Matsson, P. | Search bioRxiv

bioRxiv2026

Lipid-ASO therapeutics exhibit differential tissue targeted delivery upon systemic or local CNS administration

Antisense oligonucleotides (ASOs) are a powerful therapeutic modality, but their full potential is hindered by pharmacokinetic properties that affect tissue and cellular delivery. Lipid conjugation is increasingly used to modulate ASO's biodistribution and promote extrahepatic activity, yet lipid dependent effects on in vivo functional delivery, particularly in the central nervous system (CNS), remain less explored. Here, we performed a side by side in vivo comparison of cholesterol, palmitic acid (C16:0), docosanoic acid (C22:0), and eicosapentaenoic acid (C20:5) conjugated to a fully phosphorothioated 3 10 3 LNA gapmer ASO targeting the Malat1 long non coding RNA. Lipid-ASO conjugates were administered systemically or locally in the brain of mice and evaluated for tissue level and cellular level distribution by imaging, qPCR and single-cell RNA sequencing, simultaneously annotating cell origin and global transcriptional changes within the cell. Following systemic administration in mice, lipid conjugation improved overall multi organ efficacy compared to unconjugated ASO, but with pronounced tissue specific differences. Single cell sequencing of liver and heart transcriptomes revealed lipid dependent cellular uptake patterns and transcriptional responses distinct from administration of unconjugated ASO. After intracerebroventricular administration, selected fatty acid conjugates enhanced silencing in deep brain regions such as the striatum, whereas cholesterol conjugation impaired functional delivery despite increased CNS retention. Light-sheet microscopy showed restricted parenchymal penetration of cholesterol ASOs compared with broader but heterogeneous distribution of palmitic acid conjugate. Together, these findings demonstrate that lipid identity critically determines ASO efficacy, productive cellular uptake, and regional CNS engagement, emphasizing the need for context specific lipid design in ASO therapeutic development.

pharmacology and toxicology↗

bioRxiv2025

Conformational dynamics in the membrane interactions of bispecific targeted degrader therapeutics

Proteolysis targeting chimeras (PROTACs) offer vast new therapeutic opportunities, however their physicochemical properties are difficult to combine with optimal cell permeability and exposure at the target sites. We have systematically analyzed a dataset of more than 3500 PROTACs to investigate how the choice of ubiquitin E3 ligase ligands, linker design, and global molecular properties can be optimized to achieve the desired cell permeability and intracellular exposure. We find that conformational flexibility leads to environment-dependent shielding of polar functions and improved interactions with cell membranes, but that at the same time extended, linear conformations within the membrane are beneficial. Linker composition was a major factor in determining the folding propensity. Collectively, our results suggest that strategies to rationally design linkers and to shield polarity selectively within the protein-of-interest (POI) ligand and/or E3 ligand domains, rather than more extensive folding, may be beneficial in the design of permeable and effective PROTACs.

pharmacology and toxicology↗