Specific changes in exosomal cargo and transmembrane proteins are reported to be significantly related to AD onset and progression, and they can provide useful and valuable biomarkers. Exosomes can carry Aβ1–42, Tau and its phosphorylated forms (e.g., p-Tau
thr181 and p-Tau
ser396), lncRNAs, miRNAs and other proteins that constitute molecular hallmarks of AD and could help distinguish AD or mild cognitive impairment (MCI) patients from healthy individuals [
146,
147]. As the pathology progresses from preclinical stages through MCI to dementia, the concentrations of these molecules in biological samples change, along with increased expression of other risk factor-associated molecules which are involved in neuroinflammation (C1q), autophagy system dysregulation (cathepsin-D) and metabolic disorders (IRS-1 and p-IRS-1) [
148]. Of all, neuron-derived exosomes (NDEs) present the highest diagnostic relevance compared to total exosomes, especially when considering p-Tau levels [
129]. In addition, combination of multiple markers (CSF p-Tau and Aβ) may increase the sensitivity and specificity. For instance, Aβ1–42 levels in neuron-derived blood exosomes in patients with preclinical AD are significantly higher than those from healthy control subjects, but significantly lower than those from AD patients [
141]. Along with protein analysis, characterization of exosomal miRNA profile can provide accurate insights into the pathogenesis and progression of the disease. Alterations of exosomal miRNAs from other body fluids, including plasma and CSF, have been reported by several studies. For example, analysis of CSF-derived exosome samples reveals alterations of miR-16-5p, miR-125b-5p, miR-451a, miR-605-5p, miR-9-5p and miR-598 in AD patients when compared to healthy subjects [
149]. Many of these miRNAs have been shown to be implicated in AD pathogenesis [
150]. Moreover, analysis with deep sequencing techniques of exosome-enriched plasma fractions reveals increased expression of 20 plasma exosomal miRNAs in AD patients. Among these 20 miRNAs, a panel of 7 (miR-185-5p, miR-342-3p, miR-141-3p, miR-342-5p, miR23b-3p, miR-338-3p, and miR-3613-3p) is highly valuable for predicting AD status with high accuracy using a machine learning model [
151]. Finally, more exosome-based biomarkers for both preclinical and clinical AD are emerging. Indeed, the blood exosomal levels of BACE1-antisense transcript (BACE1-AS), NDE APP, APPα, and APPβ are reported to be significantly higher in AD patients compared to control subjects [
143,
152]. Of note, free plasma proteins such as phospho-Tau217 and GFAP have recently been reported to be useful biomarkers of early AD, even before neuroimaging alterations can be detected [
153,
154]. These results provide a wider panel of biological biomarkers for AD compared to other neurodegenerative disorders. Interestingly, many clinical trials have been started in the recent years to investigate the clinical relevance of EVs as new biomarkers for diagnosis or drug response in AD. In 2017, the University Hospital in Lille started to recruit participants to analyze levels of Tau in EVs derived from AD patient CSF (ClinicalTrials.gov Identifier: NCT03381482). In 2019, the University of Oxford began to study if the drug JNJ-40346527 can block the colony stimulating factor-1 receptor which is responsible for the regulation of microglial cells, in order to change the activity or the number of activated microglial cells in the brain. To find evidence of this change, both free protein biomarkers and the number of EVs are monitored. In the same year, the National Institute of Aging in Baltimore started a clinical trial to investigate the ability of the molecule empagliflozin, which is an antidiabetic drug, to elevate ketone levels and boost neuronal health, thus delaying the onset and progression of cognitive impairment. To this aim, Egan and colleagues isolated from plasma both total and neuronal-origin EVs to analyze if an increase in ketone bodies may upregulate IGF-1 and insulin cascades in non-diabetic individuals. These studies, only partially concluded, suggest a novel approach to identifying non-invasive diagnostic and prognostic biomarkers of neurodegeneration and provide novel insights into the clinical tools available to follow the progression of AD compared to other neurodegenerative disorders.