Circulating brain-enriched microRNAs as novel biomarkers for detection and differentiation of neurodegenerative diseases.

BACKGROUND: Minimally invasive specific biomarkers of neurodegenerative diseases (NDs) would facilitate patient selection and disease progression monitoring. We describe the assessment of circulating brain-enriched microRNAs as potential biomarkers for Alzheimer's disease (AD), frontotemporal dementia (FTD), Parkinson's disease (PD), and amyotrophic lateral sclerosis (ALS). METHODS: In this case-control study, the plasma samples were collected from 250 research participants with a clinical diagnosis of AD, FTD, PD, and ALS, as well as from age- and sex-matched control subjects (n = 50 for each group), recruited from 2003 to 2015 at the University of Pennsylvania Health System, including the Alzheimer's Disease Center, the Parkinson's Disease and Movement Disorders Center, the Frontotemporal Degeneration Center, and the Amyotrophic Lateral Sclerosis Clinic. Each group was randomly divided into training and confirmation sets of equal size. To evaluate the potential of circulating microRNAs enriched in specific brain regions affected by NDs and present in synapses as biomarkers of NDs, the levels of 37 brain-enriched and inflammation-associated microRNAs in the plasma of all participants were measured using individual qRT-PCR. A "microRNA pair" approach was used for data normalization. RESULTS: MicroRNA pairs and their combinations (classifiers) capable of differentiating NDs from control and from each other were defined using independently and jointly analyzed training and confirmation datasets. AD, PD, FTD, and ALS are differentiated from control with accuracy of 0.89, 0.90, 0.88, and 0.83 (AUCs, 0.96, 0.96, 0.94, and 0.93), respectively; NDs are differentiated from each other with accuracy ranging from 0.77 (AUC, 0.87) for AD vs. FTD to 0.93 (AUC, 0.98) for AD vs. ALS. The data further indicate sex dependence of some microRNA markers. The average increase in accuracy in distinguishing ND from control for all and male/female groups is 0.06; the largest increase is for ALS, from 0.83 for all participants to 0.92/0.98 for male/female participants. CONCLUSIONS: The work presented here suggests the possibility of developing microRNA-based diagnostics for detection and differentiation of NDs. Larger multicenter clinical studies are needed to further evaluate circulating brain-enriched microRNAs as biomarkers for NDs and to investigate their association with other ND biomarkers in clinical trial settings.

Universal screening test based on analysis of circulating organ-enriched microRNAs: a novel approach to diagnostic screening.

Early disease detection leads to more effective and cost-efficient treatment. It is especially important for cancer and neurodegenerative diseases, because progression of these pathologies leads to significant and frequently irreversible changes in underlying pathophysiological processes. At the same time, the development of specific screening tests for detection of each of the hundreds of human pathologies in asymptomatic stage may be impractical. Here, we discuss a recently proposed concept: the development of minimally invasive Universal Screening Test (UST) based on analysis of organ-enriched microRNAs in plasma and other bodily fluids. The UST is designed to detect the presence of a pathology in particular organ systems, organs, tissues or cell types without diagnosing a specific disease. Once the pathology is detected, more specific, and if necessary invasive and expensive, tests can be administered to precisely define the nature of the disease. Here, we discuss recent studies and analyze the data supporting the UST approach.

Analysis of organ-enriched microRNAs in plasma as an approach to development of Universal Screening Test: feasibility study.

BACKGROUND: Early disease detection with a minimally invasive screening test will significantly increase effectiveness and decrease the cost of treatment. Here we propose a framework of a novel approach - Universal Screening Test (UST) for the detection of pathological processes in a particular organ system, organ, or tissue by RT-qPCR analysis of circulating cell-free miRNAs in plasma. As the first step towards assessing the feasibility of this concept, the present study was designed to analyze whether the same microRNAs (miRNAs) can detect various diseases of a particular organ system. METHODS: RNA was extracted from plasma using Trizol treatment and silica binding. Levels of miRNAs were measured by single target RT-qPCR. The following innovations have been tested and proven effective: (i) the use of organ system/organ/tissue-enriched miRNAs; (ii) the use of miRNAs associated with broad disease categories, such as cancer and inflammation, in combination with the organ-enriched miRNAs; and (iii) the use of "miRNA pairs" for selecting miRNA combinations with the highest sensitivity and specificity. RESULTS: Here we report biomarker miRNA pairs effectively differentiating (i) patients with pulmonary system diseases (asthma, pneumonia and non-small cell lung cancer) and gastrointestinal (GI) system diseases (Crohn's disease, stages I/II esophageal, gastric and colon cancers) from controls, each with 95% accuracy; (ii) patients with a pathology of the pulmonary system from patients with a pathology of the GI system with 94% accuracy; and (iii) cancer patients (stages I/II esophageal, gastric, colon cancers, or non-small cell lung cancer) from patients with inflammatory diseases (asthma, pneumonia, or Crohn's disease) with 93%-95% accuracy. CONCLUSIONS: The results obtained in the present study, along with the data reported by us and others previously, are encouraging and lay the ground for further investigation of the described approach for UST development.

Plasma microRNA biomarkers for detection of mild cognitive impairment: biomarker validation study.

A minimally invasive test for early detection and monitoring of Alzheimer's and other neurodegenerative diseases is a highly unmet need for drug development and planning of patient care. Mild Cognitive Impairment (MCI) is a syndrome characteristic of early stages of many neurodegenerative diseases. Recently, we have identified two sets of circulating brain-enriched miRNAs, the miR-132 family (miR-128, miR-132, miR-874) normalized per miR-491-5p and the miR-134 family (miR-134, miR-323-3p, miR-382) normalized per miR-370, capable of differentiating MCI from age-matched control (AMC) with high accuracy. Here we report a biomarker validation study of the identified miRNA pairs using larger independent sets of age- and gender- matched plasma samples. The biomarker pairs detected MCI with sensitivity, specificity and overall accuracy similar to those obtained in the first study. The miR-132 family biomarkers differentiated MCI from AMC with 84%-94% sensitivity and 96%-98% specificity, and the miR-134 family biomarkers demonstrated 74%-88% sensitivity and 80-92% specificity. When miRNAs of the same family were combined, miR-132 and miR-134 family biomarkers demonstrated 96% and 87% overall accuracy, respectively. No statistically significant differences in the biomarker concentrations in samples obtained from male and female subjects were observed for either MCI or AMC. The present study also demonstrated that the highest sensitivity and specificity are achieved with pairs of miRNAs whose concentrations in plasma are highly correlated.

Circulating cell-free microRNA as biomarkers for screening, diagnosis and monitoring of neurodegenerative diseases and other neurologic pathologies.

Many neurodegenerative diseases, such as Alzheimer's disease, Parkinson disease, vascular and frontotemporal dementias, as well as other chronic neurological pathologies, are characterized by slow development with a long asymptomatic period followed by a stage with mild clinical symptoms. As a consequence, these serious pathologies are diagnosed late in the course of a disease, when massive death of neurons has already occurred and effective therapeutic intervention is problematic. Thus, the development of screening tests capable of detecting neurodegenerative diseases during early, preferably asymptomatic, stages is a high unmet need. Since such tests are to be used for screening of large populations, they should be non-invasive and relatively inexpensive. Further, while subjects identified by screening tests can be further tested with more invasive and expensive methods, e.g., analysis of cerebrospinal fluid or imaging techniques, to be of practical utility screening tests should have high sensitivity and specificity. In this review, we discuss advantages and disadvantages of various approaches to developing screening tests based on analysis of circulating cell-free microRNA (miRNA). Applications of circulating miRNA-based tests for diagnosis of acute and chronic brain pathologies, for research of normal brain aging, and for disease and treatment monitoring are also discussed.

Plasma microRNA biomarkers for detection of mild cognitive impairment.

Early stages of many neurodegenerative diseases, such as Alzheimer's disease, vascular and frontotemporal dementia, and Parkinson's disease, are frequently associated with Mild Cognitive Impairment (MCI). A minimally invasive screening test for early detection of MCI may be used to select optimal patient groups in clinical trials, to monitor disease progression and response to treatment, and to better plan patient clinical care. Here, we examined the feasibility of using pairs of brain-enriched plasma microRNA (miRNA), at least one of which is enriched in synapses and neurites, as biomarkers that could differentiate patients with MCI from age-matched controls. The identified biomarker pairs fall into two sets: the "miR-132 family" (miR-128/miR-491-5p, miR-132/miR-491-5p and mir-874/miR-491-5p) and the "miR-134 family" (miR-134/miR-370, miR-323-3p/miR-370 and miR-382/miR-370). The area under the Receiver-Operating Characteristic curve for the differentiation of MCI from controls using these biomarker pairs is 0.91-0.95, with sensitivity and specificity at 79%-100% (miR-132 family) and 79%-95% (miR-134 family), and p〈0.001. In a separate longitudinal study, the identified miRNA biomarker pairs successfully detected MCI in majority of patients at asymptomatic stage 1-5 years prior to clinical diagnosis. The reported biomarker pairs also appear useful for detecting age-related brain changes. Further testing in a larger study is necessary for validation of these results.