AI-Validated Brain Targeted mRNA Lipid Nanoparticles with Neuronal Tropism
Mor Sela, Gal Chen, Haim Kadosh, Tomer Kagan, Raneen Nicola, Sally Turutov, Yuval Richtman, Lin Zhige, Mia R. Albalak Menasherov, Shaked Kagan, Tzur Schroeder, Patricia Mora‐Raimundo, Reaam Kablan, Egor Egorov, Anas Odeh, Tasneem Abu-Raiya, Inbal Abutbul‐Ionita, Inbar Freilich, Galoz Kaneti, Ibrahim Knani, Yehuda Arav, Yael Leichtmann‐Bardoogo, Keshet Tadmor, Jeny Shklover, Tommaso Patriarchi, Dganit Danino, Peleg Hasson, Uri Ashery, Amit Zeisel, Ben M. Maoz, Tal Laviv, Kira Radinsky, Avi Schroeder
Abstract
High Resolution Image Download MS PowerPoint Slide Targeting therapeutic nanoparticles to the brain poses a challenge due to the restrictive nature of the blood–brain barrier (BBB). Here we report the development of mRNA-loaded lipid nanoparticles (LNPs) functionalized with BBB-interacting small molecules, thereby enhancing brain delivery and gene expression. Screening brain-targeted mRNA-LNPs in central nervous system (CNS) in vitro models and through intravenous administration in mice demonstrated that acetylcholine-conjugated LNPs achieved superior brain tropism and gene expression, outperforming LNP modifications with nicotine, glucose, memantine, cocaine, tryptophan, and other small molecules. An artificial intelligence (AI)-based model designed to predict the BBB permeability of small-molecule ligands showed strong alignment with our experimental results, providing in vivo validation of its predictive capacity. Cell-specific biodistribution analysis in Cre-reporter Ai9 mice showed that acetylcholine-functionalized LNPs preferentially transfected neurons and astrocytes following either intravenous or intracerebral administration. Mechanistic studies suggest that acetylcholine-LNP uptake is mediated by the functional engagement of acetylcholine receptors (AchRs) followed by endocytosis, which synergistically enhances intracellular mRNA delivery. Moreover, acetylcholine-LNPs successfully crossed a human BBB-on-a-chip model, enabling transgene expression in human iPSC-derived neurons. Their effective penetration and transfection in human brain organoids further support their potential activity in human-based systems. These findings establish a predictive and modular framework for engineering CNS-targeted LNPs, advancing precision gene delivery for brain disorders.