Abstract More than 6 million Americans are living with Alzheimer’s disease (AD). AD is a neurodegenerative disease driven by abnormal accumulation of amyloid-β (Aβ) in the brain. Currently, we have very limited therapeutic approaches to treat AD. Aducanumab, designed to bind and inhibit toxic Aβ in the brain, is the only disease- modifying drug approved by The FDA, however, the clinical outcomes of Aducanumab were not satisfied. Therefore, there is a great need to develop novel therapeutic strategies against AD. Recently, RNA therapeutics were shown to inhibit disease gene expression, thereby reducing or abolishing diseased protein production. To realize RNA therapeutics, a key challenge is delivering the RNA across the blood brain barrier (BBB) which is a structural and functional roadblock for over 98% of small molecule drugs and nearly all large-molecule therapeutics. The goal of our proposed study is to develop novel DNA-inspired nanomaterials for the delivery of RNA therapeutics that penetrate the BBB for AD treatment. This study is built on our R01 project studying these nanomaterials for RNA delivery. We have successfully developed a family of delivery vehicles called Janus base nanopieces (NPs) which are rod-shaped nanoparticles formed by bundles of DNA-mimicking Janus base nanotubes (JBNTs) with RNA cargoes incorporated inside via charge interactions. Our recent publications have shown NPs can effectively deliver RNA therapeutics that inhibit disease gene expression. Moreover, we serendipitously found NPs had an excellent ability to penetrate BBB. Our preliminary results showed NPs were ~11 times more efficient than lipid nanoparticles to deliver a fluorescence-labeled RNA into a mouse brain. In addition, previous studies have found that Aβ accumulation can be prevented by downregulation of β- and γ- secretases or upregulation of a CX3CL1 peptide. These targets are likely to result in potent therapeutics. Combining all the information together, we hypothesized that we are able to develop and optimize NPs to deliver therapeutic RNAs across BBB into the brain to treat AD. To test the hypothesis, we will accomplish two specific aims: 1) To develop a library of Janus base NPs for RNA delivery and assess their delivery efficiency to penetrate BBB in vitro and in vivo. 2) To evaluate treatment outcomes of the developed NPs in an established AD mouse model. Completion of the proposed study will lay a solid foundation for the development of RNA therapeutics against AD and provide valuable preliminary results for future grant applications to realize a novel disease-modifying RNA therapeutic to treat AD.