This project aims to generate blood tests for: 1. mild cognitive impairment (MCI), and 2. conversion risk from MCI to full-blown Alzheimer's disease (AD). We have reported that systemic footprints, increases in key micro- RNA (miR) levels, are identifiable in AD blood samples; here we suggest that this is also true for MCI, the pre- AD phase. Our recent results suggest two stages of cognitive decline continuum: Stage 1, for key microRNAs increased from circulating blood of normal elderly controls (NEC) to that of MCI patients, but not further to AD; and Stage 2, for specific circulating blood microRNA increases from MCI to AD. This led us to suggest our mo- del: Stage 1 circulating microRNAs as MCI-specific biomarkers and Stage 2 circulating microRNAs as biomark- ers for conversion of MCI to AD. Prototypes of these biomarkers are: Stage 1, miR-181b increases from NEC to MCI levels, but not further in AD plasma samples; Stage 2, increased miR-34a & miR-34c levels, observed only from MCI to AD, but not from NEC to MCI plasma. These results suggest that the MCI cognitive decline may start with increased Stage 1 microRNAs' expression, then Stage 2 increases add insult to injury; the for- mer being MCI-specific biomarkers, and the latter specific for AD and/or converter patients. Our goal is then to establish two sets of stage-specific biomarkers, based on: a. highly reproducible data for identified blood micro- RNAs; b. training datasets for next-stage development for FDA approval; and c. validated specificity for de- mentia due to AD, in contrast to other types of dementia. Toward this end, Phase I will study 30 archived sam- ples each of MCI, NEC, and AD blood, to attain: Aim 1, establishing the reproducibility of our protocol for opti- mal blood microRNA assays, and identifying lead microRNAs' origins by determining concordance and/or dis- cordance among the above three microRNAs' increases in three circulating blood compartments: peripheral blood mononuclear cells (PBMC), plasma/serum, and its derivative, exosomes; and Aim 2, identifying other members of four lead array-profiled and PCR-validated microRNA families: miR-34, -181, 200, & Let-7, to expand our pool of candidate Stage 1 and Stage 2 biomarkers. In Phase II, Aim 1 expands to a larger cohort of 200 MCI patients' samples, to establish a Stage 1 biomarker training dataset tracing 21 microRNAs of these 4 families; Aim 2 establishes a repertoire of Stage 2 biomarkers, in archived specimens from 22 MCI subjects in our study, 12 of whom have converted to AD within the past decade, by comparing freshly collected with archived blood samples from converters versus non-converters by qPCR assays of the 21 listed miRNAs, and array profiling to identify additional novel ones; and Aim 3 establishes the identified biomarkers as specific for AD in contrast with non-AD dementias, i.e. Frontotemporal (FTD), Lewy body (LBD), and Parkinson's disease (PD) with dementia (PDD). Success of this project will provide blood miRNA-based diagnostics for MCI, and predict conversion from MCI to AD, unprecedented tests to monitor disease onset, progression, and drug efficacy for novel therapies to retard the decline of MCI to bona fide AD.