Polyanion Chemistry Engineers Ternary RNA Nanoparticle Structure/Function from the Inside-Out.

Area of Science:

• Biomaterials Science
• Nanotechnology
• Drug Delivery Systems

Background:

• Lipid nanoparticles (LNPs) are common for nucleic acid delivery, but polymeric alternatives like ternary polyelectrolyte nanoparticles (TNPs) offer potential for targeted delivery.
• Understanding the role of polyanion chemistry in TNP stability, protein binding, and transfection efficiency is crucial for developing advanced delivery systems.

Purpose of the Study:

• To engineer hydrophobic polyanions that provide TNPs with a negative surface charge and enhanced extracellular stability for targeted nucleic acid delivery.
• To systematically investigate how PEG architecture and polyanion chemistry influence TNP structure and function.

Main Methods:

• Synthesis of chemically diverse PEGylated polyanions to coat self-amplifying RNA (saRNA) polyplexes (PP).
• High-throughput stability assays and Small Angle Neutron Scattering (SANS) for structural studies.
• Molecular Dynamics (MD) simulations and in vitro cell studies for functional analysis.

Main Results:

• PEG5k-bl-polyanion5k formulations resulted in small, pH-responsive core-shell TNPs.
• A lead formulation (TNP5) with balanced hydrophobicity and charge density demonstrated effective extracellular stability and intracellular unpackaging.
• MD simulations indicated polyanions control TNP function by managing water exclusion and protein binding.

Conclusions:

• Polyanion engineering is key to controlling TNP structure and function for efficient RNA delivery.
• The study establishes a framework for high-throughput engineering of pH-responsive nanoparticles to overcome biological barriers in RNA delivery.
• Chemically diverse polyanions offer a tunable platform for developing targeted nucleic acid delivery systems.