PROJECT SUMMARY There are hundreds of studies of Alzheimer’s disease, Alzheimer’s disease-related dementias, and other neurodegenerative diseases comparing post-mortem (PM) human brain tissues obtained from human brain repositories. Current approaches that compare multiple, separately measured, -omic profiles studies introduce variability from using multiple samples. Thus, a multi-omics approach that can utilize a single biospecimen is needed. We have developed a penta-omic extraction method for frozen tissue and propose a proof-of-concept study. This effort would create the first penta-omic database, utilizing normal PM human brain tissue from the NIH NeuroBioBank. A new penta-omic simultaneous metabolomic, proteomic, lipidomic, DNA, RNA extraction method called SiMPL-DREx will be applied to a single tissue sample which will minimize the heterogeneity associated with testing multiple samples. SiMPL-DREx has added value because it more efficiently uses small volume, highly requested brain tissue. PM tissue selection is generally based entirely on a single quality control measure, RNA integrity number (RIN), obtained from a single tissue sample, usually the occipital pole (OP). Of the four -omic macromolecules extracted with SiMPL-DREx, RNA is the most labile primarily due to the universal distribution of RNase in the body. Tissue with RIN >7 is highly desirable for genomic and transcriptomic studies, yet how the overall quality of the metabolome, proteome, and lipidome (MPL) varies as a function of RIN has never been fully investigated. Tissues with RIN<7.0 are rejected for most -omic studies but may have sufficient quality for MPL studies. The primary post-mortem factor affecting -omic quality is autolysis which is dependent on body temperature. The time to be placed in cold storage is often not easily determined so RIN serves as a surrogate. This study will assess the association of RIN and measures of -omic quality in OP from unaffected donors with three RIN values (9-10, 6-7, 3-5) having an immediate death (agonal duration <1hr). The overarching goal of this study is to validate the penta-omic extraction method and to identify markers of tissue quality for different -omics outcomes that are independent of RIN. To this end, we will pursue two specific aims. In aim 1, SiMPL-DREx will be validated against existing single-omic extraction methods. In aim 2A, we will examine association of RIN category with DNA integrity number (DIN), protein integrity number (PIN) and pH to identify potential additional measures of tissue quality. In aim 2B a new measure, lipid integrity number (LIN) based on lipase activity is defined and will be tested for independence from RIN. The metabolome is unique by contributions from macro- and micro-molecular degradation, and therefore individual metabolites will be examined for association with RIN as potential markers of quality. If successful, this study will establish a more efficient method for multi-omic tissue ex...