UC San Diego

The accumulation and aggregation of misfolded proteins (amyloids) is a common hallmark of many neurodegenerative diseases (ND). Among the ND, Alzheimer’s Disease (AD) is the most prevalent. The diagnosis of AD, when a patient is showing clinical symptoms, is typically too late for the efficacies of many AD treatments. However, on a molecular level, the deposition of amyloids in the brain appears years before clinical symptoms. The two pathological hallmarks of AD are the aggregation of beta-amyloid (Aβ) plaques and tau neurofibrillary tangles (NFTs). Historically, research has focused on detecting Aβ; however, recent evidence shows that tau better correlates with AD progression resulting in a shift to targeting tau. This thesis can be divided into two main projects. Project 1 documents the development of aryl cyano amide (ARCAM) derivatives (Probes 1-7) for detecting Aβ and tau in solution and tissue and encompasses Chapters 2-4. Project 2 discusses the utility of Fluorescence Lifetime Imaging (FLIM) for improving confidence in amyloid probe staining ambiguity and encompasses Chapters 5-6. Chapter 2 describes the systematic investigation on the effect of π-network on amyloid selectivity with recombinant amyloids in solution. Fluorescence lifetime measurements were performed to further study these probes and determine their potential utility for detecting amyloids in biological fluids. Chapter 3 is a preliminary study on the ability of amyloid-binding probes to detect amyloids in cerebrospinal fluid (CSF) and discriminate between healthy and AD patients. Additional CSF samples are needed to make any definitive claims; however, fluorescence probes show promise as an additional method for detection of amyloids in CSF. Chapter 4 tests the ability of our probes to detect insoluble amyloid aggregates in transgenic mouse models and confirmed AD patients. Molecular docking studies provide insight towards the binding of our probes in amyloids and can potentially be a useful tool for future rational design of tau-specific probes. Project 2 encompasses Chapters 5 and 6 and explores the utility of Fluorescence Lifetime Imaging (FLIM) for improving confidence in probe staining ambiguity. With the development of more sensitive confocal microscopes and increased autofluorescence in aged human brains, distinguishing autofluorescence from non-classical amyloid aggregates in fluorescent probe staining can be complex. Chapter 5 focuses on screening probes and performing FLIM experiments in AD patients while Chapter 6 is a preliminary study focusing on using FLIM to assist with probe staining ambiguity in Parkinson’s Disease (PD) and patients with comorbidity.