High-Resolution Spatial MIST Technology for Functional Proteomic Study of Neuroinflammation in Alzheimer's Disease

Summary Alzheimer’s disease (AD) is the most common form of dementia and is a looming crisis in the US. Despite substantial progress made in AD research, the molecular and cellular processes governing neurodegeneration are still not well understood, and AD therapies have not resulted in significant benefits to patients. The traditional

hallmarks of AD include amyloid beta aggregation and neurofibrillary tangle deposition, while recently inflammation, an innate immune response in the brain, emerges as a third hallmark. Inflammation particularly occurs near epicenters of amyloid beta plagues and neurofibrillary tangles, and it involves complicated cellular

interactions that synergize with the progression of neurodegeneration. Understanding the molecular mechanisms of the functional roles of the cells in neuroinflammation and its influences on neurons is the key to searching for effective therapeutic targets. Due to the nature of high complexity and spatial heterogeneity, recent

research has vastly turned to next generation sequencing and transcriptomics tools in neurodegeneration studies. These results on gene expression will still need protein-level validation since proteins carry out most of cellular functions and biochemical processes. The current multiplexed protein assays on tissue samples are either labor

intensive and low coverage or in low spatial resolution. In this project, we aim to develop a spatial proteomics technology with cellular resolution to fill the technological gap and timely address the most imperative issues in AD mechanisms. This technology is built upon a multiplex in situ tagging (MIST) array that measures ~200 AD

relevant proteins from single neurons in our preliminary study. With ~10-100X higher multiplexity than other spatial protein tools, our spatial MIST will measure most important regulatory proteins and markers in spatially localized cells of brain sections. Two specific aims we propose include (1) Optimize the experimental techniques

in spatial MIST for detecting 280 key proteins in signaling and regulation of whole mouse brain slices, and (2) Profile the molecular features of cells near Aβ accumulation and tau enriched regions by spatial MIST during AD progression. The completion of this project will generate an enabling technology and method widely accessible

in the AD research community to investigate AD pathogenesis from a new, clinically relevant perspective. This technology will lay the foundation for future mechanistic studies of AD development and identification of potential therapeutic targets.