Aschmann, D. | bioRxiv Science

bioRxiv2026

Stereochemical identity of lipid nanoparticles modulates protein expression via internal lipid organization

Stereochemistry plays a crucial role in how molecules interact with complex physiological environments, affecting pharmacokinetics, pharmacodynamics, efficacy, and toxicity. Although these effects are well studied for small-molecular drugs, they are largely overlooked for supramolecular assemblies used in drug delivery. Even for lipid nanoparticles (LNPs)--the most advanced RNA delivery platform--stereochemical effects are rarely investigated and, when considered, are typically limited to the ionizable lipid rather than the overall stereochemical identity of the LNP. Here we separate the ionizable lipid cKK-E12 into its two stereoisomers (trans: R,S/S,R; cis: R,R/S,S), which are normally used as a mixture. LNPs containing the cis isomer exhibit improved physicochemical properties, stability, and protein expression. By systematically varying the stereochemistry of the ionizable lipid, phospholipid, and cholesterol, we reveal stereochemistry-dependent differences in uptake and protein expression across six cell lines and in vivo in zebrafish embryos and mice. AI-assisted cryo-TEM analysis and SAXS link enhanced protein expression to structural differences, demonstrating control over internal lipid phases (lamellar and inverse hexagonal), influencing sample uniformity, and identifying stereochemical identity as a key determinant of functional RNA delivery.

pharmacology and toxicology↗

bioRxiv2026

A narrow thermodynamic design window governs selective membrane permeabilization and antiviral activity of amphipathic peptides

Designing molecules that selectively target therapeutically relevant membranes, such as viral envelopes, while sparing host cells is challenging: these membranes closely resemble host bilayers, so selectivity must exploit subtle lipid composition and curvature differences and demands precise tuning of affinity and hydrophobicity, yet curated sequence-specificity data are scarce. Here we show that selective membrane permeabilization and membrane-selective activity of amphipathic peptides are governed by a narrow thermodynamic design window defined by membrane curvature affinity and molecular hydrophobicity. Using a physics-driven generative workflow combining evolutionary molecular dynamics and a transformer predictor (PMIpred), we systematically explored and thermodynamically mapped peptide sequence space de novo without reliance on natural templates or experimental training data. Across four design generations we synthesized and experimentally characterized 43 peptides. Mapping functional activity onto a low-dimensional free-energy landscape reveals a confined thermodynamic "sweet spot" separating weak membrane binding from excessive hydrophobic association and cytotoxicity. Peptides operating within this regime efficiently permeabilize model membranes while maintaining low cellular toxicity. Antiviral activity against Zika virus and HIV-1 emerges in the same region but depends sensitively on membrane lipid composition. Quantitative thermodynamic design rules emerge for membrane-active peptides, illustrating how low-dimensional free-energy landscapes can guide the engineering of selective interactions at soft-matter interfaces.

biophysics↗