# Project 3: Development of multiplex assays for clinical monitoring of disease

> **NIH NIH U19** · UNIVERSITY OF WASHINGTON · 2021 · $608,188

## Abstract

Abstract
The pathological hallmarks of Alzheimer’s disease are the neurofibrillary tangles and amyloid plaques that form
in gray matter regions of the cerebral cortex. Misfolded and aggregated proteins reside within those
pathological features, which may to be central to the neuronal death that causes regional atrophy, dementia,
and ultimately death. The cerebrospinal fluid that bathes the central nervous system is the fluid most proximal
to the disease and has been the focus of biomarker discovery for more than 30 years. There are two important
gene products that are present in cerebrospinal fluid and have been shown to be predictive of disease activity
and cognitive decline in patients that present with mild cognitive impairment: (1) the amyloid precursor protein,
which gives rise to Aβ(1-42) and other fragments, and (2) tau, which has many phosphorylation sites. The
deposition of Aβ(1-42) as amyloid aggregates appears to lead to the death of adjacent neurons. The resulting
injury increases the amount of tau protein released into the cerebrospinal fluid. From previous biomarker
studies of Aβ proteins and tau, it seems likely that the entire disease process, from Aβ(1-42) deposition to cell
death, is orchestrated differently than in other neurodegenerative diseases. As a result, it is a compelling
hypothesis that novel biomarkers, which may be mediators of disease, are present in the cerebrospinal fluid
and could add more information over the current biomarkers used in the diagnosis of Alzheimer’s disease.
There have been previous attempts to use discovery proteomics to identify proteins that are differentially
expressed in cerebrospinal fluid in patients with Alzheimer’s disease compared with controls and there have
been few efforts to expand upon these studies in a clinically meaningful way. However, some of these efforts
have identified the presence of additional fragments of amyloid precursor protein [i.e., in addition to Aβ(1-42)]
and other proteins, which suggests that post-translational changes may be important in the formation of
amyloid, the dysfunction of neuronal synapses, and the subsequent death of neurons.
In this project entitled, “Identification of mass spectrometric targets and development of multiplex assays for
disease management,” our objective is to identify the post-translational modifications that lead to altered
LP/EV concentration, molecular weight, and stability in cerebrospinal fluid. Part of the reason that the
clinical translation of Aβ(1-42), tau, and phosphorylated tau as cerebrospinal fluid biomarkers took decades
was that reproducible assays were not available early on. As a result, another overarching goal of this project
is to develop precise, transferable, validated targeted proteomic assays to quantify proteins in
cerebrospinal fluid that can be used to investigate disease mechanism and predict poor outcomes.

## Key facts

- **NIH application ID:** 10105265
- **Project number:** 5U19AG065156-02
- **Recipient organization:** UNIVERSITY OF WASHINGTON
- **Principal Investigator:** ANDREW N HOOFNAGLE
- **Activity code:** U19 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $608,188
- **Award type:** 5
- **Project period:** 2020-02-15 → 2025-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10105265

## Citation

> US National Institutes of Health, RePORTER application 10105265, Project 3: Development of multiplex assays for clinical monitoring of disease (5U19AG065156-02). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10105265. Licensed CC0.

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