Neuron‐derived extracellular vesicles blood‐based diagnosis of synuclein TDP43 autophagy and synaptic dysfunction pathologies

Abstract

AbstractBackgroundStratifying dementia into different subgroups based on specific pathologies can facilitate the development of more effective treatments for subgroups of individuals with dementia. Effective stratification relies on reliable biomarkers. Neuron‐derived extracellular vesicles (NDEs) are lipid structures released by neurons into the extracellular space, and some get into the bloodstream. NDEs carry proteins and RNA that reflect their cell of origin and thus provide a promising biomarker platform.MethodNeuroDex developed two methods. ExoSORT is an optimized NDE isolation platform by immunoaffinity, where NDEs are captured with antibodies against the neuronal‐specific markers GAP43 and NLGN3. LuminEV is an intact extracellular vesicle (EV) immunoassay based on the Luminex platform. Antibodies against a potential biomarker, general EV marker, and IgG (negative control) are multiplexed to generate a robust assay.ResultThe vesicular nature and size of NDEs were confirmed by electron microscopy and nanoparticle tracking. EV markers were ascertained by western blot and proteomic analyses, and capture specificity was demonstrated by significantly elevated neuron‐specific proteins and mRNA levels compared to procedural control (IgG). Capture efficiency (∼80%) and precision (CV < 20%) demonstrated recovery of spiked NDEs. Measurements of αSYN following ExoSORT with an MSD assay and on EVs with the LuminEV generated a separation with 83% sensitivity and 86% specificity (45 Parkinson and 30 healthy controls plasma samples). LC3, an autophagy marker, was also measured following ExoSORT and was significantly (P<0.03) elevated in PD and Alzheimer’s disease (AD) (P<0.02, N = 30) plasma samples. TDP43 was also significantly (P<0.004) higher in AD plasma samples following ExoSORT. The synaptic marker GLUR2 was measured by the LuminEV assays and was significantly (P<0.001) reduced in AD and PD plasma samples. Its level was also significantly (P<0.03) correlated with the patient’s cognitive score.ConclusionImmunoaffinity isolation of EVs from plasma samples according to their cell of origin is a promising biomarker platform. We demonstrate the NDEs isolation selectivity, specificity, and precision. We also show the platform’s potential to identify pathologies like αSYN, TDP43, autophagy, and synaptic dysfunction; this reflects the potential for patient stratification.