Efficient delivery of nucleic acids to cartilage remains a major translational challenge due to the dense extracellular matrix and limited cellular internalisation within joint tissues. Although lipid nanoparticles have achieved clinical success in hepatic delivery, rational chemical design principles for extrahepatic targeting remain insufficiently defined.1 Benchmark polymer–lipid systems derived from poly(ethylenimine) (PEI), such as 7C1, have demonstrated endothelial and bone marrow delivery in vivo, yet their heterogeneous polymer architecture limits mechanistic understanding of structure–performance relationships.2 Here, we report a chemically defined library of polymer–lipid hybrid nanoparticles engineered from poly(amidoamine) (PAMAM) dendrimers. Generations G0–G2 were functionalised with C12 or C15 alkyl chains to generate structurally controlled ionisable polymer–lipids. These materials were formulated with helper lipids and PEG–lipids to produce nanoparticles with tunable N/P ratios and surface shielding densities. Physicochemical characterisation revealed monodisperse assemblies across the library.
Functional performance was evaluated using eGFP plasmid as a reporter system. A clear generation-dependent trend emerged, with G0C12 and G0C15 formulations achieving >40% eGFP expression while maintaining cytocompatibility. Optimal transfection correlated with nanoparticles exhibiting hydrodynamic diameters of 70–100 nm and zeta potentials between +30 and +40 mV following complexation, defining a quantitative performance window for intracellular delivery. Importantly, these chemically defined formulations demonstrated comparable or improved activity relative to benchmark 7C1 systems under matched conditions. Increasing dendrimer generation reduced transfection efficiency despite comparable nanoparticle formation, highlighting the critical role of polymer architecture in governing delivery performance.
To assess translational relevance, selected high-performing formulations were further evaluated in chondrocyte models using antisense oligonucleotides. G0C12 nanoparticles achieved the highest antisense uptake in chondrocytes, whereas free antisense oligonucleotides remained predominantly associated with the extracellular cartilage matrix in ex vivo cartilage models, indicating limited cellular internalisation in the absence of nanoparticle engineering.
Collectively, this work establishes quantitative structure–property–function relationships linking dendrimer generation, hydrophobic substitution, nanoparticle size, and surface charge to nucleic acid delivery efficiency. These findings provide chemically grounded design principles for engineering next-generation polymer–lipid nanoparticles for intra-articular nucleic acid therapeutics.