PROJECT SUMMARY Alzheimer’s disease (AD) is a severe neurodegenerative illness that destroys cognitive abilities causing memory impairment, difficulties with speech and language, behavioral changes, functional decline, and significant impairment. AD affects an estimated 13.8 million people in the United States and 50 million worldwide, imposing a significant economic burden and global health crisis. While many researchers are working towards developing a cure for the disease, there is currently no objective, point-of-care diagnostic test for the early diagnosis of AD, significantly limiting screening efforts for clinical studies and delaying treatment for patients suffering from early AD pathology. The current standard methods for AD diagnosis involve expensive PET imaging methods that irradiate the patient and cerebrospinal fluid biomarker tests, which are invasive and require a lumbar puncture. Recently, exosomes (30-150 nm extracellular vesicles) have been identified as a possible tool for AD diagnosis, attributed to their ubiquitous presence in biofluids, their ability to pass through the blood-brain-barrier, and their rich library of AD-relevant physiological information present in the molecular cargo they carry, making them prime candidates for use as biomarkers. However, the clinical application of exosomes is hindered by slow, inefficient techniques for exosome isolation and the absence of standardized exosomal biomarker detection and analysis. Thus, an automated, highly sensitive, fast, and efficient system that can isolate exosomes from biofluids and analyze the miRNAs and proteins they contain will significantly improve early-stage AD diagnosis efforts. In this R01 project, we will develop an Automated High-purity Exosome isolation-based AD diagnostics system (AHEADx) to address the limitations of current technologies. The proposed AHEADx platform includes two units: (1) a rapid (<1 min), high-yield (>90%), and high-purity (>90%) acoustic Bessel beam-based separation unit to isolate and enrich exosomes from whole blood, and (2) a rapid (<6 mins), highly sensitive photonic PCR and immuno-PCR (~1 copy/µL for nucleic acids and ~5 copies/µL for proteins, respectively) utilizing a plasmonic nanopillar array to enable on-chip thermocycling and multiplexed exosomal screening of combined panels of AD-relevant biomarkers. Our rapid and precise AHEADx platform will provide a simple, minimally invasive liquid biopsy to detect molecular AD biomarkers with ultrahigh accuracy and sensitivity in early-stage AD patients, allowing for an effective diagnostic method of AD screening for earlier treatment before the onset of severe symptoms of the disease and enable long-term studies on AD development and progression. Additionally, the proposed technology will accelerate the discovery of new exosomal miRNA and protein AD biomarkers and help to elucidate the mechanisms in which exosome trafficking and transport contribute to AD pathology. With these advantages, our AHEADx platform can potentially exceed current clinical standards in AD diagnostics, addressing a significant need in the field and providing a compelling platform for earlier, accurate, and sensitive detection of AD and long-term studies on the effects of novel AD therapeutics.

Project Narrative Alzheimer’s disease (AD) is a devastating neurodegenerative disease with no cure, imposing a major healthcare and economic strain worldwide. Standardization for an easy-to-use and reliable biomarker-based diagnosis of AD is lacking, resulting in invasive, high-cost tests of variable accuracy that may put the patient at risk. Thus, there is a significant need to develop highly accurate, sensitive, and minimally invasive diagnostic methods to promote improved treatment strategies and standardize AD diagnosis. The proposed project aims to create a liquid biopsy platform for AD that is capable of fully automated, rapid, precise exosome isolation and ultrasensitive, accurate exosome analysis from blood/plasma, realizing a point-of-care platform for early-stage AD diagnostics and treatment.