 cases per 100 000 people. NDD prevalence as a whole is high Old age, familiar history of NDD, psychiatric disorder onset in midlife, and diabetes represent conditions identifying asymptomatic subpopulations with higher probability to develop NDD

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DIFFICULTIES OF BLOOD BIOMARKERS DISCOVERING

The biomarker must be able to cross the blood–brain barrier Low concentration of CNS-derived proteins in the blood (blood:CSF volume ratio causing a substantial dilution) Biomarker expressed in peripheral tissues (e.g., Aβ is also expressed in blood platelets and several other tissues) The high amount of other proteins in blood (e.g., albumin and immunoglobulins) may interfere in the assays Blood may also contain endogenous antibodies directed against the non-human monoclonal antibodies of the assay, which may cause flawed results The analyte of interest may undergo proteolytic degradation by various proteases in plasma (tau, which is stable in CSF but has a very short (∼10 h) halflife in blood) The technical issues are mainly a question of sensitivity and antibody specificityStandard immunochemical methods have often been insufficiently sensitive to allow the reliable quantification of CNS-derived molecules in the blood Lack of standardization of pre-analytical protocols

Zetterberg

H, et al. Journal of Neuroscience Methods 2018;

Hampel

H, et al. Nature

Rev

Neurol

2018;

Andreassos

U, et al. Alzheimer &

Dementia

2016

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ULTRASENSITIVE TECHNOLOGIES TO FACILITATE RESEARCH ON BLOOD BIOMARKERS FOR NDD

With an average of 1000x greater sensitivity than current immunoassays, analyte concentrations can be efficiently measured in blood

Digital ELISA

(Enzyme Linked Immunosorbent Assay) based on

single molecule arrays (Simoa) has improved sensitivity of traditional ELISA from picomolar (10-12 M) to femtomolar (10-15 M)Digital ELISA counts signal generated from single immunocomplexes formed on superparamagnetic beads confined in arrays of femtoliter-sized wells in which fluorescent molecules are highly concentrated.

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The composite biomarker showed very high areas under the AUROC in both data sets (discovery, 96.7%,

n= 121 and validation, 94.1%, n= 111) with an accuracy near to 90% when using amyloid-β-PET as a gold standard. Plasma biomarkers correlated with amyloid-β-PET burden and CSF Aβ1–42 levels.

Nakamura A, Nature 2018

POTENTIAL CLINICAL UTILITY OF PLASMA AMYLOID-β BIOMARKERS IN PREDICTING BRAIN AMYLOID-β BURDEN

AUROC for each biomarker when predicting individual Aβ+/Aβ- status in CN,MCI,ADD

High-performance plasma amyloid-β biomarkers by immunoprecipitation coupled with mass spectrometry

Biomarker performance was validated in a blinded manner using independent data sets (Japan and Australia) and involved an established large-scale

multicentre

cohort

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Plasma tau was analyzed with the Human Total Tau kit (research use only grade, Quanterix, Lexington, MA) on the Simoa HD-1 analyzer (CE marker)

Plasma tau

was

higher in AD and associated with significantly greater decline of MMSE over time, slightly greater decline of hippocampal volume over time, greater ventricular volume at baseline and greater increase in ventricular volume over time, and greater decline in FDG-PET over time

Mattsson N, Neurology 2016

Plasma tau partly reflects AD pathology, but the overlap between normal aging andAD is large, especially in patients without dementia. Despite group-level differences, these results do not support plasma tau as an AD biomarker in individual people

PLASMA TAU IN AD

Longitudinally followed patients with AD (n = 179), MCI (n = 195), and HC (n = 189)

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8 studies on AD/MCI/SCD; 5 studies on ALS; 5 studies on FTD; 4 studies on PD/atypical parkinsonisms; 3 studies on sCJ; 2 studies on HD; 2 VD

PLASMA/SERUM NFL IS ASSOCIATED WITH NDD DIAGNOSIS

Hansson O, et al.

Neurology

2017

Steinacker P, et al. Scientific Report 2016

Steinacker P, et al.

Neurology

2018

Steinacker P, et al.

Neurology 2017

Wylke

C et al. JNNP 2016

PD and atypical parkinsonisms

FTD: bvFTD, PPA

CJ

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8 studies on AD/MCI/SCD; 5 studies on ALS; 5 studies on FTD; 4 studies on PD/atypical parkinsonisms; 3 studies on sCJ; 2 studies on HD; 2 VD

PLASMA/SERUM NFL IS ASSOCIATED WITH NDD DIAGNOSIS

Byrne LM, et al. Lancet

Neurology 2017

Wylke C et al. JNNP 2016

Feneberg

E, et al. Neurology 2018

Duering

M, et al. Journal of Stroke 2018

Vascular dementia

ALS

HD

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Plasma NFL

moderately

correlated with CSF NFL

Plasma NFL differentiated ADD vs controls, with an AUROC of 0.87. The AUROCs were 0.87 to 0.90 for CSF NFL, Aβ42, t-tau, p-tauPlasma Tau differentiated ADD vs controls with AUROC of 0.78

PLASMA NFL IS ASSOCIATED WITH AD DIAGNOSIS

Ultrasensitive assay (Simoa) was used to measure plasma NFL concentration in 193 HC, 197 MCI, and 180 ADD

Mattsson N, JAMA neurology 2017

Plasma NFL

correlated with

poor cognition

and

AD-related atrophy

(at baseline and longitudinally)

and with

brain

hypometabolism

(longitudinally)

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Byrne L et al. Lancet

neurol 2017

Premanifest HD

Baseline plasma NfL at is predictive of disease onset after 3-year follow-up

ALS

survival

Baseline plasma NfL is predictive of death

Lu C et al. Neurology 2015

PSP

survival

Baseline plasma NfL is predictive of death

Kaat LD et al. Park rel Dis 2018

Genetic

FTD survival

Meeter LH et al. Anna Cli Trans Neuro 2016

Baseline plasma NfL is predictive of death

Genetic

pre AD

Preische O et al. Nature Med 2019

Baseline plasma NfL increase overtime in carriers

PLASMA/SERUM NFL IS ASSOCIATED

WITH NDD PROGRESSION

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CONCLUSIONS

BIOMARKERS of AD:CSF Aβ42, t-tau, p-tauCANDIDATE BIOMARKERS of PD:CSF Aβ1-42, t-tau, p-tauCANDIDATE BIOMARKERS of FTD:CSF (plasma) progranulinCANDIDATE BIOMARKERS of NEURODEGENERATION:CSF (plasma) NFL

Pathophysiological biomarkers (preclinical, prodromal, diagnostic, prognostic)

Screening of neurodegenerative diseases, and disease progression monitoring

Predictive of cognitive decline

(prodromal, diagnostic, prognostic)

Screening of genetic FTD due to

GRN

mutations

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CONCLUSIONS

Clinical

Phenotype (cognitive profile, cognitive network disruption)Staging (Asymptomatic at risk, SMC, MCI, Dementia)Rate of disease progression (MCI stable, MCI converters)Pathophysiological mechanisms (Liquid biopsy) ???Clinical Diagnosis???

COGNITIVE EVALUATION

BIOMARKERS

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GRAZIE !!!

Cellular Biochemistry and Molecular Biology Department of PharmacyUniversity of Pisa

Claudia Martini Maria Letizia TrincavelliSimona DanieleDeborah PietrobonoRebecca Piccarducci

Neurology Unit, Department of Clinical and Experimental MedicineUniversity of Pisa

Ubaldo BonuccelliGabriele SicilianoGloria TognoniFilippo Sean GiorgiRoberto CeravoloValentina NicolettiCristina PagniSimona CintoliCaludia RadicchiLuca Tommassini

Linda

Giampietri

Paolo Libertini

Alessandro Galgani

Daniela Frosini

Lucia

Petrozzi

Annalisa Lo

Gerfo

Lucia Chico

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CSF AND BLOOD BIOMARKERS FOR AD

Core CSF

biomarkers

Blennow K, Neurol Ther 2017

CSF Aβ42/ Aβ40 and Aβ42/ Aβ38: - Good correlation with amyloid-PET (better prediction than Aβ42 alone)- Recognize HC with low Aβ production- Recognize AD with high Aβ peptides production

Lewckzeb P, JAD 2017

CSF Aβ42/ Aβ40: - Good correlation with amyloid-PET (better prediction than Aβ42 alone)Aβ oligomersCSF t-tau/ Aβ42; CSF p-tau/Aβ42: - Predictive of amyloid-PET deposition better than any single biomarker

Janelidze S, Ann Clin Transl Neurol 2016

Fagan AM, Arch Neurol 2011

Seeburger JL, JAD 2015

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CSF AND BLOOD BIOMARKERS FOR AD

Blennow

K,

Neurol

Ther 2017

Candidate biomarkers

While plasma and CSF t-tau levels correlate poorly, the correlation is very tight between plasma and CSF NFL

Inflammatory markers: - CSF YKL-40: AD>MCI>HC, predictive value MCI converters - CSF TREM2 (?) - IL-6, TNF-alpha, IL-1beta, TGF-beta, IL-12 > AD vs HC

Baldacci F, Exp Rev Proteom 2017

Swardfager W, (metanalysis), Biol Psich 2010

Synaptic markers: - CSF Neurogranin (dendritic): specific, diagnostic, prognostic also in HC - CSF SNAP-25, synaptotagmin (vesicle, presynaptic):: AD and MCI > HC) - CSF t-alpha-syn (presynaptic) :AD and MCI > PD and HC

Brinkmalm A, Mol degen 2014

Berge G, BMC neurol 2016

Korff A, JAD 2013

Ohrfelt A, Alz Res Ther 2016

Neuronal calcium sensor protein: - CSF VILIP: AD>HC, cognitive decline progression in MCI

Axonal markers: - CSF NFL, plasma NFL: AD and MCI > HC, not specific associated with cognitive impairment, prognostic value

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Beta-amyloid (Aβ) sequelae in the human eye in AD

POTENTIAL OCULAR AD BIOMARKERS

Shah

TM, Molecular Psychiatry 2017

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HOMOGENOUS AND SYSTEMATIC DATA COLLECTION

INITIAL POPULATION

«BIG-DATA»

DENSITY-BASED

CLUSTERING

(no a priori N of clusters)

DATA-DRIVEN CATEGORIZATION«SMART-DATA»

Baldacci F, Methods in Molecular Biology 2018

TOWARDS A NOVEL CLASSIFICATION OF THE

SPECTRUM

OF NDs

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STATE OF COMPLETION OF BIOMARKERS DEVELOPMENT IN AD

Frisoni GB, Lancet

Neurol 2017

Sufficient evidence of analytical validity (phase 1 of a structured framework adapted from oncology: Pepe MS, Phases of Biomarker Development for Early Detection of Cancer, Journal of the National Cancer Institute, 2001) is available for all biomarkers, but their clinical validity (phases 2 and 3) and clinical utility (phases 4 and 5) are incomplete

Proof of concept(preclinical)

AD diagnosis (cross-sectional)

MCI-AD progression (longitudinal retrospective)

MCI-AD progression (longitudinal prospective)

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Abbreviations: A

β42,

amyloid‑

β42; ALS, amyotrophic lateral sclerosis; APOE, apolipoprotein E; APP, amyloid precursor protein; bvFTD, behavioural variant frontotemporal dementia; DJ1, parkinson protein 7; DLB, dementia with Lewy bodies; EOAD, early-onset Alzheimer disease; FUS, fused in sarcoma; GBA, glucocerebrosidase; GRN, granulin; HP‑tau, hyperphosphorylated-tau; LOAD, late-onset Alzheimer disease; LRRK2, leucine-rich repeat kinase 2; MAPT, microtubule-associated protein tau; PARK2, parkin RBR E3 ubiquitin protein ligase; PCA, posterior cortical atrophy; PD, Parkinson disease; PINK1, PTEN induced putative kinase 1; PPA, primary progressive aphasia; PS, presenilin; PSP, progressive supranuclear palsy; SNCA, α‑synuclein; SOD1, superoxide dismutase 1; TARDBP/TDP‑43, TAR DNA-binding protein 43; TOMM40, translocase of outer mitochondrial membrane 40 homologue; TREM2, triggering receptor expressed on myeloid cells 2; UPS, ubiquitin proteasome system; VCP, valosin containing protein.

CEREBROVASCULAR DISEASE

Pievani M, Nat Rew Neurol 2014, modified

OVERLAPPING AMONG DEMENTIAS AND NEURODEGENERATIVE DISEASES

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AD PATHOPHYIOLOGY

Patogenesi

Complex

pathophysiological

mechanisms

A

β amyloid cascade hypothesis

Oxidative

stress

Neuroinflammation

Age,

genetic

and

environmental factors

Cerebrovascular risk factors

Aβ amyloid peptide accumulation

Tau protein hyperphosphorilation

Endothelial damageBBB permeability

Insuline-resistence

Iron metabolism alterations



-

syn and other protein depositions

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POTENTIAL DIAGNOSTIC VALUE OF RED BLOOD CELLS A-SYNUCLEIN HETEROAGGREGATES IN AD

A

D

C

B

E

F

Baldacci

F, Daniele S, et al.

Molecular

Neurobiology

N = 39 AD/MCI

biomarker

+

N = 39

age

and

sex

matched

controls

ROC curves to evaluate the utility of RBC biomarkers concentrations in discriminating AD/MCI patients from HC.

α-

syn

-tau heterodimers AUROC = 0.76 (95% CI 0.65 – 0.87,

P

< 0.001); α-

syn

-Aβ

1-42

heterodimers AUROC = 0.72 (95% CI 0.61 – 0.84,

P

= 0.001); AUROC α-

syn

= 0.64 (95% CI 0.51 – 0.77,

P

= 0.036)

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PLASMA P-TAU181 NOVEL BIOMARKER FOR AD

Mielke

MM, A&D 2018 in press

Plasma t-tau and pTau181 > in ADD than HC.

Plasma pTau181 was more strongly associated with both Aβ and tau PET than t-tau.Plasma pTau181 was a more sensitive and specific predictor of elevated brain Aβ than t-tauBiomarker of AD pathophysiology and as a noninvasive screener for elevated brain Aβ

HC=172 MCI=57 ADD=40

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TOWARD A BIOLOGICAL DEFINITION OF AD

NIA-AA A&D 2018

A (

amyloid) = CSF Aβ, Amyloid-PETT (tau) = CSF p-tau, tau-PETN (neurodegeneration) = CSF t-tau, hyppocampal volume, FDG-PET

A/T/N SYSTEM

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BIOMARKERS DEFINITION

The National Institutes of Health (NIH) defines a biomarker as “a characteristic that is objectively measured and evaluated as an indicator of normal biological processes, pathogenic processes or pharmacological responses to a therapeutic intervention” (2001).

In other words, is a tool to aid in the study of human diseases by confirminga diagnosis and tracking disease progression, and may help toidentify specific therapeutic targets. In general biomarkers involve measurements of biological samples (fluids and biopsy) or measurements using brain imaging techniques (structural, functional)

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Molinuevo JL, Actha Neuropathologica, 2019

ANATOMY OF AD BIOMARKERS – LIQUID BIOPSY (Pathological mechanisms implicated in AD and associated fluid biomarkers)

A

β38

amyloid beta 38,

A

β40

amyloid beta 40,

A

β42

amyloid beta 42,

AD

Alzheimer’s disease,

BACE1

β-

site amyloid precursor protein cleaving enzyme 1,

hFABP

heart-type fatty acid-binding protein,

IP

-

10

interferon-

γ-

induced protein 10,

NF

-

L

neurofilament light,

P

-

tau

phosphorylated tau,

SNAP

-

25

synaptosome-associated protein 25,

TDP

-

43

transactive response DNA-binding protein 43,

TREM2

triggering receptor expressed on myeloid cells 2,

T

-

tau

total tau,

VILIP

-

1

visinin

-like protein 1