Early Detection of Alzheimer’s Disease using microRNAs

Abstract

Background: MicroRNAs (miRNAs) represent a class of ∼22nt short non-coding RNAs that have been identified as a sophisticated layer of post-transcriptional regulation, governing many cellular, inflammatory and vascular processes. This characteristic, together with the observation that miRNAs are frequently secreted into the extracellular space and stable in blood and other body fluids, make them specific, robust and above all non-invasive biomarkers that could qualify to augment the amyloid and tau framework. miRNAs have been used for the detection of alzheimer's disease (AD) in its early forms (e.g., mild cognitive impairment (MCI) due to AD) and to distinguish AD from other dementias. However, to date no miRNA panel has been translated into a clinical test. Objectives: With funding from the alzheimer's Drug Discovery Foundation (ADDF) Hummingbird Diagnostics (HBDx) is developing a blood-based microRNA (miRNA) biomarker intended to inform the diagnosis and prognosis of alzheimer's disease (AD). The project objective is two-fold: a) to evaluate the diagnostic performance of a miRNA panel in the following deeply phenotyped cohorts: i) amyloid-positive MCI due to AD subjects (prodromal AD), ii) amyloid-negative cognitively unimpaired control subjects, iii) amyloid-positive cognitively unimpaired subjects (preclinical AD), iv) amyloid-negative MCI (due to non-AD conditions) subjects. And, b) to compare baseline small RNA profiles with longitudinal profiles (≥2 years) and clinical outcome data. HBDx seeks to explore biomarkers that can be utilized to characterize clinicopathologic heterogeneity and could lead to the discovery of identifiers of disease prognosis (e.g. rapidly progressive AD). Methods: Through collaboration with the European Prevention of alzheimer's Dementia (EPAD) consortium, we have analyzed 3,302 blood samples of 1,895 patients from over 20 European sites. To ensure a simple, robust, and reproducible platform, the IVD-certified PAXgene Blood RNA System (PAXgene) was used for the collection, lysis, and subsequent RNA stabilization of whole blood samples to enable "pipetting-and cell sorting-free" sample collection in the clinic. Analytical processes, including RNA extraction, library preparation, and next-generation sequencing (NGS) were optimized to measure a whole blood, immune enriched, small RNA expression profile. We analyzed 1,895 prospectively enrolled individuals ≥50 years of age, 1540 were amyloid negative (81%) and 355 were amyloid positive (19%). Abeta-1-42/P-tau-181 > 0.024 was used as the threshold. We deployed 100-fold cross validation of a linear regression classifier to construct and validate small RNA feature models to evaluate their utility as biomarkers for amyloid positivity as well as for MCI with or without amyloid positivity. Results: We generated small RNA feature models and report a median diagnostic receiver operating area under the curve (AUC) of 0.64 for amyloid positivity (Abeta-1-42/P-tau-181 > 0.024). Amyloid positive MCI (CDR 0.5) individuals could be discerned from cognitively unimpaired (CDR 0) amyloid negative individuals with an AUC of 0.75. In a cross-sectional analysis of amyloid positive individuals, we could predict MCI (CDR 0.5) with an AUC of 0.76. Deconvolving the signature into its blood cell and plasma origin, revealed that ∼38% of features used for the analysis were found in plasma whilst ∼62% originated from circulating immune cells. Conclusion: These data suggest the potential of a small RNA-based blood test as a viable complement to the AT(N) framework for the management of individuals at risk for AD.