MicroRNAs (miRNAs) are recently-discovered small regulatory RNAs that play fundamental roles in neurobiology. Preliminary results indicate that decreased expression of a particular miRNA, miR-107, may contribute to Alzheimer's disease (AD) pathogenesis through a metabolic pathway. These are the first published data pertaining to miRNAs in a specific pathway that may contribute to AD. We will test a specific strategy to target this pathway for AD therapy. Hypothesis #1: MiR-107 expression is decreased very early in AD, which increases BACE1 expression, and hence increases the amount of neurotoxic Abeta peptides in AD patients' brains. Hypothesis #2: Bezafibrate treatment decreases AD-type pathology by increasing levels of miR-107.
Specific Aim #1 : Characterize fully the regulation of BACE1 by miRNAs. Sub-Aim a. A novel technique will test directly whether BACE1 mRNA is a miRNA target. This biochemical approach involves co- immunoprecipitation using our monoclonal anti-Argonaute antibody. Sub-Aim b. Tissue culture studies will be performed to evaluate exactly which parts of the BACE1 mRNA 3'UTR constitute miRNA targets. Sub-Aim c. `Knock-in' and `knock-down' techniques will be used to alter miR-107 levels specifically in human cultured cells, to determine the effects of miR-107 expression changes on the levels of BACE1 protein, C99 polypeptide, and A2 peptide. Experiments will be performed initially on H4 and SH-SY5Y cells.
Specific Aim #2 : Characterize the impact of miR-107 on glucose metabolism and correlate the expression of miR-107 and other miRNAs with AD pathology in situ Sub-Aim a. Tissue culture studies will be performed to assess how pharmacological treatments that alter metabolism affect miR-107 expression, and to evaluate how cellular changes in miR-107 influence the levels of specific metabolic intermediaries. Sub-Aim b. Human brain in situ hybridization will be used to understand how miR-107 expression relates to pathological hallmarks of AD and non-AD dementia.
Specific Aim #3 : Evaluate bezafibrate for increasing miR-107 levels and decreasing BACE1 protein and A2 peptide(s) formation in vitro and in vivo. Bezafibrate is an orally-administered, well-tolerated medication. Sub-Aim a. Preliminary results in cultured cells demonstrated that bezafibrate causes increased miR-107 expression and also induced down-regulation of BACE1 protein. The specific mechanism of bezafibrate action will be characterized using experiments in which the levels of miR-107 are manipulated. Sub-Aim b. Bezafibrate will be administered to mice - APPNLh/NLh x PS1P264L/P264L humanized APP knock-in mutants - that are an excellent model of AD-type amyloidogenesis (1), to demonstrate in vivo the efficacy of bezafibrate in modulating miR-107, BACE1, A2 levels, and AD-type neuropathology.
MicroRNAs are recently-discovered molecules that serve fundamental functions in the human brain. This research demonstrates for the first time that a particular microRNA may play an important role in Alzheimer's disease. A research program is proposed which exploits this new discovery, in order to develop and evaluate a novel therapy for patients at risk for Alzheimer's disease. ? ?
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